2007 Summer Student Research Program Reception
Thursday, July 19th, 2007 2007 Summer6:00 p.m. – 7:30 p.m. Student University High Memorial Library ResearchChildren’s Hospital Oakland Symposium Research Institute August 10, 2007
Program Guide
The National Institutes of Health Short Term Training for Minority College Students Grant #5 T35 HL07807
NHLBI Comprehensive Sickle Cell Center Training of High School Students in Sickle Cell Disease Research Grant #1 P60 MD000222 2007 CHORI Summer Research Program Research 2007 CHORI Summer
August 10, 2007
This summer we were provided another opportunity to offer a research internship program for high school and undergraduate students. We had an outstanding class of students working in clinical (8) and laboratory (37) research settings. As can be seen from the personal statements and research abstracts, the contribution and experience gained by each participant is remarkable. Students came from a broad range of backgrounds and experiences. All were provided unique research projects, coordinated with mentors willing to teach and train their student.
This year, we initiated a partnership with the University of New Mexico (UNM), Albuquerque, which involves training two of their undergraduate students in research with mentors from UNM. The goal of this partnership is to provide off-site research opportunities for students who may not be able to spend time away from their families, in particular to recruit Native American students.
We are grateful for the funding we received for the summer program from the NHLBI, as part of a Short Term Training to Increase Diversity in Research program, as well as from the Summer for Sickle Cell Training Program, the Elizabeth Nash Foundation, and Williams Jenkins, M.D.
I would like to thank the mentors who dedicated time and effort to train the students and to infuse the excitement and pleasure of working in basic, translational, or clinical research. They have been the pillars supporting this program. The staff and scientists who took the time to make the lecture series informative and engaging were outstanding. Dr. Barbara Staggers, Dr. Vasanthy Narayanaswami, Diane Kohlman and Phillip Bollinger are to be particularly acknowledged for their scientific, technical and administrative assistance and for guiding students as required.
Again, congratulations and thanks to all members of the CHORI family for making the Summer Student Research Internship program a success in 2007.
Bertram H. Lubin, MD President, CHORI Director of Medical Research Children’s Hospital & Research Center at Oakland
5700 Martin Luther King, Jr. Way Oakland, CA 94609 510-450-7600 Director: Betram H. Lubin, M.D. President & Director of Medical Research at CHORI
Co-Director: Co-Director: Barbara Staggers, M.D. Vasanthy Narayanaswami, Ph.D. Director, Adolescent Medicine Assistant Scientist at CHORI
MENTORS (Children’s Hospital & Research Center at Oakland) Tariq Ahmad, MD Janet King, Ph.D. Bruce Ames, Ph.D. Ronald Krauss, MD Atamna Hani, PhD Frans Kuypers, Ph.D. Padmavathi Bandhuvula, Ph.D. Edward Lammer, M.D. Suruchi Bhatia, M.D. Guangming Li, PhD Dario Boffelli, PhD David Martin, M.D. Phillip Bollinger Gregory Moe, Ph.D. Lucia Carbone, Ph.D. Vasanthy Narayanaswami, Ph.D. Giorgio Cavigiolio, Ph.D. Janelle Noble, Ph.D. Susan Conrad, M.D. Michael Oda, Ph.D. Kitty de Jong, Ph.D. Jennifer Olson, M.D. Pieter de Jong, Ph.D. Babak Oskouian, PhD Deborah Dean, M.D., M.P.H. Donald Reason, Ph.D. Joseph Dhahbi, PhD Robert Ryan, Ph.D. Ervin Epstein, M.D. Julie Saba, M.D., Ph.D. Horst Fischer, Ph.D. Kathy Schultz, Ph.D. Trudy Forte, Ph.D. Stuart Smith, Ph.D., D.Sc. Ellen Fung, MD Po Lin So, Ph.D. Ethan Geier Barbara Staggers, M.D. Ward Hagar, MD Shiori Tamamizu-Kato, Ph.D. Carolyn Hoppe, M.D. Jean Tang, Ph.D. . Beate Illek, Ph.D. Patsy Wakimoto, Ph.D. David Iovannisci, Ph.D. Gordon Watson, Ph.D.
MENTORS (University of New Mexico) Mark Burge, Ph.D. Alberta Kong, Ph.D.
SELECTION COMMITTEE Bertram Lubin, M.D. Frans Kuypers, Ph.D. Barbara Staggers, M.D. Tomas Magana, M.D. Vasanthy Narayanaswami, Ph.D. John Matsui, Ph.D. Caroline Kane, Ph.D. Laurie Schumacher, M.D.
PROGRAM ADMINISTRATION Bertram Lubin, M.D. Phillip Bollinger Barbara Staggers, M.D. Diane Kohlman Vasanthy Narayanaswami, Ph.D. Tom McElderry Blia Yang
2007 CHORI SUMMER STUDENT RESEARCH PROGRAM CURRICULUM
Orientation: June 11, 2007 There will be an all-day orientation for summer interns on Monday, June 11, 2007, from 10:00 am until 3:00 pm. Lunch will be served. Agenda to include:
• Introduction and overview by Director and Co-Directors • Keynote lecture • Explanation of curriculum • Obtaining badges and tour of CHORI
Safety Training: June 12, 2007 The mandatory Safety Training with CHORI Safety Officer, Miriam Fang will be held on Tuesday, June 12 from 9:30 – 12:00. The student will be required to complete this training BEFORE beginning their project. There will be no lunch served.
Make-up Orientation, Safety Training and Photographs: June 18, 2007 Starting at 9:00 A.M., Phillip Bollinger will present his IT talk to all students. At 10:00 am, all students will adjourn to the main courtyard for group and individual photos. From 10:30 A.M. to approximately 1:30 P.M. a make-up orientation and safety training will be presented those students who missed the June 11 and/or June 12 meetings. Lunch will be served to these students.
Research Project: June 12 to Aug 10, 2007 The student will conduct research with assigned mentor. The details of project and research plan are left entirely to the individual mentor and each summer intern will follow the procedures, and schedule laid out by their respective labs.
Submit Written Research Plan: June 29, 2007 Students must submit to the Scholars Program Coordinator a written Research Plan outlining their project. The Research Plan should be 2-3 pages long and include: (a) Statement of hypothesis (b) Specific aims (c) Background (d) Methods (e) Anticipated outcome of project
Students will work closely with their mentor in the preparation of these reports, and mentors should review and approve the reports before submission. Figures, flow charts and schematics may be used to illustrate the research plan. The written report will be sent to: [email protected], and must include student’s name, mentor’s name and the title of the project.
Weekly Meetings: Research Ethics and Project Discussions (Wednesdays at 12:00 noon) Students are required to attend weekly meetings to hear presentations on Ethics in Research and Responsible Conduct of Research, and to discuss their scientific research projects. This is designed to aid them in discussing their projects with their peer group, and to help them to obtain an overall perspective of their progress. Lunch will be served.
Weekly Lectures: Current Topics in Health and Disease (Tuesdays at 4:00 pm) Students are required to attend weekly lectures delivered by CHORI and CHRCO faculty members. The lectures will cover various scientific topics that are currently being researched at CHORI and CHRCO.
2007 CHORI Summer Student Symposium, August 10, 2007 A one-day symposium will be held on Aug 10, 2007, where all students are required to participate. The students will submit an abstract concisely summarizing their work, which will be considered for an oral or a poster session for the Symposium. Abstracts are due on July 27, 2007. A committee comprised of the Director, Co-Directors and other leading members of the CHORI scientific community will review the abstracts for the Symposium. Students will work closely with their mentors in the preparation of abstracts and presentations. Family members, teachers, lab members and friends are welcome to attend. A catered breakfast and lunch will be provided for all attendees on the Symposium day. Details of the Symposium and the scientific sessions will be available by Aug 3, 2007.
The Symposium will comprise of about 15 oral presentations (10 min each, with 5 min discussion), and a poster session during which the presenters will be on-hand to explain their research project. An abstract book, which will include the Symposium program, personal statements, and the research project abstracts, will be presented to each student.
A certificate of participation in the CHORI Summer Student Research Program will be awarded to those who successfully complete the program.
Summary of important dates:
June 11, 2007 Orientation: 10:00 A.M. – 3:00 P.M. June 12, 2007 Safety Training: 9:30 A.M. – 12:00 P.M. June 18, 2007 Make-up orientation and safety training: 9:00 A.M. – 2:00 P.M. June 29, 2007 Written Research Plan due July 6, 2007 Personal Statement for Program Guide due by 5:00 P.M. July 27, 2007 Abstract for Program Guide due by 5:00 P.M. August 10, 2007 Summer Student Research Symposium
Please address any additional questions and concerns to: [email protected]
2007 Schedule of Weekly Lectures: Current Topics in Health and Disease The lectures will be held in the Little Theater at CHORI on Tuesdays at 4:00 pm
June 11 Monday June 11 at 10:15 am Frans Kuypers, PhD Senior Scientist, Children’s Hospital Oakland Research Institute Center for Sickle Cell Disease and Thalassemia “Membrane Biology and Hemoglobinopathies: Translational Research at CHORI” June 19 Renee Wachtel, MD and Paige Nues Director, Developmental & Behavioral Pediatrics Coordinator, Katie’s Clinic for Rett Syndrome Children’s Hospital Oakland “Rett Syndrome”
June 26 Bertram H. Lubin, MD President, Director of Medical Research Children’s Hospital Oakland Research Institute “The Social Life of Stem Cells”
July 10 Janet King, PhD Scientist, Children’s Hospital Oakland Research Institute Center for Nutrition and Metabolism “Childhood Obesity and What Can We Do About It?”
July 17 Edward Lammer, MD Associate Scientist, Children’s Hospital Oakland Research Institute Center for Genetics “Birth Defects Research in California”
July 24 Esteban Burchard, MD, MPH Assistant Professor, Departments of Biopharmaceutical Sciences and Medicine University of California, San Francisco “Importance of Race/Ethnicity in Biomedical Research and Clinical Practice” July 31 Jean Tang, MD, PhD Clinical Instructor, University of California San Francisco Postdoctoral Fellow, Children’s Hospital Oakland Research Institute, Center for Cancer Research “Training for a Career in Translational Research: One Example From Dermatology and Cancer Biology”
2007 SCHEDULE OF WEEKLY MEETINGS: RESEARCH ETHICS AND PROJECT DISCUSSIONS
The lectures will be held in the Little Theater/Board Room at CHORI at 12:00 noon Lunch will be provided
Wednesday, June 20
Carolyn Kane Ethics in Research and Responsible Conduct of Research
Wednesday, June 27
Summer Student Research Project Discussions
Wednesday, July 11
Carolyn Kane Ethics in Research and Responsible Conduct of Research
Wednesday, July 18
Summer Student Research Project Discussions
Wednesday, July 25
Summer Student Research Project Discussions
Wednesday, August 1
John Matsui Ethics in Research and Responsible Conduct of Research
2007 Summer Student Research Program Reception
Thursday, July 19th, 2007 6:00 p.m. – 7:30 p.m. University High Memorial Library Children’s Hospital Oakland Research Institute
Program
Bertram H. Lubin, M.D. President, Children’s Hospital Oakland Research Institute Program Director
Barbara Staggers, M.D. Director, Adolescent Medicine, Children’s Hospital Oakland Program Co-Director
Vas Narayanaswami, Ph.D. Scientist, Children’s Hospital Oakland Research Institute Program Co-Director
Brad Barber Senior Vice President & Chief Development Officer Children's Hospital & Research Center Foundation
Lucille Day, Ph.D. Director, Hall of Health
Mina Nikanjam Graduate Student Researcher, Children’s Hospital Oakland Research Institute
Jennifer Brakeman, Ph.D. Faculty Member, Head-Royce School
Student Affiliated Laboratory Tours
10 11 SYMPOSIUM SCHEDULE
12 13 2007 Summer Student Research Symposium Friday, August 10, 2007 CHORI Theater
8:30 – 9:00 Check-in and Continental Breakfast 9:00 – 9:15 Introduction by Program Directors Bertram H. Lubin, M.D. – Director Barbara Staggers, M.D. – Co-Director Vasanthy Narayanaswami, Ph.D. – Co-Director
Oral Presentations Session 1 Syed Askree, Ph.D., Postdoctoral Fellow Chairs: Dominique Johnson, California State University Hayward 9:15 – 9:30 Blia Yang, University of California, Berkeley 104 Mentor: Vasanthy Narayanaswami, Ph.D., Phillip Bollinger CHORI Summer Research Program: Looking Back
9:30 – 9:45 Chris Cox, California State University Sacramento 40 Mentors: Babak Oskouian, Ph.D., Julie Saba, M.D., Ph.D. Leveraging Sphingolipids in the Treatment of Leukemia
9:45 – 10:00 Ramine Cromartie, Stanford University 42 Mentors: Guangming Li, Ph.D., Bruce Ames, Ph.D. Differential Distribution of 25-Vitamin D in Human Plasma and Leukocytes
10:00 – 10:15 Ben Goldenson, University of California, Los 54 Angeles Mentors: Ronald Krauss, M.D. Effect of Lipoprotein Substrate, Microenvironment and Fatty Acid Source on Lipolytic PPAR a Activation
14 10:15 – 10:30 Nadia Kaaid, San Francisco State University 68 Mentor: Donald Reason, Ph.D. Binding and Neutralization Ability of Human Antibodies Directed Against Bacillus anthracis Toxins
10:30 – 10:45 Samuel Black, Albany High School 34 Mentors: Kitty de Jong, Ph.D., Frans Kuypers, Ph.D. Phosphatidylserine Exposure on Normal and Sickle Human Erythrocytes by Oxidative Damage
10:45 – 11:00 Break Session 2 David Killilea, Ph.D., Assistant Staff Scientist Chairs: Sua Yang, University of California, Berkeley 11:00 – 11:15 Amanda Harris, University of New Mexico 60 Mentor: Alberta Kong, Ph.D. Comparison of RT3 Triaxial Accelerometer and 7-day Physical Activity Recall in Hispanic and American Indian Adolescents
11:15 – 11:30 Spenser Perloff, University of California, Davis 86 Mentor: Ed Lammer, M.D. Optimization of the Thymidylate Synthase Genotyping for Case Control Birth Defect Studies
11:30 – 11:45 Griselda Velasquez, University of California, 102 Berkeley Mentors: Robert Raphael, M.D., David Martin, M.D. Detection of hMLH1 Somatic Hypermethylation by Methylation-Specific PCR and Quantitative Methylation-Specific PCR (Q-MSP)
15 11:45 – 12:00 Myles Fisher, California State University Chico 50/36 and Amelia Brazil, The College Preparatory School Mentor: Barbara Staggers, M.D. Depression and High Risk Behavior in Adolescents
12:00 – 12:15 William Green, University of Pennsylvania 58 Mentors: Vladimir Serikov, Ph.D., Frans Kuypers, Ph.D.. Human Placenta Tissue is a Novel Niche for Hematopoietic Stem Cells
12:15 – 12:30 Marena Trinidad, Benecia High School 100 Mentor: Carolyn Hoppe, M.D. Genetic Risk Factors for Stroke in Adult SCA Patients
12:30 – 12:45 Miguel Martinez-Romo, University of California, 84 Berkeley Mentors: Tariq Ahmad, M.D., Ginny Gildengorin, M.D. Glycemic Control Among Diabetes Patients Living in the Bay Area
12:45 – 1:15 Lunch in the CHORI Main Courtyard
1:15 – 3:30 Poster Session in the CHORI Library and Adjoining Hallways (1:15-2:15 odd numbers at poster; 2:15-3:15 even numbers at poster)
#1 Emily Adney, Occidental College 22 Mentor: Dario Boffelli, Ph.D. Characterization of the Peroxisome Proliferator- Activated Receptor Gamma Protein in Dyslipidemia
16 #2 Olayemi Akintunde, University of California, 24 Berkeley Mentors: Shiori Tamamizu-Kato, Ph.D., Vasanthy Narayanaswami, Ph.D. Oxidative Stress Modification of Apolipoprotein E, Apolipoprotein A-1 and Apolipoprotein B-100
#3 Ahmed Alghrouz, University of New Mexico 26 Mentor: Hani Atamna, Ph.D. Developing Methods for Isolating Amyloid-b-heme Complex From Brain Homogenate of AD Patients
#4 Andrea Banuelos, University of California, 28 Berkeley Mentor: Janelle Noble, Ph.D. HLA-A Genotyping for African-American Control Group
#5 Kate Barber, Stanford University 30 Mentors: Beate Illek, Ph.D. Pyocyanin-induced H2O2 Production in Cystic Fibrosis Airways
32 Sophia Blachman-Biatch, College Preparatory #6 High School Mentor: Stuart Smith, Ph.D., D.Sc. Expression, Purification and Mutagenesis of Human Acyl Carrier Protein
#7 Sam Cartmell, Yale University 38 Mentor: Ward Hagar, M.D. Factors Associated with Death in Patients with Sickle Cell Disease and Pulmonary Hypertension
17 #8 Nadia Dhahbi, Alameda High School 44 Mentors: Joseph Dhahbi, Ph.D., David Martin, M.D. Affects of Senescence on DNA Methylation
#9 Tyronda Elliott, San Francisco State University 46 Mentor: Ellen Fung, M.D. Bone Health in Chronic Diseases of Childhood
#10 Melanie Fish, Piedmont High School 48 Mentor: Barbara Staggers, M.D. Standardized Comprehensive Care for Adolsescent Sickle Cell Patients
#11 Ivan Friedman, International High School 52 Mentor: Gregory Moe, Ph.D. Sialyl Transferase Gene Expression in Three Human Cancer Cell Lines
#12 Wendy Gordillo, San Francisco State University 56 Mentor: Patsy Wakimoto, Ph.D. Latinos and Type 2 Diabetes
Sejal Hathi, Norte Dame High School #13 62 Mentor: Deborah Dean, M.D., M.P.H. Genotyping of the ompA Gene of Chlamydia Trachomatis Among Commercial Sex Workers in Ecuador
#14 Daniel Hausrath, Head Royce High School 64 Mentor: Trudy Forte, Ph.D., Robert O. Ryan, Ph.D. Novel Molecular Targets for Treating Glioblastoma Multiforme Tumors
18 #15 Dominique Johnson, California State University 66 Hayward Mentor: Gordon Watson, Ph.D. Biomarkers for a Genetic Deficiency in Cholesterol Synthesis (Smith-Lemli-Opitz Syndrome)
#16 Ragini Kathail, Pomona College 70 Mentor: Janelle Noble, Ph.D. Control Data for Class I HLA-B in an African- American Population
#17 Andrew Lee, Duke University 72 Mentor: Julie Saba, M.D., Ph.D. Sphingolipids and the AKT Pathway
#18 Jacqueline Leon, University of California, 74 Berkeley Mentors: Susan Conrad, M.D., Suruchi Bhatia, M.D. Factors at Diagnosis of Type 1 Diabetes Predict Outcome at 2 Years
#19 Aaron Levine, Piedmont High School 76 Mentors: Lucia Carbone, Ph.D., Pieter de Jong, Ph.D. Chromosomal Rearrangements in Gibbons
#20 Olivia Linney, Head-Royce High School 78 Mentor: Janelle Noble, Ph.D. Contribution of Human Leukocyte Antigen C Locus to the HLA-Associated Disease in African American Population
#21 Erin Mamuyac, Piedmont High School 80 Mentors: Kitty de Jong, Ph.D., Frans Kuypers, Ph.D. Phosphatidylserine Exposure in Normal Erythrocytes by the Addition of Zinc and Zinc Chelators
19 #22 Yessica Martinez, San Francisco State University 82 Mentors: Giorgio Cavigiolio, Ph.D., Ethan Geier, Michael Oda, Ph.D. Determination of ApoA-I’s Helical Turn Orientation by FRET Analysis
#23 Mallery Quetawki, University of New Mexico 88 Mentor: Mark Burge, M.D. Disordered Sleep in Obese Adolescents Wth and Without Impaired Glucose Tolerance
#24 Jesse Sandberg, Duke University 90 Mentors: Po-Lin So, Ph.D., Ervin Epstein, M.D. Inhibition of Hedgehog Signaling Pathway in BCC Skin Tumors
#25 Chia Teoh, San Francisco State University 92 Mentors: Jennifer Olson, M.D., Suruchi Bhatia, M.D. Timing of Estrogen Replacement Therapy in Girls with Turner Syndrome
#26 Ashleigh Thompson, Piedmont High School 94 Mentors: Julie Saba, M.D., Ph.D., Padmavathi Bandhuvula, Ph.D. Screening the Inhibitory Effect of Some Compounds on SPL Activity
#27 Lauren Tom, University of California, Los Angeles 96 Mentors: Edward Lammer, M.D. Maternal Smoking, Genetic Variation of Glutathione S-Transferases, and Risk of Gastroschisis
20 #28 Nora Trice, Lick-Wilmerding High School 98 Mentor: Robert Ryan, Ph.D. Environmental Stability of Amphotericin B Nanodisks
Blia Yang, University of California, Berkeley #29 104 Mentors: Vasanthy Narayanaswami, Ph.D., Phillip Bollinger CHORI Summer Research Program: Looking Back
#30 Sua Yang, University of California, Berkeley 106 Mentor: Patsy Wakimoto, Ph.D. Core Issues for the Hmong in Food Shopping and Food Safety
Amanda Yee, Alameda High School #31 108 Mentors: Janet King, Ph.D. Glucose Clearance and Energy Expenditure in Pregnant Obese Women
#32 Zhe Zhang, Stanford University 110 Mentors: Jean Tang, Ph.D., Ervin Epstein, M.D. Exploring the Effect of Vitamin D for Prevention of BCC in Mice and Humans
21
Additional Poster Session Participants
#33 Melody Liao, Veronica Ng, May Bhetraratana, Tina Liu
Mentor: Lucille L. Day, Ph.D. Hall of Health, Children’s Hospital & Research Center at Oakland, Oakland, CA 94609 Health and Biomedical Science for a Diverse Community
3:15 – 4:00 Certificate Presentation and Cake in the CHORI Main Courtyard
22 Emily Adney Volunteer
Occidental College Sophomore
Mentor: Dario Boffelli, PhD
Honestly, I am not one of those individuals who has always dreamed of becoming a doctor or a scientist. While I did challenge myself in science, I went through high school dabbling a little of everything: I intensified my passion for ballet, focused on my talents in math and leadership, and thrived in learning the French language. My interest in biology was somewhat sparked while attending the California State Summer School for Mathematics and Science for the summer of 2003. I continued onto the school I had fallen in love with, Occidental College. The day I attended my first college biology class, I knew that was the concentration for me. Now, I have two study majors in French language and biology with an emphasis in cell and molecular biology. Occidental’s outstanding biology professors have inspired me and I have grown to be very interested in both health and the intricacies of the human genome. The many links between genetics and human disease is a field in which I aim to gain expertise.
Working as an intern at CHORI this summer has given me the opportunity to meet a variety of amazing students and doctors. Most importantly, I have been able to work closely with Dr. Weinsheimer, who has involved me in Dr. Boffelli’s lab, where I am receiving the hands-on experience that I crave. Now that I have been given a glimpse into a lab research environment, I have an enhanced infatuation with learning in depth about biology, especially genetics. I would like to move onto graduate school and someday earn a Ph.D. I have formulated new dreams of someday working along side other bright scientists to further research and to contribute discoveries of our own.
23 Characterization of the Peroxisome Proliferator-Activated Receptor Gamma Protein in Dyslipidemia
E. Adney, D. Boffelli & S. Weinsheimer Children’s Hospital Oakland Research Institute, Oakland, California 94609
Introduction Peroxisome proliferator-activated receptor gamma (PPARG) is a member of the nuclear hormone receptor superfamily that controls many cellular and metabolic processes. PPARG is expressed in several cell types including adipocytes and macrophages and has a role in adipocyte differentiation, lipid metabolism and inflammation. As a transcription factor, PPARG regulates the expression of genes involved in promoting fat storage and reducing serum lipid levels. PPARG is activated by certain ligands including free fatty acids and eicosanoids, and forms a heterodimer with the retinoid X receptor. PPARG plays a critical role in the regulation of intracellular lipid levels. Characterization of PPARG in dyslipidemia will further define the role this protein has in the regulation of lipid levels. Objective To characterize the PPARG protein expression in lymphocytes from individuals at the tails of plasma lipid distribution, as defined by low and high levels of the LDL4a subfraction. We hypothesize that the PPARG protein can be detected in lymphocytes, and that PPARG protein levels may differ quantitatively depending on the lipid profile. We aim to test the specificity of a commercially available PPARG polyclonal antibody. Methods Immortalized lymphocytes from individuals with low (n=5) and high (n=5) levels of LDL4a subfraction were cultured and nuclear protein extracts were isolated. Nuclear protein was separated by molecular weight by SDS polyacrylamide gel electrophoresis and transferred to nitrocellulose membrane for Western analysis. A goat polyclonal PPARG antibody (Santa Cruz, sc-1984) and mouse anti-goat secondary antibody conjugated to HRP (sc-2354) was used to detect the PPARG protein by chemiluminescence. Results The PPARG protein (molecular weight ~67 kDa) was detected in lymphocytes from individuals with extreme low and high lipid profiles, yet there was no detectable difference in protein expression in these two phenotypes. The primary PPARG antibody (sc-1984) recognizes the PPARG protein, with some non- specific binding to additional nuclear proteins. Conclusion Immortalized lymphocytes from individuals with extreme low and high lipid profiles express PPARG, and can be useful for future PPARG studies. The primary PPARG antibody (sc-1984) detects the PPARG protein and a few non-specific proteins.
24 Olayemi Akintunde Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Third Year
Mentor: Vasanthy Narayanaswami, PhD Shiori Tamamizu-Kato, PhD
My involvement with the CHORI summer program has been a constant source of inspiration and motivation for my future plans. Working in a research lab, combined with my personal relationships with people who have various illnesses, has shown me just how vital research is. Despite the fact I have always associated physicians with healing people, I now understand that researchers are also working toward healing but their work transcends the individual patient. Scientists have the opportunity to help many people by pushing back the boundary of the unknown and developing new treatments for diseases.
My appreciation and respect for scientists has grown tremendously after being exposed to a research environment. It takes a lot of dedication, intelligence and heart to set up an experiment and know why it works, interpret data and to return to a problem after it has failed and then to be able to look at it from a different perspective. This tenacious spirit that I have repeatedly witnessed in the lab has encouraged me to be just as persistent in my own life. I would like to thank Vasanthy Narayanaswami and Shiori Tamamizu-Kato for their never ending support, patience and kindness.
25 Oxidative Stress Modification of Apolipoprotein E, Apolipoprotein A-1 and Apolipoprotein B-100
O. Akintunde, S. Tamamizu-Kato & V. Narayanaswami Children’s Hospital Oakland Research Institute, Oakland, CA 94609 Introduction Apolipoprotein E (apoE), an anti-atherogenic protein, plays a crucial role in cardiovascular disease by regulating plasma cholesterol levels and lipoprotein metabolism. By assisting in the transportation of very low-density lipoproteins (VLDLs) and high density lipoproteins (HDL), apoE is able to remove excess amounts of cholesterol and triglyceride from the blood and into the liver for processing. ApoE is able to remove plasma lipoproteins by acting as a ligand for cell surface localized lipoprotein receptors and proteoglycans. Specific lysine residues on apoE play a crucial role in lipoprotein receptor and proteoglycans interaction. Acrolein, a chemical found in cigarette smoke and generated as a metabolite of age-related oxidative stress, is highly reactive with lysine residues on proteins. Objective The objective of my research was to investigate if second hand smoke exposure leads to oxidative modification of plasma apoE by acrolein. Methods Rat plasma was treated with acrolein, and the lipoprotein fractions separated by density gradient ultracentrifugation. Immunoprecipitation and/or Western blot analyses of the lipoprotein and lipid-free fractions were carried out using anti-apoE and anti-acrolein- lysine antibodies. In addition, isolated lipoprotein fractions were treated with acrolein, and treated as above. Finally, Chinese hamster ovary (CHO) cells were treated with unmodified or acrolein-modified lipoproteins to monitor the uptake of apoE-containing particles by receptor mediated endocytosis. The uptake was monitored by immunofluorescence microscopy. Results Ex vivo exposure of plasma or isolated lipoprotein fractions to acrolein leads to oxidative modification of apoE. The lipoprotein binding ability of apoE, a critical function in facilitating receptor recognition, appears to be altered by acrolein. These results are consistent with earlier in vivo and in vitro observations from our lab that oxidative modification by acrolein causes structural and functional impairment of apoE. Conclusions Information obtained from this study will tell us if acrolein modification of plasma apoE can potentially lead to increased plasma cholesterol and triglycerides levels, which are established risk factors for developing heart disease. Thus, second hand smoke exposure may predispose individuals towards developing heart disease.
26 Ahmed Alghrouz Volunteer
University of New Mexico Second Year
Mentor: Hani Atamna, PhD
My name is Ahmed Alghrouz and I’m an international student from Palestine. I came to the U.S. in the summer of 2005 and I study at the University of New Mexico. My main goal is to get a degree in biology and go to medical school. That has always been the future that I look for. Coming from Palestine, where medicine is an essential service that people lack, gives me the courage and energy to pursue what I want. There are a lot of conflicts in Palestine and many people need medical care, so my ambition is to support the development of medical care and help as many people as I can in my country and other countries as well. Being a doctor is an honor that I really want to achieve.
I like sciences generally, but I like biology and chemistry more. Participating in this research program at CHORI has enriched my knowledge which will definitely help me do better in reaching my goal. Working in the lab and dealing with experiments that focus on real life issues has been very interesting and it taught me that I should try hard to learn and discover new things rather than just following what others found.
Helping people is my main goal and participating in this project, even with the little I could provide, teach me what it means to work for people’s health and it’s a very nice feeling. I will continue working in such projects in order to develop my abilities and gain more health-related knowledge. I will also keep working towards achieving my goal of being a productive and helpful person in the society.
27 Developing Methods for Isolating Amyloid-β-heme Complex From Brain Homogenate of AD Patients
A. Alghrouz & H. Atamna Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Alzheimer’s disease (AD) is a neurodegenerative disorder that affects about 4 million Americans and is the leading cause of late-onset dementia worldwide. The core biochemical impairments that lead to AD include excessive accumulation with age of a small protein in the brain called Amyloid-β (A-beta) and mitochondrial aberrations. This core change leads to energy deficiency, oxidative stress, and adverse metabolic consequences in nerve cells of the brain. However, the mechanism of mitochondrial aberrations by Aβ is not known. Research from Atamna’s lab has concentrated on elucidating this mechanism. Objective: The objective of the pilot experiments that I performed was to find a method to isolate or demonstrate Aβ-heme complex protein in brain homogenate from a number AD patients. Methods: The main methods used were sucrose gradients, ultracentrifugation, SDS-PAGE, Western blotting, HPLC, and enzyme assays. The criteria we used at this early stage to assess the presence of Aβ-heme in the different fractions was the presence of Aβ, peroxidase activity, and heme. Results: A protein fraction separated at sucrose gradients of 60 and 50% show enrichment in Aβ, peroxidase activity, and heme content. Future experiments will be directed at adding additional steps of purifications to demonstrate the presence of the Aβ-heme complex in these fractions. Conclusion: More specific and strict criteria to identify Aβ-heme will be used in the follow up experiments.
28 Andrea Banuelos Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Third Year
Mentor: Janelle Noble, PhD
I am privileged to be the first in my family to pursue a college education let alone have the opportunity to become involved in an endeavor such as research. I thank CHORI for the opportunity to participate in research.
From an early age I have known that I want to pursue a career in medicine. I grew up in an underserved primarily Latino community in Los Angeles, CA. As the oldest daughter in my family, one of my roles was to translate for my mother during medical visits. I developed a keen interest in medicine from these experiences observing doctors and I have always thought that I would return to my community to serve as their doctor and advocate. For I feel that in the process of attending their health I hope to also close the language and cultural barrier that often exists between patients and doctors in my community. Now more than ever I plan to pursue that goal for my family has experienced first-hand the loss of my brother due to lack of adequate healthcare in our community.
The research bug did not hit me until recently when I realized that there are so many diseases affecting our communities we do not understand well. Some diseases we know a lot about but there is still more we can learn about them. I have learned that the results found in a laboratory can affect the way medicine works for it may provide perspective on how to treat diseases and may help a physician better understand treatments. I now understand that with good research comes good medicine and I feel that I can help my community by conducting research. For that reason I plan to participate in research from this point on until my path in the medical field allows me to.
Currently I am attending UC Berkeley majoring in American Studies-Children with Disabilities. I plan to attend a post-bac program after my graduation in December and continue my path to medicine. There are many obstacles I have encountered throughout my undergraduate career but I believe that when there is a will there is a way and I look forward to all the experiences I may encounter on my way to medicine.
29 HLA-A Genotyping for African-American Control Group
A. Banuelos, J. Johnson, R. Kathail, O. Linney, J. Noble. Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: HLA (Human Leukocyte Antigen) is a protein found on the cell surface which is the human form of MHC (Major Histocompatibility complex). MHC is a set of genes coding for proteins involved in antigen presentation to the immune system. There are two main classes, HLA-Class I and class II. There are three HLA loci in Class I: A, B and C. For this study, HLA-A will be analyzed. HLA antigens are found to be the most polymorphic in humans-which may represent a greater variation in allele frequencies in a given population. Several autoimmune diseases are associated with HLA-A, one being Type I Diabetes which affects about 5% of the US population. In past studies it has been found that HLA accounts for about 50% of the genetic susceptibility to Type I Diabetes. There is little literature and research performed on HLA for the African-American ethnicity. For that reason the Noble Lab will genotype 1000 Blood Spot Cards from African American Newborns in order to create a control group for HLA-A Allele frequencies. This may then be used in comparison studies with African- American patients and in cross-ethnic comparisons. Objective: The overall objective of the project is to create a control group of 1000 African-American HLA-A genotypes. 1000 blood spot cards have been collected from African American newborns and can be used as a control group because they can effectively represent the general population. This information will then be made available as a comparative group from which differences in genotype may help determine predisposing or protective HLA-A alleles for several autoimmune diseases. For this project 92 out of the 1000 samples have been genotyped for HLA-A allele frequencies. Methods: 1000 African-American blood spot cards were sorted. Each card was labeled with an 8 digit numerical code which was used to help identify and organize the blood spot cards into plate grids. We then proceeded to blood spot card punching where using a hole puncher, each of the blood spot cards was punched into a bar-code labeled tube with the same corresponding numerical code which was followed by DNA extractions using a combination of GenVault reagents for DNA recovery and Qiagen reagents for DNA purification. We extracted DNA from 12 samples at a time. This was followed by a PCR (Polymerase Chain Reaction) which is a process of DNA replication through a series of heating and cooling. The two final steps included an SSOP (Sequence Specific Oligonucleotide Probe) which determines the probe binding pattern for each sample and analysis by Score a program that calculates the allele frequencies for each sample.
30 Kate Barber Funded by the Elizabeth Nash Foundation Stanford University First Year Mentor: Beate Illek, PhD
I’ve always been one to become curious about anything unknown. While taking physiology in my junior year of high school, I realized how amazingly intricate and phenomenal the human body is, and couldn’t wait to investigate it further.
My internship at CHORI has been particularly meaningful because of my research on Cystic Fibrosis. I have a friend I’ve known since kindergarten who is afflicted with CF, and her positive attitude and determination are truly inspiring. I hope to be able to make a contribution in the fight for a cure for Cystic Fibrosis.
While I don’t know exactly what major I’ll pursue at Stanford University over the next four years, I’ve taken this internship as an opportunity to explore scientific and medicinal research in daily life. I’ve loved hypothesizing and experimenting with incredibly knowledgeable doctors and researchers; it has been a fabulous way for me to explore medicine and determine how I want to become involved in it. Between my newfound scientific experience and my various passions of foreign languages, writing, and history, I hope to help connect patients, researchers, doctors, and the public.
I’d like to thank Dr. Lubin and Dr. Narayanaswami for providing this amazing opportunity, and Dr. Illek and Dr. Fischer for always being thoughtful and inspiring! Finally, I’d like to dedicate my research to my incredible physiology and biology teacher, Paul Yriberri, who recently passed away. His lively spirit lit up the life of everyone who knew him; without his friendship and inspiration I’d never have come this far!
31 Pyocyanin-induced H2O2 Production in Cystic Fibrosis Airways
K. Barber, C. Schwarzer, H. Fischer & B. Illek Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction Cystic Fibrosis (CF) is a hereditary disease that afflicts 70,000 people worldwide. CF results from a mutation on the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which affects the transmembrane transfer of chloride ions on the apical membrane of airway epithelial cells, leading to chronic bacterial infection. Antibiotic treatment prevents bacterial infection in pediatric CF cases, but by age 18, around 80% of CF patients are infected with the Pseudomonas aeruginosa bacterium. Pyocyanin (PCN), a small molecule virulence factor produced by this bacterium, plays a role in CF by its ability to redox cycle with NADPH and thus produce excess hydrogen peroxide (H2O2). Infected CF patients have significant levels of PCN in their airways (up to 100µ M). While scientists
have recently determined that airway epithelial cells produce small amounts of H2O2 on their own (via NADPH oxidases), high amounts of H2O2 can be cytotoxic. In addition to
producing harmful amounts of H2O2, PCN may also affect CFTR chloride transport, which is sensitive to reactive oxygen species. Objective The objective of my research was to mimic bronchial infection by exposing cells to PCN. We investigated how PCN affects
H2O2 production in both wildtype CFTR- and mutant ∆F508 CFTR-expressing human bronchial epithelial cells. We hypothesize that CF cells are more sensitive to pyocyanin
treatment and generate more H2O2 than normal cells. Methods Human bronchial epithelial cells expressing the wildtype CFTR and cells with the CF allele were plated onto Costar
96-well plates and H2O2 production was measured using the Amplex Red assay. The cells were then given either (i) phosphate-buffered saline (PBS) + glucose solution (ii) PBS +glucose+diphenylene iodonium (DPI, an NADPH oxidase blocker), or (iii) PCN at concentrations of 100 to 0.78 µM using serial dilutions +/- DPI. Amplex Red fluorescence was measured (excitation, 530 nm; emission, 590 nm) every 20 min. for 3 h. Results We
found that PCN increases H2O2 prodcution in both cell lines in a concentration- and time-
dependent manner. In wildtype CFTR-corrected CF cells, the rates of H2O2 production
were highest at mid-level PCN concentrations (12.5-50 µM) while CF cells’ H2O2 rates had a positive correlation with the PCN concentration. On average, CF cells produced H2O2 at a rate of 42.3 nM/min in a PCN concentration of 12.5 µM and at 92.5 nM/min in a PCN
concentration of 100 µM. Conversely, CFTR-corrected cells produced H2O2 at a rate of 101.6 nM/min in 12.5 µM PCN but only 60.0 nM/min in 100 µM PCN. DPI blocked 35% of
H2O2 in CFTR-corrected cells and 22% in CF cells. In cell free experiments, DPI blocked
20% of H2O2 generation in the prescence of NADPH and PCN. Thus, although we aimed to use DPI to distinguish physiological and chemical oxidation processes, we determined that it blocks both processes. Conclusion Based on these studies, we conclude that
PCN stimulates H2O2 production in human bronchial epithelial cells with and without the CF allele. Additionally, as CFTR may be involved in functional glutathione release, we
attribute the H2O2 peak at the mid-level PCN concentrations in CFTR-corrected CF cells to glutathione. We speculate that CF cells cannot buffer H2O2 production at high levels of PCN infection and are thereby less able to protect themselves against oxidative stress.
32 Sophia Blachman-Biatch Volunteer
The College Preparatory School Sophomore
Mentor: Stuart Smith, PhD, DSc
My name is Sophia Blachman-Biatch and I will be a high school junior this Fall at the College Preparatory School in Oakland, California. I am an academically focused 16 year old who enjoys science, history, research, socializing, singing in choirs and doing aerial arts.
I believe protecting and enhancing the health of children is critical to the survival of the world. I love spending time with children, and they enjoy spending time with me because I treat them with respect and know how to have fun with them. So after looking into possible summer jobs and programs that would combine these interests, I concluded that my best choice was the Children’s Hospital Oakland Research Institute (CHORI) Summer Research Program and this experience has exceeded my expectations.
Working in Dr. Stuart Smith’s laboratory focusing on fatty acid biosynthesis has proven to be an incredible experience; I have learned much that I expect to be useful in the future, and I am enjoying contributing to the research. Although I am unsure what I will ultimately study and where I will go, I believe it will have something to do with medicine and improving peoples’ health. If I have time I hope to continue working at CHORI during the coming school year and I hope to be at CHORI next summer.
I want to thank Dr. Saloni Pasta for teaching me biochemistry and lab techniques and for her patience with all my questions. Everyone in Dr. Smith’s laboratory has been kind, helpful and patient and I have learned more than I could have imagined at our laboratory team meetings, at the CHORI lectures and just in conversations. I would like to thank everyone who enabled me to participate in this marvelous experience.
33 Expression, Purification and Mutagenesis of Human Acyl Carrier Protein
S. Blachman-Biatch, S. Pasta & S. Smth Children’s Hospital Oakland Research Institute, Oakland, CA 94609
I have been working with the human acyl carrier protein (ACP). This protein domain is a component of the fatty acid synthase (FAS) system, which is a single multifunctional polypeptide involved in the de novo synthesis of fatty acids. Animal FAS contains 6 additional enzymatic domains besides ACP; the malonyl transferase, β-ketoacyl synthase, ketoreductase, dehydratase, enoyl reductase and thioesterase. The ACP carries the growing acyl chain to each of the six other enzymatic domains where reactions to make a saturated fatty acid occur. This process is important for energy storage and fat production and a better understanding has potential of developing new treatments for cancer and obesity realted illnesses. I began the process of expressing and purifying human ACP fragment by growing E.coli containing plasmid carrying the human ACP and inducing protein expression with IPTG. After lysing the cells, I proceeded with two different chromatography steps - Ni-NTA column chromatography, protein separation based on the presence of a His-tag, and anion exchange chromatography, separation based on charge. I checked purification levels throughout, on a 15% SDS-PAGE gel. Finally, I concentrated my protein and quantified it by the BCA method. ACP is active only in its holo-enzyme form once a phosphopantetheine (PSH) has been attached to a serine residue, by a phosphopantetheine transferase (PPTase). Since the activation of ACP and fatty acid synthesis involves interaction of the ACP domain with at least 7 different enzymes, I performed mutagenesis on the human ACP fragment to establish, which amino acids are the important ones that participate in the ACP interaction. By looking at the crystal structure of human ACP with the PPTase, we can see that certain amino acids, mainly hydrophobic, play a key role in this interaction. To confirm this theory we mutate the hydrophobic amino acids of the human ACP to Alanine. The mutations include – I2137A, V2143A and L2152A from the coil between helix 1 and 2; L2157A, M2158A and V2160A in helix 2; and V2176A in helix 3. Changes are made to the DNA through primers carrying the mutation by the quick-change site-directed mutagenesis method. Presence of mutations will be confirmed by sequencing reaction.
34 Sam Black Funded by the NIH/NHLBI
Albany High School Junior
Mentor: Kitty de Jong, PhD Frans Kuypers, PhD
Almost all my life I have been interested in helping others. This interest of mine has led me to peruse a career in medicine. As a high school student I recently developed an interest more specifically in research. Research struck me as an excellent opportunity to help impact a multitude of people in a positive manner: to have a significant impact upon the daily lives of everyone that is troubled by the investigated disease. It would be an amazing accomplishment, that someday I would hope to achieve, to discover something that could affect the lives of people with terminal diseases. Working at CHORI has exposed me to what working in research is really like. Most high school students are only exposed to research in their high school science classes, and these results are always expected and predetermined. By working at CHORI I have been given the opportunity to explore the unknowns of research. Investigating something that has not already been done a million times fills me with a sense of genuine accomplishment. Hopefully, my research at CHORI will help uncover some of the unsolved mysteries of sickle cell disease. Working at CHORI has been a fun and challenging experience that has truly affirmed my interest in perusing a career in medicine. I am exceedingly grateful that I was given this extremely unique opportunity.
35 Phosphatidylserine Exposure on Nornal and Sickle Human Erythrocytes by Oxidative Damage
S. Black, K. de Jong & F. Kuypers Children’s Hospital Oakland Research Institute, Oakland CA 94609
Introduction Within normal erythrocyte membranes there exists an asymmetrical distribution of phospholipids. Phosphatidylserine (PS) is usually found on the inner monolayer of an erythrocyte membrane. The flippase maintains this asymmetrical distribution by transporting PS from the outer monolayer to the inner monolayer using ATP. The scramblase, when activated, reduces phospholipid asymmetry by randomizing phospholipid placement within the membrane. Sickle cell disease causes flippase failure and scramblase activation leading to a small percentage of erythrocytes with increased PS exposure. PS exposure is a signal to the macrophage for cell removal, which can result in anemia. The scramblase becomes activated when Ca++ is introduced within an erythrocyte. Objective PS exposure will be measured in normal and sickle human erythrocytes after the introduction of organic peroxides. Within this setup the effects of Ca++ and a variation of incubation times will be examined. Additionally, permeabilized cells’ fluorescence will be compared to that of normal erythrocytes to check for erythrocyte absorption of the fluorescent compounds.Methods Normal and sickle human erythrocytes were exposed for 15 minutes at 37ºC to cumene hydroperoxide (97 µM to 741 µM) or tert- butyl hydroperoxide (250 µM to 1200 µM). The oxidized erythrocytes were labeled with Alexa 633–Annexin V, to identify exposed PS and then analyzed by flow cytometry. The detection of AV labeling in the FL4 channel was chosen to avoid autofluorescence in the FL1 channel caused by oxidation. Additionally, the times of oxidant incubation was varied from 5 to 30 minutes. Peroxide treated cells were evaluated for permeability when loaded with fluorescein or allophycocyanin-conjugated streptavidin (APC-SA). As a positive control for permeabilized cells, erythrocytes were fixed with gluderaldehyde and formaldehyde and then permeabilized with triton. Results It was found that as concentrations of organic peroxides increase the percentage of PS exposing cells also increases. Exposure to additional 2mM Ca++ during oxidation did not influence PS exposure. Sickle cells were found to have greater PS exposure than normal erythrocytes at lower organic peroxide concentrations, but lower PS exposure at higher organic peroxide concentrations. PS exposure was time dependant, reading its peak at 15 minutes in presence of 600 µM CuOOH and 1000 µM tBuOOH. Permeabilized erythrocytes, but not oxidized cells, had higher levels of fluorescence for both fluorescein and APC-SA. Conclusion Organic peroxides at high concentrations cause significant PS exposure to normal and sickle erythrocyte membranes in absence or presence of Ca++. The oxidant–treated cells were also found to not leak or absorb the fluorescent compounds. Sickle cells have oxidative damage that might contribute to the enhanced PS exposure found in these cells.
36 Amelia Brazil Volunteer
The College Preparatory School Junior
Mentor: Barbara Staggers, MD
My name is Amelia Brazil, and next year I will be a senior at The College Preparatory school. I have many academic and extracurricular interests, including singing, dancing, English, and Spanish, but my summer with the CHORI student program has greatly expanded my interest in science. I had always been a bit apprehensive about some sciences because of all the chemicals and calculations, but this year is school, I found my biology class to be really interesting and fun, and I especially enjoyed the units we did on the different organ systems of the human body. Learning about adolescent medicine this summer has been really fascinating, and I have been exposed to so many new things, including areas of medicine and social work. I have really enjoyed my time in all of the Teen Clinics and I have learned an amazing amount about adolescent medicine. Besides adolescent medicine, I have also learned about may other current topics in medicine through the weekly lectures on topics that I might otherwise never have been exposed to. I am so grateful to be able to participate in this great program, and after my last year of high school next year, I hope to somehow integrate my love of science with my other interests throughout college and in a career.
37 Depression and High Risk Behavior in Adolescents
M. Fisher, A. Brazil & B. Staggers Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction Depression is a serious health issue that affects many Americans today. Since depression is a precursor to many other health problems, including suicide, the third leading cause of death in adolescents, early and effective screening for warning signs of depression is especially important for adolescents. Adolescent physicians currently ask their patients a series of questions that comprise the H.E.A.D.S. survey, which includes questions about depression. If certain high risk behaviors were known to correlate significantly with adolescent depression, then physicians would be able to watch out for those specific behaviors and screen patients exhibiting them more comprehensively for depression. Objective The main goal of this project is to determine if there is a significant correlation between teens participating in high risk behavior and teens with depression. Finding specific links between high risk activities and depression will allow physicians to screen more effectively for warning signs associated with depression. This study will encourage a more complete assessment of depression in the H.E.A.D.S. assessment. Methods We shadowed doctors at the Children’s Hospital Teen Clinics for six weeks and asked patients to fill out the Center for Epidemiological Studies Depression Scale for Children (CES-DC). With the results, we were able to assess whether or not the patient was exhibiting possible signs of significant depression. Then we looked up the patient’s file and H.E.A.D.S. survey. Noting whether or not the patient was participating in the high risk behaviors being examined: alcohol use, not regularly exercising, fighting, having a GPA under 2.0, and tobacco use, we analyzed the relationship between these behaviors and depression. Results Our results showed that adolescents who showed possible signs of significant depression were more likely to participate in all five high risk behaviors than adolescents who did not show such signs. The specific high risk behavior with the strongest correlation to adolescent depression is not exercising regularly. While 36.1% of adolescents showing possible signs of significant depression did not report regular exercise, 18.75% of adolescents not showing such signs reported no regular exercise. The two behaviors with the weakest correlation to depression are alcohol abuse and having a GPA under 2.0. Conclusion Based on our study, we can conclude that adolescents who abuse alcohol, do not exercise regularly, fight, earn a GPA of under 2.0, or use tobacco are more likely to exhibit possible signs of significant depression than adolescents who do not participate in these behaviors. Our results will now be incorporated into the H.E.A.D.S. assessment.
38 Samuel Cartmell Volunteer
Yale University First Year
Mentor: Ward Hagar, MD
I am currently a rising sophomore at Yale, planning to major in Molecular, Cellular and Developmental Biology with a concentration in Biotechnology. After having completed the first semester of my freshman year of college, I found myself no closer to knowing exactly what I wanted to pursue professionally or academically. While I always had an interest in science, I had no experience putting the concepts I learned in school into practice. As such, I felt I didn’t have a solid grasp of what a career in science might be like. Then I was made aware of CHORI, and what seemed like a great opportunity to gain some experience in research. CHORI has proved to be just that, and not even halfway through the program, I feel like I’ve gained a solid grasp on what clinical medical research involves, and I’m extremely thankful for the opportunity CHORI has provided. CHORI has been an invaluable experience--beyond building upon my understanding of the concepts I’ve learned in school, I’ve learned a great deal about the field.
39 Factors Associated with Death in Patients with Sickle Cell Disease and Pulmonary Hypertension
S. Cartmell & W. Hagar Children’s Hospital & Research Center at Oakland, Oakland, CA 94609
Introduction Sickle Cell Disease is a genetic disorder of the hemoglobin affecting approximately 80,000 – 100,000 people in the US. Patients with Sickle Cell can face numerous, and often unique, health problems, in addition to a lower life expectancy. One such problem is Pulmonary Hypertension (PH), as defined by a tricuspid regurgitant velocity ≥ 2.5 m/s, which approximately 30% of adult patients develop. In adult patients, the development of PH is strongly correlated with a reduction in life expectancy often characterized by sudden death. While the association between the onset of PH and patient death is well established, the exact mechanism is unknown. Liver disease has been reported as a predicting factor, possibly playing a role in the death of patients with PH. Objective Since little is known about what ultimately causes death, it was objective of my research to compile and analyze existing data to determine possible predicting and contributing factors to PH and patient death. Additionally, it was my objective to develop a prospective study to monitor these factors and establish a natural history of PH in sickle cell patients. We were especially interested in liver, kidney, and lung function in relation to the development of PH. Methods With IRB approval, data was collected from several different sources, including echocardiograms, pulmonary function tests, and laboratory tests (blood and urinalysis). Transfusion history was also considered. Data, obtained electronically and manually, was entered into a excel database. The data was then read from the excel files into STATA, an analytical statistics program. The data was then put into a survival models for analysis. Results Preliminary results are compelling. PH in pediatric patients differs in clinical manifestation and presentation in adults. Factors found to be associated with PH in children included asthma or obstructive disease, history of sepsis, and history of acute chest syndrome (ACS). In contrast, in adults, neither sepsis nor asthma/obstructive disease was associated with PH, and ACS was a protective factor for patient survival with a univariant protective hazard rate of 0.23 (CI 95% 0.085 – 0.62, p=0.004). Additionally, a history of liver disease was just above significance with an adjusted hazard rate of 5.1 (p=0.073). Survival analysis, limited to adults, shows a univariant hazard rate of 17.3 (95% CI 4.9 – 60.4, p<0.00005) in patients with PH compared to those without the disease. Conclusions PH appears to differ in children and adults. It should be further examined to confirm possible risk factors, such as liver disease, identified in this study, and to establish a natural history of the disease.
40 Chris Cox Funded by Saba Lab
California State University Sacramento Post-baccalaureate
Mentor: Babak Oskouian, PhD Julie Saba, MD, PhD
When I was ten years old I was diagnosed with ALL leukemia and was treated under the care of Dr. Feusner, Dr. Wu, and Dr. Hastings at Oakland Children’s Hospital. Thanks to the excellent care I received by the physicians and nurses, I have been in remission for over thirteen years. It has been my dream to become a pediatric oncologist and help other children who are fighting childhood disease. I hope that my work this summer, performing cancer research with Dr. Saba, will increase our understanding of cancerous cell lines that will ultimately lead to better cancer treatments.
41 Leveraging Sphingolipids in the Treatment of Leukemia
C. Cox, B. Oskouian & J. Saba Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: About 33 percent of cancers in children ages 0-14 years are leukemia. Current treatment for leukemia is invasive and non-specific. Sphingolipids are natural lipids found in the membranes of all human cells. They contribute to the functions of the cell membrane and give rise to lipid signaling molecules. Some of these signaling molecules produce cell death and account for the main mechanism by which chemotherapy and radiation kill cancer cells. In contrast other molecules, such as S1P, have been shown to stimulate the proliferation and survival of cancer cells. Our lab has found that S1P levels are greater in colon cancer polyps than in other areas of the colon, affirming the hypothesis that colon cancer cells use elevated levels of S1P to multiply and grow. Similarly, many studies link S1P accumulation to leukemia cell development, survival and resistance to chemotherapy. These findings suggest the balance of growth-inhibitory versus growth- stimulating sphingolipids could be responsible for determining whether a cancer cell lives or dies. We would like to know if this is also true for leukemia cells. Hypothesis: Altering intracellular sphingolipids could be leveraged to improve leukemia responses. This could be accomplished by either adding cytotoxic sphingolipids or removing growth- stimulating sphingolipids such as S1P. Objectives: 1) Characterize the sphingolipid pathway in leukemia cells; 2) Evaluate the effect of different sphingolipid compounds on the survival and signaling events in leukemia cells. Methods: We propagated several kinds of lymphocytic and myelocytic leukemia cell lines. Then we treated the cells with different concentrations of sphingolipids and then evaluated the effect on cell survival and apoptosis. Next, we harvested the cells from the culture and determined the amount of S1P lyase in these cells, which required separating the membrane (where the lyase resides) from the cytoplasm. This allowed us to probe specifically the S1P-Lyase in the membrane without any background interference. We used a western blot technique to identify S1P-Lyase levels in the cells. These results were obtained by probing for the S1P-Lyase enzyme using a western blot technique. These results can then be analyzed to determine relative levels of S1P-Lyase. Similar methods were used to explore AKT activity and cell death proteins in the leukemia cells. Results: The experiments have not yet been completed. Conclusions: Understanding the ways in which leukemia cells proliferate is a critical step in the development of better, less invasive treatments for leukemia. Results that affirm our hypothesis it would be a major step in using anew signaling pathway to treat leukemia.
42 Ramine Cromartie Funded by the NIH/NHLBI/Professional Student Short Term Program Stanford University Second Year Mentor: Guangming Li, PhD Bruce Ames, PhD
In the fall, I will be entering my third year in the Chemical Engineering program at Stanford University. I enjoy learning about the world around me and studying how its different components interact. Since childhood, I frequented the area’s science museums to build my knowledge and continually spark my curiosity in various topics in science. Mathematics is another academic interest of mine. Over the years, I have been confronted with newer, more difficult problems and have loved the challenge of implementing mathematical skills and models to solve them. With these interests in math and science, it seemed natural for me to gravitate toward further study in an engineering field. Like scientists, engineers seek to answer “how” questions, but they do so through design and process analysis. I hope that this training will provide me with a unique perspective toward both clinical and scientific research.
Through my participation in community outreach programs and community service organizations, I have realized another passion- community health. I feel especially drawn toward the physical, mental, and spiritual treatment of the teenage population. Born and raised in the Bay Area, I have been blessed with many great opportunities for personal and professional enhancement. I have been exposed to many great individuals who have helped me cope with the challenges of my adolescence. I would like nothing more than to do the same for future generations. I recognize my obligation to give back to the community that has given me so much. After graduation, I plan to attend medical school. I aspire to have a career that integrates my interests in science, technology, medicine, and public health.
This summer, I had the fortune of working with Dr. Bruce Ames lab group in the Nutrition and Metabolism Center. I would like to thank Dr. Guangming Li for his mentorship and experimental instruction and the Summer Program staff for the discussions and lectures that emphasized the social aspects of research. I am very appreciative of CHORI’s efforts in my development into not only a proficient scientist, but also a socially conscious one.
43 Differential Distribution of 25-Vitamin D in Human Plasma and Leukocytes
R. Cromarie, G. Li & B. Ames Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Vitamin D is a seco-steroid hormone that is responsible for efficient calcium and phosphorus absorption. In humans, the precursor for vitamin D, 7-dehydrocholesterol, reacts with the UV B rays of sunlight to form pre-vitamin D3. Pre-vitamin D3 then isomerizes into the metabolite cholecalciferol, or Vitamin D3. Vitamin D is stored in the adipose tissue and enters blood circulation by forming a binding complex with Vitamin D Binding Protein
(VDBP) and albumin. Vitamin D is hydroxylated in the liver by vitamin D3-25-hydrolase to produce 25-hydroxyvitamin D. The metabolite may undergo further hydroxylation by renal enzymes to produce 1,25-dihydroxyvitamin D. 25-hydroxyvitamin D has a higher binding affinity with VDBP, and it is the major circulating form of vitamin D in the plasma. It is used to evaluate vitamin D sufficiency status since its level is less affected by sun exposure, diet and vitamin supplementation, in comparison with other vitamin D levels in the plasma. Ames proposes a triage allocation of scarce micronutrients. In such instances, biological functions essential for the short-term survival will be favored in emergent circumstances and receive the bulk of scarce micronutrients. Consequently, biological functions that may be required to sustain long-term health may receive disproportionately lower amounts of micronutrient concentrations. In this research project, we investigated whether Vitamin D is differentially allocated in different components of human blood, with various biological functions, in healthy individuals as a beginning step to test this hypothesis. Objective: To determine the concentration of 25-hydroxyvitamin D and blood plasma and white blood cells. Methods: Red blood cells, leukocytes, plasma and platelets were isolated from blood donated by volunteers for Vitamin D analysis. Isolation of leukocytes was conducted according to the protocol used in our laboratory. Plasma and white blood cells were stored in an -80ºC freezer until analysis. Vitamin D3 concentration was determined from plasma and leukocyte samples by EIA assay, according to kit instructions. Results: The 25-hydroxyvitamin D in the plasma was determined from 23 healthy human subjects. The corresponding concentration for leukocytes from each subject has not yet been determined. However, preliminary data from one healthy subject indicates the 25-hydroxyvitamin D level in leukocytes is indeed low, compared with its concentration in plasma. Conclusion: In a normal health state, 25-hydroxyvitamin D was shown to preferentially allocate in human plasma.
44 Nadia Dhahbi Volunteer
Alameda High School Junior
Mentor: Joseph Dhahbi, PhD David Martin, MD
My name is Nadia Dhahbi and I am going to be a senior at Alameda High next year. As a kid growing up in Southern California I was surrounded by all kinds of desert wildlife. I have always had a fascination for reptiles and arachnids. I have never been afraid of any living thing and I am a risk taker. I like living life never knowing what is going to come around the corner. I prefer to live life to its fullest and see how inspiration hits me.
My mother is a very intelligent woman and excelled in school. She is now a clinical psychologist and has a Masters in Business Administration. My mother has installed the courage I have inside me again and is always pushing me in the right direction. She is always looking for opportunities to help me grow as a person and to help me get into a good college.
A few months ago the CHORI summer student program was recommended to me by my father, and I found it to be the perfect opportunity to broaden my horizons on the subject of Biochemistry. My father is a biochemist and studies the effects of Senescence, he also worked in the field of life expansion for a while. When I was younger he would let me watch him work in the lab work with agarose gels, viewing cells under the microscope, and much more. He also talked to me in a non-condescending way, explaining his research and the purpose of every experiment I had the opportunity to observe. Because of my fathers attitude in showing me the ways of science I became very interested in his field of work. I want to be just like my father, he has always been my hero. He has influenced me in such a big way and because of this program I have the opportunity to work side be side with him as a fellow researcher, I am very thankful and hope to continue my project next year. I am excited to see what is in store for me.
45 Effects of Senescence on DNA Methylation
N. Dhahbi, J. Dhahbi, D. Martin & W. Magis Children’s Hospital Oakland Research Institute, Oakland , CA 94609
Introduction: DNA methylation is critical for normal development and becomes altered during aging and age-associated diseases such as cancer. Global hypomethylation and CpG island hypermethylation progressively accumulate with age, causing changes in gene expression that may directly contribute to aging. The Avy allele of the agouti mouse has an intracisternal A particle (IAP) inserted upstream of the coding region; the IAP may be methylated and silent, or unmethylated and active. These properties make Avy an ideal cellular model system for measuring modulations of DNA methylation. The potential hypomethylation of the IAP may trigger the transcription of the agouti gene in senescent fibroblasts. Objectives: 1. To assess the effects of cellular senescence on cytosine methylation in the Avy locus. 2. To evaluate the effects of any potential change in the methylation state of the Avy locus on its activity state. Methods: Tail biopsies are minced and digested. The digest will be filtered, washed with DMEM and centrifuged. The cell pellet is resuspended in 3 ml of DMEM and cultured in a 6-well dish. Cells are passaged when they are confluent at 1 x 105 cells/cm2 flask surface area at each passage. One half of cells are harvested from the early passages and frozen. The remaining half of the cells are cultured until they manifest the flat senescent morphology and show a constant decline in cell number over time. These late-passage cells are harvested and frozen. DNA is extracted from the young and senescent fibroblasts using the Qiagen Epitect kit. DNA is treated with sodium bisulphate and used in PCR. PCR products are cloned into pGEM-T and transformed into E.coli, and plasmid DNA from individual colonies is sequenced. Results: Young fibroblasts display dense CpG methylation at the IAP in the Avy allele. After several passages, when fibroblasts reached senescence, the Avy IAP displays significantly less CpG methylation in comparison to young fibroblasts. This confirms the view that methylation is affected by aging. The senescence-induced de- methylation of retroelements could interfere with transcription of genes in their vicinity, a mechanism that may contribute to aging.
46 Tye Elliott Funded by the NIH/NHLBI/Professional Student Short Term Program San Francisco State University Fourth Year Mentor: Ellen Fung, MD
I am a pre-medical, post-baccalaureate student at San Francisco State University. I completed my B.S. of psychobiology at the University of California, Los Angeles. In my current career at SFSU, I am a very active member of the health professions society on campus.
Becoming a medical doctor and serving underserved communities is something that I aspire to do. Ultimately, I want to affect the health consciousness of the underserved on a large scale. I plan to further my knowledge of the disparities in healthcare and the cultural and social conditions that predispose people in underserved communities to certain illnesses in order to create educational and intervention programs. With my breadth of experiences I hope to matriculate at UCLA David Geffen School of Medicine in the upcoming year.
47 Bone Health in Chronic Diseases of Childhood
T. S. Elliott & E. B. Fung Children’s Hospital & Research Center at Oakland, CA 94609
Introduction: Osteoporosis is a disease characterized by decreased bone mass and strength leading to increased fracture risk. Failing to attain optimal bone mass during childhood increases the risks of developing osteoporosis later in life. Dual-energy x-ray absorptiometry (DXA) measures bone mineral density (BMD) and Z-scores are calculated from standard reference data to predict relative risk. Objective: The aims of this project are 1) to determine which group of children cared for at CHRCO has the greatest prevalence of low BMD, 2) identify factors that predict low BMD in these patient populations and 3) evaluate the relationship between BMD severity, referral characteristics and clinical management. Methods: The Bone Density Clinic’s (BDC) database was retrospectively queried for pediatric patients scanned within the last 2 years from five common referrals (hematology, nephrology, pulmonology, gastroenterology, and orthopedics). Information regarding anthropometrics, BMD Z-score from the spine, hip or whole body, calcium intake and supplement use, medication, fracture history, and family history of osteoporosis were gathered and analyzed using STATA, v. 9. Low bone mass was defined as a BMD Z- score <-2.0. Additionally, a brief 10 question survey was created using Surveymonkey. com. 34 CHRCO providers from the 5 disciplines were asked to complete the online survey of which 16 (47%) responded. Results: 138 patients (67 male) between 4.4 and 20.9 years were scanned between June 2005 and July 2007. 21 patients had follow-up scans within this time period. There were no differences in age or gender distribution by diagnosis. On average, calcium intake was 950 ± 600 mg/day and decreased with age. Despite the majority with low calcium intake, only 33% of patients took calcium supplements. Thirty-five percent of patients had one or more sites with a BMD Z-score <-2. The prevalence of low bone mass was highest in hematology patients; 64.5% of patients characterized with low bone mass had either Thal or SCD. With the exception of asthma, each group had at least 19% of patients with abnormally low BMD Z-scores. In univariate analysis, factors related to low BMD were increased age, family history of osteoporosis, and consumption of >2 servings of dairy/day (all p<.02), whereas weight was protective. Of the 21 with follow-up scans, only 8 showed Z-score improvements of which 5 had increased calcium intake. Based on the survey, more than 66% of providers refer patients to the BDC between 12 and 15 years old, primarily based on diagnoses. Almost all respondents (94%) recommend calcium and/or vitamin D supplementation in patients who receive a low BMD report. Conclusions: Though low bone mass is common in all pediatric referrals to the BDC at CHRCO, it is most prevalent in patients with hematological disorders. In children with chronic disease, bone accrual is slower compared to healthy children, therefore Z-score decreases with age. Unfortunately, many referrals are for older patients who may already have significant bone deficits and calcium supplementation is uncommon. These results suggest that when possible, patients should be referred at a younger age such that modifiable factors such as nutritional adequacy, calcium/vitamin D supplementation, and weight bearing activity may be implemented.
48 Melanie Fish Funded by NIH/NHLBI
Piedmont High School Junior
Mentor: Barbara Staggers, MD Justin McKone
I have always had a love for medicine and medical science. I grew up in a medical family. My father is a radiologist; my mother is a critical care nurse. From as far back as I can remember, medical terms, medical issues, and related topics were frequently discussed in my household. That familiarity, along with a natural curiosity about such matters, has sparked my interest in the field of medicine. I found it to be fascinating and always wanted to learn more about medicine and medical science.
In school, I have been very active in pursuing my interest in science. As a junior, I took three science courses––AP Environmental Science, Honors Chemistry, and Anatomy & Physiology. Next year, as a senior, I plan to take AP Biology, AP Physics, and Sports Medicine. By the time I graduate, I will have taken eight science classes (including Integrated Sciences I & II as a freshman and sophomore). Because many fields of science interest me, I have tried to explore as many of them as possible throughout my high school career. Through the CHORI Summer Research Program, I have been able to bridge the gap between my textbook knowledge and real-life experiences. Working in the Teen Clinic has given me insight into the lives of teens from an objective point-of-view, which in turn has given me a new perspective on my own life as a teenager. This has been an extremely rewarding experience, and I would love to continue doing similar work in the future. Now, more than ever, I can see myself going to medical school and pursuing a career in the field of medicine.
49 Standardized Comprehensive Care for Adolescent Sickle Cell Patients
M. Fish, J. McKone & B. Staggers Children’s Hospital & Research Center at Oakland, Oakland, CA 94609
Introduction: The Comprehensive Sickle Cell Center at the Children’s Hospital Oakland outpatient clinic treats a total of approximately two hundred sickle cell patients between the ages of thirteen and twenty-one, many of whom require monthly blood transfusions. Although these patients are receiving world-class treatment for sickle cell, they do not have adequate access to sensitive services. A large portion of these teens are participating in high-risk behaviors and putting themselves in dangerous positions. Although Children’s Hospital Oakland has a nationally recognized Division of Adolescent Medicine at off- site locations, these doctors who specialize in high-risk adolescent behavior are seeing only a few of the teen sickle cell patients. Objectives: Our objective is to improve the psychosocial care of patients with sickle cell disease and to ensure that they are receiving adequate services for high-risk as well as normal teenage issues. Method: We designed a bi-annual questionnaire for the teenage sickle cell patients, which will enable the treating physicians to maintain a current psychosocial profile. We also taught the physicians at the Comprehensive Sickle Cell Center how to implement the HEADDSS assessment, a psychosocial assessment that screens for depression and high-risk behaviors. We established a protocol for referring the most high-risk patients to a specific Teen Sickle Cell Clinic at the Division of Adolescent Medicine’s Teen Clinic on Telegraph Avenue. Once identified as high-risk (using the tools stated above) these teens will be referred to the clinic to be provided with the specific sensitive services they require. The patients who attend the Teen Sickle Cell Clinic will have better access to birth control, gynecological services, counseling, routine check-ups, etc. Onsite social workers will also be readily available to the patients. Anticipated Outcome: Starting in September there will a be a monthly or bi-monthly (depending on the need) dedicated Sickle Cell Teen Clinic held at the Teen Clinic on Telegraph Avenue. Patients will attend the Sickle Cell Clinic in the morning and shuttle to the outpatient clinic at the hospital site in the afternoon for transfusions and other sickle cell-related services. By coordinating the times for the Teen Clinic and Hematology appointments, the amount of school or work the patients miss will be minimized. Our goal is to provide identification, communication, and access, all of which will enable these patients to receive standardized comprehensive care.
50 Myles Fisher Funded by the NIH/NHLBI/Professional Student Short Term Program California State University Chico Second Year
Mentor: Barbara Staggers, MD
My name is Myles Fisher and I will be starting my third year at Chico State University in the Fall. I started college as a physics major, but in reality I had no idea what I wanted to major in, and ultimately what to do with my life. Trying to figure out what I wanted to do with my life I decided to apply to a medical internship and last summer, the summer of 2006, I participated in an internship called Faces for the Future, which is run through Children’s Hospital Oakland and Research Institute. During the internship the other students and I were exposed to many different fields with in the medical profession. Although there were many great things about the FACES program the surgical unit really caught my attention and interest. At this point I had become enthusiastic and excited about a possible career in medicine and later during that summer I was able to shadow a vascular surgeon at UCSF for a few days. This was a truly amazing experience and after witnessing multiple surgical operations I became convinced that I wanted to go to medical school and eventually become a surgeon.
This summer I am studying under my mentor Dr. Staggers and am conducting a study that compares teen depression and high risk behavior. Working at a teen clinic in Oakland with Dr. Staggers and the other M.D.s has really opened my eyes to adolescent medicine within our community. This CHORI internship has been such a great opportunity for me to grow and develop my perspective on the medical profession. I’d like to thank everyone who has dedicated their time and effort to make this a great experience for myself as well as the other students.
51 Depression and High Risk Behavior in Adolescents
M. Fisher, A. Brazil & B. Staggers Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction Depression is a serious health issue that affects many Americans today. Since depression is a precursor to many other health problems, including suicide, the third leading cause of death in adolescents, early and effective screening for warning signs of depression is especially important for adolescents. Adolescent physicians currently ask their patients a series of questions that comprise the H.E.A.D.S. survey, which includes questions about depression. If certain high risk behaviors were known to correlate significantly with adolescent depression, then physicians would be able to watch out for those specific behaviors and screen patients exhibiting them more comprehensively for depression. Objective The main goal of this project is to determine if there is a significant correlation between teens participating in high risk behavior and teens with depression. Finding specific links between high risk activities and depression will allow physicians to screen more effectively for warning signs associated with depression. This study will encourage a more complete assessment of depression in the H.E.A.D.S. assessment. Methods We shadowed doctors at the Children’s Hospital Teen Clinics for six weeks and asked patients to fill out the Center for Epidemiological Studies Depression Scale for Children (CES-DC). With the results, we were able to assess whether or not the patient was exhibiting possible signs of significant depression. Then we looked up the patient’s file and H.E.A.D.S. survey. Noting whether or not the patient was participating in the high risk behaviors being examined: alcohol use, not regularly exercising, fighting, having a GPA under 2.0, and tobacco use, we analyzed the relationship between these behaviors and depression. Results Our results showed that adolescents who showed possible signs of significant depression were more likely to participate in all five high risk behaviors than adolescents who did not show such signs. The specific high risk behavior with the strongest correlation to adolescent depression is not exercising regularly. While 36.1% of adolescents showing possible signs of significant depression did not report regular exercise, 18.75% of adolescents not showing such signs reported no regular exercise. The two behaviors with the weakest correlation to depression are alcohol abuse and having a GPA under 2.0. Conclusion Based on our study, we can conclude that adolescents who abuse alcohol, do not exercise regularly, fight, earn a GPA of under 2.0, or use tobacco are more likely to exhibit possible signs of significant depression than adolescents who do not participate in these behaviors. Our results will now be incorporated into the H.E.A.D.S. assessment.
52 Ivan Friedman Volunteer
International High School Junior
Mentor: Greg Moe, PhD
My name is Ivan Friedman and I am about to enter my senior year at the International High School.
Since I can remember, sciences have played a significant role in my life in and outside of school. As a child, before going to sleep, I would read through “How Things Work” books, or my National Geographic magazines. In school, I always seemed to excel more in the sciences than other subjects because of a youthful interest and curiosity for the world and how it “works”.
The older I grew, the more I knew I’d want to pursue a career in science. Whether my job involved physics, chemistry, or biology, I wanted to be some form of scientist exploring the ways and the reason the world functions. Though at first being a doctor didn’t appeal to me, a particular event in my life changed my perspective. In sixth grade I broke my arm and was rushed, of course, to Children’s Hospital in Oakland. Although I was much younger, I remember being impressed by the doctors’ work. It was then that I realized that the best way for me to put my scientific curiosity to use was to help sick or injured people. I decided medicine was the perfect path for me.
I applied for this internship not only with the hope of experiencing the world of medicine first hand, but also to deepen my understanding of the field. Until recently, when I thought of doctors, I thought of hospitals rather than research laboratories. This internship will show me more of what goes on “behind the scenes” in medicine.
53 Sialyl Transferase Gene Expression in Three Human Cancer Cell Lines
I.Friedman, P. Plested & G. Moe Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction. Sialic acids are negatively charged sugars that are added to proteins and lipids to modify molecular interactions. Human cancers frequently display aberrations in both the amount of and type of sialic acid modifications. Our laboratory has discovered that a monoclonal antibody, SEAM 3, binds to an antigen (SEAM 3-reactive antigen or S3RA) that is highly expressed in many human cancers but is not detected in the corresponding normal cells. The epitope recognized by SEAM 3 is poly alpha (2à8) N-acetyl neuraminic acid, which also is called polysialic acid (PSA), containing a small fraction of de-N-acetylated residues. SEAM 3 binding to S3RA expressed by cancer cells results in arrested cell growth and induction of programmed cell death (apoptosis). Currently, little is known about how S3RA is synthesized or what glycoconjugates might be modified with it. There are six sialyl transferases that have been identified in human cells that can synthesize alpha (2à8) glycosidic bonds between N-acetyl neuraminic acid residues. Only two of the sialyl transferases, ST8siaII, ST8siaIV, have been shown to synthesize PSA. ST8siaII and ST8siaIV are both highly expressed during embryonic development but ST8siaII is not expressed or expressed at low levels in adult tissues. Objective. The goal of this project is to determine which of the six known sialyl transferase genes are expressed in three different human cancer cell lines known to express S3RA. Methods. mRNA was isolated from human Jurkat T-cell leukemia, CHP-134 neuroblastoma and SK-MEL-28 melanoma cancer cells using a High Pure mRNA Isolation kit (Roche Applied Science). One or, in some cases, two validated PCR primer sets for each sialyltransferase gene were used to produce and amplify cDNA fragments using a OneStep RT-PCR kit (Qiagen). The PCR products were analyzed by agarose gel electrophoresis. PCR experiments were repeated multiple times using two or more mRNA preparations to ensure consistency. The beta-actin PCR product served as a control for the amount of mRNA added to the reaction and human brain mRNA was used as a positive control for the predicted PCR product for each transferase gene. Results. CHP-134 cells expressed mRNA for all of the ST8sia genes except for ST8siaVI, which was ambiguous for all three cell lines and the brain mRNA control. SK- MEL 28 cells expressed mRNA for ST8sia I, II, and IV and Jurkat cells were positive for expression of only ST8siaI and IV. Conclusion. It is likely that S3RA is derived from PSA synthesized by ST8siaIV since S3RA is highly expressed in Jurkat cells but of the two PSA sialyl transferases only ST8siaIV is expressed. The result is of interest because ST8siaII is most active in producing embryonic PSA antigens and tumors often have embryonic sialic acid expression profiles as opposed to those observed in adult tissues. Since the glycoconjugates modified by ST8siaII and ST8siaIV are different, the result suggests that S3RA is not an embryonic antigen that might explain its apparent absence from normal adult tissues. Thus, S3RA may result from an abberant modification of a normal adult PSA glycoconjugate.
54 Ben Goldenson Volunteer
University of California Los Angeles Second Year
Mentor: Ronald Krauss, MD
For as long as I can remember, I have been intrigued by the ways things function. Various natural phenomena from eclipses to magnetism and volcanoes to the human nervous system have piqued my interest. In high school, 11th grade, I took my first formal biology class, AP biology. I was immediately drawn to the subject. The elegance in which complicated tasks like DNA replication and signaling in the nervous system were handled, continue to amaze me. Another aspect of the class that was extremely interesting and novel to me was the experiments which accompanied information presented in the biology text. Certain experiments such as Oswald Avery’s genetic material experiment and the Meselson-Stahl semi-conservative DNA replication experiment were so logical that the conclusions were nothing less than beautiful. At this point biology was my favorite subject, and I am majoring in biology as a junior at UCLA. I am deeply interested in biological research as well.
I aspire to be a scientist or researcher in the field of biology or medicine. My experiences in research now will assist me in achieving my goal of becoming a skilled and knowledgeable researcher. Basic scientific research is crucial, but the knowledge that the work I do can be directly applied to helping people is deeply satisfying to me. Research on diseases and other biological/medical conditions is the area of inquiry most interesting to me.
Interning at CHORI has been a great experience. I have been exposed to cutting-edge research techniques in a beautiful lab and have learned a tremendous amount about a field I had not known much about before. My research project on lipoprotein metabolism is extremely fascinating and important due to its relation to the obesity epidemic in America. The experiments may lead to a cure for several diseases, but no matter the results, I am grateful for and will greatly benefit from my experience working at CHORI.
55 Effect of Lipoprotein Substrate, Microenvironment and Fatty Acid Source on Lipolytic PPAR α Activation
B. Goldenson, M. Ruby & R. Krauss Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction Peroxisome proliferator activated receptor (PPAR) α, a nuclear hormone receptor, regulates lipid metabolism and inflammation. The discovery that fatty acids bind and activate PPAR α has led it to being viewed as a general lipid sensor. Recent work suggests that lipoprotein-derived lipolysis products act as natural PPAR α ligands, in preference to free fatty acids (FFA) from other sources. A better understanding of endogenous PPAR α activation may allow more effective agonists to be developed and deleterious side effects to be minimized. Objectives We aim to assess the lipolysis of very- low density lipoproteins (VLDL) as a pathway generating PPAR α ligands. These studies will determine the effect of lipoprotein substrate, the lipolytic microenvironment on PPAR α activation and compare the efficiency of lipoprotein-derived FFA to fatty acids from other sources. Methods Lipoproteins were fractionated from plasma by ultra-centrifugation. PPAR α activation was assessed using a standard cellular reporter system. Briefly, bovine aortic endothelial cells were transfected with a chimeric PPAR α receptor driving a luciferase reporter and beta-galactosidase as an internal control. Cells were stimulated with lipase, lipoproteins, and albumin as indicated. FFA measurements were made in cell culture media using a Wako enzymatic kit. Student’s T-test was used to determine significance. Results VLDL treated with lipoprotein lipase (LpL), but not VLDL or LpL alone, significantly activated PPARα (25.7 fold; p=.0001). Enzymatic inhibition of LpL by orlistat or Apo CIII prevented the PPAR α activation (95% and 40% inhibition respectively; p<.05,). PPAR α activation was linearly correlated with FFA concentrations in cell culture media (R2=.81; p<.0001). There was no significant difference in the relationship between PPAR α activation and FFA concentration between VLDL–derived FFA and albumin- bound oleic acid. However, similar FFA concentrations achieved by addition of human plasma failed to activate PPAR α. Addition of a physiological concentration of albumin to VLDL-LpL significantly reduced PPARα activation (p<.0001). Conclusion All treatments displayed a strong correlation between FFA concentration and PPAR α activation. PPAR α activation was strongly affected by enzymatic inhibition of LpL, microenvironment, and fatty acid source. These studies support the role of lipoproteins, and not plasma FFA, as circulating pools of PPAR α ligands. Elucidating the mechanisms of endogenous PPAR α activation may lead to improved efficacy of current treatments and novel therapeutic targets.
56 Wendy Gordillo Funded by the NIH/NHLBI/Professional Student Short Term Program San Francisco State University Third Year Mentor: Patsy Wakimoto, PhD
Coming from a monolingual household and being the oldest child, I had the responsibility of being the translator whenever there was a medical situation. I had to help my mother translate when she had to take any of my siblings to the hospital. Being a nine-year old child just learning the English language, I had to try my best to tell my mother what the physician was telling her, but at the same time felt confused and frustrated when the pediatrician used medical terminology that I did not understood. These types of personal experiences made become interested in the medical field.
As a high school sophomore at Skyline High School, I was given the opportunity to become part of the FACES for the future program at Children’s Hospital where I shadowed physicians in different departments. From the program, I learned that medicine is truly what I want to do. I also appreciate the importance of being bilingual. Currently I am a third year student majoring in Cell and Molecular Biology and minoring in Raza studies at San Francisco State University.
Last summer I was given the tremendous opportunity to participate in the program, where I conducted community-based research. I learned that community-based research interest me, as a type of research which helps to better understand the real problems that impact our communities. Medicine is the field that truly interests me. I hope to become a pediatric pulmonologist and serve our community as well as to conduct community-based research.
I’d like to thank CHORI for giving me the tremendous opportunity to become part of the Summer Student Research Program. I also want to thank Patsy Wakimoto for giving me the support and tools necessary to understand more about community-based research in our community.
57 Latinos and Type 2 Diabetes
W. Gordillo and P. Wakimoto Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Latinos are the largest minority group in the U.S. Diabetes is a serious health concern and a growing epidemic for Latinos. National data indicates that Latinos are 1.7 times as likely to get Type 2 Diabetes than non-Hispanics whites. Objective: The objectives are to: 1) examine the attitudes, beliefs and issues surrounding diabetes. 2) examine knowledge about diabetes, and 3) to provide informational resources that are available in their own communities. Methods: The study sample was Latino adults with Type 2 Diabetes. Institutional Review Boards at Children’s Hospital Oakland Research Institute and La Clinica de La Raza approved the research study. Participants were recruited from Alameda County Public Health Department, La Clinica de La Raza, and personal contacts. Two methods, focus groups and individual interviews, were used to examine how much the patients knew about diabetes, relation to nutrition and physical fitness, cultural beliefs and use of traditional remedies. A questionnaire was developed which consisted of neutral, open-ended questions about their experience as Latinos with Type 2 Diabetes. The questionnaire contained questions from previous research studies as well as questions that would address the objectives above Individual interviews lasted approximately 30 minutes. The focus groups lasted approximately 90 minutes and were composed of eight to twelve subjects. The participants shared their personal experiences and knowledge. Transcripts and notes were reviewed and analyzed; common themes, major points and notable comments were identified. Results: Preliminary results (n=35) indicate that there is a clear understanding that diabetes is a disease that is progressive and that complications can be minimized through diet, physical activity and taking medications as prescribed by physicians. An interesting result was that although information is presented in Spanish, there is a lack of providing cultural appropriate information about diet. Another important result was that diabetes affects the patient’s life in many different aspects because they began to experience many physical and emotional changes that affect the body negatively. Also another aspect that is affected, but positively, was that many diabetic patients become closer to their family because they want to prevent their family members from getting diabetes. Conclusion: The results indicated that there is a need for exposure to further information, such as culturally sensitive information about diet and resources such as healthcare insurance and accessibility to glucose meters.
58 William Green Volunteer
University of Pennsylvania First Year
Mentor: Vladimir Serikov, PhD Frans Kuypers, PhD
After finishing my freshman year at the University of Pennsylvania I have had my share of epiphanies and realizations about what goes on outside of life as a child. I have recognized that at this point in my life what I do today lays the stepping stones of a path that leads me to tomorrow. Coming to this realization is not easy, as it makes many, or at least me, scared of taking a wrong step.
Let’s back up now. Ever since third grade I excelled at math and science, which has lead me to the engineering school at Penn. However, halfway through the year I began to doubt whether engineering was meant for me. To me it seemed as though applied science was a step behind the raw sciences of chemistry and physics, as those disciplines are the foundation for engineering. I began to think that hard research following the scientific method might interest me more, or might just stimulate my mind in a way that I hadn’t yet experienced. Moreover, being involved with scientific inquiry that leads to the creation, not just learning, of knowledge is unparalleled inspiration, that one cannot find hardwiring circuit boards. So far I have gotten that opportunity here at CHORI, being involved with stem-cell immuno-staining that can allow me to make hypotheses on one of the most controversial medical fields, which could lead to a new life-saving discovery. Exciting, right? What kind of 18 year old gets that type of an amazing opportunity? And while I don’t know if I ultimately want to be a doctor or a researcher, I could never know until I try it out.
59 Human Placenta Tissue is a Novel Niche for Hematopoietic Stem Cells
W. Green, V. Serikov & F. Kuypers Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: All blood cells are generated from hematopoietic stem cells (HSC). Transplant of HSC is an important tool used to cure blood disorders. When HSC in pathologic bone marrow need to be replaced, as in Sickle Cell Disease and Leukemia patients, donor cells are required that match the patient immunologicaly. Finding an appropriate bone marrow stem cell donor is extremely difficult, and cord blood (CB) has become a viable source of HSC. While CB has many advantages, the limited volume of blood collected from the cord and placenta after delivery equates to a limited number of HSC. While HSC from this resource have successfully been used in transplant for young children, the total number of HSC is often too low for an adult patient. Placenta may provide a source of HSC to boost the efficacy of CB. Murine placenta seems a niche for HSC, but such studies on human tissue are lacking. Objective: The goal of the project is to determine if HSC are abundant in human placenta by immuno-florescent staining, and if so, evaluate the total number of HSC in this resource. Methods: Immuno-fluorescent staining was performed on placental tissue that had previously been prepared on slides. Tissue was stained with primary antibodies that bind to a variety of HSC markers, such as CD-34. A secondary staining of these primary antibodies and a fluorescent labeling of nuclei generated fluorescent cells. As cells fluoresce at certain wavelengths depending on the fluorophore used, a composite view of cells positive for different markers could be made. Cells positive for HSC markers as compared to nuclei were counted to enumerate the total number of HSC in placental tissue. Results: It was found that HSC make up 0.2±0.03 % of all cells present in placenta, and the total estimated number of HSC (CD34+) cells in the whole placenta averaged 2.5x107 cells. Compared with the .1x107 CD34+ cells found on average in the CB from the same placenta, these data indicate that placental tissue is a valuable resource that can provide an order of magnitude more HSC than CB alone. In addition, cell markers indicated the presence of embryonic like stem cells in placenta. Conclusion: The placenta is a plentiful source of human HSC. These cells can be extracted from fresh or frozen placenta and augment CB as a source of stem cells for transplant. Further studies will be needed to establish the optimal conditions that will allow the storage and extraction of these HSC to offer this resource, in addition to CB, for transplant in adults, not just children.
60 Amanda Harris Funded by the NIH/NHLBI/Professional Student Short Term Program University of New Mexico Third Year Mentor: Alberta Kong, PhD
I am Amanda Harris. This fall I will continue my education at the University of New Mexico. I will be entering my fourth year of undergraduate studies with an intended major of Biology and a minor in Native American Studies. This is my second summer with the CHORI Summer Student Program. I am very fortunate to have been given the opportunity to return to the program, but also very appreciative to have been able to continue my understanding and learning of research in my home state this summer. With the two summers I have spent with CHORI, I have experienced many new things and gained valuable knowledge that I will use in the future to help me obtain my goals and dreams.
61 Comparison of RT3 Triaxial Accelerometer and 7-day Physical Activity Recall in Hispanic and American Indian Adolescents
A. Harris & A. S. Kong University of New Mexico, Albuquerque, NM 87131
Introduction: Obesity and type 2 diabetes are increasing chronic medical conditions facing adolescents. These conditions disproportionately affect minority populations, particularly Hispanics and American Indians. Physical activity is paramount in the prevention and intervention of these health problems. Objective: To compare the RT3 accelerometer and the 7-day Physical Activity Recall (7dPAR) in estimating energy expenditure and physical activity in adolescents of primarily Hispanic and American Indian descent for an obesity intervention. Methods: Eighteen participants (ages 16-18 years, 61% females, 44% Hispanic, 44% American Indian) were referred to the study from a job training center. Height and weight were obtained at the center’s health clinic along with programming the accelerometer. Participants were instructed to wear the accelerometer on the right hip for 7 days, except for during showers, swimming, and sleeping. On the seventh day, accelerometers were retrieved, data downloaded and the 7dPAR was administered to participants to recall the previous week of physical activity. Participants with at least 4 complete days of accelerometer recording and who completed the 7dPAR interviews were analyzed. Relationships of the two instruments with regards to average daily energy expenditure, minutes of moderate physical activity (defined as 4-6 METs) and minutes of vigorous physical activity (defined as ≥ 6 METs) were determined using Spearman rank correlation. Results: Mean BMI was 28.5 (±8.2 SD) for the 18 participants. The RT3 and 7dPAR had comparable mean daily energy expenditure (2,731.5 vs 2,971.9 kcal/day, p = 0.15). Spearman correlation was high and statistically significant for average daily energy expenditure between the RT3 and 7dPAR, r = 0.88 (p < 0.001). Mean minutes of moderate physical activity collected by the 7dPAR was 99.1 (±103.4 SD) minutes and by the RT3 was 44.7 (±21.3 SD) minutes; r = 0.56 (p = 0.02). While mean minutes of vigorous physical activity collected by the 7dPAR was 38.1 (±54.5 SD) minutes and by the RT3 were 56.0 (±44.9 SD); r = - 0.20 (p = 0.44). Conclusion: These data suggest that energy expenditure is highly correlated between the RT3 accelerometer and 7dPAR. However, minutes of moderate and vigorous physical activity were inconsistent between the RT3 and 7dPAR. Use of both instruments will be necessary to assess the effects of the obesity intervention on physical activity.
62 Sejal Hathi Volunteer
Notre Dame High School Sophomore
Mentor: Deborah Dean, MD, MPH
Ever since I was a little girl, I have been enthralled and stimulated by science: Since perhaps junior high school, I have voraciously devoured my science textbooks, science newspaper articles, and journals- from Science to Nature to Scientific American- to plumb the depths of my curiosity and enrich my knowledge about biology and the natural world. Reading about scientists’ creative work and increasingly sophisticated and penetrating experiments goaded me with the desire to perform research of my own and thereby contribute to the community in some viable way. Despite all the science fair experiments I did in school, my yearning to experience a deeper research experience was not quenched, until I found out about CHORI’s Student Research Program. Here, I am delighted to say, I have fulfilled my childhood’s dreaming and discovered the essence behind a career in basic science laboratory research. CHORI has accorded me the opportunity I desperately needed to understand the reasons behind the facts I learned in school, the “wherefore” and “how” behind the bland textbook theory- while simultaneously teaching me to apply those plethora of biology, chemistry, and physics lessons I learned in new and innovative ways. CHORI has deepened my love for science and convinced me of my now insuperable resolution to pursue a career in medicine by granting me a more lucid picture of my prospects and the multiple specialties that medicine entails.
63 Genotyping of the ompA Gene of Chlamydia Trachomatis Among Commercial Sex Workers in Ecuador
S. Hathi, A. Zen & D. Dean Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Chlamydia Trachomatis (CT) is an obligate intracellular bacterium and the human pathogen principally responsible for sexually transmitted diseases in the world. Because the majority of these infections are subclinical and non-acute in presentation, up to 50% of adolescent and young adult women with CT become reinfected or develop persistent infections, increasing their risk of contracting the sequelae like tubal factor infertility and ectopic pregnancy. We must improve our understanding of the epidemiologic, molecular, and genetic factors leading to CT infection and persistence to design appropriate interventional strategies. Adolescent and young adult women form a significant target population in which to analyze CT infection rates, transmission patterns, and types of recurrence strategies because of their documented insouciance toward using disease prevention techniques. Objective: By determining the prevalence of chlamydial strain types in the target population, we can define the CT genetic profile for each patient further and thereby determine associations with clinical findings, demographics, reinfection, and persistence to compile valuable data toward appropriate intervention. We hypothesize that of the 78 equivocal/inhibitory samples, 50% will be true positives, giving us a prevalence of 15.28%. Methods: The prevalence of CT among a population of chiefly female commercial sex workers will be determined using the commercial PCR Amplicor test on both cervical swabs and urine per the manufacturer’s instructions. Those samples that are Amplicor positive will be cultured and subjected to ompA genotyping and MLST to facilitate an understanding of CT transmission patterns. In preparation for DNA sequencing, the chlamydial DNA from all samples will undergo extraction through Roche Hi Pure, amplification through polymerase chain reaction (PCR) and gel electrophoresis, and finally, purification through gene cleaning. The DNA samples will be then sequenced for ompA, the major outer membrane protein of chlamydia, to determine the serovar, using forward (CT40UF) and reverse (VF3) ompA primers. Results will be analyzed for multiple sequence alignment, comparing the clinical C. trachomatis strain with the reference laboratory C. trachomatis strain for the particular serovar. Results: Endeavoring the most sensitive approach to resolving the intermediate and inhibitory samples takes time and is still ongoing. We are also in the process of genotyping the true positives and will genotype additional samples as they are determined positive, but must first essay different protocols to optimize our PCR amplification product. Conclusion: Strain typing and serotyping are essential for defining and differentiating the molecular epidemiology of the various chlamydial disease states like urethritis, cervicitis, pelvic inflammatory disease, and recurrent as opposed to persistent infections. By determining the prevalence of CT infections and their strain types and correlating these data with clinical disease phenotypes and co-infection STD’s, this study will contribute valuable data for eventually developing a rational vaccine to prevent chlamydial STD’s. However, further studies are needed to more precisely type CT and understand how these genetic profiles relate to reinfection, and persistence. 64 Daniel Hausrath Volunteer
Head-Royce High School Senior
Mentor: Trudy Forte, PhD Robert Ryan, PhD
My name is Daniel Hausrath. I am eighteen and in June of this year I graduated from the Head-Royce School in Oakland, California. In a month or so I will be starting my first semester at Yale. In high school I was involved in student government and volunteer work at Rebuilding Together Oakland. Outside of school I enjoy just kicking it with friends, listening to music and watching terrible movies.
I’ve always been interested in science and hope to eventually become a doctor. My interest in medicine goes back almost as long as I can remember. This longstanding love of science and medicine propelled me through my various high school science and math classes and last year landed me in the CHORI Summer Student Program where I worked in the de Jong lab under Dr. Lucia Carbone. I worked with Dr. Carbone on a comparative genomics project investigating major differences between the gibbon and human somatic chromosomes. I cannot say enough good things about the people I met last summer. I had a great experience working on that project, and it inspired me to return for both my senior project (a month long internship at the end of senior year required for graduation at Head-Royce) and this summer’s research program. This summer I decided to try working on a different project, so as to learn yet more procedures and techniques.
I am working in the Forte/Ryan lab under Dr. Forte and Mina Nikanjam on developing a targeted drug-delivery system to fight glioblastoma multiforme tumors. I want to thank Dr. Ryan and Dr. Forte for choosing me/helping counsel me/letting me work in their lab, Mina for teaching me everything I needed to know to work on my project, and probably more, and Son for helping me with pretty much everything.
65 Novel Molecular Targets for Treating Glioblastoma Multiforme Tumors
D. Hausrath, M. Nikanjam, K. Beckman, S. Ho & T. Forte Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Glioblastoma multiforme (GBM) is an extremely aggressive type of brain tumor (WHO Grade IV astrocytoma) with a mean survival time of one year following diagnosis. The tumor grows rapidly and is difficult to completely resolve with surgery due to tentacle-like extensions branching off from the main tumor mass. Surgical resection of the tumor is followed by post-operative chemotherapy and radiation. A new focus in brain tumor therapy has been the identification of molecular targets specific for tumor cells. Cell receptors that are particularly numerous on tumor cells but are relatively scarce on healthy brain cells will be good targets for therapeutics. Genes that have been suggested to be upregulated in other cancers include the low-density lipoprotein receptor (LDLR), transferrin receptor (TFR2), ephrin A2 receptor (EphR A2), epithelial growth factor receptor (EGFR), folate receptor (FOLR) and ErbB-2/Her2 receptor. Each of these receptors is known to mediate endocytosis, (i.e. cell uptake). Synthetic particles consisting of a specific receptor ligand and a core containing anti-cancer drugs can potentially be directed into tumor cells via the receptor. Objectives: The objective of this project was to develop a method for assessing levels of unique receptors on cancer cells using real-time polymerase chain reaction (RT-PCR). We are specifically interested in identifying receptors that are upregulated on GBM cells as compared to healthy brain tissue. Methods: Gene expression levels were assessed in SF-767, SF-763, and U-251 glioblastoma cell lines, which are known to possess 288,000, 950,000, and 128,000 LDL receptors per cell, respectively. Cells were grown in culture, plated onto six-well plates, and grown to confluence, and RNA was extracted for RT-PCR. The Ambion RiboPure RNA isolation kit was used to isolate RNA, and the ABI High Capacity cDNA reverse transcription kit was used to create a cDNA library from the RNA. ABI TaqMan probes (for EphR A2, Transferrin, LDLR, FOLR, EGFR, Her2 and Beta-Actin) were used to perform a real time PCR absolute quantification assay on an ABI 7900 machine. Ct number (point of most rapid change in amplification), as an indication of gene expression, was determined and compared between genes and cell lines. Results: LDLR gene expression levels were observed to increase as LDL receptor numbers increased, thereby validating the assay. Transferrin gene expression was higher than LDLR expression in all three cell lines. Ephrin A2 receptor expression levels were higher than both LDLR and Transferrin in all three cell lines. Conclusions: An assay to assess gene expression was developed and validated. As the medical community endorses more personalized medicine, it will be extremely useful to be able to test a sample of tumor tissue from a patient for a small number of different receptor-targets, and customize their treatment according to what will be most effective for them.
66 Dominique Johnson Funded by the NIH/NHLBI/Professional Student Short Term Program California State University Hayward Third Year Mentor: Gordon Watson, PhD
I was born in San Francisco, but spent most of my childhood in a small town by the name of Fairfield. I spent my freshman year of college at Dominican University of California, but soon realized that my happiness was to lead me elsewhere. As a sophomore in 2005, I transferred to California State University, Hayward where I am majoring in Biology with a minor in Biomedical Laboratories. I have been fortunate to be apart of the CHORI Summer Program for the two summers now and hope to return in the future. My interest in biomedicine has been cultivated by my involvement at CHORI. Being challenged intellectually in a welcoming environment has leaded me to return this summer. Furthermore, I was motivated by my mentor Gordon Watson, Ph.D., who aided me in guiding me throughout my first year and Vas Narayanaswami, Ph.D. I remember distinctly my first day in the lab at CHORI. I was fascinated by the fact that instead of using bulbs with Pasteur pipettes and burners which were considered old fashioned, I was able to work with modern lab equipment. This clearly set apart my hands on experience in school laboratories versus that if CHORI’s and gave me the realization that this would be an opportunity in which would be a stepping stone into my future.
Being apart of the summer program in 2006 I was able to also attend the 21st National Conference on Undergraduate Research a second time with my abstract conducted under the guidance of my mentor on “The Role of Caloric Restriction in the Prolongation of Life and Mimetic Use of Metformin”. My involvement at CHORI has lead me to many other avenues and friendships with other interns. I am grateful for this experience and blessed to be surrounded by such a wonderful mentor, who has taught me to never be afraid to ask any question and for providing a nurturing environment. More so, I would like to thank my loving parents, grandparents, brother, and my boyfriend for always encouraging me.
67 Biomarkers for a Genetic Deficiency in Cholesterol Synthesis (Smith-Lemli-Opitz Syndrome)
D. A. Johnson & G. L. Watson Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Smith et al. in 1964 reported three unrelated boys with multiple malformations including distinct facial features, mental retardation, microcephaly, and hypospadias. This syndrome was called RSH syndrome from the names of the first three patients, but is often referred to as SLOS after the names of the authors. At diagnosis, children with SLOS syndrome have cholesterol levels less than the normal and have high levels of 7- dehydrocholestrol (7-DHC). It is apparent that the deficiency of cholesterol is caused by abnormally low levels of 7-dehyrocholestrol reductase (7-DHCR), which is responsible for the final step in cholesterol production in many cells. It is important in embryonic development by interacting with proteins that control early development of the brain, limbs, and genital tract. Objective: The objectives of my research were to breed and test mutant mice to get experimental mice with SLOS genotype and establish biomarkers for SLOS phenotype using microarrays for mRNA expression pattern. Methods: (I) Breeding; (a) First model: ΔDHCR7/ + bred to wild type (C57BL/6) mice to produce heterozygote (- / +) because homozygous (- / -) mutants do not survive. Since only one-half of the progeny will carry the mutation, genotyping is necessary. (b) Second model: T 93M/ T 93M are viable and can be bred as homozygotes. Therefore all the progeny of this model will be ( - / - ). (c) Compound heterozygotes Δ DHCR7/ + bred to T 93M/ T 93M. Only fifty percent of the progeny will be ΔDHCR7/ T 93M (SLOS Phenotype) and the other fifty percent will be T 93M/ + (normal phenotype). (II) Genotyping; (a) Extract DNA from mice and follow the appropriate steps for a PCR protocol. (b) PCR (c) Gel Electrophoresis. (III) Microarrays; (a) Extract RNA from compound heterozygotes and controls (b) Check RNA in agarose gel (RNA must be intact, not degraded (c) Tag mRNA with fluorescent dye (d) Using microarrays from Ilumina, hybridize RNA to extensive set of oligo-DNAs (e) Record results with microarray reader and transfer data set by computer (f) Analyze data set using algorithms provided by Ilumina. Results: We were successful in expanding the SLOS mouse breeding colony and generated compound heterozygotes by cross breeding. The expected genotypes were identified by PCR. Preparation and analysis of mRNA is ongoing. Conclusion: SLOS mice are difficult to raise; their impaired health and delayed development makes them fragile and many litters die shortly after birth. Nevertheless, we produced enough mutant mice to provide RNA samples that can be used to determine altered patterns of gene expression.
68 Nadia Kaaid Funded by the NIH/NHLBI/Professional Student Short Term Program San Francisco State University Senior Mentor: Donald Reason, PhD
My name is Nadia Kaaid and I am working in Dr. Don Reason’s lab and studying the interactions of human antibodies against Bacillus anthracis toxins. My reason for participating in the CHORI program for the second year is ultimately because I learned so much and had an interesting time doing it last summer. I feel that CHORI is in some way an oasis for me because within the program I don’t have to feel worried about being graded or a failed experiment, not that I get too many failed experiments, but it happens. Everyone that I have met here are super nice and helpful in answering my many questions and the seminar speakers and fantastic. Being a part of the program also gave me the opportunity to take a tour of the clinical lab and clinical research lab at the hospital building. I will be starting the Clinical Laboratory Scientist Internship Program at SFSU in the fall so I was glad I got a chance to talk to clinical scientist and see how they run their daily tasks.
I would like to thank the whole Reason lab for welcoming me into the lab and making me feel part of the team.
69 Binding and Neutralization Ability of Human Antibodies Directed Against Bacillus anthracis Toxins
N. Kaaid, A. Ullal, J. Liberato & D. Reason Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introductions: Protective Antigen (PA) is one of the three toxic factors made by Bacillus anthracis bacterium, which causes anthrax. The current Anthrax vaccine, Biothrax, includes
PA as its main component. PA induces a polyclonal antibody response from the immune system of a vaccinated host. The other two factors, Lethal Factor (LF) and Edema Factor
(EF) are dependent upon PA for entry into the cell. During the process of intoxication, PA83 binds to a cell surface receptor, and the cell associated proteolytic enzyme furin cleaves
to into two segments, PA63 and PA20. PA63 combines with other PA63 molecules to form a heptamer on the cell surface. This serves as a binding site for the other LF and EF. The pore is transported into the cell, and the delivery of LF and/or EF cause cell death. Since the role of PA is critical to the entrance of the other two toxins into the cell, and since PA is also crucial to the functioning of the vaccine, understanding the interaction of PA binding to specific antibodies will provide information on the mechanism by which neutralization occurs. Objective: To study antibody clones isolated from donors vaccinated against the disease anthrax to determining the location of binding, to test for neutralization abilities, and understand their mechanisms of action. Methods: Antibody ability to bind to PA was determined by conducting PA-specific Enzyme-linked Immunosorbent Assays (ELISA). Dot blots using proteolytic and engineered fragments of PA was used as the method to identify which fragment(s) a specific antibody bound to. Neutralization was determined by the use of the RAW 264.7 cell line in a PA-mediated killing assay. These cells carry PA receptors and a lethal dose of PA/LF was added with dilutions of antibodies to test the percentage of neutralization. Flow cytometry was used to look for antibody-mediated blocking of PA binding. Results: Previous studies in the lab have shown that the majority of antibodies elicited by vaccination bind to the portion of PA that is cleaved off and has no major role in toxicity. In this study neutralizing epitopes were found throughout the PA, however, the most frequent and most efficient determinants were located in the PA63 portion of the molecule. Preliminary results indicate that PA20-specific clones may function by
blocking proteolytic cleavage of PA83. No clones have yet been shown to block PA binding to the cell surface receptor. Conclusion: The PA-specific antibody response elicited by vaccination with BioThrax is biased towards clones that react with non-neutralizing or weakly neutralizing determinants on PA. This factor may partially explain the relatively poor performance of this vaccine.
70 Ragini Kathail Volunteer
Pomona College Second Year
Mentor: Janelle Noble, PhD
I am a rising sophomore and a potential molecular biology major at Pomona College in Claremont, California. I chose to volunteer at CHORI this summer to better understand the life and job of a research scientist and to get a sense of completing a research project from beginning to end, which interests me more than does medical school.
Last semester, I worked at Pomona’s Asian-American Resource Center, and next year, I am planning to be part of the Asian-American Mentoring Program, which provides guidance to self-identified Asian and Pacific Islander first-year students. In the same vein, I am especially interested in eventually conducting research in a field which has relevance to the Asian-American population.
I would like to thank Dr. Janelle Noble for giving me the opportunity to work in her lab, as well as Jameel Johnson and Julie Lane for answering all of my incessant questions. And, of course, I want to thank Olivia Linney and Andrea Banuelos, the two other summer students with whom I worked, for their cheerful optimism and their dedication.
71 Control Data for Class I HLA-B in an African-American Population
R. Kathail, J. Johnson, A. Banuelos, O. Linney & J. Noble Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction The human leukocyte antigen (HLA), located on the short arm of chromosome 6, is the human version of the major histocompatibility complex (MHC), which regulates the immune response to foreign antigens and, by extension, affects the occurrence of various autoimmune and infectious diseases. There are two main classes of HLA. Within class I, there are three loci, termed A, B, and C, and within class II, there are three additional loci, termed DR, DP, and DQ. The HLA region as a whole has a high degree of genetic polymorphism which, when combined with one of the highest levels of linkage disequilibrium (the non-random association of alleles at more than one locus) within the human genome, makes it difficult to determine the contribution of a given allele to disease risk. Previous research has shown that HLA genes account for approximately 50% of the genetic susceptibility to Type 1 Diabetes (T1D). Objective Since most of the previous HLA research has been on Caucasian, Asian, or Hispanic samples, the overall objective of this project is to create control data for all HLA loci from approximately 1000 African- American blood samples. For the summer, the objective is to produce background allele frequencies for 92 samples at the class I HLA-B locus. Methods In 1998, the Noble Lab received 4000 blood spot cards collected through the California Newborn Screening Program from four ethnicities: Caucasian, Asian, African-American, and Hispanic. At the Noble Lab, the African-American cards were previously genotyped at their DR and DQ loci. For this summer project, these cards were first sorted into their respective ethnic groups, and the African-American cards were then organized into 96-sample plates, including four negative controls per plate. Each blood spot card was hole-punched several times to provide dried blood from which to extract DNA. A protocol was developed to recover and purify this DNA, and a polymerase chain reaction (PCR) was performed to amplify it into a usable quantity. This amplified DNA was run on a hybridization-based “linear array” assay, and the results were interpreted to provide genotypes for each sample at the HLA-B locus. Results The 92 samples from plate 3 were extracted and genotyped. The allele frequencies will be compared to HLA-B frequencies available for the Caucasian population. Conclusion Further work will yield genotypes for all the samples at each HLA locus. These allele frequencies can be compared against the genotypes of African- American patients with T1D, as well as other low-frequency autoimmune disorders that have a genetic association to the HLA region.
72 Andrew Lee Volunteer
Duke University Third Year
Mentor: Julie Saba, MD, PhD
My interest in science began at the age of two, starting off with feeding the wildlife in Golden Gate Park and roaming the Lawrence Hall of Science. I will be entering my junior year at Duke University in Durham, North Carolina. I expect to graduate in 2009 with a major in biology and a minor in economics. This past year, I studied molecular and genetic biology and physiology, and I conducted research in an immunology lab at the Durham VA Hospital. These experiences have equipped me with the fundamental tools to make the best of my experience at the Children’s Hospital Oakland Research Institute.
I have hoped to pursue a career in medicine for many years now, and my time at CHORI has helped me realize just how inseparable science is from medicine. My time working with Dr. Saba and Dr. Fyrst has been invaluable, providing insight into the unpredictable and yet methodical nature of science. The combination of my recent experiences of theoretical learning in the classroom, hands-on scientific work at CHORI, and direct encounters with medicine have given me a deeper understanding and appreciation for both science and medicine, and I’m sure my experiences in the future, both at CHORI and elsewhere, will only further increase my passion for these fields.
73 Sphingolipids and the AKT Pathway
A. Lee, H. Fyrst, & J. Saba Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Sphingolipids are a class of lipids with incredibly diverse and crucial functions, including proliferation, apoptosis, and regulation of cell survival. Two sphingolipids, ceramide and a novel sphingadiene, promote apoptosis, which is programmed cell death. In contrast, sphingosine-1-phosphate (S1P) promotes cell proliferation and inhibits apoptosis. S1P levels in cancerous cells are often elevated, indicating that sphingolipids play a role in cancer. One aim in cancer treatments is to make cancerous cells more susceptible to apoptosis, while preventing apoptosis in healthy cells during treatment. Therefore, sphingolipids are of great interest in developing cancer therapies. Sphingolipids may affect cells through the Akt pathway, a growth pathway responsible for protein synthesis, cell growth and proliferation, and inhibition of apoptosis. Objective: The objective of my research was to examine the effects of sphingolipids on the Akt pathway in HT29 colon cancer cells and to determine if sphingolipids deactivate the Akt pathway by dephosphorylating Akt. Methods: HT29 cells were incubated overnight with 20 μM of sphingosine, dihydrosphingosine, sphingadiene, ceramide, and sphingadiene- ceramide. The cells were incubated with insulin for 1 hour, after which Western blot analysis was used to probe for Akt and Akt-P. For the in vitro translation assay, cytosol was isolated from lipid-treated HT29 cells. ATP, GTP, creatine kinase, and mRNA for the luciferase gene were added, and luminescence was measured. In the Caspase 3 assay, HT29 cells were treated with 10 μM LY294002 for 24 hours, then treated with 37.5 μM lipid and 10 μM rapamycin for 18 hours. The cells were treated with caspase reagent, and absorption was measured. Results: Western blot analysis revealed that the levels of Akt- P are significantly lower in HT29 cells treated with the sphingadiene. Although baseline levels of Akt-P were lower in lipid-treated cells, a decreased response to insulin was not observed. The Caspase 3 assay revealed that the sphingadiene increases levels of apoptosis in HT29 cells, and Akt pathway inhibitors LY294002 and Rapamycin compound this effect. The in vitro translation assay revealed decreased levels of protein synthesis in lipid treated cells, particularly with the sphingadiene. Conclusion: We can conclude that sphingolipids affect HT29 cells through the Akt pathway, preventing phosphorylation of Akt, thereby reducing protein synthesis and growth and inducing apoptosis. Furthermore, the sphingadiene is particularly potent in blocking the Akt pathway. Further research needs to be done to determine the mechanism by which sphingolipids affect the Akt pathway.
74 Jacqueline Leon Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Third Year
Mentor: Susan Conrad, MD Suruchi Bhatia, MD
Like most individuals I envision my contribution to the world by helping people. I grew up with the idea that a job in the medical field would do just that.
During my academic years I became interested in the sciences. During high school and college taking science classes has not been an easy adventure. With the demands and expectations of both institutions, especially U.C. Berkeley, the culture of science is not a joke. As the oldest child and from a family that does not have a science background studying the sciences did I ever feel so alone. CAL not only tests your knowledge of the material but your commitment in wanting to be part of the science community. Fortunately with the Biology Scholars Program I assimilated into the culture of science and not let a “simple” institution like CAL detour me away from my dream.
Along the way I developed passions for organic chemistry and biology. In these classes I was exposed to research. The field of research seemed like an exciting subfield to get myself involved into. For the second time in my life my dream started to become endangered. How can I become a scientist and be a physician at the same time? It is either one or the other.
Before participating in the summer research program at CHORI my ideas of research was limited to working at a bench, working with pipettes, and running PCR gels. However, CHORI has given me the opportunity to see first hand that that is not the case. I was fortunate to conduct clinical research with Susan Conrad, MD. Her mentorship along with the weekly seminars was so valuable because they taught me three important things. First, one can be a clinician and a scientist at the same time. Secondly, clinical research is just as valuable as bench research. Lastly that scientists and physicians are as valuable to society in terms of helping people by their research they conduct or the prescription they prescribed. CHORI has allowed me to continue on my dreams on becoming a physician and a scientist knowing that doing both are possible.
75 Factors at Diagnosis of Type 1 Diabetes Predict Outcome at 2 Years
J. Leon & S. Conrad Children’s Hospital & Research Center at Oakland, Oakland, CA 94609
Introduction Type 1 diabetes is a condition with potential significant long-term complications. Numerous studies show that the risk of complications tremendously decreases with better blood glucose control. However, there is little data available to help identify patients early in their disease course who may be at higher risk for problems. Objective To determine if factors at diagnosis of type 1 diabetes predict outcome at 2 years. Specifically, the objective was to determine if the following factors were associated with HbA1c at 2 years: initial HbA1c, pH, blood glucose, length of hospitalization, insurance status, age, and race. Methods The study was a retrospective chart review. It was approved by the CHRCO Institutional Review Board. The population included subjects from the CHRCO Diabetes Center that were diagnosed with type 1 diabetes between June 2004 and May 2005. Subjects were included if they were followed regularly at the Diabetes Center for the first 2 years after diagnosis. Data collected from patient’s medical charts included: date of diagnosis, age at diagnosis, initial blood glucose, pH, length of hospitalization, insurance status, race, HbA1c at 2 years, and number of DKA episodes in first 2 years. Data was entered into an Excel spreadsheet. Statistical analysis was done using Pearson’s Product-Moment Correlation and Student t-test. Results There were 68 subjects included in the study. The mean age of subjects at diagnosis was 8.5 +/- 3.8 years. They were hospitalized for a mean duration of 3.9 +/- 1.9 days. Mean HbA1c at diagnosis was 10.8 +/- 1.8% and at 2 years was 8.7 +/- 1.5%. Both initial HbA1c and length of hospitalization were significantly associated with HbA1c at 2 years (p<0.05). There was a significant difference in patients who had private insurance compared with patients with public or no insurance in the following: HbA1c at diagnosis (10.4 vs. 11.4%, p<0.05) and length of hospitalization (3.4 vs. 4.6 days, p<0.05). There was also a significant difference in Caucasian subjects compared to non-Caucasian subjects in the following: HbA1c at diagnosis (10.1 vs. 11.6%, p<0.05), length of hospitalization (3.3 vs. 4.6 days, p<0.05), and HbA1c at 2 years (8.3 vs. 9.4, p<0.05). Conclusions There are factors identifiable at diagnosis associated with 2-year outcome in our subjects with type 1 diabetes. There are also differences in presentation and outcome that are associated with insurance status and race. Early identification of those at risk for worse outcome could result in prompt intervention and hopefully improvement in prognosis.
76 Aaron Levine Volunteer
Piedmont High School Junior
Mentor: Lucia Carbone, PhD Pieter de Jong, PhD
I have always been intrigued by, and interested in, science. To me, one of its most appealing features is its ability to offer explanation for phenomena both common and obscure in the surrounding world. Textbooks and science classes, however, can satiate only a portion of this interest; the window into the natural world in this scenario is limited to the content of the book and the insight of the teacher. Much that is taught through these sources is done so with daunting certainty, not to be debated or discovered, but merely to be learned and taken for granted. I therefore feel privileged to participate in unique research, to contribute to the understanding of areas of science that remain elusive and unknown. I would like to thank Drs. Lubin, Narayanaswami, de Jong and especially Dr. Carbone for allowing me the opportunity to work with them at CHORI. It has been a great summer.
77 Chromosomal Rearrangements in Gibbons
A. Levine, L. Carbone & P. de Jong Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: In most cases, the Eutherian (Placental Mammalian) karyotype has been very stable during evolution, sustaining few changes over the past hundred million years. Humans typify this stability, having experienced roughly one inter-chromosomal repositioning every 10 million years. Among all mammals, there seems to be a striking similarity between the arrangement and content of the chromosomes. There are, however, exceptions, one of the most enigmatic of which is the family Hylobatidae (Gibbons or Small Apes). Although Small Apes, Humans, and Great Apes are all members of the same superfamily (Hominidae), diverging from a common ancestor only 15 million years ago, the Gibbon karyotype is fraught with chromosomal rearrangements. The Gibbon karyotype has undergone over 100 chromosomal rearrangements in the past 15 million years, a time span in which only 1.5 such rearrangements would have been expected. In fact, Karyotypes in the Hylobatidae family itself also vary widely between species and serve to split the family into four genera: Hylobates (2n = 44), Bunopithecus (2n=38), Symphalangus (2n = 50), and Nomascus (2n = 52) (Muller et al). Objective: The objective of my project is to determine which chromosomal breakpoints existed in the ancestor of the Gibbons, and, to some extent, elucidate the mechanisms through which the Gibbons attained their puzzling karyotypes. Furthermore, we hope to determine whether the vast number of chromosomal rearrangements found amongst the Small Apes is due to an anomalous occurrence in the Hylobatidae ancestor or if the rearrangements happened independently of one another during several, distinct events. Methods: The primary method we employed to identify which breakpoints are species-specific and which are shared with other species, is fluorescence in situ hybridization (FISH). Bacterial Artificial Chromosomes (BACs), predetermined to contain breakpoints of interest in the white- cheeked gibbon, are labeled in either Cy3 or FITC and hybridized onto slides containing chromosomal metaphases spreads from the other Gibbon species. Using this data, we can determine which breakpoints are common among the Gibbon genera, and, consequently, whether each breakpoint is ancestral or genus-specific. Results: So far, we have successfully used FISH on 23 white-cheeked gibbon BACs to determine if the breakpoints they span are syntenous with the other species. Of these, we have determined that five rearrangements are shared among the four species, indicating that they are ancestral, and that seven are present only in the white-cheeked gibbon, indicating rearrangements specific to that species. The other 11 BACs provided data concerning the order in which the four gibbon genera evolved. Conclusion: The data obtained through our research can be utilized to fully or partially construct a Gibbon phylogenetic tree. Additionally, the data can show which rearrangements were more prevalent at the various stages of Gibbon evolution. Lastly, through additional PCR and sequence analysis, the mechanisms and factors that spawn chromosomal rearrangements can be better understood.
78 Olivia Linney Volunteer
Head-Royce High School Senior
Mentor: Janelle Noble, PhD
My name is Olivia Linney and this fall I will be a freshman at Stanford University. Outside of the lab, I enjoy time with friends, dance, music, and pretty much all activities outdoors. Academically, for as long as I can remember, I have always been drawn to the sciences. It is practically instinct for me to question how and why things work, and because there are so many important but unanswered questions in biology, I have always been particularly captivated by this area of science.
In May, I spent a month with the endocrine department at CHO before beginning the summer program at CHORI and feel very fortunate to have gotten to see both the research and clinical aspects of diabetes care. My younger brother was diagnosed with type I diabetes at age 11, so being able to participate in research being done for this area of medicine has been particularly gratifying for me. I have learned so much from everyone at Noble Lab whom I have worked with this summer. In particular, I wish to thank my PI, Janelle Noble, for such a wonderful experience, Jameel Johnson for training me and putting up with me even when I labeled his hyb strips upside down, Julie and Shaily for answering my questions, and last but not least my coworkers Andrea and Ragini for always being so efficient and enjoyable to work with all summer long.
79 Contribution of Human Leukocyte Antigen C locus to HLA-associated Disease in African American Population
O. Linney, J. Johnson, R. Kathail, A. Banuelos & J. Noble Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction HLA is the human form of major histocompatiblity complex (MHC), and its proteins play a critical role in immune recognition. The two gene clusters for HLA (class I and class II) are located on the short arm of chromosome six. While class II (DP, DR, DQ) is almost exclusively found on immune cells, class I (A, B, C) exists on the surface of all somatic cells, in varying concentrations depending on cell type. HLA class I proteins display antigen for CD8 cytotoxic T cell recognition. Because diversity is so crucial to defending the body against a wide range of pathogens, HLA loci are among the most polymorphic regions found in the human genome. Since autoimmune disorders like type I diabetes (T1D) are caused by an immune system that wrongly recognizes its own tissues as foreign, genotyping HLA loci has proved to be incredibly useful in helping determine genetic risk for autoimmune disorders. For example, it has been found that the HLA proteins account for approximately 50% of the genetic risk for T1D. Objective To create a control set of allele frequencies of the C locus for the African American population to be used in studies of HLA-associated disease. Methods The entire study will use 1000 self-identified African-American newborn bloodspot cards. DNA was extracted from 92 of these samples. The HLA-C region was amplified using PCR. Sequence-specific- oligonucleotide-probe “linear array” assays were run, and genotypes were assigned to the results to determine the allele frequencies of the group. Conclusion The control set of data generated for HLA-C will be useful in studying any type of low-frequency HLA- associated disease. Although, statistically speaking, a few of the babies who provided the blood samples may develop T1D, because the sample size is so large (1000 once the study is completed), the control data should not be significantly affected. There have been many studies examining the HLA region in Caucasians, but until this point, few studies exist for the HLA-loci in African-Americans. However, by studying different groups, it is possible to gain a better picture of what is going on for all ethnicities. The HLA region exhibits some of the highest rates of linkage disequilibrium found in the human genome (the alleles from two loci are found together far more frequently than can be attributed to chance). This makes it very difficult to discern which of the alleles is protective for or predisposing against a particular disease when they are almost always found in tandem. However, by studying other ethnicities that have different allele combinations at these two loci, the effects of one allele can be distinugished from the other by cross-ethnic comparison, knowledge which is beneficial to everyone.
80 Erin Mamuyac Funded by the NIH/NHLBI
Piedmont High School Junior
Mentor: Frans Kuypers, PhD
With the first wield of a scalpel in my th7 grade science class, I unknowingly launched myself into what was to become an intense passion: medicine. I have always been intrigued by the study of human anatomy as well as biology. Ever since the summer of sophomore year I have spent my summers exploring different aspects of the medical profession. I know that all I’m absorbing are tiny bits and pieces of the medical community, but each glimpse only seems to push me to discover more. After interning in the angiography department at John Muir Hospital, scrubbing in on a brain surgery, and participating in the FACES Summer Medical Academy at Children’s Hospital in Oakland, I decided to look into research. Within the first few days of working in the lab at CHORI, I became fascinated with the work that they were doing and the potential impact that it could have on the millions of people suffering from certain diseases. It is an amazing feeling knowing that however small your contribution to the research community, you are a part of the global movement to save lives. After this summer, I know that I will come away feeling more enthusiastic about a career in medicine than when I started research. Somehow, all of my summer internships have given me this same feeling.
81 Phosphatidylserine Exposure in Normal Erythrocytes by the Addition of Zinc and Zinc Chelators
E. Mamuyac, K. de Jong & F. Kuypers Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: The study of red blood cell membrane function and anatomy has become crucial to understanding various diseases such as sickle cell anemia and thalassemia. The cell membranes of erythrocytes are asymmetrical, as certain phospholipids have an affinity for either the inner or outer monolayer. Phosphatidylserine (PS) is a phospholipid located in the inner monolayer, because PS exposure in the outer monolayer is a signal to that cell destruction is in process. Asymmetry is maintained by the phospholipid translocases “flippase” and “scramblase.” When activated, the translocase “scramblase” disrupts the asymmetry by inducing a redistribution of phospholipids across the bilayer. Scramblase is activated by protein kinase C (PKC) and an intracellular increase in Ca++. The presence of Zn bound to PKC plays a critical role in the protein’s activity. We hypothesized that an increase in Zn availability would stimulate scrambling. The addition to the cell of a Zn chelator is predicted to disable this bond, in turn crippling PKC. Objective: The aim was to investigate the effects of varying Zn levels and certain Zn chelators on PS exposure induced by the addition of Ca and a Ca-ionophore in normal erythrocytes. Methods:
Washed red blood cells (RBCs) were treated with different CaCl2 concentrations and Ca-ionophore. Phenanthroline and TPEN were the designated Zn chelators. The control
group always consisted of erythrocytes in 2mM CaCl2 without ZnCl2 or Zn chelator, and
sample groups had increasing concentrations of ZnCl2 or Zn chelator. Samples were taken every 10 minutes for 30 minutes. To identify PS exposure, RBCs were labeled with Annexin (Alexa488-AV or Alexa633-AV) and then analyzed by flow cytometry. Results: The experimental findings largely contradicted anticipated outcomes. The addition of
ZnCl2 did not appear help scrambling and actually caused inhibition, which was complete at 100 µM ZnCl2. Though it was anticipated that TPEN would cause inhibition, it seemed to stimulate scrambling. Higher concentrations of TPEN, above 250 µM, finally caused inhibition. Phenanthroline did present scrambling, although possibly not by chelation of Zn. Scrambling in the presence of a higher Ca++ concentration was also inhibited by phenanthroline, indicating that this is not just a Ca-chelator. Conclusion: Zn is not a scrambling stimulator and instead interferes with Ca-induced scrambling. Chelation of Zn reduces scrambling but only at concentrations much higher than necessary for Zn chelation, indicating that other mechanisms play a role. Current investigations are examining the effects of Zn and Zn chelators on Ca-influx.
82 Yessica Martinez Funded by the NIH/NHLBI/Professional Student Short Term Program San Francisco State University Fourth Year Mentor: Ethan Geier Giorgio Cavigiolio, PhD Michael Oda, PhD
As a young girl I always knew I wanted to be a scientist. Everything in my life was science. My bedtime stories were straight from Discover magazine. They filled my head with planets and dinosaurs. More importantly, my family’s health influenced my decision to work in research. Collectively my family suffered from allergies, chronic fatigue, high blood pressure, diabetes, breast cancer, colon cancer, arthritis, dementia, and asthma. I always felt that it was my responsibility to help them and everyone else. Growing up., my parents didn’t want me to waste my creativity on cartoons and video games, so I spent most of my time reading and watching public television. Children dieing from starvation and diseases always caught my attention. In 1993, I took a life changing trip to El Salvador. I saw how little people had and how happy they were, even though they suffered from diseases like polio. Since then, I’ve dedicated my life to serving others through volunteering in organizations such as Friends of Animals, F.I.L.A.D., a non-profit health insurance agency, and Arts Benicia. I believe that one person can make a difference in the live of others and our planet. I’ve always known that biology was the way to accomplish my goals. At first, I wanted to be a botanist and go live in the jungles researching plants for medicine, but then I learned about genetics. I feel that genetic research is the key in developing cures for diseases like AIDS and protecting endangered species. I just graduated from San Francisco State University with a bachelors of science in Cell and Molecular Biology. This is my second summer with Children’s Hospital of Oakland Research Institute and I am very happy with the opportunity I’ve been given.
83 Determination of ApoA-I’s Helical Turn Orientation by FRET Analysis
Y. Martinez, E. Geier, G. Cavigiolio & M. Oda Children’s Hospital Oakland Research Institute, Oakland, CA 94609, USA
Introduction Apolipoprotein A-I (ApoA-I) is the main protein responsible for the reverse cholesterol transport, which delivers cholesterol from peripheral tissues back to the liver. ApoA-I’s properties enable high density lipoproteins (HDL) to exert its antiatherogenic character and participate in lipid metabolism. Indeed, plasma concentrations of apoA-I are the best indicators of cardiovascular disease susceptibility. However, lipid free apoA-I structure in solution has yet to be determined. Objective The known lipid free structure of apoA-I consists of an N-terminal four-helix bundle followed by a more flexible C-terminal domain. We will test if fluorescence resonance energy transfer (FRET) is sensitive enough to discriminate the changes in distance between a fixed position on one α-helix and amino acids at different angular positions around an aligned α-helical turn. In case angular orientation is a major contributor to differences in distance measurements, FRET experiments will be performed with residues on the same α-helical face. Methods From the X-ray structure of apoA-I, we selected two positions aligned on two parallel-arrayed α-helices: 108 and 174. To determine the effect of differences in distances due to the angular orientation around one helical turn, we placed donor-acceptor FRET couples at positions 108-174, 108-173, 108-172, and 108-171. The wild-type tryptophan at position 108 was the donor fluorophore. Positions 174-171 were mutated to cysteine and labeled with 5-((((2-iodoacetyl)amino)ethyl)amino)naphthalene-1-sulfonic acid (I-AEDANS) as the acceptor fluorophore. FRET analysis of the four double mutants W108/R174AEDANS, W108/Q173AEDANS, W108/R172AEDANS, and W108/L171AEDANS was performed on a Horiba-Jobin-Yvon Fluoromax-4. Results Fluorescence of AEDANS was affected by the polar environment in a fashion consistent with the hydrophobicity pattern predicted by Edmunson helical wheel analysis and the crystal structure orientation. Blue shift in AEDANS peaks of residues 171 and 174 confirmed their orientation towards the interior of the helical bundle. Red shift for residues 172 and 173 was consistent with solvent exposure on the hydrophilic side of the α-helix. The FRET dependent reduction in fluorescence intensity of tryptophan 108 (donor) was similar for all the four mutants surveyed. Conclusion We conclude that the FRET technique may not be sensitive enough to resolve differences in distances in the range of 4-15 Å, such as the ones depending on the angular orientation of residues around one α-helical turn. However, AEDANS is a reliable probe for reporting the hydrophobicity environment of a specific residue.
84 Miguel Martinez-Romo Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Fourth Year
Mentor: Tariq Ahmad, MD Ginny Gildengorin, MD
My interest in the biological and medical sciences stems mainly from my personal background. I was born in Aguascalientes, Mexico and at the young age of 6, my family moved to the Bay Area. Living in parts of the East Bay such as Richmond and Pittsburg opened up my eyes that there are great disparities in health in the Latino and African- American community, as well as other minority groups. I noticed that some diseases seemed to be unfairly more common in these communities, such as diabetes, which is a disease my own mother is inflicted with and which claimed the life of my paternal grandmother. Upon my arrival at the University of California, Berkeley, I realized that one of the causes for these disparities was that minority groups are hugely underrepresented in the biological and medical sciences; it didn’t take me long to realize in my freshman year that I was one of few Latino students in my science classes. It is because of these reasons that I am passionate about pursuing the biological and medical sciences, to help alleviate the under representation in biology and medicine that I have witnessed, as well as to help bridge the gap in healthcare that these under-representations have helped to create. The CHORI Summer Research Program is a great opportunity for me to enrich myself in expanding my knowledge of research, in particular clinical research. Because of the program, I am beginning to understand the importance of research in the medical sciences, and how that research can be translated to help patients, as well as to prevent diseases in the first place. This upcoming fall, I will be entering my fifth and final year at the U.C. Berkeley, majoring in Molecular Environmental Biology. After I graduate, I plan to take a couple of years off from academia and then apply to medical school.
85 Glycemic Control Among Diabetes Patients Living in the Bay Area
M. A. Martínez-Romo, G. Gildengorin & T. Ahmad Children’s Hospital & Research Center at Oakland, Oakland, CA 94609
Background: Poor glycemic control and body mass index (BMI) are related to increased risk of diabetes complications. Objective: Investigate whether patients’ socio-economic status, geographic location, and primary language impact HbA1c and BMI percentiles (%iles) among Type I (T1DM) and Type II (T2DM) diabetes patients. Methods: The Children’s Hospital & Research Center at Oakland (CHO) has a diabetes population of over 1100 children. Each child’s most recent HbA1c, BMI%’ile, ethnicity, primary language, insurance, zip code, and city of residence were obtained. Associations of the above descriptors with HbA1c and BMI%’ile were performed separately for T1DM and T2DM patients. Insurance (public vs. private) and income levels (<$40k, $40k-$69,999, $70k-$99,999, or > $100k) were used as measures of socio-economic status. Geographic factors included distance from CHO (< 20 miles vs. > 20 miles) and location (east of Caldecott Tunnel vs. west). Cultural impact was measured by primary language (English vs. Non-English) and ethnicity (African-Americans (A.As), Asians, Caucasians, or Latinos). Analysis of variance models with Scheffe’s adjustment for multiple comparisons were used to compare HbA1c and BMI%iles between each group. Chi-square tests were used to compare differences in the groups of the proportion of HBA1c above the recommended ADA levels. Results: In T1DM patients, the HbA1c levels were significantly higher for those on public insurance as opposed to private insurance (9.1% vs. 8.3%, p<0.0001). No difference was found between the T2DM insurance groups. The lower income groups, <$40k (8.7%) and $40k-$69,999 (8.9%), had significantly higher HbA1c values than the $70k-$99,999 (8.3%) and > $100k (7.8%) groups, for T1DM patients. The HbA1c was significantly higher for the lower income group (<$40k) then higher income levels (p = 0.026) for T2DM. Distance from CHO did not effect HbA1c or BMI%ile for both T1DM or T2DM. T1DM who live west of the Caldecott Tunnel had a significantly higher mean HbA1c (p=0.011) than those east of the tunnel. Language had no significance on HbA1c or BMI%ile for either group. A.As had a significantly higher HbA1c compared to all the other ethnicities (p<0.05) for T1DM only. The BMI %ile among T2DM was significantly greater for both A.A.s and Latinos compared to Asians (p=0.010 & p=0.009). Conclusions: A.As and Latinos have higher BMI%iles than Asians with T2DM. Also, T1DM patients with a lower socio-economic status and of certain ethnicity are at risk for poorer glycemic control, while the greatest risk factor for poor glycemic control in T2DM was an income of <$40k. The reasons for these disparities should be the subjects of further studies.
86 Spenser Perloff Volunteer
University of California Davis First Year
Mentor: Ed Lammer, MD
Unlike many of my friends, my interests have been in flux my entire life. My first thought was that I would become an aerospace engineer and design the next generation of airplanes. However, I found a strong pull to the unknowns of genetics. I have loved pondering the questions in genetics ever since taking Mr. Chugh’s biotechnology course in my junior year of high school. The next year, I followed up on this newfound passion by taking AP Biology. As I was applying to college, the litmus test for every institution was whether or not they had a good genetics program. My goal in life is to discover ways of curing genetic disorders such as Rett’s Syndrome and Tay Sacs. I look forward to learning more about what my goal entails as I go through this internship and the rest of my college career.
As a one time patient of Children’s Hospital, I can think of no better place to learn about medical mysteries. Following my experience with a ruptured AVM in the eighth grade and the subsequent years of follow up visits, I decided to give back to Children’s by getting involved in their Teens for Kids program. In the TFK program, I produced concerts and worked fundraisers to raise money for the clinics at Children’s Hospital. I hope that this internship at CHORI will make just as big a difference in a child’s life.
87 Optimization of the Thymidylate Synthase Genotyping for Case Control Birth Defect Studies
S. Perloff, D. Iovannisci, K.Schultz, L. Tom & E. Lammer. Children’s Hospital Oakland Research Institute, Oakland CA 94609
Introduction: Structural birth defects affect one in every thirty third child born and folate deficiency has been associated with several birth defects. Our laboratory is currently evaluating polymorphisms with the genes regulating folate metabolism for their possible association with birth defects such as orofacial clefts and Spina Bifida. One such gene encodes Thymidylate Synthase (TS). Together with folate, the TS enzyme converts dUMP into dTMP which is necessary for DNA replication and repair. Within the TS gene promoter, there is a tandem repeat polymorphism. The most common allele is a two-copy 28bp tandem repeat of 56 nucleotides associated with decreased expression and the second most common allele is a three-copy tandem repeat of 84 nucleotides associated with greater expression. Objective: In the preliminary studies, the genotyping protocol was not well optimized which obscured the interpretation of the results. Many gels were not readable, and those that were, often contained extra bands, confused the results. A significant problem with amplifying repeats is that often the template and extended strands can partially dissociate and loop out leading to either shorter or longer copies than intended. My goal for this project is to improve the genotyping method employed for this gene. Methods: In this study, have used a variety of techniques to evaluate the genotyping protocol. These include the design of better primers to shorten the amplicon, using different polymerases with proof reading functionalities or with better processivity through GC rich regions, varying the thermal cycler protocol for the specific primers, inclusion of additives in the PCR mixture such as DMSO to prevent secondary structures from forming, and varying the gel electrophoresis conditions. We extracted DNA from dried blood spots, amplified it using PCR, and performed gel electrophoresis ofthe PCR products. We then scored the gels to determine the genotype (homozygous two repeats, homozygous three repeats, or heterozygous for both). Results: The greatest improvement was obtained for the TS assay through the design of new PCR primers that reduced the sizes of the amplicons corresponding to the two most common alleles 215 and 243 nucleotides to 88 and 116 nucleotides. The smaller amplicons not only reduced the presence of stutter bands, but also resulted in better separation of the two alleles. Some additional improvement was obtained by employing a high fidelity polymerase. Conclusion: In contrast to results in the literature, the inclusion of DMSO did not improve TS genotyping.
88 Mallery Quetawki Funded by the NIH/NHLBI/Professional Student Short Term Program University of New Mexico Third Year
Mentor: Mark Burge, PhD
My name is Mallery Quetawki and I am from Zuni, New Mexico. I am about to enter my 4th year this fall at the University of New Mexico in Albuquerque, as a Biology and Art studio major. This is my 3rd year as an intern with the CHORI intern program. This summer I took a different approach to the program by staying at UNM to do my research. I am working with Dr. Mark Burge from UNM Hospital. Our project is a sleep architecture study on obese pre-diabetic young adults. I am very grateful that CHORI had given me an opportunity to have a head start here in my home state. I am surely on my way to a bright future in the field of medicine. This program has assured me that this is what I want to do for a living. I appreciate all that CHORI has done for me and hope that this connection will continue beyond my undergrad years. I would like to thank Dr. Lubin, Vas, and Diane for everything that they have done to make sure this summer across the states went well. Thank you.
89 Disordered Sleep in Obese Adolescents With and Without Impaired Glucose Tolerance
M. Quetawki & M. Burge University of New Mexico, Albuquerque, NM 87131
Introduction With socioeconomic changes in the life of Americans and other urbanized countries, sedentary lifestyles have developed into the weight gain of today’s youth. Data from New Mexico suggests an even higher rate of obesity in our state. With increasing BMI, the risk for metabolic dysfunctions, such as Type 2 diabetes, also increase. Obesity has also been found to parallel Obstructive Sleep Apnea Syndrome, which is characterized by pauses in breathing during sleep. According to previous studies, overweight Hispanic males are at a greater risk for developing pre-diabetes or the metabolic syndrome. The metabolic syndrome is a cluster of conditions that may increase diabetes and cardiovascular risk. Recent studies have looked into sleep deprivation and apnea as a possible lead in the development of insulin resistance. Conversely, it is also possible that insulin resistance may cause sleep deprivation, obstructive sleep apnea and other sleep disturbances. As of lately, there have only been a handful of sleep architecture studies in obese and pre- diabetic adolescents based on gender, BMI, ethnicity and age. Objective We propose to perform a prospective study designed to compare sleep quality among obese Hispanic young adults with and without pre-diabetes. We hypothesize that obese pre-diabetic young adults will have more measurable sleep disturbances than those without pre- diabetes. Methods This pilot study will distinguish between the sleep architecture of 3 distinct groups of Hispanic young adults age 18-21. The study subjects will be matched according to BMI, age and gender. 12 obese subjects with pre-diabetes and 12 obese subjects without pre-diabetes will be needed for this study. Obesity will be measured as a BMI above the 85th percentile according to CDC guidelines. 12 normal weight individuals without pre-diabetes will also be recruited as our control group. An overnight sleep study will be performed at the UNM General Clinical Research Center. An accommodation night will be done before the actual study where data will not be gathered. The second night will consist of full data collection with a multi-channel polysomnography apparatus that will be attached to the patient during both nights. The apparatus is a tool that takes comprehensive records of the biophysiological changes that occur in sleep. This apparatus will include the following: electroencephalography, electromyography, electrocardiography, nasal and oral airflow, snore, rib cage movement, abdominal movement and oximetry (SaO2 and heart rate). Abnormal events will be scored according to American Academy of Sleep Medicine guidelines and established UNM Sleep Center Protocols. Results and Conclusion This study is still in the processes of data collection. We hope that the significant findings, if any, will aid in the intervention and treatment of obesity and pre- diabetes among today’s Hispanic youth and young adults. We also hope that our findings will inspire further sleep study research on this particular topic.
90 Jesse Sandberg Volunteer
Duke University Fourth Year
Mentor: Po Lin So, PhD Ervin Epstein, MD
Early in life, playing outside, getting dirty and trying to find different types of bugs and animals got me interested in the vast diversity of life on earth. I always wondered why certain animals had certain abilities and how their characteristics function in their bodies. As well as having an interest in outdoor activities, I also readily enjoyed playing with Linkin Logs and Legos, building numerous random structures. It seemed then only logical that upon matriculating into Duke University that I would major in Biomedical Engineering, a combination of both life science and design. Over the course of my college career, I became more and more interested in medicine after volunteering in the Duke hospital and conducting research on pediatric radiology. Now as an upcoming senior, I am focused on finishing my biomedical engineering degree as well as all pre-med requirements. In addition to being a student at Duke, I have also been a student athlete as a member of the Duke Varsity Swim Team, writer for the Duke Engineer Magazine and a volunteer for the Duke/Durham community.
The hands on laboratory experience gained this summer by working for Dr. Epstein and Dr. Po Lin So on Basal Cell Carcinoma (type of skin cancer) has helped me bridge my academic knowledge to a real world application. Ironically, this research really relates to me because of my continued exposure to the sun due to my numerous summer activities including swimming for a competitive USS (year round) swim team and working as a swim coach to three hundred kids at Orinda Country Club. Overall this summer has been a lot of fun and helped me further my desire to work in the medical field.
91 Inhibition of Hedgehog Signaling Pathway in BCC Skin Tumors
J. Sandberg, P.-L. So & E. Epstein Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Basal cell carcinoma (BCC) is the most prevalent somatic skin cancer, affecting approximately 750,000 Americans per year. Approximately 1 in 50,000 individuals in the United States are born with the autosomal dominant genetic disease, Basal Cell Nevus Syndrome (BCNS), which predisposes them to develop many BCCs at an early age. The causative genetic lesion in BCNS has been mapped to the tumor suppressor gene PATCHED1 (PTCH1), and affected individuals inherit a defective copy of PTCH1. Wildtype PTCH1 protein, a 12-transmembrane receptor for the ligand, Hedgehog (Hh), represses the Hh signaling pathway via its repressive indirect interaction with SMOOTHENED (SMO), a positive regulator of Hh signaling. This repression is reversed when Hh binds to PTCH1. Both sporadic and inherited forms of BCC have been shown to be caused by overactive Hedgehog (Hh) signaling The Hh signaling pathway is normally required for embryonic and postnatal development of structures such as the brain and hair follicle. In postnatal animals, active Hh signaling is required to promote the growth of a new hair follicle (anagen phase). Without the hedgehog signaling pathway, hair growth is stalled at telogen (resting phase).Objective: We will investigate whether tazarotene inhibits the growth phase of hair follicle in vivo, which normally requires Hh pathway activation, by inhibition the expression of Gli1. Methods: A 1 inch x 2 inch region of hair was depilated on each mouse using Nair hair removal cream. Two groups of mice five male C57BL/6J and five female C57BL/6J mice in each group were used. Both groups were treated once a day for seven days a week for two weeks. The first test group was the negative control group (vehicle cream) and the second test group was the independent variable test group (Tazarotene Cream). After two weeks of applied treatment, skin biopsies from each mouse were obtained, sectioned and a standard Immunohistochemistry protocol was followed to test for Gli expression. Results and conclusion: Preliminary results suggest that tazarotene does inhibit hair growth on mice through the hedgehog signaling pathway. Upon visual observations of the mice after treatment, the hair on the control mice were growing back but not on the tazarotene treated mice. As of this point, the skin biopsies previously taken are still being sectioned so the histology of the skin samples have not been performed. However, upon arrival, we expect that there will be down regulation of Gli1 protein, confirming our belief that tazarotene inhibits the hedgehog signaling pathway.
92 Chia Teoh Funded by the NIH/NHLBI/Professional Student Short Term Program San Francisco State University Postbaccalaureate
Mentor: Jennifer Olson, MD Suruchi Bhatia, MD
I am currently a post-baccalaureate student at San Francisco State University, majoring in Biology. My family came to America 15 years ago, thus making my family the first generation here in United States. Growing up in California as immigrants, I truly endured the trials and tribulations that any new family in America had to go through. One great struggle was that my father’s first job did not provide us with health care benefits. As a result, my family and I strived to stay healthy and tried our best to avoid getting sick. Another great struggle was the language barrier. In fact, this communication barrier in our family made it even more difficult for us to receive the proper care that we so greatly needed. From this life experience, I not only understand the difficulties that many disadvantaged children face today in getting the right treatment but was one of the reasons why I decided to go into the field of medicine.
During my undergraduate years at the University of California, Berkeley I not only fulfilled the prerequisites to become a Molecular Cell Biology major, but I also discovered many other resources to help me get closer to my goal. One resource was my voluntary job with hospitals where I was able to get a glimpse of what being a doctor is all about. I continued my learning through an informal post-baccalaureate program at SFSU where I was able to continue to broaden my science education and take numerous classes where I didn’t have a chance to take at Cal. One of the classes I took at SFSU was Endocrinology which fascinates me. This summer, CHORI has given me an opportunity to work with Dr. Olson at Children Hospital Oakland who specializes in pediatrics endocrinology and diabetes. We are looking at females who are diagnosed with Turner’s syndrome. The main objective of this study is to determine the timing, dose, form and route of estrogen replacement in girls with Turner’s syndrome as well as other forms of primary and secondary ovarian failure within the current practice of six pediatric endocrinologists at Children’s Hospital and Research Center at Oakland. Thanks to CHORI, this clinical research has furthered my interest and opened up my eyes in the world of medicine. I know that my goals of becoming a doctor is now possible and reachable if I put my mind to it and continue to keep the determination in my heart to achieve it.
93 Timing of Estrogen Replacement Therapy in Girls with Turner Syndrome
C. Teoh & J. Olson Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Background: Turner Syndrome is caused by a missing or partial absence of one of the two X chromosomes normally found in females. It is a genetic disorder that affects female development (1 out of 2000 females) and the major cause of primary ovarian failure in females. Girls with Turner’s syndrome are of short stature, lacking ovarian development, have webbed neck, and are prone to cardiovascular problems. Growth hormones are recommended to increase final height and estrogen replacement which is necessary for both feminization and for prevention of osteoporosis. The timing of estrogen replacement is crucial because estrogen affects the result of growth, where high doses of estrogen will inhibit growth and low doses of estrogen helps to stimulate growth. Objective: The purpose of this study is to determine the timing, dose, form and route of estrogen replacement in girls with Turner Syndrome as well as other forms of primary and secondary ovarian failure within the current practice of six pediatric endocrinologists at Children’s Hospital and Research Center at Oakland. Methods: The method involves a retrospective chart review of subjects with Turner Syndrome and other forms of primary or secondary ovarian failure who have been treated at Children’s Hospital and Research Center at Oakland within the last 10 years. The sample size was estimated by the number of patients with Turner Syndrome and other forms of primary or secondary ovarian failure between the ages of 10-18 years. There are seven variables that were viewed and analyzed between Turner Syndrome and other forms of primary or secondary ovarian failure patients including: a) initial BMI b) average BMI after 6 month -1 year of estrogen c) differences between initial and average BMI after 6 month – 1 year of estrogen d) current age as of July 2007 and e) age started on estrogen f) dosage and g) final height after estrogen (same 2 points in height at least 6 months after estrogen). Result: There are 62 cases of patients with Turner Syndrome and 51 cases of patients with other forms of primary or secondary ovarian failure. There was no significance (p>0.05) between the two groups in six of the variables that were analyzed : a) initial BMI b) average BMI after 6 month -1 year of estrogen c) differences between initial and average BMI after 6 month – 1 year of estrogen d) current age as of July 2007 and e) age started on estrogen f) dosage. However, there was a significance (p<0.05) in the final height after estrogen between the two groups. Conclusion: Based on these studies, we concluded that the association between the timing on estrogen and final height was not significant (p=.727). In addition, the correlation between dosage and final height was not significant as well. Therefore further studies need to be conducted, and patients with these disorders who have not yet been started on estrogen may be enrolled in additional studies.
94 Ashleigh Thompson Funded by the NIH/NHLBI
Piedmont High School Senior
Mentor: Padmavathi Bandhuvula, PhD Julie Saba, MD, PhD
Although many people claim to have had an interest in science since birth, I cannot say that the same was true for me. While I always enjoyed my science classes and loved participating in science fairs at my school, my true interest in science developed later on. During the spring of my sophomore year in high school, one of my close friends was diagnosed with non-Hodgkin’s type Lymphoma. That fall I decided to train and run a half- marathon with the intention of raising money in her honor for the Leukemia and Lymphoma Society. Throughout the training process I met many incredibly strong individuals who had successfully battled these cancers and also had the opportunity to learn a tremendous amount about those diseases and the treatments that are currently available for them.
I would like to think that my running made even a small improvement in people’s lives. More importantly, I was so inspired by the experience that I knew that I wanted to do something that would continue to make a difference in the lives of others. I found a way to couple my interest in science with my desire to make that difference. In the summer of 2006, I had the chance to intern at a small Biotechnology company in Hayward. There I took part in the development of a pharmaceutical to cure drug-induced alopecia. I was already on the way to achieving my goal of making a difference!
I am so indebted to the Summer Student program, as it has firmly established me on the path to make a difference. Now, more than ever, I am certain that I want to pursue a career in the sciences, as I have seen first hand the breakthroughs that have been made here at CHORI. I have been so fortunate, not only to have had the experience of working in this incredible program, but also to have had the opportunity to work alongside a group of unparalleled mentors who have gone out of their way to encourage me and to educate me in the laboratory environment.
I recently graduated from Piedmont High School and in the fall I will be attending Johns Hopkins University in Baltimore, Maryland, where I plan to pursue a degree in Biomaterials Engineering.
95 Screening the Inhibitory Effect of Some Compounds on SPL Activity
A. Thompson, P. Bandhuvula & J. Saba Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Sphingosine-1-phosphate (S1P) is a naturally occurring lipid that is present in most human cells and in the bloodstream. S1P is important for many biological functions and activates signals that stimulate cell survival, migration, and proliferation. S1P is irreversibly degraded by the enzyme SPL, an integral membrane protein that
resides in the endoplasmic reticulum and catalyzes the cleavage of S1P at the C2-3 bond. In most cases, the effects of SPL can be accounted for by its ability to regulate S1P levels. SPL is downregulated in human colon cancer and intestinal adenomas of the Min mouse, suggesting that SPL may be a useful marker and potential therapeutic target in intestinal tumorigenesis. SPL plays many diverse roles in biology and raises the possibility that SPL may be a useful target for immunomodulatory and anti cancer therapy. Screening for small molecule inhibitors that act on SPL enzyme is important in understanding the repercussions of the inhibition. Additionally, as the current way to screen SPL inhibitors, a regular fluorescent assay, although a sensitive process is still time consuming and a modification to detect SPL inhibition from hundreds of compounds using a fluorescent plate reader would be helpful. Objectives: The objectives of this experiment are 1) to test inhibitory effect of compounds on S1P lyase enzyme activity using HPLC based SPL assay and 2) to develop a high throughput assay to test the inhibitors using a fluorescent plate reader. Methods: Fluorescent SPL assay: A regular fluorescent assay using NBD-labeled S1P was performed with the inhibitors PE, 11-cis, PA, SC, and NEM. The reaction was started by the addition of Ad-SPL cell extract and continued for 30 minutes at 37°C. The products were extracted and separated and then injected onto the HPLC under a gradient established system. The NBD signals were then detected by fluorescent detection and quantified by integrating peaks. Fluorescent SPL assay using plate reader: The SPL assay is done as the regular fluorescent assay mentioned above except that the products are not separated from the reaction mixture. After the reaction was stopped the SPL product formed in the plate reader is labeled with 2,4-DNPH and then read in a plate reader. Results: Significant inhibition was observed from the following inhibitors; PA: 36.33%, PE: 20.66%, SC: 8.82%, and 11-cis: 17.11% although no inhibition was detected from NEM. After making a series of standards for the plate reader assay, work is still being done to standardize the assay, which promises to provide a quick and straightforward way to screen hundreds of inhibitors at a time.
96 Lauren Tom Funded by Genetics Research Fund/ Lammer Lab University of California Los Angeles Third Year Mentor: Ed Lammer, MD
My name is Lauren Tom and I will be a fourth year Ecology, Behavior, and Evolution major at the University of California Los Angeles. This is my first year interning at CHORI and I feel very fortunate to be a part of this program. Growing up in Berkeley, I always wondered what was inside that big pink building on Martin Luther King Jr. Way. I never imagined that someday I would be working there. Working in the Lammer Lab has been a great experience. On a daily basis, I have had the chance to do things that, up until this summer, I had only read about in textbooks. I have had the opportunity to take part in ongoing research on gastroschisis, a birth defect that is on the rise, especially in California. I would like to thank my fellow lab members, Dave, and Kathy for their help and patience and Ed for hiring me and letting me work on this wonderful project. As my final undergraduate year approaches, I have been getting a lot of questions regarding what will come next. Sorry, no new information here. I still do not know what it is that I would like to do once I graduate. However, I plan on traveling to Japan this summer and, in the fall, I will be going to Africa to study animal behavior at the Mpala Research Center in Kenya. Hopefully, these trips will guide me towards something that I will enjoy doing in the future, whether it is research, graduate school, or something entirely different.
97 Maternal Smoking, Genetic Variation of Glutathione S-Transferases, and Risk of Gastroschisis
L. Tom, K.Schultz, D. Iovannisci & E. Lammer Children’s Hospital and Research Center at Oakland, CA 94609
Introduction: Gastroschisis is a birth defect that is characterized by the external development of internal abdominal organs and its prevalence has been increasing, especially in California. Infants with gastroschisis require immediate surgery and often suffer from respiratory problems, morbidity, and mortality. In our study, we are investigating the role of GSTM1 and GSTT1 genes to determine if they are associated with the risk of gastroschisis. GSTM1 and GSTT1 are glutathione S-transferases that are important detoxification enzymes. These enzymes conjugate glutathione to potentially harmful reactive intermediates (such as 4-aminobiphenyl and 2-naphthylamine that originate from tobacco smoke), inactivating them and making them more water-soluble and more easily excreted from the body. The most common polymorphism for both GSTM1 and GSTT1 is the deletion of the entire gene. Approximately 50% of the general population is homozygous null for GSTM1 and 15% is homozygous null for GSTT1. Objective: The goal of this study is to determine how the presence or absence of GSTM1 and GSTT1 gene sequences, in combination with maternal smoking, affect an infant’s risk of being born with gastroschisis. Methods: We are using polymerase chain reaction (PCR) endpoint analysis to amplify DNA extracted from blood spotted onto newborn screening cards. The extracted DNA is added to a mix containing primers designed to co-amplify the GSTM1, GSTT1, and Albumin genes. Since every individual contains at least one copy of the Albumin gene, the presence of the Albumin band serves as a positive PCR control for individuals that may be homozygous null for both GSTM1 and GSTT1. The mix containing the DNA is placed inside a thermocycler and amplified. The amplified samples are run on a 1.5% agarose gel, stained with ethidium bromide, and photographed under ultraviolet light. The resulting data will be sent to the California Birth Defects Monitoring Program (CBDMP) and combined with maternal interview data concerning smoking behavior during pregnancy. The combined data will be used to calculate the risk of gastroschisis given the genotype of the infant and the level of exposure to tobacco smoke. Results: A total of 227 blood spots have been genotyped for this study. Our preliminary findings show that the homozygous null deletion occurs at a frequency of 41.0% for GSTM1 and 16.3% for GSTT1. 6.61% of the samples analyzed were homozygous null for both GSTM1 and GSTT1. These frequencies are similar to the reported literature values. Conclusion: Statistical analysis is currently underway. However, our model predicts that, in the presence of maternal smoking, infants who lack GSTM1 and GSTT1 will experience a higher risk of gastroschisis than infants with normal levels of GSTM1 and GSTT1 activity born to a mother who does not smoke.
98 Nora Trice Volunteer
Lick-Wilmerding High School Junior Mentor: Robert Ryan, PhD
My name is Nora Trice and I am entering my senior year at Lick-Wilmerding High School in San Francisco. At school, I’m a member of the tennis and track teams. Outside of school, I tutor children ages 8-18 at a writing workshop in the Mission called 826 Valencia, and I’m also a member of a Teen Panel designed to help parents with adolescent issues. I’ve spent the past several summers taking courses on various college campuses. However, I’ve been waiting for my chance to participate in the CHORI student program since the summer before my freshman year, when my brother participated for the first time. After seeing his enthusiasm and watching his oral presentation at the August symposium, I knew that I wanted to have a similar experience.
My experience working in the lab has been far more rewarding than I could’ve imagined. Although I have not yet applied to colleges or selected a major, I am now aware of so many more possibilities in the scientific fields. I was able to see the many processes that are involved in conducting research, and experience firsthand the beginning stages of a possible career path. I feel very fortunate, and I’d like to thank Dr. Lubin for this opportunity. I’d also like to thank Dr. Ryan, Megan Tufteland, and the entire Ryan lab for welcoming me and teaching me everything I needed to know for a successful summer.
99 Environmental Stability of Amphotericin B Nanodisks
N. Trice, R.O. Ryan & M. Tufteland Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Amphotericin B (AMB) is a hydrophobic, antifungal drug often used to treat systemic fungal infections. To overcome the solubility barrier, we’ve encapsulated AMB into lipid and protein-based particles called nanodisks (ND), which serve as a delivery vehicle. The ND contain three elements: the phospholipid, the protein, and the drug. The protein used in this project is apolipoprotein A-1, which has the ability to solubilize lipids. The primary lipid is the zwitterionic dimyristoylphosphatidylcholine (DMPC). Encapsulation into ND allows water solubilization of AMB as well as retention of its ability to bind ergosterol, the primary sterol in the fungal cell membrane. AMB is able to create a pore in ergosterol containing membranes that leads to the leakage of ions, resulting in fungal cell death. Objective: The stability of AMB ND to standard environmental conditions, including sunlight, air, and temperature, was tested. These studies were designed to evaluate the potential of AMB ND as a biofungicide. Methods: Two samples of AMB ND were incubated outside and two samples were kept inside in a dark environment. In each environment, one sample was sealed with nitrogen and one was exposed to air. Four parallel control samples of fungizone (an AMB detergent micelle complex) were also tested. After 48 hours, a yeast growth inhibition assay was performed whereby 20µl of a saturated overnight culture of Saccharomyces Cerevisiae was used to inoculate 5ml YEPD media in the presence of increasing amounts of specified AMB formulations. Cultures were grown for 16 hours at 30°C and the extent of culture growth monitored by measuring sample turbidity at 600nm. Results: In monitoring AMB stability, we found that after 48h, ND exposed to light were equally effective at killing yeast as unexposed ND or fungizone, despite a drastic decrease in color intensity. Fungizone experienced the same decrease in color however lost all biological activity after light exposure. The ND samples exposed to air were identical to those sealed with nitrogen, as were the Fungizone samples. Conclusion: Integration of AMB into ND confers protection from degradation of this antibiotic by environmental factors. Thus, AMB nanodisks may be useful biofungicides and may have application beyond systemic fungal infections.
100 Marena Trinidad Funded by the NIH/NHLBI
Benecia High School Junior Mentor: Carolyn Hoppe, MD
To those familiar with me at Benicia High School, my name, Marena Trinidad, is synonymous with overachiever, and personifies a quiet character too gung ho about science. People often inquire as to what could possibly be my impetus, and comment upon the fact I always have a textbook under my nose. However, I wouldn’t abandon my interest in science for anything in the world. In fact, if it weren’t for my habit of lugging around schoolbooks, my doctor at Children’s Hospital of Oakland would never have discovered my desire to become a research scientist and told me about the summer program at CHORI. I am extremely grateful to have had this experience. It has been my most fulfilling and enjoyable one yet. Moreover, working with my mentor, Dr. Hoppe, and the people of Noble Lab has made me more determined than ever to help others as CHO has helped me.
101 Genetic Risk Factor for Stroke in Adult SCA Patients
M. Trinidad & C. Hoppe Children’s Hospital & Research Center at Oakland, Oakland, CA 94609
Background/Intro: Stroke is a devastating complication of sickle cell anemia (SCA), affecting both children and adults with an incidence of 24% by age 45 years. Although the incidence of ischemic stroke is highest in young children, hemorrhagic stroke predominates in adolescents and young adults, and is the leading cause of death due to stroke in SCA. Ischemic stroke is a result of progressive narrowing and obstruction of the large intracranial vessels. Persistent vascular injury with rupture of collateral blood vessels results in hemorrhagic stroke. The cause of stroke in SCA is not known. Although adult stroke may differ from childhood stroke in SCA, many of the pathways leading to vessel injury may be similar. Several studies have reported genetic associations with stroke in SCA, but these studies have focused almost exclusively on children. We previously reported an association between the TNF G(-308)A polymorphism and stroke risk in children with SCA and found that the TNF (-308)GG genotype conferred an increased risk of ischemic stroke due to large vessel disease. Hypothesis and Specific Aims: As stroke in adults and children with SCA is similarly characterized by large vessel disease, we postulate that the TNF G(-308)A polymorphism is a risk factor for stroke in adults with SCA. The objective of this pilot study is to extend our findings in children to adult SCA patients with stroke. Methods: Informed consent was previously obtained and DNA collected and stored on 53 adult patients followed at the Comprehensive Sickle Cell Center at CHRCO. Corresponding clinical and neuroimaging data were used to classify patients into case and control groups. Case subjects were defined as having had a first clinical stroke or new MRI/MRA findings of large vessel (LV) disease after the age of 13 years. Control subjects had no history of stroke and/or absence of LV disease on MRI/MRA. Genotyping for the TNF(-308) polymorphism was carried out using a PCR-based linear array assay. Genotype frequencies were compared between LV-positive case and LV-negative control subjects with the Fisher’s exact test for 2x2 contingency table analysis. Anticipated Results: 1. Briefly describe study population: sample size (53), number of cases and controls, mean age of cases/controls 2. Compare TNF (-308) GG genotype frequencies between cases and controls. 3. Are findings consistent with previous results in children? ie.homozygosity for the common TNF G(-308) allele is found in excess in the LV-positive case group. Conclusions: Wait for results.
102 Griselda Velasquez Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Second Year Mentor: Robert Raphael, MD David Martin, MD
In many countries people have no access to social health protection -- affordable health insurance or government-funded health services.(WHO) Growing up in the Mission District of San Francisco and visiting family in Mexico helped me develop an awareness of the health inequities prevalent in both communities where access to health is either ‘illegal’ or non-existent. With this consciousness in mind, I have felt responsible to catalyze for change in the healthcare system and invest in the health of underserved communities across political borders.
The importance of biomedical research lies in the power that it holds to have a positive impact in the lives of people worldwide who may not necessarily have access to health care but can benefit from research being conducted on diseases devastating their regions. My intention is to contribute, if only in a small way, to the investment that goes into research in order to give back to communities battling life threatening diseases through biomedical research internships and eventually medicine. Currently, I work with other students in a Diabetes awareness and prevention program whose focus is to provide education to the community and run a course at Cal so that students can learn the necessary components of diabetes with the purpose of training them to become health educators. In addition I am a part of Biology Scholars Program and work as an Assistant Publicist for Hesperian Foundation, a non-profit publisher of health books and guides.
Epigenetic research at CHORI has challenged me to think differently about inheritance and I have learned to analyze and interpret data accurately and patiently. Most importantly, I have learned to strategize and decide which steps to take next in bench work in order to find answers to the questions and hypotheses posed. Thanks to the Martin lab, especially Dr. Raphael, for all your support and patience.
103 Detection of hMLH1 Somatic Hypermethylation by Methylation-Specific PCR and Quantitative Methylation-Specific PCR (Q-MSP)
G. Velasquez, D.K. Martin & R. Raphael Children’s Hospital Oakland Research Institute, Oakland, CA 94609, USA
Introduction: Epigenetics is defined as the study of mitotically or meiotically heritable changes in gene expression and function that are independentof DNA sequence. The epigenetic state mediates transcriptional activation or silencing of a gene and these phenomena are caused by modification of DNA (DNA cytosine methylation). Methylation of CpG islands (CpG rich regions) are associated with transcriptional silencing while unmethylated CpG islands are associated with active transcription. Epigenetic silencing can result in the loss of alleles of a gene causing an “epimutation.” Epimutation can cause abnormal activity of a gene that should be silent. The study of the occurrence of epimutations in cancers cells has shown that hypermethylation of tumor suppressor genes is common. Specifically, hypermethylation of the DNA mismatch repair gene hMLH1 has been observed in colorectal tumors exhibiting MSI. The study of germline epimutation in hMLH1 has the potential to show that epimutation of this mismatch repair genes underlies apparent sporadic tumors. Thus, it is possible that epimutation plays an important role in cancer and can increase our understanding of oncogenesis. Objectives: (1) Ask if epimutation of hMLH1 is present in peripheral blood cells of healthy individuals using a highly sensitive quantitative methylation-specific PCR (Q-MSP) to assay for epimutation in unselected blood donors. (2) Test the hypothesis that somatic mosaicism for the hMLH1 epimutation is associated with the risk of developing an MSI tumor. Normal lymphocytes of sporadic MSI tumor patients with matched microsatellite-stable colorectal cancer controls will be compared. Methods: Bisulfite modification of the DNA where unmethylated cytosine nucleotides are converted to uracil and methylated cytosines are not converted. Quantitative methylation-specific PCR amplification of the hMLH1 promoter follows using methylation specific primers selectively amplifying only methylated sequences to determine the frequency of methylated alleles among the samples. Real time PCR uses a fluorescent signal to detect the product. The amount of template DNA is correlated with earlier amplification and samples are calibrated with a standard curve. The samples are compared to a standard curve to estimate the amount of methylated copies. Samples are then sent to Berkeley DNA Sequencing Labs and are further analyzed for proportions of methylation in the sequence of each sample. Results and Conclusions: In testing for hMLH1 epimutation in normal peripheral blood cells 0/15 adult samples, 0/28 cord blood samples and 0/7 pediatric blood samples have detectable hMLH1 methylation. The study is ongoing as is the development and improvement of Q-MSP assay.
104 Blia Ashley Yang Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Third Year Mentor: Phillip Bollinger Vasanthy Narayanaswami, PhD
I am an incoming senior at UC Berkeley majoring in Integrative Biology. This summer I had the wonderful opportunity of working with two great mentors, Vas Narayanaswami and Phillip Bollinger, in evaluating and creating an alumni database for the summer program.
This project is different in many ways from the research being conducted by other program participants, but I believe it holds much significance in its contribution to the summer program. This database can potentially serve as the foundation for networking among past summer program participants where students can share experiences and interests in scientific research. More importantly, this project will highlight the differences the CHORI Summer Research Program is making in providing research opportunities undergraduate students.
This will be my second summer as a summer program participant. Working on my project has strengthened my appreciation for the CHORI Summer Research Program. Working with mentors who are devoted to providing wonderful experiences at CHORI and who are willing to approach scientific research with a different mentality is encouraging for me as a student. My aspirations in obtaining a health and research profession where I can work in underserved communities has been renewed.
I would like to thank the Biology Scholars Program (BSP) at UC Berkeley who, in more ways than they know, have provided direction in the creation of this database and have, time and time again, relentlessly supported women and minorities entering the field of science. To Vas, Phil, Diane, and all the mentors thank you for your dedication to the program. There is a dire need for diversity in the field of science and what you are doing, your partnership with the CHORI Summer Program, makes it possible for us.
105 CHORI Summer Research Program: Looking Back
B. A. Yang, P. Bollinger, B. Lubin & V. Narayanaswami Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: The CHORI Summer Research Program is a nine week program that was initiated in 1994 with the objective of providing undergraduate and high school students research experiences in the clinical or basic science areas. The program is open to students nationwide. The students are paired with mentors, who are scientists and clinicians from CHORI and CHRCO. Objective: The overall objective of my project is to assess if the CHORI summer program is effective in providing research experiences and stimulating interest in pursuing biomedical/public health professions among a diverse student population. We will conduct a two-part qualitative and quantitative evaluation of: 1) the current 2007 program students, and, 2) the past program participants. The goals are: (i) to contact the past participants, (ii) to create a survey and an alumni database, and, (iii) to determine the influence of the program on the students’ educational and career goals. Finally, we will assess the ethnic and gender representation of the participants and the focus of the research areas. Methods: The past students were contacted by e-mail, telephone, and through a web-based search. An alumni database was initiated using the software, FileMaker Pro. The students were asked to complete a survey that included their biographical and educational information, and their current educational and career pursuits. Results: There were a total of 419 students (81% undergraduate and 19% high school) who participated in the program from 1994 to 2007. A majority of the students (about 60%) participated in basic research. About 65% of the participants were women. The program represented an ethnically diverse group of undergraduate students from across the US, with 40% of the students from the Bay Area. 198 students could not be reached because of lack of contact information. Of the 221 students contacted, 80 completed a survey for a response rate of 36.20%. Data from the program participants is being received on a rolling basis. A qualitative evaluation of the comments received reveals that the students benefited tremendously from their summer research experience. A majority (77%) of the students who responded to the survey are pursuing a career in the public health and biomedical areas. NIH funded students, who are also included in this general study, are evaluated separately. Conclusion: Tracking the past participants and obtaining their survey response seem to be the major challenge and limitation for the program evaluation. Nonetheless, the CHORI Summer Research Program appears to have a tremendous impact in influencing and reinforcing the educational and career pursuits of a diverse population of students in the direction of medical professions, biomedical research and public health programs.
106 Sua Yang Funded by the NIH/NHLBI/Professional Student Short Term Program University of California Berkeley Fourth Year Mentor: Patsy Wakimoto, PhD
My name is Sua Yang. I was born and raised in Sacramento, California. The past 22 years has been an extraordinary but tough adventure. Growing up, I envisioned myself attending a good school, meeting wonderful people and somehow working in the health field. Being at UC Berkeley has aided in the process of my development into a mentally, emotionally, and physically stronger person. I have encountered countless inspiring opportunities of which I have been able to actively engage myself in, both within my own and the local community. These opportunities include being a Biology Scholars Program (BSP) student, leading a de-cal titled ‘Prepare to Achieve a College Education’ (PACE), working with the Undergraduate Research Apprentice Program (URAP) and being privileged enough to work with CHORI’s summer internship for the past three summers. All these opportunities have strengthened my vision and encouraged my objective of making a difference in everything I do. Furthermore, working with high school students, attending meetings with health care professionals and medical school representatives as guest speakers, and closely working with Hmong students and families is what continues to truly feed my adrenaline and commitment to the underrepresented. Regardless of differences between ethnic groups, time and geographic location, language and cultural differences persists in creating barriers. I have personally witnessed this dilemma as a child and in the Hmong community today with nutrition and diabetes/obesity awareness. I strongly believe that differences in language and cultural perspectives should not have to negatively affect one’s health and illnesses. As a health care professional, I hope to one day serve as the link in bridging the two. I am thankful for those who have once again given me the opportunity to pursue my research, particularly with the Hmong community and their health. Being at CHORI has opened many doors, including the door to my aspirations. I have found amazing friends and peers who share this genuine goal and passion of working with underrepresented ethnic minority groups, especially by placing oneself in the community of interest. I hope to one day share my vision and opportunities with those who are dreaming the same dream. Thank you, Patsy, Lisa, and Mary for your kindheartedness and devotion to the Hmong people. Thank you, Eric and the BSP staff for your faith and commitment in students like myself.
107 Core Issues for the Hmong in Food Shopping and Food Safety
S. Yang & P. Wakimoto Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: There are 170,000 Hmong scattered in the U.S.. In California, the largest numbers of Hmong are located primarily in the Central Valley. The Hmong have an oral tradition; most can not read or write both Hmong and English. For many Hmong refugees, there exists a major challenge of accessing healthy yet reasonably priced foods. Additionally, cultural practices, including methods of food preparation and food storage have left many at risks. Differences in language and unfamiliarity with nutritious foods pose as obstacles. Objective: To conduct an assessment of the needs of Hmong families in the areas of accessibility to healthy, affordable foods and food safety knowledge and skills. Specific Aims: 1) To examine the needs around food shopping, saving money and nutritional guidance, 2) To develop educational messages around food shopping, saving money, and nutritional guidance, 3) To provide Hmong families with the skills and knowledge needed to access food and eat healthily, and 4) To conduct an educational session, using the above mentioned messages. Methods: A questionnaire was created and administered to 34 Hmong adult participants at locations such as the Hmong community church and Hmong clinical services offices. Data gathered served as baseline information in constructing messages around promoting awareness and understanding of food safety practices and how they support healthy families, including information on food shopping, saving money, and eating healthily. Selected health promotion messages will be field tested and delivered during a workshop–format, educational session. Pre- and post-tests (knowledge-based) will be used to evaluate the effectiveness of messages delivered. Results: Many participants were unable to ‘locate foods and or prices’. Specifically, people were unable to identify items and distinguish healthy from unhealthy foods. Although 19 participants recognized ‘unit pricing’, only 7 actually used the system. Many indicated overbuying, overspending, and not knowing what to purchase for home meals as being the most difficult while food shopping. Only 2 participants suggested using a shopping list to help solve these issues. Conclusion: To successfully address the challenges and develop effective messages in overcoming barriers to purchasing healthy foods economically, cultural food patterns, beliefs about health, and language issues must be considered. Additionally, effective educational practices must include non-traditional approaches such as interactive demonstrations, visual graphics and multi- media as alternatives to print materials.
108 Amanda Yee Funded by the NIH/NHLBI
Alameda High School Junior Mentor: Janet King, PhD
As a young competitive figure skater, subject to the risks and injuries of a demanding physical regimen, I was exposed to the medical world more than most children. Although I fortunately have never had serious injuries that required surgery, my childhood was marked by multiple trips to different types of doctors for check-ups, MRI scans, and sports therapy. As I navigated the hallways of hospitals, I’d glimpse brief snapshots of the lives of different kinds of doctors and the technologies at their disposal. The physicians’ breadth of knowledge mesmerized my young mind. I cannot pinpoint the precise moment when I knew I wanted to pursue a career in medicine, but for as long as I can remember, I’ve answered the inveterate question, “what do you want to be when you grow up,” simply with “doctor.”
Now that I’m sixteen, my passion for medicine, a discipline of answers to the world’s incessant questions, remains as strong as it was when I was eight, but has become more focused and informed. Last summer, I worked as a Patient Ambassador at The University of California San Francisco’s Comprehensive Cancer Center, and as a Galaxy Explorer at Chabot Space and Science Center. Whether in traversing through the maze of hospital hallways or exploring the complexities of the universe, my interest in science has guided my academic goals, plans, class-work and summer internships. Additionally, I have a personal interest in researching the causes of diabetes because I have a family history of, and thus a greater susceptibility to, this chronic disease. My research on gestational diabetes and its correlation with a low glycemic diet here at CHORI has been more than ideal.
This fall, I am considering spending my senior year at the University of Southern California through their Resident Honors Program and Thematic Option Program as a neuroscience major. Outside the world of academia, I am also a competitive figure skater, dancer, musical theatre fanatic, and Lead Mentor Youth Attorney at Alameda County’s McCullum Youth Court.
Also, I’d like to thank Dr. Janet King, Jessica DeHaene, Gaby Hernandez, and Pamela Meija for all their understanding and patience. 109 Glucose Clearance and Energy Expenditure in Pregnant Obese Women
A. Yee, J. King & J. DeHaene Children’s Hospital Oakland Research Institute, Oakland, CA 94609
Introduction: Pregnancy poses several changes and challenges to metabolism. During the first half of pregnancy, insulin sensitivity is increased to facilitate fat storage and glucose clearance from the blood. During late pregnancy, insulin sensitivity declines and postprandial blood glucose is cleared more slowly to conserve glucose for fetal growth. Obese women are prone to enter pregnancy with a certain degree of insulin resistance already leading to a heightened insulin resistance, impaired glucose tolerance and Gestational Diabetes Mellitus. Most likely, glucose clearance from the blood is related to the need of energy by the mother and fetus; however, the relationship between blood glucose clearance and energy expenditure among obese pregnant women has not yet been studied. We hypothesize that obese women with a slower glucose clearance from the blood (measured using a 100g oral glucose tolerance test) have a lower energy expenditure and are less efficient at switching from fat to carbohydrate as the source of fuel, after consuming a 100g oral glucose load. Objective: To evaluate the association between glucose clearance and carbohydrate or fat utilization (respiratory quotient) and total energy expenditure (indirect calorimetry) among obese pregnant women at 20 weeks
of gestation. We will determine how Respiratory Quotient (RQ) values (Vol CO2 expired
/ Vol. O2 consumed) changes as the body switches from a catabolic (fasting) state to an anabolic (fed) state. Methods: Twenty pregnant women with a body mass index (BMI) >25 and body fat percentage >35%, without a previous history of diabetes or any other chronic diseases were invited to participate in the study. Women who consented attended the GCRC at SFGH after a 10hr overnight fast at 20 weeks of gestation. Resting energy expenditure (REE) was measured by indirect calorimetry for 30 min. using a Deltatrac II Metabolic Monitor. Fasting blood sample was drawn and women were given a 100g glucose solution to drink. Additional blood samples were drawn at 60, 120, and 180 minutes post- drink to evaluate glucose clearance. REE was measured at 30, 90, and 150 minute time points. Results: Mean glucose values were 78.11±1.48 mg/dL at baseline; 134.4±5.2mg/ dL at 60 min; 120.8±5.7mg/dL at 120min and 109±5.2mg/dL at 180min. RQ/REE mean values were 0.85±0.01 / 1615.2±48.4 kcal at baseline; 0.93±.012 / 1768.23±52.4 kcal at 30 min; 0.92±.006 / 1730.97±53.53 kcal at 90min; and 0.93±.005/1733.77±49.5 kcal at 150min, respectively. A linear regression model was used to predict changes in REE and RQ using glucose concentrations as the independent variable. Both RQ and REE were positively associated with the change in glucose (r2 = 0.52, p<0.001 and r2=0.468 p<0.001, respectively). Conclusion: Our results suggest that both RQ and REE could be influenced by the concentrations of glucose in the blood. Possibly changes in dietary glucose could have an effect as well, allowing modification of REE and RQ in this group. Further analysis controlling by other variables are pending.
110 Zhe Zhang Funded by the NIH/NHLBI/Professional Student Short Term Program Stanford University First Year Mentor: Jean Tang, PhD Ervin Epstein, MD
My name is Zhe Zhang and I’ll be a sophomore at Stanford University this fall. Returning for my second year at CHORI, I had a chance to reflect on my experiences that I had last year and I could tell a noticeable difference, mostly in myself. Last year, I had just graduated high school and was heading off to college, and I felt that I might be interested in science, medicine, and engineering. I had a small perspective of the world and the roles that every thing plays. However, after both participating in the program and attending college, I returned with much growth. After taking classes, meeting and talking to bunches of interesting people, and exploring and looking at all the interesting opportunities, I came back with not only interests in the previous, but also in economics, math, and political science. I had expanded my understanding and thinking of the world and how all things interconnect. This brings me to my goal this summer to think about all of my interests and ambitions, and to find possibilities to continue my involvement in them. This summer at CHORI, I wanted a chance to continue my interest and involvement in science and medicine because I hadn’t very much at school. Although I have developed more interests, I do not want to give up science, so my goal is to find a path where I can explore and engage in many different things. CHORI has been a great place where I see how many different aspects are involved at one place and how many different things are available. It has been a great place to learn, explore, and imagine. Thanks to my mentor, Jean Tang, for everything and even just mentoring is a great action in its own. Thanks also to Yaroslav Gelfand, Po Lin So, Ervin Epstein, and Jesse Sandberg, who all contributed greatly to making this experience what it was, and thanks as well everyone involved in setting up and working hard on running this program.
111 Exploring the Effect of Vitamin D for Prevention of BCC in Mice and Humans
Z. Zhang, J. Tang, Y. Gelfand, P.-L. So & E. Epstein Children’s Hospital Oakland Research Institute, Oakland, CA
Introduction: My project was an exploration into translational research by becoming involved in all aspects and actions of the lab. I first began an observational clinical study on vitamin D and basal cell carcinoma (BCC) tumors based on recent in vitro findings of vitamin D’s ability to block the hedgehog pathway that creates the tumors. I also became involved in a basic research project that explores how to prevent BCC tumors in mice. BCC tumors develop due to an inappropriately activated hedgehog pathway, commonly due to a mutation in the Ptch gene. Our laboratory has generated Ptch mutant mice to find new drugs to inhibit BCC. We also have patients who are mutated for Ptch (Basal Cell Nevus Syndrome) who would benefit from new therapies. Vitamin D has been shown to inhibit the hedgehog pathway despite a Ptch mutation. Objectives: To investigate whether vitamin D can prevent BCCs in mice and in humans by inhibiting the hedgehog pathway. Methods: For the observational clinical study, I performed a chart review of 26 Basal Cell Nevus patients that were previously enrolled in a clinical trial. I extrapolated data on their BCC tumor numbers and sunscreen use, and I will compare them to the vitamin D levels in the patient’s frozen serum. This allows me to determine if higher vitamin D levels can help reduce tumor development. Another way examine vitamin D’s impact on the hedgehog pathway is to test if vitamin D can prevent BCCs in mice. I helped to genotype mice to determine which mice contain the correct transgenes to be enrolled in the preclinical trial. We genotyped mice for 3 genetic changes: Ptch+/-, K14 Cre recombinase, and p53 flox/ flox. The Ptch+/- allows development of BCC tumors and the Cre recombinase will delete p53, an important tumor suppressor gene, in K14 basal keratinocytes. These Ptch+/-, K14- Cre, p53 flox/flox mice have reliably developed numerous BCC after ionizing radiation, and serve as an important model to test new anti-BCC agents. We shave the dorsal skin of the mice and topically apply the test drug. The mice are then observed for several months for BCC tumor development. Results: I have successfully genotyped transgenic mice for enrollment in the mouse preclinical trial of vitamin D for BCC prevention and we are currently monitoring for tumor development. I have learned to perform PCR, gel electrophoresis, and drug application. For our clinical research project, I have performed a chart review to generate a cohort of 26 BCNS patients with key clinical information. I am using this cohort to examine if there is association between age, sunscreen use, smoking, and the number of BCC tumors at initial entry into the study. I have collected previously frozen serum samples for vitamin D level measurements. If our hypothesis that vitamin D can prevent BCC tumors is true, I expect that patients with a low BCC tumor burden would have high vitamin D levels.
112 For more information about this program, please contact:
The Summer Student Program Children’s Hospital & Research Center at Oakland 5700 Martin Luther King, Jr. Way Oakland, CA 94609 [email protected]
Or, sign on to: http://www.chori.org/Education/Summer_Internship_Program/announcements.html
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