Research and Development Service (151)

Department of Veterans Affairs

VA Medical Center 601 Highway 6 West

Iowa City, Iowa

Ph: 319-339-7151 www.research.iowa-city.med.va.gov

TABLE OF CONTENTS:

Investigator Title Page

Lee-Ann H. Allen, Ph.D. Interaction of Helicobacter Pylori 1 with Neutrophils

Zuhair K. Ballas, M.D. Cancer Center Support Grant- Flow 1 Cytometry Core

Zuhair K. Ballas, M.D. [Lymphoma Specialized Program of Research 2 (SPORE)] Core 4 –Immunology Core

Zuhair K. Ballas, M.D. Oligodeoxynucleotids as Cancer Immuno- 2 Therapeutic Agents

Zuhair K. Ballas, M.D. Regulation of Human Immune Responses 3

Gail A. Bishop, Ph.D. Regulation of B Lymphocyte Function 4 and B Cell Transformation

Kyle Brown, M.D. Redox Regulation of Hepatic Stellate 4 Cell Activation

A. Brent Carter, M.D. Regulation of Alveolar Macrophage 5 Function in Asbestosis

Mark W. Chapleau, Ph.D. Differential Mechanisms of Baroreflex 5 Dysfunction in Atherosclerosis and Aging Mediated by Reactive Oxygen Species

Mark W. Chapleau, Ph.D. Baroreflex and Autonomic Dysfunction in 6 Atherosclerosis and Hypertension

Research in Progress 2005, VAMC Iowa City, Iowa: Page I Mark W. Chapleau, Ph.D. Oxidant Stress-Induced Autonomic/ 7 Baroreflex Dysfunction in Heart Failure

Joseph J. Cullen, M.D. Role of Antioxidant Enzymes in 7 Pancreatic Cancer

G.M. Denning, Ph.D. Pathogenesis of Pseudomonas 8 aeruginosa-Associated Airway Disease

Gerald F. DiBona, M.D. The Neural Control of Renal Function 9

Joseph S. Dillon, M.D. Dehydroepiandrosterone, a Novel 9 Regulator of Vascular Function

Bradley S. Dixon, M.D. P27Kip Regulation by G protein 10 Receptors in Vascular Cells

Elizabeth H. Field, M.D. CD4+CD25+ Regulation of the IL-2/IL-2R 10 Autocrinel-Paracrine Pathway

F. Jeffrey Field, M.D. Regulation of Intestinal SREBPs by 11 Sterols and Fatty Acids

Kenneth A. Follett, M.D., A Comparison of Best Medical Therapy and 11 Ph.D. Deep Brain Stimulation of Subthalamic Nucleus and Globus Pallidus for the Treatment of Parkinson’s Disease

Sue E. Gardner, Ph.D., The Diagnostic Validity of Three 12 RN Quantitative Swab Techniques

Research in Progress 2005, VAMC Iowa City, Iowa: Page II

Douglas R. Geraets, 80+ Hemorrhagic Cohort 12 Pharm.D. Study

Douglas R. Gereats, The Home INR Study (THINRS) 13 Pharm.D.

Theresa Gioanni, Ph.D. Molecular Determinants of Endotoxin 13 Interaction with Toll-Like Receptors

Thomas H. Haugen, M.D., Viral Gene Expression and Replication in 14 Ph.D. Papillomavirus-Associated Neoplasia

Patrick W. Hitchon, M.D. Motion and Intradiscal Pressure Adjacent to 15 Segment Implanted with Maverick Artificial Disc

Patrick W. Hitchon, M.D. Indications of Anterior and Posterior 15 Cervical Instrumentation

Daniel G. Hug, B.S., Photobiology and Photo Medicine 16 Ph.D. of Urocanic Acid

Gary W. Hunninghake, Human Lung Responses to Respiratory 17 M.D. Pathogens

Gary W. Hunninghake, Role of P13K in Sepsis-Induced Acute 17 M.D. Lung Injury

Gary W. Hunninghake, Regulation of Alveolar Macrophage Function 17 M.D.

Gary W. Hunninghake, Molecular and Cellular Biology of the Lung 18 M.D.

Gary W. Hunninghake, Modulation of Acute Lung Injury 18 M.D.

Research in Progress 2005, VAMC Iowa City, Iowa: Page III Gary W. Hunninghake, Iowa Multidisciplinary Clinical Research 18 M.D. Career Development Program

Jacob Ijdo, M.D., Ph.D. Intracellular Survival of Anaplasma 18 Phagocytophilum in Neutrophilsc

Peter Kaboli, M.D., Randomized Control Trial of Structured 19 M.S. Pharmacy Care in Older Veteran Outpatients

Randy H. Kardon, M.D., Predicting Rehabilitation of Visual 19 Ph.D. Loss in Compressive Optic Neuropathy

Ulla C. Kopp, Ph.D. Afferent Renal Nerves: Role of 20 Neurotransmitter

David J. Kusner, M.D. Regulation of Phagocyte Antimicrobial 21 Activity by Phospholipase D

Steven R. Lentz, M.D. Vascular Dysfunction in 21 Ph.D. Hyperhomocysteinemia and Atherosclerosis

David M. Lubaroff, Ph.D. Summary of VA Research Projects 22

Donald Lund, Ph.D. Effects of a Common Human 23 Gene Variant of Extracellular Superoxide Dismutase on Endothelial Vasomotor Function after Endotoxin in Mice

Rama Mallampalli, M.D. Regulation of Cytidylyltransferase 24 In Fetal Rat Lung

Rama Mallampalli, M.D. TNF-Lipoprotein Control of a Surfactant 24 Enzyme

Research in Progress 2005, VAMC Iowa City, Iowa: Page IV

Rama Mallampalli, M.D. Transcriptional Regulation of a Surfactant 25 Enzyme

Rama Mallampalli, M.D. The CCT Promoter to Drive High-Level 25 Expression of CFTR

James B. Martins, M.D. Purkinje Origin of Ischemic Ventricular 26 Tachycardia

Stephen E. McGowan, Septal Fibroblast Proliferation and 27 M.D. Migration during Alveolar Growth

William M. Nauseef, M.D. The Cell and Molecular Biology of 27 Human Neutrophils

Christine L. Oltman, Ph.D. Role of Arachidonate 12- 28 Lipoxygenase in Porcine Coronary

Christine L. Oltman, Ph.D. Progression of Vascular Dysfunction 29 with Metabolic Syndrome: Role of Oxidative Stress

Gary E. Rosenthal, M.D. Center for Research in the 29 Implementation of Innovative Strategies in Practice—CRIISP

James Rossen, M.D. Clinical Outcomes Utilizing 30 Revascularization and Aggressive Drug Evaluation

Anne Sadler, Ph.D., R.N. Sexual Violence Exposures and Women 31 Veteran’s Gynecologic Health

Research in Progress 2005, VAMC Iowa City, Iowa: Page V

Mary S. Vaughan Estimating the Magnitude 31 Sarrazin, Ph.D. of Unmeasured Risk in VA Patients

Warren Schmidt, M.D., A Multi-center Trial to Evaluate the 32 Ph.D. Epidemiology, Natural History, and Treatment Response of Hepatitis C in the United States Veterans Population

John P. Schneider, M.D. The Effect of Clinical Practice 33 Guidelines on Costs

Konrad Shulze, M.D. Mechanics of Stomach and Intestine 33 FRCP

William L. Sivitz, M.D. Dr. Sivitz’s Research Project 34

Jack Stapleton, M.D.; GB Virus C-HIV Interactions 34 Jinhua Xiang, M.D.

John B. Stokes, M.D. Long-term Regulation of Sodium 35 Channels

Gail Takahashi, Ph.D.; A Long Term Follow-Up of 35 Janet Shanks, Ph.D.; Patients in the NIDCD/VA Hearing Micheal Crum, Ph.D.; Aid Clinical Trial Richard Wilson, Ph.D.; Barbara Peek, Ph.D.; Stephen Fausti, Ph.D.; Gabrielle Saunders, Ph.D.; Charles Martinez, Ph.D.; Douglas Noffsinger, Ph.D.

Gail Takahashi, Ph.D. Clincial vs. Real World Performance 36 with Telecoils

Research in Progress 2005, VAMC Iowa City, Iowa: Page VI Gail Takahashi, Ph.D. Perceived Hearing Difficulties and 36 Hearing Aid Satisfaction in Veterans Vs. Significant Others

William T. Talman, M.D. Parasympathetic Influences on Cerebral 37 Blood Flow

Christie P. Thomas, M.D. Regulation and Function of Nedd4-2 Isoforms 38 in Epithelial Na+ Transport

Bonnie Wakefield, Ph.D. Evaluating Telehealth Home Care 38 R.N. for Elderly Veterans with Congestive Heart Failure

Michael Wall, M.D. Mechanisms of Perimetric Variability 39

Neal Weintraub, M.D. Inflammatory Mechanisms in Vascular 39 Disease

Jerrold Weiss, Ph.D. Neutrophils and Bacterial Phospholipid 40 Degradation

Jerrold Weiss, Ph.D. Microbicidal Activity of Leukocytes: 41 Active Factors

John Wemmie, Ph.D. Exploring the Role of ASIC1 in Fear 42

Mary E. Wilson, M.D. Protective Immunity to 42 Leishmania chagasi

Mary E. Wilson, M.D. Immunogenetics of Leishmania 43 chagasi Infection

Mary E. Wilson, M.D.; The Major Surface Protease 43 John E. Donelson, Ph.D. of Trypanosomatid Protozoa

Research in Progress 2005, VAMC Iowa City, Iowa: Page VII Mary E. Wilson, M.D. Interactions of Leishmania chagasi 44 with Host Macrophages

Mary E. Wilson, M.D.; Training in Mechanisms of Parasitism 44 John E. Donelson, Ph.D. NIH/NIAID T32 (Training Grant)

Mary E. Wilson, M.D.; International Maternal and Child Health 45 Jeffrey Murray, M.D. Research and Training NIH Fogarty Foundation T32 (Training Grant)

Mary E. Wilson, M.D.; NIH/NIAID Tropical Medicine Research 45 Edgar Carvalho, M.D., Center P50: Pathogenesis of Leishmaniasis: Ph.D. Host, Parasite and Vector Project 1: “Host and Parasite Factors in Mucosal Leishmaniasis”

Patricia L. Winokur, M.D. Antibiotic Resistance in Food Animals and 45 Humans

Catherine Woodman, Fibromyalgia in Persian Gulf War Veterans: 46 M.D. A Family Study

Chaoqun Yao, M.D., Expression of the Major Surface Protease 47 Ph.D. of Leishmania Chagasi

Mark A. Yorek, Ph.D. Pathology of Type 2 Diabetes 48

Mark A. Yorek, Ph.D. Vascular Disease in Diabetic 49 Neuropathy

Mark A. Yorek, Ph.D. Effect of Angiotensin Converting Enzyme 49 Inhibitor, and/or Angiotensin 2 Receptor Antagonist on Diabetic Neuropathy

Asgar Zaheer, Ph.D. Glia Maturation Factor as a Signaling 50 Molecule

Research in Progress 2005, VAMC Iowa City, Iowa: Page VIII

Nicholas ZavaZava, M.D., Immune-Privilege and Tolerogenicity 50 Ph.D. of Stem Cells

Nicholas ZavaZava, M.D., Potency of Embryonic Stem Cells to Induce 51 Ph.D. Graft Tolerance

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the same time promotes damage to normal stomach tissue. Additional data indicate that NADPH oxidase targeting is affected by Principal Investigator: one or more factors on the surface of H. Lee-Ann H. Allen, Ph.D. pylori and that enzyme targeting is manipulated by alterations in signaling Project Title: pathways that include protein kinase C, Interaction of Helicobacter Pylori phosphoinositide 3-kinase and tyrosine with Neutrophils kinases of the src family. Understanding how H. pylori alters the immune response may lead to new treatments for this infection and thereby reduce the significant Summary: impairment and death associated with peptic ulcers and stomach cancer. Helicobacter pylori is a type of bacteria that lives in the stomach of about half of all MeSH Terms: neutrophil, ulcer, humans. It has been known for over ten inflammation, phagocytosis, reactive years that infection with these bacteria oxygen species causes inflammation and in some cases this progresses to stomach ulcers, intestinal ulcers or more rarely to stomach cancer. The human immune system responds to H. Principal Investigator: pylori and these bacteria cause a Zuhair K. Ballas, M.D. pronounced accumulation of white blood cells called neutrophils at the site of Project Title: infection. Normally, neutrophils are able to Cancer Center Support Grant ingest and kill microorganisms and this Flow Cytometry Core function of neutrophils is essential to normal functioning of the immune system. Interestingly, during H. pylori infection neutrophils accumulate in proximity to the Summary: infecting bacteria but are not able to kill these microbes and the infection persists for many years. We are exploring at the The Flow Cytometry Facility is a shared molecular level how H. pylori evades killing resource available to all UICC investigators. by neutrophils. Toward this end we have The overall goal of the Flow Cytometry shown that ulcer-causing strains of H. pylori Facility is to provide investigators with the are eaten by neutrophils and strongly ability to analyze and sort cell populations activate the cells to produce toxic reactive using multiple parameters for a broad range oxygen species (ROS) including superoxide of research applications. Consultation is anions and hydrogen peroxide. Under provided as needed with the Director, normal circumstances, these ROS are Technical Director, and other support concentrated in the vicinity of the ingested personnel. Specifically, the facility: 1. bacterium where they contribute directly to provides advice and technical expertise for analysis and/or separation of cell bacterial killing. However, we have shown populations based upon their ability to bind that H. pylori disrupt targeting of the NADPH fluorochrome-conjugated antibodies. 2. Aids oxidase enzyme in neutrophils which investigators in the use of the FACS IV and generates the ROS. Consequently, the Epics 753 flow cytometers for analysis ROS do not accumulate nearby ingested (FACS IV and Epics) and sorting (Epics) for bacteria but rather are generated in the cell populations. 3. Trains investigators and extracellular space where they can damage their laboratory personnel in the use of the normal stomach tissue. By this unusual FACSCAN for analysis of cell populations. 4. mechanism H. pylori escapes killing and at Trains and assists investigators in the use of

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the facility’s computers for the analysis and proposed in the UI/MC Lymphoma SPORE. output of data accumulated during the flow More straightforward immunologic assays of cytometry studies. 5. Aids investigators in will be done in the individual investigators’ the use and/or development of new laboratories. As new approaches are fluorochromes for analysis of cell developed, the Immunology Core will serve populations. to spearhead the development of these assays even for the experienced MeSH Terms: immunity, host defense, investigators. Moreover, if one of the immunology, NK cells, T cells, flow projects finds a need for an immunology cytometry, diagnostic immunology assay or procedure, the Core will be ready to help the investigators test their modified hypothesis in an efficient, accurate, and timely manner. This proposal describes the Principal Investigator: various assays and procedures that will be Zuhair K. Ballas, M.D. offered. A governance structure for evaluation of new requests for the use of the Project Title: Core and for evaluating the efficacy of the [Lymphoma Specialized Program of Core itself is detailed as well. Research (SPORE)] Core 4 –Immunology Core MeSH Terms: immunology, NK cells, T cells, flow cytometry, tumor immunology, diagnostic immunology

Summary:

Principal Investigator: Many of the research projects outlined in Zuhair K. Ballas, M.D. the UI/MC Lymphoma SPORE, including both preclinical and clinical experiments, require sophisticated correlative Project Title: immunological studies. Many of these Oligodeoxynucleotides as Cancer methodologies are complex and require a Immunotherapeutic Agents considerable investment in time, personnel, and funds in order to establish these techniques in any one laboratory. Moreover, Summary: many of these methodologies require rigorous quality controls and, preferably, Innate immunity, by virtue of its pattern should employ Good Lab Practice (GLP). recognition receptors, is perfectly poised for Utilizing new methodology and, more the recognition of danger signals. Bacterial importantly assuring reproducibility and infection is one such danger signal. It is now credibility of any given assay is a major well established that bacterial DNA can burden for any individual laboratory. The serve as a danger signal to mammalian Immunology Core was designed to greaty hosts. Fragments of bacterial DNA, or facilitate the adoption and successful synthetic oligodeoxynucleotides (ODN), that execution of new methodology. Ideally, an contain a particular motif, called CpG, have Immunology Core should be in a laboratory been established as potent stimulators of that practices GLP, which employs innate immunity and certain components of extensive quality controls and which has a adaptive immunity as well. We have well-defined procedure for troubleshooting demonstrated that some CpG ODN are each and every assay. These characteristics potent stimulators of natural killer (NK) cells are examined before the College of in humans and in mice. We have identified American Pathologists (C.A.P.) can certify a two classes of CpG ODN which differ in their laboratory. The proposed Immunology Core immune effects. CpG-X are potent fulfills the above criteria and several of the stimulators of NK cells; CpG-B are potent proposed assays are already approved by stimulators of cytokine secretion and of B C.A.P. The immunology core will be a cell activation. This proposal is designed to resource for the more demanding assays

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delineate the different effector mechanisms MeSh Terms: immunology, NK cells, flow responsible for the anti-tumor properties of cytometry CpG-X and CpG- B invitro and in vitro. Futher impetus is given to the importance of these investigations by our recent results in a phase I human study demonstrating that Principal Investigator: CpG ODN augment NK and antibody- Zuhair K. Ballas, M.D. dependent cell mediated cytotoxicity (ADCC) in lymphoma patients. Methods: Project Title: Murine lymphocytes will be used in vitro. In Regulation of Human Immune Responses vivo, the ability of ODN to accelerate tumor rejection will also be examined as above. Research Plan: 1. evaluate the contribution of various immune effector mechanism to Summary: the anti-tumor effect of CpG ODN. A. Evaluate the contribution of NK, NKT, T The immune system is believed to be cells, B cells and cytokines B. determine the important in combating infections and in relative contribution of the killing vs the protecting against cancer. However, we now cytokine secreting capacity of NK cells to the know that the immune system is regulated anti-tumor effect C. determine the by a number of checks and balances within parameters that would allow the prediction the body. It has recently become apparent of which CpG motif works for which tumor D. that a mere assessment of lymphocyte Determine the synergy of COX-2 inhibitors numbers is not sufficient. One can have the with CpG in tumor immunotherapy Analyze right number, but the function may be the apparent paradox of dendritic cells (DC): skewed towards a detrimental outcome. DC are needed for NK stimulation by CpG With the proliferation of FDA- approved but DC are susceptible targets to CpG- immunomodulators one can now offer activated killer lymphocytes E. Examine the innovative therapy. Before one can consider relative contribution of co-stimulatory such therapy however, one should be able molecules expression vs cytokine secretion to document the functional status of by various DC subsets with particular lymphocytes. This is done by obtaining emphasis on the role of IFN=\-X/B and its blood from an individual; serum and receptors F. Determine the CpG motif, lymphocytes will be purified from one cytokines, precursor and effector cells donated sample. Lymphocyte function will needed to induced CpG-activated killers be examined, including proliferation, killing capable of killing DC, as well as NK- activity, cytokine pattern secretion. The resistant tumors. Findings: We have recently serum will be used to measure cytokine shown that CpG ODN are very effective as levels. Normal donors will be used in order monotherapeutic agents for two murine to establish a reference range. Patients will tumors: B16 melanoma and EL4 lymphoma. be asked to participate in order to determine CpG-X was effective in the B16 but not the an immunological profile for each clinical EL4 model while CpG-B was effective in the presentation. Objective: Determine novel EL4 but not the B16 model. Moreover, it immunological abnormalities in humans. appeared that NK cells were necessary and Research plan: Assess lymphocyte sufficient for the therapeutic effect in B16 responses in vitro and measure serum while NK cells were necessary but not cytokine levels in normals and patients. sufficient for EL4. More recent findings Methods: Obtain serum and heparinized suggested that the location of the tumor may blood. Measure serum cytokines by ELISA. also play an important role in determining Stimulate lymphocytes with antigens and the optimal CpG motif as well as the relative mitogens; assess their proliferation and their contributions of NK and T cells. These ability to secrete various cytokines. Findings: findings have a high clinical impact as they Reference ranges have been established for will guide the design of phase II/III studies in several tests we can now start running patients with lymphoma and melanoma. patient samples as part of the diagnostic workup for immune system disorders. For the purpose of quality control, we will

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continue to need to obtain samples from lymphomas. As >90% of the world normal donors in order to determine whether population is latently infected with EBV by to accept or reject the results of the patient adulthood, this malignancy is a major samples. medical complication in patients whose immune systems are suppressed by HIV MeSH Terms: immunology, NK cells, T infection, or treatment with post-transplant cells, flow cytometry, diagnostic anti-rejection drugs. We are in the process immunology, immunodeficiency of learning the details of how the LMP1 abnormal signaling pathway differs from the normal CD40 signaling pathway. It is hoped that our findings will be applicable to design Principal Investigator: of more effective therapy for B cell Gail A. Bishop, Ph.D. lymphoma and other immunological disorders. Project Title: Regulation of B Lymphocyte Function and B MeSH Terms: immunology, Cell Transformation rheumatology, lymphomal, virology, antibody response, B cells, transmembrane signaling, autoimmunity, Summary: molecular biology.

My laboratory studies the regulation of B lymphocytes, the white blood cell which Principal Investigator: produces antibodies. In addition to their Kyle Brown, M.D. critical role in antibody production, B lymphocytes serve to make foreign Project Title: substances recognizable to the other major Redox Regulation of Hepatic Stellate Cell type of lymphocyte, T cells, which mediate Activation cellular immunity. The process of B cell activation must be very precisely regulated, as production of antibodies to normal "self" components can lead to autoimmune Summary: diseases such as rheumatoid arthritis, Systemic Lupus Erythematosis, and Hepatic stellate cells are normal residents of diabetes. Uncontrolled B cell activation can the liver. During liver injury, these cells also lead to the development of lymphomas become “activated” and begin to produce and leukemias. The focus of my research is large amounts of connective tissue proteins B cell membrane molecules which transmit that result in the formation of scar tissue. regulatory signals to the inside of the cell to Over time, the accumulation of excess scar control its activation. One of the major tissue results in cirrhosis. Understanding molecules we study is called CD40, and this how liver injury causes stellate cell activation receptor delivers signals critical to antibody and how activation leads to connective production, the development of the recall tissue production may lead to treatments to response (which is mimicked by prevent or slow the development of cirrhosis vaccination), and the ability of B cells to in patients with chronic liver diseases for stimulate T cells. People with defects in the which no other treatment is currently CD40 signaling pathway suffer from an available. Several lines of evidence have immunodeficiency disease which results in suggested that free radicals (a type of highly susceptibility to recurrent infections and reactive molecule) may play a role in these decreased lifespan. Interestingly, a viral events. We hypothesized that stellate cells protein produced by the Epstein-Barr virus may themselves have a mechanism to mimics the CD40 signaling pathway in B produce a certain type of free radical that cells, but in an uncontrolled fashion. This sends signals that make the cells behave in protein, called LMP1, is necessary for the an activated fashion. This mechanism development of EBV-associated involves a family of enzymes called NADPH

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oxidases. These enzymes have multiple macrophages to TNF-α release and reactive biological functions including important roles oxygen species generation is not known. in defense against bacteria and other We have found that adequate steady-state pathogens. They have also recently been levels of hydrogen peroxide (H2O2) are recognized to be important in disease states necessary for TNF-α production in such atherosclerosis. We have confirmed macrophages. These steady-state levels of that human stellate cells express multiple H2O2 are also dependent on the activity of components of NADPH oxidases using RT- peroxide-removing enzymes, such as PCR. Identification of NADPH oxidases in catalase. Thus, it appears that the balance stellate cells provides an opportunity to between oxidant-producing enzymes and determine the importance of these enzymes antioxidant enzymes regulates TNF-α in stellate cell activation. If NADPH production in macrophages. Although the oxidases play an important role in this studies in this proposal relate to asbestosis, process, drugs that inhibit these enzymes they are novel studies that also provide may be useful slowing or preventing important basic clues to understand cirrhosis. macrophage cytokine gene regulation and the functional difference between mature MeSH Terms: cirrhosis, fibrosis, free and immature macrophages. radicals, NADPH oxidase, oxidative stress, superoxide anion MeSH Terms: macrophages, cytokines, inflammation, respiratory diseases

Principal Investigator: A. Brent Carter, M.D. Principal Investigator: Mark W. Chapleau, Ph.D. Project Title: Regulation of Alveolar Macrophage Function Project Title: in Asbestosis Differential Mechanisms of Baroreflex Dysfunction in Atherosclerosis and Aging Mediated by Reactive Oxygen Species Summary:

Asbestosis, which is an important cause of Summary: pulmonary fibrosis, results from a high occupational exposure to asbestos Aging and atherosclerosis are both (minimum fiber dose of 25-100 associated with decreased baroreflex fibers/ml/year). Although standards sensitivity. Impairment of this important regarding handling asbestos have changed, blood pressure regulating mechanism can it is estimated that asbestos-associated predispose individuals to fainting, high blood deaths in the United States may exceed pressure, and/or possibly dangerous heart 200,000 by the year 2030. The lungs of rhythms. Previous studies have suggested patients with asbestosis contain a mixture of that decreased baroreflex sensitivity in both mature and immature macrophages. This aging and atherosclerosis is simply an mixture results from the recruitment of blood indirect result of structural changes in the monocytes to the sites of fiber deposition. large arteries that contain the baroreceptor Alveolar macrophages obtained from the nerve endings. The structural changes in lung of patients with asbestosis the arteries create less distensible, stiffer spontaneously release cytokines (TNF-α), blood vessels and are relatively difficult to and asbestos fiber spontaneously catalyze reverse. the generation of reactive oxygen species. Studies indicate that inhibition of TNF-α and Aging and atherosclerosis are also both reactive oxygen species prevent the associated with oxidative stress. In this development of pulmonary fibrosis. The project we test the hypothesis that oxidative relative contribution of mature and immature stress in aging and atherosclerosis may

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impair baroreflex sensitivity independent of the structural vascular changes, and that the Principal Investigator: impairment may be reversible with Mark W. Chapleau, Ph.D. appropriate antioxidant treatment. Our studies indicate that baroreflex control of heart rate (conscious mice), and baroreflex Project Title: control of sympathetic nerve activity and Baroreflex and Autonomic Dysfunction in blood pressure (anesthetized mice) are Atherosclerosis and Hypertension impaired in both young mice with high blood cholesterol, and old mice with normal cholesterol. The impaired reflex function is partially reversed by administration of an Summary: antioxidant drug in the drinking water, and by increasing endogenous expression of antioxidant molecules at the site of Patients with cardiovascular disease commonly have both high levels of baroreceptor innervation using gene transfer. Interestingly, preliminary findings cholesterol in the blood and high blood suggest that the source of oxidative stress pressure (hypertension). Both high cholesterol and hypertension impair the may differ in aging and atherosclerosis. Oxygen-derived free radicals generated in baroreceptor reflex (an important blood the arterial wall in atherosclerosis appear to pressure regulating mechanism) and alter how the autonomic nervous system controls inhibit baroreceptor sensitivity in a paracrine manner, i.e. through cell-to-cell cardiovascular function− but the transmission. In contrast, in aging, oxidative mechanisms are not well understood. In stress in the baroreceptor neurons addition, the majority of previous basic themselves may impair their function. The research studies have examined the effects effects of oxygen-derived free radicals on of these two disorders separately. Very little baroreceptor sensitivity appear to result from is known regarding the combined effects of modulation of potassium and sodium ion high cholesterol and hypertension on channels on the baroreceptor nerve regulation of blood pressure and heart rate. endings. By breeding together mice with high Atherosclerosis is extremely common in cholesterol (apolipoprotein E deficient mice) elderly humans. Therefore, our results may and mice with hypertension (renin- provide new insights into the interactions angiotensinogen transgenic mice), we between effects of aging and atherosclerosis created a new type of mice that have both on baroreflex and autonomic nervous high cholesterol and high blood pressure. system function with important clinical We discovered that brief treatment of either implications for patients. group of mice (high cholesterol or hypertension) with an antioxidant drug for MeSH Terms: aging, atherosclerosis, only one week improves baroreflex pressoreceptors, sympathetic nerve sensitivity and autonomic nervous system activity, nodose ganglia, oxygen derived control of the heart. Our results also free radicals, potassium channels, indicate that in general, the combination of sodium channels high cholesterol and high blood pressure does not dramatically worsen the blood pressure regulation, at least in young mice. The combination of high cholesterol and hypertension did reduce the magnitude of the changes in heart rate that normally occur between day and night. The results of our study indicate that oxidative stress (the production of oxygen free radicals) is an important cause of decreased baroreflex sensitivity and abnormal cardiovascular regulation in mice with high cholesterol and hypertension. The

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rapid improvement in function after the baroreflex/autonomic dysfunction seen beginning the antioxidant therapy suggests in heart failure. An important goal of the that the defect is reversible, and that study is to determine whether treatment of antioxidants may provide benefit to patients the mice with different types of antioxidants with cardiovascular disease. improves baroreflex and autonomic function in the mice with heart failure. The MeSH Terms: blood pressure, preliminary results of our investigation cardiovascular disease, pressoreceptors, suggest that oxidative stress may alter sympathetic nerve activity, antioxidants, function by acting at multiple sites including oxygen free radicals, atherosclerosis, the baroreceptor nerves that sense changes hypertension in blood pressure in large arteries, the central nervous system (brain), and the sympathetic nerves that influence heart and kidney function. Based on our results and the work of others, we believe that improved Principal Investigator: baroreflex and autonomic function during Mark W. Chapleau, Ph.D. antioxidant therapy has the potential to reduce the incidence of abnormal heart rhythms and sudden cardiac death in patients with high cholesterol and Project Title: hypertension. Oxidant Stress-Induced Autonomic/Baroreflex Dysfunction in Heart MeSH Terms: myocardial infarction, heart Failure failure, pressoreceptors, sympathetic nerve activity, antioxidants, sudden cardiac death, atherosclerosis, hypertension Summary:

High levels of cholesterol in the blood and Principal Investigator: high blood pressure greatly increase the risk Joseph J. Cullen, M.D. and incidence of myocardial infarction (heart attack) and subsequent development of Project Title: heart failure. Chances for survival in heart Role of Antioxidant Enzymes in Pancreatic failure are poor with about half of the Cancer patients dying from irregular heart rhythm and sudden cardiac death. Decreased baroreflex sensitivity and abnormal autonomic nervous system control of the Summary: heart is associated with the irregular heart th rhythm and death, and is predictive of these Pancreatic cancer is the 4 most common adverse outcomes. cause of cancer death in the U.S. and Therefore, we have begun experiments to increasing in incidence. NAD(P)H:quinone investigate the effects of heart attack and oxidoreductase (NQO1), which detoxifies heart failure on baroreflex sensitivity and quinone xenobiotics, serves as a protective autonomic control of the heart in normal mechanism against reactive oxygen species healthy mice and, most importantly, in mice (ROS), and is elevated 12-fold in pancreatic with high cholesterol and high blood cancer. Using NAD(P)H, NQO1 directly pressure. The effects of a heart attack and reduces the quinone form to the subsequent development of heart failure on hydroquinone form, thus bypassing the baroreflex sensitivity and autonomic control reactive semiquinone intermediate. have not been previously studied in animals Dicumarol inhibits NQO1, causing ROS with high cholesterol and hypertension. We production when reactive semiquinones propose that the presence of high generate a redox cycle, resulting in cholesterol and hypertension will exaggerate superoxide formation. The objective of this

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application is to determine if NQO1 can be used as a specific target for therapeutic Principal Investigator: purposes in pancreatic cancer. The central G.M. Denning, Ph.D. hypothesis of the application is that dicumarol, a drug that selectively inhibits the two-electron reductase NQO1, will be an Project Title: effective agent for use against pancreatic Pathogenesis of Pseudomonas aeruginosa- cancer cells that specifically overexpress Associated Airway Disease this bioactivating enzyme. Our hypothesis has been formulated on the basis of strong preliminary data demonstrating that Summary: dicumarol induces cytotoxicity and oxidative stress in pancreatic cancer cells that Pseudomonas aeruginosa (P. aeruginosa) is overexpress NQO1. The rationale for the a bacterium that causes both acute and proposed research is that if dicumarol chronic airway disease in susceptible patient induces cytotoxicity in pancreatic cancer populations. Among the VA patient cells that overexpress NQO1, this would population, P. aeruginosa is a common represent a new and innovative approach to cause of hospital-acquired pneumonia the treatment of pancreatic cancer. We plan (mortality rate of greater than 75%). When to test our hypothesis and accomplish the P. aeruginosa colonizes the airway, the objective of this application by pursuing the bacterium secretes numerous products, following three specific aims: many of which can alter the host immune 1) Determine if dicumarol selectively and inflammatory responses. Our studies induces cell death in human are designed to determine how P. pancreatic cancer cells that are aeruginosa and its secretory products cause known to overexpress NQO1. an excessive inflammatory response that 2) Determine if superoxide and/or leads to airway damage in the P. hydrogen peroxide mediate the aeruginosa-infected lung. For these studies, cytotoxicity and metabolic oxidative we use primary human cells, laboratory and stress seen with dicumarol clinical strains of the bacterium, as well as treatment in pancreatic cancer cells. an animal pneumonia model. In addition, we 3) Determine if the therapeutic efficacy have been studying the ability of of dicumarol can be enhanced by antioxidants and other chemicals to inhibit increasing levels of superoxide the pro-inflammatory effects of the and/or hydrogen peroxide. bacterium. Understanding the mechanisms The proposed work is innovative because it by which the bacterium causes airway investigates if dicumarol inhibition of NQO1 damage and identifying interventions that can be used for therapeutic purposes in prevent this damage will contribute to pancreatic cancer. Additionally, it takes improved treatment for patients with P. advantage of techniques in free radical aeruginosa-associated pneumonia. biology, which are readily available in our laboratory. These results will be significant MeSH Terms: pseudomonas aeruginosa, because the proposed experiments will bacterial pneumonia, inflammation, provide evidence supporting the rational cytokines, antioxidants design of combined modality therapy for pancreatic cancer. It is our expectation that the results will advance the field of free radical biology and facilitate the discovery and development of targeted therapeutics, while developing and validating preclinical models of human pancreatic cancer

MeSH Terms: pancreatic cancer, reactive oxygen species dicumarol, NAD(P) H: quinone

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medications to decrease their risk of vascular Principal Investigator: events. Because of the high prevalence of vascular disease a search for novel, cost Gerald F. DiBona, M.D. effective, protective agents is justified.

Plasma DHEA levels fall sharply with aging. Project Title: Atherosclerotic diseases are inversely The Neural Control of Renal Function associated with concentrations of DHEA. DHEA has beneficial effects on vascular function in animals, and use in humans Summary: improves vascular function. Until recently, no mechanism of action was identified for the The kidney receives an abundant supply of beneficial vascular effects of DHEA. This nerves from the nervous system. These lack of understanding hindered further nerves control important aspects of overall evaluation of DHEA in vascular diseases. kidney function. They regulate the amount of We recently characterized a vascular DHEA blood flow the kidney receives, the balance receptor, which causes increased nitric oxide between absorption and secretion of fluid production in blood vessels. Nitric oxide is that determines the final quantity and quality crucial for healthy blood vessels. Thus, of the urine produced and the secretion of DHEA may represent a novel agent that important hormonal substances that can benefits human vascular function. influence blood pressure. Increases in Our long range goal is to develop new drugs activity in the nerves going to the kidney to improve cardiovascular illness in the VA negatively influence overall kidney function. population. The objective of this application In diseases such as high blood pressure and is to determine how DHEA improves heart failure, this results in excessive vascular function. Our hypothesis is that retention of salt and water leading to DHEA acts on a receptor in endothelial cells excessive collection of fluid in the legs, to enhance nitric oxide production. Our abdomen and lungs. This research project specific aims are as follows: seeks to define the exact mechanisms Aim #1: Isolate, sequence and clone the whereby increased nerve activity to the vascular DHEA receptor that we have kidneys causes these abnormalities in pharmacologically characterized. overall kidney function. It also seeks to Aim #2: Determine what signals DHEA define the exact mechanisms whereby nerve initiates within endothelial cells to cause an activity to the kidneys is increased in increase in nitric oxide. diseases such as high blood pressure and Once we have isolated the DHEA receptor heart failure. and defined its signaling system we will be able to screen for more potent variants of MeSH Terms: kidney, nerves, DHEA that will activate the DHEA receptor hypertension, heart failure, sodium. pathway. These agents will then be used in human trials to see how well they improve

vascular function. Furthermore, we will be able to design compounds to specifically Principal Investigator: target aspects of this pathway and evaluate Joseph S. Dillon, M.D. the role of this steroid in human vascular complications. We will be able to evaluate Project Title: the changes in the DHEA hormone and Dehydroepiandrosterone, a Novel Regulator receptor throughout the aging process, and of Vascular Function evaluate whether mutations in the DHEA receptor gene are associated with vascular dysfunction.

Summary: MeSH Terms: dehydroepiandrosterone, prasterone, cell surface receptor, G- Heart and vascular diseases are major protein, vascular endothelium, nitric causes of illness in the VA population. oxide Increasing numbers of veterans are taking

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will lead to new targets for therapies to treat vascular diseases. In addition, we are developing a better understanding of the Principal Investigator: specific actions of bradykinin receptors on Bradley S. Dixon, M.D. vascular cells to ultimately test the hypothesis that pharmacological activation Project Title: of these receptors may be a potential new P27Kip Regulation by G protein Receptors therapy for vascular disease. in Vascular Cells MeSH Terms: arterial occlusive diseases; muscle, smooth; vascular; cell Summary: cycle; tumor suppressor proteins; cyclin dependent kinases; receptors, bradykinin; receptors, G-protein-coupled; Atherosclerosis is a process whereby signal transduction. arteries become thickened and narrowed so they do not provide adequate blood supply to tissues, most critically the heart and brain. This leads to clinical diseases such as heart Principal Investigator: attacks, strokes and limb amputations that Elizabeth H. Field, M.D. are the leading cause of morbidity and mortality in developed nations and are major problems affecting our older veteran Project Title: population. A central component of this CD4+CD25+ Regulation of the IL-2/IL-2R process is the proliferation of vascular Autocrinel-Paracrine Pathway smooth muscle cells within the wall of the artery that contributes to arterial narrowing. Work from this lab and others has Summary: discovered that bradykinin receptors, which belong to a large family of membrane receptors known as G protein-coupled CD4+CD25+ T cells are important cells receptors, are present on vascular smooth because they play a major role in controlling muscle cells and can inhibit the proliferation the level of activity of the immune system. A of vascular smooth muscle cells in culture proper balance of CD4+CD25+ cells is vital and the narrowing of arteries after vascular for health. Too many CD4+CD25+ T cells injury in intact animal models. We have may result in the ineffective elimination of discovered that bradykinin inhibits vascular cancer cells, viruses, such as HIV, or smooth muscle cell proliferation by parasites. On the other hand, people who upregulation of a cell cycle inhibitory protein suffer from certain autoimmune or allergic known as p27Kip. p27Kip is a critical protein diseases have too few CD4+CD25+ cells or that acts to inhibit cell proliferation by dysfunctional CD4+CD25+ cells compared inhibiting the action of enzymes (i.e. to healthy individuals. CD4+CD25+ cells kinases, which phosphorylate and change inhibit the function of conventional T cells by the activity of other proteins) that are a cell:cell contact dependent process that responsible for promoting DNA synthesis requires IL-2 and the receptor for IL-2 (IL- needed for cell proliferation. Other important 2R). The mechanism by which IL-2 and IL- G protein coupled receptors known to be 2R mediate the inhibition is poorly involved in vascular disease, such as understood. We have generated a angiotensin II also regulate the expression CD4+CD25+ hybridoma cell that exhibits of p27Kip. The goal of this research is to similar function to natural CD4+CD25+ T understand the cellular signaling pathways cells. Studies of the CD4+CD25+ inhibitory by which these G protein coupled receptors hybridoma as well as natural CD4+CD25+ T regulate the expression and function of cells suggest a model in which the p27Kip. We anticipate that a better CD4+CD25+ T cells take up IL-2 from understanding of the cellular pathways by conventional T cells via a direct cell:cell which hormones regulate cell proliferation contact process involving IL-2R (CD25). The research examines the biology of the IL-

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2/IL-2R in CD4+CD25+ cells and cholesterol synthesis, we are getting a better conventional T cells during the regulation of understanding of how dietary fat and an immune response. Understanding the cholesterol regulate these important biology of IL-2/IL-2R interactions between intestinal proteins which, in turn, regulate CD4+CD25+ regulatory cells and intestinal cholesterol trafficking and conventional T cells will help identify ways to eventually cholesterol blood levels. enhance or prevent the function of CD4+CD25+ T cells. Controlling the MeSH Terms: cholesterol, fatty acids, function of CD4+CD25+ cells would benefit intestines, Ca-Co-2 cells not only those patients with autoimmune or allergic diseases, but patients with cancer or chronic infections as well. Principal Investigator: MeSH Terms: immunology, tolerance, Kenneth A. Follett, M.D., Ph.D. CD4 cell, cytokines, receptor, IL-2 Project Title: A Comparison of Best Medical Therapy and Deep Brain Stimulation of Subthalamic Principal Investigator: Nucleus and Globus Pallidus for the F. Jeffrey Field, M.D. Treatment of Parkinson’s Disease

Project Title: Regulation of Intestinal SREBPs by Sterols Summary: and Fatty Acids

Parkinson’s Disease (PD) is a slow progressive disease, meaning symptoms Summary: become worse over time. Approximately one million people in the United States A high cholesterol level is a significant risk currently live with the illness. Parkinson’s factor for coronary heart disease. disease affects nerve cells (neurons) in the Cholesterol absorbed from our diet and part of the brain that control muscle newly synthesized cholesterol from the movement, leading to a variety of symptoms intestine contribute to the cholesterol found such as tremor, rigidity, difficulty and in our blood. Thus, the intestine is important slowness in movement and problems with in controlling how much cholesterol enters balance and coordination, which can result our body. In our research, we are in significant disability and morbidity. PD investigating the regulation of intestinal patients may have persistent disabling cholesterol synthesis by small proteins that symptoms even when treated with the best alter genes involved in cholesterol trafficking medication regimens available, or they may in the intestine. These small proteins are suffer intolerable medication side effects that called sterol regulatory binding proteins –1 limit effective treatment of the disease. and –2 (SREBPs). We are studying how Many of these individuals who have dietary cholesterol and different fats regulate symptoms that are refractory to maximum their activity. We are using several different medical management may be candidates for ways to study this, such as cell culture surgical therapy. For most of the past techniques (CaCo-2), human intestinal century, surgeons have worked to develop explants, and animal models. In brief, we and refine surgical approaches to treating have found that feeding cholesterol to PD. A growing number of studies indicate hamsters increases intestinal SREBP-1c but that deep brain stimulation (DBS) in areas of has no effect on SREBP-2. Feeding a diet the brain called the globus pallidus (GPI) or enriched in polyunsaturated fatty acids, subthalamic nucleus (STN) may relieve or decreases SREBP-1 and fatty acid reduce symptoms of PD but it is not known synthesis in the intestine but has no effect whether the clinical benefit is superior to that on SREBP-2. In cell culture experiments, achieved with comprehensive (best) medical changes in cholesterol absorption and therapy, whether the clinical benefit is

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maintained, or which brain structure (i.e., technique involves rotating a culture swab globus pallidus or subthalamic nucleus is the over the cleansed wound bed surface in a best site for stimulation. 10-point Z-track fashion. Levine’s technique DBS is a promising therapy for PD. involves rotating a culture swab over a Whether DBS is superior to comprehensive cleansed 1-cm2 area near the center of the (best) medical therapy, whether the wound using sufficient pressure to extract effectiveness of DBS differs by site or wound fluid from deep tissue layers (Levine, whether some subjects or symptoms Lindberg et al. 1976). Finally, a specimen of respond better to stimulation in one site or viable wound tissue is removed from the the other is not known. The goal of this center of the wound using sterile technique. project is to study the efficacy of DBS and to All wound specimens are processed at a compare two sites of bilateral DBS to single microbiological research laboratory determine the most effective site for surgical using serial dilution and quantitative intervention. The findings will be critically important in establishing the optimal surgical procedures. Culture findings from each treatment of the disabling symptoms of swab specimen technique are then Parkinson’s Disease. compared to wound tissue cultures to determine sensitivity, specificity, positive MeSH Terms: Parkinson’s disease, predictive value, and accuracy. Although electric stimulation therapy, subthalamic recruitment and enrollment of subjects is still nucleus, globus pallidus, drug therapy, ongoing, preliminary analyses of 78 study neurosurgical procedures wounds found that Levine’s performed best of the three swab techniques in terms of all four validity parameters. Sensitivity was .70, specificity .90, positive predictive value .79, Principal Investigator: and accuracy .81. This analysis clarified the Sue E. Gardner, Ph.D., M.D. validity of using Levine’s techniques as a measure of microbial burden in programs of Project Title: research aimed at identifying risk factors for The Diagnostic Validity of Three Quantitative complications associated with high wound Swab Techniques bioburden. Subject enrollment is ongoing.

MeSH Terms: wound infection, diabetic Summary: foot

Although swab specimens are commonly used to examine the microbiology of Principal Investigator: suspicious chronic wounds, usefulness of Douglas R. Geraets, Pharm. D. the information provided is unclear. Furthermore, nurses, who collect the Project Title: majority of swab specimens, are provided 80+ Hemorrhagic Cohort Study conflicting information regarding which collection technique represents “best practice.” The purpose of this study is to examine the diagnostic validity of three Summary: quantitative swab techniques. The study population includes male and female Most patients with the irregular heart rhythm veterans and non-veterans with greater than atrial fibrillation (AF) warrant treatment with skin deep wounds. Concurrent swab the blood clot preventing medication ® specimens are obtained from wounds using warfarin (or Coumadin ) to reduce the risk of 1) wound exudates, 2) Z-technique, and 3) stroke. The benefits of warfarin increase Levine’s technique. The first technique with advancing age due to the increased absolute risk of stroke; however, the risk of samples visible wound exudates from the major bleeding complications (requiring wound bed before cleansing. The Z-

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blood transfusions or hospitalization) also warfarin is to draw a sample of blood and rises with age. The benefits of warfarin in AF test the time it takes for the blood to clot clearly outweigh the risks among those (reported as INR) in a laboratory. Portable under age 65 who have one or more risk machines to measure the INR from a small factors for stroke, but data on warfarin’s sample of blood (obtained from a finger- benefit compared to its risk in the elderly stick) have been approved for home use (greater than 65 years) are limited or since 1997. conflicting. This three year study will compare patients This study is designed to compare the rates receiving standard warfarin monitoring (by and severity of major bleeding problems in the VA Anticoagulation Clinics) to patients individuals 80 years and older compared to doing their own testing at home using the younger individuals (in their sixties) treated portable machines. This study will include with warfarin for atrial fibrillation. For each subjects with atrial fibrillation and/or patient 80 years or older, a randomly mechanical heart valves who are expected selected patient with AF younger than 80 to be anticoagulated indefinitely. The years from the same clinic and receiving primary events that will be compared warfarin for approximately the same duration between groups will be major bleeding, will be enrolled. Information on health strokes, and death of any patient. All sites conditions, medications received, warfarin will be VA Medical Centers with dose, and lab monitoring results will be anticoagulation services (ACSs) and will collected at time of enrollment. Patients are enroll up to 3200 patients at up to 32 sites. contacted every three months for follow-up data on any new health conditions MeSH Terms: atrial fibrillation; diagnoses, major bleeding episodes, protheses, heart valve; cerebrovascular warfarin dose changes, and lab monitoring accident; warfarin; anticoagulants; results. This study will continue for a period hemorrhage; thromboembolism; drug of six years from the time the first patients monitoring were enrolled.

MeSH Terms: atrial fibrillation, cerebrovascular accident, warfarin, Principal Investigator: anticoagulants, hemorrhage, Theresa Gioanni, Ph.D. thromboembolism Project Title: Molecular Determinants of Endotoxin Interaction with Toll-Like Receptors Principal Investigator: Douglas R. Geraets, Pharm. D.

Project Title: Summary: The Home INR Study (THINRS) The ability of humans to withstand the potential dangers of bacterial invasion Summary: depends upon the existence of sensitive, specific molecular alarms in the human host. The molecules that are involved in this alarm Approximately 2 million individuals in the system are designed to recognize and U.S. are receiving warfarin (also known as ® trigger responses to small numbers of Coumadin ) that prolongs the time it takes invading bacteria. There is growing evidence the blood to clot (also known as an for the role of the increasing number of Toll- anticoagulant). Persons taking warfarin must Like Receptors (TLRs) of the innate immune be monitored closely with blood testing. Too system in recognition and response to a much warfarin may increase the risk of diverse array of conserved microbial bleeding and too little warfarin may leave the products. The molecular basis of individual at risk of getting a blood clot. The recognition remains one of the most standard way to monitor the effects of enigmatic as well as essential properties of

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TLRs. In the case of Gram-negative bacteria (GNB), detection of endotoxin, a unique glycolipid of the GNB outer surface, plays a Principal Investigator: pivotal role. How the host detects endotoxin Thomas H. Haugen, M.D., Ph.D. and how that recognition is coupled to initiation of inflammatory responses has Project Title: been intensely investigated over many Viral Gene Expression and Replication in years. These efforts have led to the Papillomavirus-Associated Neoplasia identification of several key human extracellular and cellular proteins necessary for maximum sensitivity to endotoxin: Summary: lipopolysaccharide binding protein (LBP), CD14, Toll-like receptor 4 (TLR4), and MD- 2. The requirement for all of these proteins High-risk mucosal human papillomaviruses to achieve optimal sensitivity and response (HPV) cause cervical cancer and are to endotoxin suggests that host response associated with a subset of head and neck may depend on an ordered sequential series cancers. The most common high-risk type is of endotoxin:protein and/or protein:protein HPV-16, found in ~40-60% of cervical and interactions. The long-term goal of the >80% of HPV positive head and neck proposal is to understand the molecular cancers. Despite the development, over the basis of host-endotoxin interactions that lead past decade, of cell culture techniques that to pro-inflammatory responses. permit the study of HPV life cycle in the Understanding the mechanism of endotoxin laboratory little progress has been made in recognition/response by TLR4 and related applying these techniques to HPV-16. We proteins may lend insight into how ligand have identified HPV-16 isolates that are specificity of TLR proteins is determined and capable of establishing persistent replication the molecular properties of TLR receptor and extending the lifespan of (or: complexes required for maximal host "immortalizing") primary human foreskin, responses to products of infectious agents. cervical and tonsillar keratinocytes. By More specifically the goal is to identify the adapting techniques originally developed by ligand that binds directly to TLR4 and the Meyers and Laimins for HPV-31b, we have role of CD14 and MD-2 in mediating the further developed assays that allow us to interaction responsible for TLR4 activation follow critical stages of the HPV-16 life cycle by endotoxin. Greater knowledge of the in culture: viral early gene transcription, molecular basis of host response to initial DNA amplification, viral genome microbial invasion should provide new persistence in the form of stable, insights concerning the function of normal unintegrated, replicating plasmid genomes, host defenses and the conditions that can cell immortalization, and the effects of increase susceptibility to debilitating keratinocyte differentiation on the activation bacterial infection. These new insights of viral late functions. Taken together, we should also lead to a better understanding of now have the tools for the first time to study pathological conditions characterized by the genetics of the HPV-16 life cycle. HPVs either inadequate or excessive endotoxin have a small genome encoding fewer than a signaling and potentially lead to the design dozen distinct peptide open reading frames of new therapeutic agents that attempt to (ORF). Most HPV-positive high-grade modulate in a beneficial way immune lesions and carcinomas of the cervix and the responses. head and neck contain and express two of these ORFs, the viral E6 and E7 oncogenes, MeSH Terms: CD14, endotoxin, Gram- from HPV DNA fragments integrated in the negative bacterial infections, cellular genome. These alterations invariably inflammation, lipopolysaccharide, disrupt the E2 ORF, indicating that lipopolysaccharide binding protein, disruption of the E2 regulatory circuitry is a sepsis, toll-like receptors, TLR4 critical step in the progression from infection to cancer. Using the model keratinocyte system, we have identified that multiple different forms of the E2 ORF are produced

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in cells containing persistently replicating intradiscal pressure accompanying fusion HPV-16. These individual E2 forms may contribute to the development of contribute to either positive or negative expedited degenerative disc disease levels regulation of virus gene expression and adjacent to a fused spinal segment. DNA replication in these keratinocytes. Implantation of an artificial disc (AD) in place Improved understanding of HPV-16 of fusion at the site of a diseased disc would replication control might ultimately provide theoretically restore motion to the spine, targets for treatment or diagnosis of viral resestablish stability, and maintain the infection before progression to cancer takes caliber of neural foramina, while avoiding the place. development of adjacent degenerative changes. To this end, AD implants and MeSH Terms: human papillomavirus, prosthetic nuclei have been developed, and virus replication, viral genes, viral many have had fairly long track records in proteins, cervical cancer, head and neck Europe, but still await approval in the US. cancer This study in human cadaveric spines intends to examine the biomechanical effects of the Maverick Artificial Disc (Medtronic Sofamor-Danek, Nashville, TN) Principal Investigator: on the motion and intradiscal pressure at the Patrick W. Hitchon, M.D. adjacent level. Range of motion will be measured in the intact state, after Project Title: discectomy, and after implantation of the Motion and Intradiscal Pressure Adjacent to Maverick Artificial Disc (AD). Motion and Segment Implanted with Maverick Artificial intradiscal pressure will also be measured at Disc the adjacent level. The above measurements will be repeated following posterior instrumentation with L4 and L5 pedicle screws. Summary: MeSH Terms: lumbar spine, Degenerative disc disease (DDD) neurosurgery, artificial disc, intradiscal associated with disc collapse, herniation, pressure, motion analysis osteophytes, spinal instability, and pain often entails surgical management. Current treatments for lumbar DDD generally consist of disc excision with or without fusion. Principal Investigator: Unfortunately, fusion techniques are by no Patrick W. Hitchon, M.D. means assured of success. Though the incidence of pseudoarthrosis is estimated at Project Title: less than 10%, it is by no means negligible. Indications of Anterior and Posterior Cervical Moreover, these procedures are associated Instrumentation with risks of dural or vascular tear, chronic pain, and stiffness. In addition, many patients following a lumbar fusion may go on to develop symptomatic degenerative Summary: disease rostral or caudal to the fused segments. This accelerated process of It is not always clear when anterior plus degeneration adjacent to a fused posterior cervical spine instrumentation is segment(s) has been seen in the cervical necessary. If following corpectomy the and lumbar spine, and may warrant fusion. surgeon opts to rely entirely on anterior Displacement controlled studies in cadaveric stabilization, failure will undoubtedly occur in in vitro calf and human lumbar spines, have some cases. On the other hand to avoid shown and increase in motion and those few cases of construct failure the intradiscal pressure at intact spinal levels addition of posterior instrumentation in every adjacent to instrumented segments. It is case of anterior corpectomy is unnecessary, suspected that this incrase in motion and and excessive.

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A one stage anterior decompression and posterior stabilization of the cervical spine of a variety of pathologies including trauma, Principal Investigator: tumor, infection, rheumatoid arthritis, and Daniel G. Hug, B.S., Ph.D. spodylosis was undertaken in 100 patients (McAfee and Bohlman 1995). Project Title: Supplementary anterior instrumentation was Photobiology and Photo Medicine of used in only 17 of 100 patients. After a Urocanic Acid follow-up of 2 years, there were no construct failures, or graft dislodgements. A retrospective, multicenter study of corpectomy and anterior arthrodesis was Summary: conducted in 45 patients (Vaccaro et al. It is well documented that exposure of 1998). Construct failure with anterior grafting humans to ultraviolet radiation in sunlight and plating was noted in 3 of 33 patients has some bad effects. However, these (9%) with a 2 level corpectomy, and in 6 of effects may serve as a key to discover 12 patients (50%) with a 3 level cropectomy something of fundamental medical (p<0.05). Postoperative bracing or halo importance. A natural substance found at application did not appear to influence the surface of skin in humans and mice is success or failure significantly. The urocanic acid (UCA). In UV light (i.e., association between levels of corpectomy sunlight), trans-UCA is changed into cis- and construct failure was recently UCA, which signals the immune system to documented in a retrospective review of 40 suppress itself. This process is thought to patients undergoing anterior decompression be a part of the generation of skin cancer by and plating. There were 2 failures out of 33 sunlight and also leads to increased patients undergoing a 3 level corpectomy susceptibility to infectious diseases. (6%), compared to 5 out of 7 patients (71%) Because cis-UCA is an important signal undergoing 3 level corpectomies. molecule in humans, we set out to discover Our hypotheses were that the laminectomy the enzyme which degrades cis-UCA. We destabilizes the spine and that posterior had no success in the mouse, but we instrumentation is necessary whenever discovered a new enzyme, cis-urocanic acid corpectomy is performed in the presence of isomerase, in bacteria. We confirmed that preexisting laminectomy. Currently our study the kind of bacteria which has the enzyme is has shown that a 2 and 3 level laminectomy a natural inhabitant on human skin. does not have any statistical difference from Because these bacteria destroy trans- and the intact. In addition, when adding either cis-UCA, they could be beneficial to prevent posterior or anterior fixation with then the suppression of the immune system. This addition of the other, caused no significant idea is speculative but it may be that daily decrease in motion. showers with an antibacterial soap reduces the number of beneficial bacteria. A possible MeSH Terms: cervical spine, extrapolation is that antibacterial soap may neurosurgery, laminectomy, diskectomy, have a down side. An important paper in motion analysis Cancer (January 2004) showed that second malignancies tend to accompany skin cancer which is a monitor of lifetime exposure to sunlight. African Americans with skin cancer were 3.3 times more likely to have second cancers than white Americans with skin cancer. This may be explained by a higher concentration of UCA in black skin compared to white skin. We are currently submitting a review article explaining this hypothesis. Evidence in three publications indicate black Africans have higher UCA levels on their skin than white Europeans. Thus black Africans might

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be more susceptible to sunlight-induced immune suppression. We demonstrated in mice that radioactive cis-UCA applied to skin Principal Investigator: appears rapidly in urine indicating that skin Gary W. Hunninghake, M.D. cis-UCA is internalized so it might affect the immune system. Our laboratory does basic Project Title: research which is not easily understood by Role of P13K in Sepsis-Induced Acute Lung non-chemists. Therefore I stressed the Injury possible application of this basic research to medicine, but these applications are speculative. For example, the results of this Summary: research and many other studies worldwide may lead medical authorities to recommend The purpose of these studies is to determine sun protection (e.g. sunscreens, clothing, how cells in the lung respond to inhaled hats) to prevent non-skin cancers that may particles. Many inhaled particles contain be enhanced by exposure of the skin to endotoxin and endotoxin stimulates these sunlight. cells to produce a number of molecules that

are important for the lungs response to the MeSH Terms: cancer, urocanic acid, particles. This study evaluates pathways skin, immunity, skin neoplasms, bacteria, within macrophages that are activated to ultraviolet result in the production of these inflammatory molecules.

Principal Investigator: MeSH Terms: macrophages, lung infection, endotoxin, injury, fibrosis Gary W. Hunninghake, M.D.

Project Title: Human Lung Responses to Respiratory Principal Investigator: Pathogens Gary W. Hunninghake, M.D.

Project Title: Summary: Regulation of Alveolar Macrophage Function

The purpose of these studies is to determine how cells in the lung respond to infection. Summary: The cell that is being studied is the alveolar macrophage. This is the first cell to respond The purpose of these studies is to determine to bacteria. Bacteria stimulate these cells to why normal cells that are present in the lung produce a number of molecules that are respond differently than similar cells in other important for the lungs response to infection. sites of the body like blood. This differential This study evaluates pathways within response is important because it prevents macrophages that are activated to result in the lung from developing inflammation each the production of these inflammatory time the lung is exposed to an inhaled molecules. particle. The studies postulate that blood cells must migrate into the lung to create the MeSH Terms: macrophages, lung, appropriate environment for a persistent infection, endotoxines, wounds and lung inflammation. injuries, fibrosis MeSH Terms: macrophage, lung infection, endotoxin, injury, fibrosis

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Principal Investigator: Principal Investigator: Gary W. Hunninghake, M.D. Gary W. Hunninghake, M.D.

Project Title: Project Title: Molecular and Cellular Biology of the Lung Iowa Multidisciplinary Clinical Research Career Development Program

Summary: Summary: This is a training grant that supports the research training of candidates interested in This submitted grant is to support the clinical research related to the lung. research training of faculty from seven colleges within the University of Iowa. MeSH Terms: macrophage, lung, infection, endotoxin, injury, fibrosis Principal Investigator: Jacob Ijdo, M.D., Ph.D. Principal Investigator: Gary W. Hunninghake, M.D. Project Title: Intracellular Survival of Anaplasma Project Title: Phagocytophilum in Neutrophils Modulation of Acute Lung Injury

Summary: Summary: Anaplasma phagocytophilum is a gram- The purpose of these studies is to determine negative intracellular organism that how viral infections increase the severity of exclusively infects and survives in bacterial infections. The studies show that granulocytes. We have recently viral infections block the response to crucial demonstrated that A. phagocytophilum molecules that are necessary to fight reduces the NADPH oxidase at the bacterial infections. During this window of phagosome. This reduction may allow for time, the body is very vulnerable to bacterial survival within neutrophils. We hypothesize infections that A. phagocytophilum introduces virulence factors that are responsible for the NADPH oxidase reduction. MeSH Terms: macrophage, lung, OBJECTIVES: This proposal aims to study infection, endotoxin, injury, fibrosis how these virulence factors cause NADPH oxidase reduction at the phagosome presumably leading to intracellular survival. We plan to investigate the type IV secretion system that introduces these virulence factors into the host cell. RESEARCH PLAN: We will use transposon-based mutagenesis to knock out the type IV secretion system. This transposon-based strategy is an effective way to introduce foreign DNA into the bacterial genome. Insertion of a transposon into a gene of interest will disrupt the gene of interest. Bacteria with mutated genes of

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the type IV secretion system will not be able older and taking five or more regularly to translocate the virulence factors and thus scheduled medications. They will be expected to be unable to reduce the NADPH randomized to the intervention or usual care. oxidase at the phagosome. As part of the intervention, the METHODS: We have generated a pharmacist/physician team will evaluate the transposon construct containing a rifampin patient and make recommendations to their resistance gene under an A. primary care provider in terms of optimizing phagocytophilum-specific promoter. Bacteria their medications and suggest ways to that have incorporated the transposon are prevent adverse drug events. Data will be selected by growth in the presence of collected at baseline, three months, and 12 rifampin, and mutants will be analyzed for months and will compare outcomes between transposon insertion in the genes encoding the two groups. The outcomes of interest the type IV secretion system. include adverse drug events, measured EXPECTED FINDINGS: We expect to medication appropriateness, health care identify mutants that have lost the ability to utilization, and overall satisfaction of patients survive in neutrophils, demonstrating that and their primary care providers. Through the type IV secretion system is crucial in this research project, we hope to learn how translocating virulence factors into the host to optimally manage elderly patients taking cell. Next we will focus on the nature of the multiple medications. We also hope to virulence factors themselves. identify which patients could potentially gain CLINICAL RELEVANCE: Characterization the greatest benefit from such an of the basic mechanisms of intracellular intervention so as to target this highly survival of A. phagocytophilum in neutrophils intensive intervention in practice. will further our understanding of the basic function of neutrophils, which are key MeSH Terms: polypharmacy, veterans, players in all processes of inflammation, adverse drug events. infection, and cell damage and healing.

Principal Investigator: Principal Investigator: Randy H. Kardon, M.D., Ph.D. Peter Kaboli, M.D., M.S. Project Title: Project Title: Predicting Rehabilitation of Visual Loss in Randomized Control Trial of Structured Compressive Optic Neuropathy Pharmacy Care in Older Veteran Outpatients Summary:

Summary: Veterans may suffer profound visual loss due to compression of the optic nerve by Complications resulting from medications or tumors or masses. The visual loss is usually adverse drug events, are prevalent and a caused by a block in the conduction of the major source of excess morbidity and costs. nerve impulses from the compression. In The elderly are particularly prone to many cases, vision can be restored after complications from adverse drug events. relieving the compression by surgery or The overall goal of this research project is to radiation, but some patients do not recover test the effectiveness of an intervention to and still have visual loss due to permanent improve medication use in older outpatient loss of nerve fibers in the optic nerve from veterans. This intervention involves a multi- excessive compression or compression of dimensional evaluation of a patient’s long duration. Also, some veterans have medications by a clinical pharmacist and other significant health problems that can board certified geriatrician. We will enroll put them at significant risk during surgical over 500 veterans at the Iowa City VA treatment of the compression. This study will Medical Center who are 65 years of age or determine if certain measurements of the

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optic nerve structure and function can the sensory nerves had been removed from predict which patients would benefit from the kidneys. Whereas normal healthy rats treatment versus other patients with more (and man) can ingest an increased salt permanent loss that would not be expected intake without increasing blood pressure, to benefit from treatment. The outcome will rats in which the renal sensory nerves were help determine which patients may expect to removed developed hypertension when fed get return of vision and will help advise on a high salt diet. the benefits and risks of surgery and We have also shown that activation of the radiation. Preliminary studies showed that sensory nerves in the renal pelvic wall fails the thickness of the layer of the retina to elicit an increase in urine excretion in containing the nerve fibers that eventually hypertensive rats and rats with congestive coalesce to form the optic nerve is a very heart failure. The mechanism involved in the good indicator of how many nerve fibers impaired activation of the renal sensory have been permanently lost. Optical nerves involves increased activation of the Coherence Tomography is a new technique renin angiotensin system in the kidney. that accurately images the layers of the Treating hypertensive rats and rats with retina in the living human eye and will be congestive heart failure with an angiotensin used to measure if there is loss of nerves receptor blocker, losartan, restored the leading to permanent visual loss or a normal responsiveness of the renal sensory nerves number of nerves in the retina, indicating a towards that seen in healthy rats. good prognosis for visual recovery. In Angiotensin receptor blockers are commonly addition, the study will help to evaluate used for treatment of high blood pressure. whether the treatment for compression of Our findings suggest that the improvement the optic nerve was adequate, by assessing of the responsiveness of renal sensory if the visual recovery was in proportion to the nerves to an increased salt/water intake remaining number of nerve fibers in the produced by these agents may be a retina. contributory mechanism to the beneficial effects of these drugs in the treatment of MeSH Terms: compressive optic hypertension. neuropathy, optical coherence Little attention has been placed on the role tomography, visual loss of the sensory renal nerves in the neural control of the circulation. However, increased arterial pressure in renal sensory denervated rats fed high salt diet suggests Principal Investigator: an important role of the renal sensory Ulla C.Kopp, Ph.D. nerves in the maintenance of sodium balance and thus arterial pressure. By Project Title: gaining insight into the mechanisms involved Afferent Renal Nerves: Role of in the increased arterial pressure in rats Neurotransmitter lacking intact renal sensory innervation, we will further our understanding of the renal mechanisms contributing to hypertension Summary: which may lead to better treatment of hypertension, a major health problem in

USA. Increased salt intake increases urine flow rate resulting in a stretch of the renal pelvic MeSH Terms: kidney, neurons afferent, wall. This stretch activates sensory nerves in neurotransmitters, substance P, the renal pelvic wall. Activation of these prostaglandins, hypertension, renin sensory nerves sends a signal to the brain angiotensin system that eventually results in an increase in urine excretion, a natriuretic reflex. Thus, activation of this reflex facilitates the excretion of an increased salt intake. We recently showed the importance of this mechanism when we studied rats in which

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organisms has contributed to significant Principal Investigator: increases in their global health burden. Information gained from these studies is David J. Kusner, M.D. directly relevant to our understanding of the

molecular mechanisms of tuberculosis and Project Title: S. aureus infections. These data are also Regulation of Phagocyte Antimicrobial broadly relevant to infectious (sepsis, Activity by Phospholipase D pneumonia, urinary tract and post-surgical infections) and non-infectious (myocardial infarction, stroke, acute respiratory distress Summary: syndrome, chronic obstructive pulmonary disease, arthritis) inflammatory diseases in which phagocyte activation contributes to Human defense against infection is immune defense but also damages host comprised of both the innate and adaptive tissues. The significance of this work derives immune systems. The central cellular both from the increase in our understanding components of the rapidly-activated innate of the pathogenesis of these disorders and system are phagocytic leukocytes by providing a foundation for novel therapies (macrophages, monocytes, neutrophils, and to modulate phagocyte cytotoxic responses dendritic cells). Activation of phagocytes in medically-important diseases. results in the generation of potent microbicidal compounds but also damages MeSH Terms: innate immunity, host tissues at inflammatory sites. Thus, inflammation, infection, macrophage, advances in our understanding of the tuberculosis, staphylococcus aureus, molecular mechanisms that regulate the phagocytosis activation of phagocytic leukocytes are essential to therapeutic efforts to promote beneficial immune functions and decrease harmful effects to the host. The long-term Principal Investigator: goal of this project is to characterize the Steven R. Lentz, M.D., Ph.D. biochemical mechanisms that regulate the ingestion (phagocytosis) and killing of Project Title: microorganisms. We have recently Vascular Dysfunction in demonstrated that activation of a host Hyperhomocysteinemia and Atherosclerosis enzyme, phospholipase D (PLD), is tightly coupled to phagocytosis. The specific aims of this project are to determine which isoforms of PLD regulate microbial killing Summary: and to characterize the mechanisms of this regulation. Two mammalian isoforms of Homocysteine is an amino acid that is PLD have been identified, PLD1 and PLD2, derived from methionine in the diet. Like a which exhibit differential localization and high level of cholesterol, a high level of activation. However, no information is homocysteine in the blood available on the intracellular locations, (“hyperhomocysteinemia”) is recognized as activation mechanisms and functions of a common risk factor for stroke and PLD1 and PLD2 in phagocytes. Definition of cardiovascular disease. the isoform-specific regulatory roles of PLD1 Hyperhomocysteinemia can be caused by a and PLD2 is necessary to better understand deficiency of B vitamins, genetic factors, or the biochemical pathways that govern innate kidney failure and is common in the VA immune responses and is required for patient population. Although strong translational efforts to therapeutically modify associations between phagocyte activation. We will study two hyperhomocysteinemia and cardiovascular critically-important infectious agents of events have been observed in many human disease, Mycobacterium tuberculosis epidemiological studies, it is not known and Staphylococcus aureus. The increasing whether hyperhomocysteinemia is a incidence of antibiotic resistance in these causative factor or a marker of

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cardiovascular disease. The potential benefit understanding how the immune system can of homocysteine-lowering therapy is recognize antigens on cancer cells, develop currently being tested in several large antibody and cell-mediated responses clinical trials. Using an animal model of against the antigens, and use those experimental hyperhomocysteinemia, our responses to destroy the abnormal cells. group was among the first to demonstrate These experiments have included the that hyperhomocysteinemia causes development of vaccines in which we have dysfunction of the endothelial lining cells of inserted the gene for prostate specific blood vessels. We are now working toward antigen (PSA), a protein produced by the the development of new models of prostate cancer cells, into viruses and hyperhomocysteinemia in mice. In these bacteria; as well as the use of models, hyperhomocysteinemia is produced immunostimulatory DNA to augment or by specific combinations of genetic and modulate the anti-tumor immune responses. dietary factors. We are using these animal models to examine the mechanisms of Vaccine Development: The research funded vascular injury and dysfunction caused by by a VA Merit Review Grant “Multiple hyperhomocysteinemia. By using genetic Vaccines for Therapy of Prostate Cancer” approaches in which hyperhomocysteinemia has been very successful. We have used is produced by specific alterations in multiple microbial vaccines, each with the homocysteine metabolism, these studies PSA gene inserted, for the generation of have the potential to delineate mechanisms anti-prostate cancer immune response. of vascular dysfunction in Most of the research and subsequent clinical hyperhomocysteinemia, particularly in trial has concentrated on an adenovirus/PSA association with other risk factors such as (Ad/PSA) vaccine. Immunization of mice hypercholesterolemia. This project also may with the vaccine induces strong anti-PSA, suggest novel therapeutic approaches for and hence, anti-prostate cancer immunity the prevention and treatment of vascular that protects the mice from growth of PSA- diseases associated with secreting prostate tumors. Additional hyperhomocysteinemia. Such approaches vaccines that have been tested or being might include homocysteine-lowering developed are two vaccinia (cowpox)/PSA, therapies, antioxidant or anti-inflammatory canarypox/PSA, BCG/PSA, and Listeria medications, or other pharmacological monocytogenes (Lm/PSA). Work continues approaches to target the vascular mediators on the production of a Salmonella of hyperhomocysteinemia in people at risk tymphimurium (Sal/PSA) vaccine. Data for cardiovascular disease. from our preclinical studies using these vaccines indicate a variation in the strength MeSH Terms: atherosclerosis, of the anti-PSA immunity induced and the T cardiovascular diseases, cells that predominate in the anti-tumor cerebrovascular accident, endothelium, immunity. We are investigating the use of homocysteine the different prostate cancer vaccines to be used in a “prime-boost” protocol where we are attempting to induce the highest level of tumor destroying immunity using one Principal Investigator: vaccine as the initial immunization and a David M. Lubaroff, Ph.D. different vaccine for subsequent booster immunizations. Project Title: Summary of VA Research Projects Augmentation and Modulation of Anti-Tumor Immunity. In an attempt to further augment the anti-prostate cancer immunity induced by the various vaccines we have been Summary: combining immunostimulatory DNA molecules with the vaccines. For these Our laboratory has been involved in studies we have been collaborating with Dr. research on tumor immunology, Arthur Krieg of Coley Pharmaceutical Group and formerly of the ICVAMC and the

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University of Iowa. Dr. Krieg has demonstrated that particular DNA Principal Investigator: sequences termed CpG ODN, found in Donald Lund, Ph.D. bacteria, can stimulate immune responses. Our studies have shown that the combination of the Ad/PSA vaccine with Project Title: CpG ODN induces immune responses in the Effects of a Common Human Gene Variant mice that are stronger than those induced by of Extracellular Superoxide Dismutase on the vaccine alone. In addition, the Endothelial Vasomotor Function after combination also modulates the anti-tumor Endotoxin in Mice. response such that traditional mechanisms by which the tumors are destroyed have been changed. This research, funded by a project on our VA Research Enhancement Summary: Award Program (REAP) in Prostate Cancer, is being vigorously pursued in the lab. Superoxide and oxygen radicals formed during inflammation are often harmful to Clinical Trial. We have just completed a blood vessels. Normally, Extra Cellular Phase I clinical trial of the Ad/PSA vaccine Superoxide Dismutase (ECSOD) protects at the affiliated University of Iowa. Men with the blood vessels by breaking down metastatic prostate cancer were injected superoxide into harmless by-products. with the vaccine to determine the safety of Recently, a gene variant or polymorphism of the product. The first patients were human ECSOD has been reported. This immunized with a low dose of the vaccine gene variant has a substitution of glycine for and, when no adverse reactions were noted, arginine in position 213 (R213G) of the additional patients were immunized with the heparin binding domain. The variant is next higher dose. This was repeated until common, as it has been described in 4-6 the highest dose allowed by the FDA was percent of the populations from Japan, administered to the patients. The results Sweden, and Denmark. Patients, with this from this trial demonstrated that the vaccine gene variant, have been reported to have a was safe with no serious side effects. In 2 .3-fold increase in risk of ischemic heart addition, 42% of the patients developed T disease. The purpose of this study was to cell-mediated immune responses to the determine whether gene transfer of human prostate tumor associated PSA. A ECSOD or the gene variant (ECSOD- therapeutic Phase II trial is planned. R213G) improves endothelial function during inflammation. Adenoviral vectors MeSH Terms: prostate cancer, expressing ECSOD, ECSOD-R213G, or a immunotherapy, vaccines, CpG, control were injected i.v. in mice. After 3 adjuvants days either vehicle or an inflammatory agent (lipopolysaccharide, LPS) was injected i.p. After one day, vasomotor function of aorta was examined. Maximal responses to sodium nitroprusside (10-9 to 10-5), an endothelium independent vasodilator, were similar in aorta from normal and LPS-treated mice. Relaxation to acetylcholine (10-9 to -5 10 ), an endothelium dependent vasodilator was significantly impaired (p<0.05) after LPS and a control virus (63+3%, mean±SE) compared to normal vessels (83+3%). Gene transfer of ECSOD improved (p<0.05) relaxation in response to acetylcholine (76+5%) after LPS, whereas gene transfer of ECSOD-R213G (65+7%) had no effect. Superoxide levels were increased in aorta after LPS, and the levels were reduced

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following ECSOD but not ECSOD-R213G. CT protein. Where and how calpain degrades Thus, adenoviral-mediated gene transfer of CT is unknown. This research will attempt to ECSOD reduces superoxide and improves devise new ways to stimulate CT activity and endothelial dependent relaxation in the aorta therefore elevate lung surfactant levels by after LPS. We conclude that a common delivering mutant CT enzymes that have altered polymorphism (R213G) in ECSOD greatly calpain proteolytic attack sites within their attenuates efficacy of ECSOD in protection protein sequence. The long-term goal of these against endothelial dysfunction produced by studies will be to develop the first generation of an inflammatory stimulus. mutant surfactant enzymes that can be delivered to animals and then patients that will greatly stimulate surfactant production. Principal Investigator: Rama Mallampalli, M.D. MeSH Terms: surfactant, cytidylyltransferase, tumor necrosis factor, calpain, acute respiratory distress syndrome Project Title: Regulation of Cytidylyltransferase in Fetal Rat Lung Principal Investigator: Rama Mallampalli, M.D. Summary: Project Title: TNF-Lipoprotein Control of a Sufactant Surfactant is a complex material containing Enzyme lipids and proteins that is essential in order to maintain normal lung structure and function. Surfactant replacement therapy and hormone therapy (i.e. glucocorticoid treatment) for the Summary: infant respiratory distress syndrome (RDS) has clearly been shown to save the lives of Surfactant is a complex material containing numerous babies. It has recently been lipids and proteins that is essential to maintain determined that several adult lung diseases normal lung structure and function. Surfactant including emphysema, asthma, acute lung replacement therapy and hormone therapy (i.e. injury (ARDS), and lung fibrosis all common to glucocorticoid treatment) for the infant the VA population are also associated with a respiratory distress syndrome (RDS) has clearly low levels of surfactant. In fact, pilot studies are been shown to save the lives of numerous underway with surfactant replacement in ARDS babies. It has recently been determined that and asthma. Recent discoveries in our several adult lung diseases including laboratory have shown that circulating lipids in emphysema, asthma, acute lung injury (ARDS), the blood stream that produce heart disease, and lung fibrosis all common to the VA termed low-density lipoproteins (LDL), potently population are also associated with a decrease lung surfactant production.. functional deficiency of surfactant. In fact, pilot Therefore, this research will study how studies are underway with surfactant surfactant production is decreased by LDL in the replacement in ARDS and asthma. Recent lung. studies also suggest that a key toxic substance, To date, limited information is available as to termed tumor necrosis factor (TNF α), plays an how the adult lung produces surfactant, important role in the pathogenesis of each specifically at the enzymatic or molecular level. these lung disorders. Therefore, this research The goals of this research proposal focus on will study how surfactant production might be how LDL controls the activity of a key enzyme, altered by TNF in the adult lung. cytidylyltransferase (CT). So far our data show To date, limited information is available as to that LDL triggers a process that eventually leads how the adult lung produces surfactant, to breakdown of the CT protein. This process specifically at the enzymatic, molecular level. involves a proteinase, calpain, that cleaves the The goals of this research proposal focus on

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how the adult lung controls the activity of a key what portions of a key region, termed a enzyme, cytidylyltransferase (CT). This enzyme promoter, are essential that drive the genetic is activated by fatty acids and inhibited by expression of CT in the lung. Studies will involve TNFα. This research will attempt to devise new for the first time construction of several lines of ways to stimulate CT activity and therefore transgenic mice that harbor different sizes of the elevate lung surfactant levels by delivering fatty CT gene promoter. These promoter fragments, acids packaged into very low-density when expressed in the mouse, should uncover lipoprotein particles (VLDL). We will specifically the molecular and biochemical clues as to how infuse VLDL particles in animal models to surfactant is produced and might be affected increase production of surfactant and in lung disease. Thus, knowledge gained counteract adverse effects of TNF. The long- through these basic molecular studies may term goal of these studies will be to develop ultimately help devise newer therapies for specific inhibitors or activators of enzymes surfactant deficient lung disorders such as gene involved in control of surfactant production. therapy. Thus, knowledge gained through these biochemical studies may ultimately help devise MeSH Terms: surfactant, newer therapies for surfactant deficient lung cytidylyltransferase, gene, acute respiratory disorders. distress syndrome.

MeSH Terms: surfactant, cytidylyltransferase, tumor necrosis factor, acute respiratory distress syndrome Principal Investigator: Rama Mallampalli, M.D.

Project Title: Principal Investigator: The CCT Promoter to Drive High-Level Rama Mallampalli, M.D. Expression of CFTR

Project Title: Transcriptional Regulation of a Surfactant Enzyme Summary:

Gene therapy for cystic fibrosis (CF) holds promise. Effective gene therapy requires a Summary: vector containing a DNA fragment that directs the expression of the desired protein, Surfactant is a complex material containing i.e. CFTR, in lung airway epithelial cells. lipids and proteins that is essential to maintain This DNA fragment, termed a promoter, normal lung structure and function. Surfactant must be sufficiently active and yet small replacement therapy and hormone therapy (i.e. enough to package into viral-based vector glucocorticoid treatment) for the infant systems currently under investigation in CF gene therapy. Currently available promoters respiratory distress syndrome (RDS) has clearly exhibit variable activity or are too large to been shown to save the lives of numerous package within viral vectors, such as adeno- babies. In order for the lung to produce associated virus (AAV). Recently, we surfactant, it has to increase the activity of a key generated and tested the promoter for a key enzyme, cytidylyltransferase (CT). Therefore, enzyme involved in lipid metabolism, termed this research will study how surfactant cytidylyltransferase (CCT). Our data production is controlled by CT in the adult lung. demonstrates that the CCT promoter is To date, limited information is available as to small and yet it is very active. This DNA how the adult lung produces surfactant, promoter is also stimulated by retinoic acid. specifically at the molecular level. The goals of The goal of this application is to investigate this research proposal focus on how the adult whether the CCT promoter can be used to lung controls the activity of CT at the genetic drive sustained expression of cystic fibrosis level. We will clone the CT gene and determine protein in human airway epithelia. To

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achieve this goal, we will first optimize the studied in a tissue bath allowing the heart CCT promoter further by determining the tissue to metabolize and function as if it smallest DNA fragment that can be used in were supplied by blood demonstrates gene therapy (AIM 1). We will next test abnormal electrical activity possibly whether the CCT promoter when packaged accounting for the fast rhythm in the intact into a viral-based (AAV) vector can trigger animal. The purpose of the present project the production of cystic fibrosis protein in is to identify the focus of the fast rhythm and human airway epithelial cells. We will assess subject it to various kinds of drug therapy activity of this novel DNA fragment after and then remove the tissue from the origin retinoic acid treatment to achieve very high of the rhythm distance and place it in a levels of cystic fibrosis protein in cells. We tissue bath and determine whether the will also compare the CCT promoter with response of the abnormal electricity is other existing promoters currently in use in similar to that of the intact heart. Todate a CF gene therapy. Ultimately, results from number of different drugs antagonizing these studies might uncover newer electrical channels in the heart muscle have strategies in gene therapy to target been studied including sotalol, a potassium sustained-expression of desirable proteins in channel blocker, nickel, a calcium channel cystic fibrosis. blocker, verapamil, a calcium channel blocker and lidocaine, a sodium channel MeSH Terms: surfactant, cystic fibrosis, blocker, all of which do not interfere either gene transfer the with the induced fast rhythm in the animal or in the abnormal electrical disturbance found in the tissue. However, standard beta-blocking drugs, which keep Principal Investigator: people alive after heart attack, interfere with James B. Martins, M.D. 50% of the fast rhythms that are induced in the animal and block 75% of the abnormal Project Title: electrical activity in the tissue. Moreover, a Purkinje Origin of Ischemic Ventricular part of the adrenergic nervous system, Tachycardia which is specific to the condition system lining of the heart, impairs the induction of the tachycardia in the intact animal and Summary: prevents the electrical activity in the tissue. These studies suggest that current approaches attacking or modifying the Sudden cardiac death in the United States adrenergic input to the heart importantly claims more than 350,000 lives per year. may prevent dangerous rhythm disturbances Most patients with this problem have organic owing to limited blood supply. heart disease and specifically coronary artery disease with either old heart attack or MeSH Terms: tachycardia, ventricular, new occluded blood vessel causing the ischemia, purkinje fibers, adrenergic heart muscle to function without oxygen. alpha-agonists, adrenergic beta- This project involves mapping of the origin of antagonists fast heart rhythm occurring in anesthetized dogs with a one-hour coronary occlusion. The fast heart rhythm that occurs in this setting is either due to an outer rim of the heart muscle rhythm disturbance or an inner rim of the heart muscle, which originates from one specific area and emanates out from it as the circles when a stone is thrown into a puddle of water. Our studies have shown that this specialized condition system on the lining of the heart may be the focus of these dangerous rhythm disturbances and that such foci of rhythm disturbance when

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proteins, and whether regulated cell death is Principal Investigator: involved in the repair process. To perform these studies we are using mice that lack Stephen E. McGowan, M.D. important signaling proteins or other proteins that are required for the production of these Project Title: signaling proteins. Some studies are Septal Fibroblast Proliferation and Migration performed in cultured cells and will allow us during Alveolar Growth to study the movement of individual cells on different substrata. Other studies will be performed using lung slices in culture so that Summary: we can study the effects of signaling proteins on the elongation of the air sac walls. Armed with a better understanding of My research is directed at understanding these basic processes, I am hopeful that we how the cells and proteins that form the can develop drugs and other strategies that scaffolding of the alveolar region of the lung will offer a new hope in treating emphysema grow and repair themselves. When the lung and lung fibrosis. is damaged by such agents as cigarette smoke, toxic dusts and chemicals, or MeSH terms: emphysema, elastin, harmful cells, it can repair itself under some tropoelastin, retinoids but not all circumstances. In most cases the repair is not complete, so that over time, there is a slow loss of lung function. One way to prevent the loss of lung function is to Principal Investigator: eliminate the harmful agents, but this William M. Nauseef, M.D. approach is not always successful. Therefore it is important to develop ways to reverse damage that has already occurred. Project Title: Unfortunately the currently available The Cell and Molecular Biology of Human treatments for emphysema and lung fibrosis Neutrophils cannot reverse the damage that has taken place, and in many cases cannot even successfully retard the damage that is in Summary: progress. This leads to frequent and costly hospital admissions during which drugs are The purpose of our research is to understand administered to help the veteran through an the way in which the human immune system immediate crisis, only to have him return in a responds to microbes at the earliest stages of short time with the same or even greater infection. When microbes bypass the problems. My research goal is to develop mechanical barriers such as skin, they initially our understanding of the biochemistry of the engage elements of the immune system that lung’s structural framework by analyzing are considered “innate”, meaning that they how lung cells initiate and regulate the repair are defenses with which we are born and do process. This involves studying how cells not require prior exposure to exist or to be that are responsible for the production of effective. These elements contrast with those structural extracellular proteins migrate to of the acquired immune system, including their proper location. The cells will produce antibodies and lymphocytes, which require a uniform and cooperative structural network prior exposure to be most effective. Of only if they are properly located. We are course, in most cases both schemes learning how cells signal to one another and collaborate to constitute our immune system respond to these signals in a coordinated and thus cooperate to ward off microbial fashion by examining particular extracellular invasion and protect the host from infection. signaling proteins and the cell membrane Among the elements of the innate immune receptors that respond to the proteins. This system, the most important cellular will enable us to learn how the cells grow, component is the neutrophil, the cell that replace damaged cells, how they know to predominates in “pus”. Neutrophils respond stop growing and produce extracellular to microbial invasion by crawling to the site

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of infection and ingesting the microbe, a and prostaglandin (PG) I2, a soluble process known as phagocytosis. The most factor(s) is produced by endothelial cells efficient system that neutrophils use to kill which relaxes vascular smooth muscle by microbes requires oxygen. When stimulated, + the neutrophil assembles the NADPH- activating potassium (K ) channels. This dependent oxidase, a complex factor, which is known as endothelium- multicomponent enzyme that converts derived hyperpolarizing factor (EDHF), molecular oxygen into hydrogen peroxide, contributes prominently to endothelium- which is generated in the intracellular dependent relaxation in coronary blood compartment (phagosome) in which the vessels of many species, including humans. microbe is trapped. The stimulated In conduit blood vessels, EDHF-medicated neutrophil also releases into the phagosome relaxation may be an important backup myeloperoxidase, a granule protein housed mechanism to NO, serving to maintain within the resting neutrophil. In the presence endothelial function when the activity of NO of hydrogen peroxide, myeloperoxidase is diminished, such as in atherosclerosis. In catalyzes the generation of hypochlorous resistance blood vessels, EDHF contributes acid (bleach), an extremely toxic agent that substantially to endothelium-dependent will kill the ingested microbe. vasorelaxation, suggesting that EDHF may Our laboratory studies (1) how contribute to the regulation of coronary myeloperoxidase is normally synthesized blood flow. and delivered to its normal storage site in Arachidonic acid (AA) can be converted to the neutrophil; (2) the impact of mutations in vasoactive metabolites through several the myeloperoxidase gene on the structure enzymatic pathways, including the and function of the protein (in patients with cyclooxygenase, lipoxygenase, and hereditary myeloperoxidase deficiency), (3) cytochrome P450 pathways. AA can be how myeloperoxidase-dependent responses metabolized by the endothelium through the participate in the killing of staphylococci as lipoxygenase pathway, which forms part of human defense against HPETEs at the 5, 12, or 15 carbons where staphylococcal infection; and (4) the they are then reduced to the corresponding mechanisms regulating the activity and HETEs. location of the NADPH oxidase. All of our The overall hypothesis is that the work involves the use of human neutrophils arachidonate 12-lipoxygenase pathway and addresses questions directly relevant to participates in EDHF-mediated human disease. vasodilatation of the porcine coronary microcirculation. MeSH terms: neutrophils, infection, The proposed studies will be performed innate immunity, myeloperoxidase, using methods currently available in the oxidase cardiovascular research laboratories. Porcine coronary microvascular reactivity will be studied using an isolated arteriole preparation. Internal diameters will be Principal Investigator: measured by video microscopy of arterioles Christine L. Oltman, Ph.D. mounted onto pipettes in Krebs buffer. Electrophysiologic studies will be performed Project Title: to characterize K+ channels in porcine Role of Arachidonate 12- Lipoxygenase in coronary arteriolar smooth muscle. Porcine Coronary These studies should provide important insight into regulatory mechanisms of the coronary microcirculation. Summary: MeSH terms: arachidonate 12- lipoxygenase, arachidonic acid, coronary The vascular endothelium is an important vessels, microcirculation regulator of blood flow. In addition to two previously-characterized endothelium- derived relaxing factors, nitric oxide (NO)

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the vascular complication associated with the development and progression of the Principal Investigator: metabolic syndrome. Christine J. Oltman, Ph.D.

MeSH terms: metabolic syndrome X, Project Title: zucker rats, diabetes mellitus type 2, Progression of Vascular Dysfunction with oxidative stress, coronary Metabolic Syndrome: Role of Oxidative Stress

Principal Investigator: Summary: Gary E. Rosenthal, M.D.

Metabolic syndrome is an emerging epidemic Project Title: and consists of several components Center for Research in the Implementation including, abdominal obesity, atherogenic of Innovative Strategies in Practice—CRIISP dyslipidemia, hypertension, insulin resistance ± glucose intolerance, a proinflammatory state, and a prothombotic state. All of these Summary: characteristics are significant risk factors for development of vascular dysfunction and cardiovascular disease. The pathology Hospitals, physicians, and other health care linking vascular complications with metabolic providers often fall short of using the syndrome is not well known. However, findings from research studies to improve recent studies suggest that increased health care services for veterans. While oxidative stress may contribute to vascular research studies show that the VA does a dysfunction in many diseases including better job of using research information than hypertension and diabetes. The providers in the private sector, more can still mechanism(s) by which redox regulation of be done to improve the transfer of cellular signaling contributes to vascular information from researchers to clinicians. dysfunction associated with metabolic This transfer of information is referred to as syndrome remain to be elucidated. “implementation”. The Center for Research We hypothesize that oxidative stress in the Implementation of Innovative contributes to development of cardiovascular Strategies in Practice (CRIISP) was dysfunction in metabolic syndrome and that established in June 2004 as a new VA the source of oxidative stress is non- Health Services Research and Development phagocytic NAD(P)H oxidase. To test this (HSR&D) Center of Excellence focusing on hypothesis vascular function will be implementation or transfer of information. examined in vitro in vessels from several vascular beds pertinent to the cardiovascular CRIISP’s goal is to create ways to channel system. In addition, we will determine the research information to clinicians in a more pro-atherogenic mechanisms pertinent to direct fashion in order to improve veterans’ vascular disease, and investigate enzymatic well being. Specifically, the center aims to: - 1) identify existing processes that assist or sources responsible for superoxide (O2 ) production. hinder the transfer of research information; Over 50 million Americans have the and 2) develop, test, and distribute new metabolic syndrome. The goals of treating processes that improve the quality and costs the metabolic syndrome are prevention of of medical services. cardiovascular events and development of type II diabetes mellitus. The initial step in CRIISP’s work will focus on illnesses, such curbing the metabolic syndrome epidemic as heart disease and diabetes, that are would be to identify individuals at the earliest common in veterans and cause significant possible stage of the syndrome, before suffering and disability. CRIISP will also cardiovascular disease and diabetes focus on patients who suffer from more than develops. This study will allow us to study one condition and seek to develop new

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implementation strategies that can address medicine, patient-centered care, decision several conditions at once, recognizing that support system, health services research physicians would be overwhelmed if separate implementation strategies were used for each medical condition. In addition, CRIISP will develop ways to improve care Principal Investigator: that are customized to the particular James Rossen, M.D. strengths and needs of each patient. The patient-focused emphasis of CRIISP is of Project Title: particular interest since prior work has Clinical Outcomes Utilizing shown that implementation may be more Revascularization and Aggressive Drug effective if the patient and physician both Evaluation agree on the treatment strategy.

The founding of CRIISP reflects a close partnership between VA HSR&D at the Iowa Summary: City VA Medical Center, the Veterans Integrated Service Network 23 (which Coronary heart disease remains the single- includes VA facilities in Iowa, Nebraska, most important cause of morbidity and Minnesota, and South Dakota), and mortality in the Western World. Because of researchers at the University of Iowa. The this, physicians have tried to identify the CRIISP team includes physicians and most effective clinical strategies to reduce or nurses, as well as experts in several other eliminate symptoms of angina pectoris, areas, such as psychology, public health, improve exercise performance and quality of health economics, health care organizations, life, and to improve survival. Numerous large public policy, business and sociology. clinical trials have established the CRIISP is also working to establish a importance of medical therapy in the network of health care organizations with management of coronary heart disease, vested interests in improving health care while prospective trials have established the delivery. These organizations will be roles of coronary artery bypass graft surgery important sites for testing the usefulness of versus medical therapy. Percutaneous new strategies. transluminal coronary angioplasty has been

safely and effectively used to dilate CRIISP will complement other VA Centers obstructed coronary arteries. The number of throughout the nation that are leaders in PTCAs being done has grown exponentially improving healthcare. To support these without the guidance afforded by efforts, CRIISP will sponsor several new randomized controlled trials. The activities including: a website providing COURAGE trial will be the first large-scale, measures and tools that can be used by multicenter, randomized controlled trial that researchers in designing new projects and is powered for a combined trial primary by practicing clinicians; a biannual research endpoint of all-cause mortality and nonfatal conference that will bring together national myocardial infarction in coronary heart experts to review successful strategies for disease patients. The two therapeutic improving care and high-priority areas for strategies being compared randomly are future research; and a research “Summer PCI (percutaneous coronary intervention) in Camp” for medical administrators and addition to intensive medical therapy, versus leaders. Through these activities and by a strategy of intensive medical therapy developing implementation strategies that alone. The primary hypothesis for the study improve the quality of medical services, is that PCI plus intensive medical therapy is CRIISP aims to improve veterans’ well being superior to intensive medical therapy alone. and be a national resource in the area of implementation. MeSH Terms: cardiovascular diseases,

coronary disease, myocardial ischemia MeSH Terms: quality of health care, quality of indicators, evidence based

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frequency of gynecologic health risk Principal Investigator behaviors in sexually assaulted veterans with that of their non-assaulted peers. Anne Sadler, Ph.D., R.N.

It is anticipated that our findings will improve Project Title: evidence-based clinical practice. This study Sexual Violence Exposures and could identify a population of women for Women Veteran’s Gynecologic Health whom more aggressive screening or interventions may be more appropriate. Determining the associations between The Women’s Health Act of 1992 gynecologic health, sexual violence acknowledged women sexually victimized in exposures, and gynecologic services the military as a priority-care population for utilization could guide national resource the Department of Veterans Affairs (DVA). allocation for a priority population and would Subsequently, it has become a major health have direct implications for DVA policies care provider for women whose reproductive regarding women’s health care. health may be at risk. However, the long- Furthermore, these results could help term gynecologic health or health services determine if subsequent population based or utilization of military women or sexually intervention studies are needed to address assaulted women has not been well studied. disparities in health care for women Last year (2004) the American College of veterans. Obstetricians and Gynecologists recommended decreasing the frequency MeSH Terms: rape, cervical cytology, with which the average woman has gynecologic health, women veterans, gynecologic screening. The applicability of cervical smears, sexual violence, health these preventive health recommendations services utilization for women who have been raped is unknown.

This cross-sectional research will study Principal Investigator: 1088 women veterans who have sought Mary S. Vaughan Sarrazin, Ph.D. care from the Iowa City VAMC within the preceding two years to examine the Project Title: association between sexual violence Estimating the Magnitude of Unmeasured exposures and gynecologic health behaviors Risk in VA Patients and health care utilization. Consenting participants will complete a telephone interview and medical records (both VAMC and private) will be reviewed. Summary:

The objectives are: 1) To determine if the odds of current Numerous studies over the past two gynecologic disorders are significantly decades have compared patient outcomes greater for sexually assaulted veterans in in VA and private sector hospitals. A critical comparison to non-assaulted peers. limitation of these studies is uncertainty 2) To identify if the presence and frequency about the adequacy of methods used to of cofactors known to be associated with adjust for differences in illness severity. cervical cytologic abnormalities (e.g. Users of VA services have unique smoking) is greater in sexually assaulted socioeconomic, clinical, and psychosocial veterans when compared to non-assaulted characteristics. The failure to adequately peer. account for these characteristics makes it 3) To determine if the frequency and types difficult to discern whether observed of gynecologic services used by sexually differences in risk-adjusted outcomes assaulted women veterans in comparison to between VA and private sector hospitals are that of non-assaulted peers. due to unmeasured illness or to differences 4) To identify and compare type and in the quality of care. By identifying VA

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users who are hospitalized in private sector hospitals, we can compare outcomes of patients who are VA users with outcomes of Principal Investigator: other patients treated in the same hospitals. Warren Schmidt, M.D., Ph.D. Any difference in outcomes between the two groups reflects unmeasured severity of Project Title: illness. A Multi-center Trial to Evaluate the Thus, the primary goal of this study is Epidemiology, Natural History, and identify whether VA utilization is an Treatment Response of Hepatitis C in the independent risk factor for poor outcomes. United States Veterans Population Specifically, we will compare outcomes of VA users and other patients who receive care in private sector hospitals for 10 high- Summary: volume medical and surgical diagnoses, adjusting for severity of illness. We will also identify characteristics of VA users, such as Clinical Research Studies: disability level, that are associated with poor Our clinical research involves the treatment outcomes. Using Medicare and VA of hepatitis C. Hepatitis C is a common viral administrative data, admissions to private infection in veterans and can cause chronic sector hospitals for the 10 medical and liver disease and even progress to cirrhosis surgical diagnoses in 1996-2001 will be and liver cancer. We are working with the identified. Patients who also used VA major pharmaceutical corporations services during the two years preceding the (Schering Plough and Roche) to further test private hospital admission will be identified. interferon and ribavirin therapies for Endpoints will be compared in patients who veterans with hepatitis C. These medicines did and did not use VA services prior to the are the only FDA approved treatment for private sector hospitals admission, adjusting hepatitis C and under the right conditions for observed patient risk factors. can effectively lead to permanent remission This study will provide new insights into the in 50 to 80% of patients. Our research is magnitude of risk in VA populations that has also determining whether maintenance been unmeasured in prior studies, and, in so therapy with interferon alone is a worthwhile doing will add to our understanding of the treatment to delay progression of the liver relative quality of care in VA hospitals. In disease in some patients that have addition, the study will identify specific advanced scarring and early cirrhosis. patient characteristics that explain differences in outcomes between VA users Basic Science Research Studies. and other patients (e.g., disability level). Our basic science research is studying how This information will assist in the the hepatitis C virus causes injury and development of improved risk-adjustment disease in the liver of infected patients. methods for use in future comparisons of VA Hepatitis C is a small virus that causes and private sector facilities. persistent infection, ie, it does not “go away” in the majority of patients that have the MeSH Terms: risk adjustment, veterans, disease. The virus lives in the cells of outcomes research, outcomes human liver and causes persistent scarring assessment, hospital mortality or “fibrosis” that is the major cause of cirrhosis and liver cancer. We have found that the virus may cause chronic injury to liver cells by interfering with enzyme pathways that usually protect the liver from damage to proteins and DNA. Our research is important because it is likely to identify new methods for treatment and prevention of advanced liver disease such as cirrhosis.

MeSH Terms: hepatitis C, chronic hepatitis, cirrhosis, liver disease

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structure and process factors that influence institutional efforts to translate evidence into Principal Investigator: best practice and bridge the quality chasm. John P. Schneider, M.D. The results will in turn enable clinicians, managers, and policy makers to gain a Project Title: better understanding of the resource The Effect of Clinical Practice Guidelines on implications associated with changes in Costs clinical and organizational structures and processes.

Summary: Principal Investigator: The primary goal of this project is to Konrad Schulze, M.D., FRCP determine the extent to which the adoption and implementation of clinical practice Project Title: guidelines leads to changes in the costs of Mechanics of Stomach and Intestine providing care within a healthcare system. Our main objective is to statistically evaluate the effect of two clinical practice guidelines on utilization and treatment costs in the VA. Summary: We propose to study the economic effects of two national VA clinical practice guidelines, Disorders of gastric emptying and diabetes mellitus (DM) and major depressive gastrointestinal transit are common in disorder (MDD), disseminated to VAMC Veterans, and contribute abdominal pain directors in 1998 and 1999 for system wide syndromes, reflux disease, diarrhea, and implementation. Our project has three aims. other chronic diseases. Diagnosis and First, we will create a unique database that management of these problems remains links together information on a large national empirical because little is known about the cohort of well-characterized patients in the fluid mechanical events that lead to particle 1999 Large Health Survey of Veteran dispersion, the interactions of substrate with Enrollees (LVHS) with utilization data from digestive secretions and the delivery of the VA inpatient and outpatient files. These data products of digestion to sites of epithelial will be linked with new VA cost estimates transport. We have been collaborating with provided through the VA Health Economics the team of Professor Udaykumar, Resource Center (HERC) and the VA mechanical engineering, University of Iowa, Decision Support System (DSS). The in modeling of intestinal flow. Using visual analytic file will be constructed as a panel of parameters of intestinal contractions we patients and hospitals, spanning six years recorded previously on videotape, we have from 1999 to 2004. Second, using VA chart- calculated flow parameters such as velocity review data on guideline adherence, we will vectors, pressure contours, the mixing of classify VA facilities according to the various fluid boluses and the dispersion of intensity of DM and MDD guideline particles. For work on particle dispersion, we implementation. Third, we will determine the have furthermore enlisted the collaboration extent to which the intensity of DM and MDD of Professor Jeffrey Marshall, who has guideline implementation at VA facilities is refined the patterns by which particles associated with differences in resource scatter and fluids are drawn out in sheets in utilization and treatment costs among our response to movements of intestinal wall. study cohort patients. HERC and DSS cost This generation of large surface areas estimates will be compared. Determining the between secretions and substrate is critical impact of practice guideline implementation to the process of digestion. on costs within a healthcare system will contribute importantly to a growing body of MeSH Terms: gastric emptying, health services literature on creating a gastrointestinal transit, peristalsis, “business case of quality.” The project will absorption also provide insight into organizational

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Principal Investigator: Principal Investigator: William L. Sivitz, M.D. Jack Stapleton, M.D.; Jinhua Xiang, M.D.

Project Title: Project Title: Dr. Sivitz’s Research Project GB Virus C-HIV Interactions

Summary: Summary:

Dr. Sivitz works on glucose and fat GB Virus C (GBV-C, also called hepatitis G metabolism as related to diabetes and virus) is a flavivirus, which is a close relative obesity. One important focus is on the role of hepatitis C virus. It causes persistent of mitochondria in regulating the rate of infection in humans and is very common, metabolism of fat and glucose. Within body infecting approximately 1.8% of healthy cells, glucose and fat are metabolized to blood donors and up to 40% of immune metabolic products, which are then able to compromised individuals. GBV-C has not enter mitochondria. Mitochondria represent been found to cause any known disease, the major structures within all body cells prompting the FDA to recommend that the where these products of glucose and fat are blood supply does not need to be tested for ultimately burned for energy. Dr.Sivitz’s lab this virus. Over the past several years, there is particularly interested in the role of has been substantial controversy regarding mitochondrial proteins called “uncoupling the interactions between GBV-C and HIV in proteins”. The lab is investigating whether vivo. A number of studies have these proteins alter the rate of metabolism of demonstrated a surprising survival benefit glucose and fat and how the action of these among HIV-infected patients, who are co- proteins is triggered. infected with GBV-C. These individuals The laboratory is also investigating whether delay in the progression of their HIV disease fat and glucose alter the rate of production when compared to HIV infected patients of potentially damaging substances called who are not infected with GBV-C. We found “reactive oxygen species” or ROS. The lab that co-infection of human lymphocytes with is studying how mitochondria might regulate GBV-C and HIV led to inhibition of diverse ROS production. Understanding this could HIV strains including those that utilize either help control excessive ROS production that of the two HIV co-receptors (CCR5 and occurs in states such as diabetes and CXCR4) for viral entry. The inhibition of HIV obesity and could lead to better means to was mediated by GBV-C including the protect against ROS. This is important since production of natural ligands (Rantes, MIP- ROS may damage blood vessels, nerves, 1a, MIP-1b and SDF-1) for CCR5 and and other structures and lead to the CXCR4, and by decreased the expression of complications of diabetes and obesity. CCR5 on the surface of lymphocytes. There Dr. Sivitz is also involved in clinical projects are an estimated 294,424 people in the directed at prevention of diabetes United States living with AIDS and more complications and in a project that examines than 430,000 people in the U.S. have died of the effect of the nervous system to regulate AIDS. Better understanding of the reasons the response to low blood sugar that can why HIV disease is delayed in many HIV- complicate the treatment of diabetes. infected people, and the development of novel approaches for delaying HIV disease MeSH Terms: obesity, diabetes, progression is of critical importance for mitochondria, sympathetic nerve activity, altering the destructive nature of this insulin sensitivity, insulin, glucose, fatty epidemic. To our surprise, GBV-C leads to acids decreased HIV replication in vitro, thus the study of how GBV-C leads to decreased replication seems warranted. Although very speculative, the therapeutic use of GBV-C

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for HIV must be considered as a potential including phosphorylation, and degradation novel approach to HIV therapy. At the least, of the channel components. The information determining the mechanism(s) of GBV-C from these experiments will help us better inhibition of HIV replication may lead to the understand mechanisms whereby high development of vaccine and new blood pressure is produced and thus better approaches towards treating and protecting treatments. It is also possible that this HIV infection. information will be applicable to other diseases that are often associated with MeSH Terms: GB virus, HIV, AIDS, hypertension, such as diabetes, and high coinfection, co-receptor, natural ligand blood lipids.

MeSH Terms: hypertension, ion channels, sodium channels Principal Investigator: John B. Stokes, M.D.

Project Title: Principal Investigator: Long-term Regulation of Sodium Channels Gail Takahashi, Ph.D.; Micheal Crum, Ph.D.; Janet Shanks, Ph.D.; Richard Wilson, Ph.D.; Barbara Peek, Ph.D.; Stephen Fausti, Ph.D., Gabrielle Saunders, Ph.D.; Charles Summary: Martinez, Ph.D.; Douglas Noffsinger, Ph.D. Our laboratory studies the regulation of the epithelial sodium channel. This complex of Project Title: three proteins comes together to form a A Long Term Follow-Up of Patients in the pore, or channel through which sodium NIDCD/VA Hearing Aid Clinical Trial passes. Epithelial cells line structures that separate compartments of the body, such as the intestine, the urine space of the kidney and the ducts of sweat and salivary glands. Summary: This channel plays a major role in determining how much sodium humans A research study conducted from 1996 to excrete in their urine. The balance between 1998, entitled “CSP #418: The NIDCD/VA the amount we eat and the amount we Hearing Aid Clinical Trial," compared three excrete is critical to maintain normal sodium commonly used hearing aid circuits. Each (and fluid) balance and also blood pressure. research participant used each hearing aid Several genetic forms of high blood circuit in random order over a 3-month pressure are caused by mutations in one of period, and the benefits of each circuit were the proteins that form this sodium channel. evaluated. For most of the participants, the Other kinds of mutations can cause study hearing aids helped them to hear inactivation of this ion channel and produce better, and enabled them to understand severe loss of sodium and too low blood speech more clearly even in noisy pressure. Our current work involves situations. Approximately five years have determining how agents, such as sodium now passed since that study was completed. retaining steroids, can increase the activity We are now conducting a follow-up study of this channel and thus produce sodium involving participants from the original study. retention and raise blood pressure. We are We want to know if their hearing has stayed also studying the molecular nature of agents the same, if any major changes have that produce a lower activity of this channel occurred in their lives, and if they are still and thus produce more sodium excretion in using the original study hearing aids--or any the urine and lower blood pressure. The hearing aids. If they are using hearing aids, pathways we are studying involve synthesis we want to know how they like them. If they of the individual proteins, assembly of the are not using the study aids, or any hearing proteins, activation of channel function by aids, we would like to know why they enzymes through a cascade of events changed to different aids or stopped using

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hearing aids. To this end, we are re- their own telephones at home or at work. evaluating the participants using a standard Finally, we will be gathering questionnaire audiometric test battery and measuring the data on telephone use in individuals who performance of their hearing aids. We ask a wear hearing aids. The results may help us series of questions about life changes during understand the difficulties that individuals the past five years, and about participants’ with hearing loss face when using the experiences with hearing aids. The data we telephone and may provide guidance on collect may help us to understand why some programming hearing aid telecoil settings as people do well with hearing aids and why well as counseling patients about telephone others reject hearing aids. This information use when wearing hearing aids. will help us make more informed hearing aid fitting decisions for other individuals with MeSH Terms: hearing loss, hearing aids, hearing loss. rehabilitation of the hearing impaired, telephone, treatment outcome MeSH Terms: hearing loss, hearing aids, rehabilitation of the hearing impaired, treatment outcome Principal Investigator:

Gail Takahashi, Ph.D.

Principal Investigator: Project Title: Gail Takahashi, Ph.D. Perceived Hearing Difficulties and Hearing Aid Satisfaction in Veterans Vs. Significant Project Title: Others Clinical vs. Real World Performance with Telecoils

Summary:

Summary: There are numerous self-report questionnaires that can be used to assess Many hearing aids are equipped with the impact of a hearing impairment. Self- telecoils, which allow users to use their report questionnaires can also be used to hearing aids on the telephone without measure the perceived benefit and generating acoustic feedback. The satisfaction associated with hearing aid use. capability of programming the overall gain This study will examine the perceived and frequency response of the telecoil is hearing difficulties as well as hearing aid available on some hearing aids. However, benefit and satisfaction using two self-report at present, there is no standardized protocol questionnaires: the Hearing Handicap for setting these parameters. In addition, Inventory for the Elderly (HHIE-S) and the setting the telecoil response is problematic International Outcome Inventory for Hearing in a clinical setting because there are many Aids (IOI-HA). The purpose of this study is factors that influence real world speech to compare veterans’ perceptions of the communication via the telephone, such as social and emotional impact of their hearing the acoustic characteristics of the listener’s loss to the perceptions of their significant telephone as well as the talker’s telephone. others. We will also be examining the This study seeks to compare word effectiveness of hearing aids as reported by recognition scores obtained using default veterans compared to the effectiveness telecoil settings determined by a proprietary perceived by their significant others. The algorithm developed by the hearing aid results may help us in counseling veterans manufacturer to scores obtained using and their significant others about the impact clinically optimized programmed telecoil of hearing loss and the benefit of hearing settings. We are also interested in aids. examining whether word recogniton scores obtained using a clinic telephone are similar to those obtained when participants use

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the projections and the preganglionic cells MeSH Terms: hearing loss, hearing aids have morphological characteristics of rehabilitation of the hearing impaired, excitatory synapses. Our data suggest that patient satisfaction, treatment outcome an excitatory amino acid may serve as a questionnaires transmitter at those synapses and excitation of the pontine cells leads to cerebral vasodilatation as a result of the release on cerebral vessels of nitric oxide derived from Principal Investigator: neuronal nitric oxide synthase. Through our William T. Talman, M.D. studies we seek to define the histochemistry of synapses formed by SSN neurons and Project Title: projections from the NTS and to study Parasympathetic Influences on Cerebral pharmacological mechanisms that underlie Blood Flow neuro-genic cerebral vasodilatation as a result of stimulation of the pathway. Each of these issues will be important as we Summary: approach our long-term goal. to determine if stimulation of the barorecep-tor/pontine parasympathetic pathway may lead to Some arteries have within them specialized neural protection during acute cerebral nerve endings called baroreceptors that play ische-mia. The proposed studies will further an acknowledged role through the baroreflex our hypothesis through two specific in regulating tone with peripheral arteries. objectives. First we will seek to determine if There has been controversy about the role vasodilatation elicited by activation of of the baroreflex in regulating cerebral baroreceptor/SSN pathways depends on vascular tone. Our studies have shown that local synthesis of nitric oxide by neuronal interruption of either arterial baroreceptor nitric oxide synthase and second we will nerves or the nucleus tractus solitarii (NTS) seek to determine if projections from NTS where those nerves terminate has similar make glutamatergic synapses on pontine effects on cerebral blood flow (CBF). preganglionic neurons in SSN. The Specificaly, those lesions lead to significant combined studies will further our blunting of vasodilatation seen when the understanding of a potentially important cerebral circulation is exposed to very high neural mechanism for regulating cerebral blood pressures that exceed the capacity of circulation and provide the foundation for cerebral blood vessels to autoregulate CBF. pharmacological studies of transmission in Likewise, the dilatation, referred to as SSN. In that published studies suggest that “breakthrough”, was inhibited by systemic interruption of parasympathetic nerves to the administration of inhibitors of nitric oxide cerebral vessels may increase neuronal synthesis or by interrupting parasympathetic damage produced by cere-bral ischemia, nerve fibers that release nitric oxide on our proposed studies may, likewise, have cerebral arteries. Interruption of the same implications for protecting the brain during parasympathetic input to cerebral vessels stroke. has been shown to increase the size of cerebral infarctions pro-duced in a stroke MeSH Terms: cerebral blood flow, model. Our findings led to the hypothesis glutamate, mechanoreceptor, medulla, that the arterial baroreflex modulates neurotransmitters, nitric oxide, cerebrovascular resistance through parasympathetic, pons, solitary tract connections to the preganglionic parasympathetic innervation of nitroxidergic neurons in the pterygopalatine ganglia. We have now established that the NTS projects to and synapses with parasympathetic preganglionic neurons in the pontine superior salivatory nucleus (SSN) at a site whose stimulation leads to cerebral vasodilatation. Synapses formed between

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the renal collecting duct, study the function Principal Investigator: and localization of the expressed protein and begin to examine the basis for cell- Christie P. Thomas, M.D. specific gene expression and identify the

mechanism of PKC stimulation of selected Project Title: Nedd4-2 transcripts. Altered expression or Regulation and Function of Nedd4-2 regulation of hNedd4-2 gene expression Isoforms in Epithelial Na+ Transport could lead to abnormal Na+ handling by the kidney and other epithelia with resulting effects on blood pressure and effective Summary: circulating volume.

+ MeSH Terms: epithelial amiloride Na transport in the collecting duct of the sensitive Na+ channel, kidney collecting kidney occurs via the hetero-multimeric + ducts, gene expression, ubiquitin protein epithelial Na channel (ENaC). The level of ligase channel activity is regulated at distinct transcriptional and post-transcriptional steps including pathways that regulate insertion and retrieval of the assembled channel to Principal Investigator: and from the cell surface. Mutations that Bonnie Wakefield, Ph.D., R.N. disrupt an endocytosis motif in ENaC + subunits result in unregulated Na Project Title: reabsorption and an inherited form of severe Evaluating Telehealth Home Care for Elderly hypertension. Ubiquitin ligases with multiple Veterans with Congestive Heart Failure WW domains appear to be involved in regulating ENaC removal from the cell surface, thus exerting a tonic inhibition on Na+ transport. It is now know that Nedd4-2, Summary: a ubiquitin ligase, reduces ENaC-dependent Na+ transport. Furthermore, Nedd4-2 is Congestive heart failure (CHF) is one of the expressed in the kidney cortex and in most common reasons for hospitalization in epithelia derived from kidney cortex, patients aged 65 years and older. Many suggesting that Nedd4-2 may function as a hospitalizations for CHF are potentially regulator of ENaC function in the collecting preventable if warning signs such as weight duct. gain and shortness of breath are recognized The principal goal of this project is to and treated early. Home-based nurse care determine the expression pattern, function management programs have reduced and regulation of hNedd4-2 transcripts and hospital admission rates for patients with its encoded protein variants in the collecting CHF by up to 50%. Using advanced duct and in a lung epithelial cell line with technology it is now possible to provide care regulated Na+ transport. We have evidence management services directly to patients’ that the hNedd4-2 has a complex gene homes more efficiently. The purpose of this structure with multiple 5' exons where randomized controlled clinical trial is to transcription is probably initiated in a tissue compare the effectiveness and resource use specific manner, and data to show that a of two telehealth interventions to traditional diverse array of transcripts are created. As care provided for recently discharged a consequence of this structural outpatients with CHF. Intervention patients organization, variant protein isoforms are receive telephone or videophone follow-up translated. Additionally, at least one of for 90 days after hospital discharge. these transcripts is regulated by activation of Outcomes of interest include readmission PKC and by a MAP kinase, establishing a rates, quality of life, self-efficacy, satisfaction putative mechanism by which these with care, hospital days, urgent care visits, pathways can regulate Na+ transport. In this and survival rates. Telehealth care will proposal we will determine which 5' and enable earlier detection of key clinical which WW domain variants are expressed in symptoms, triggering early intervention and

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thus reducing the need for hospitalization. visual field change is the single most difficult Reduced hospitalization will result in problem in visual testing today. We decreased overall costs, and improved anticipate identifying a method that allows outcomes for veterans with CHF. efficient and accurate determination of visual field change. Identification of a superior MeSH Terms: telemedicine; heart failure, method would (1) reduce the number of congestive; home care services; examinations needed, thereby reducing the telephone costs of medical care; (2) minimize misdiagnosis, unnecessary testing and even unnecessary surgery that results from mistakenly interpreting fluctuation of the Principal Investigator: visual field as progression or improvement; Michael Wall, M.D. (3) allow earlier disease intervention and (4) reduce the costs of clinical trials. Project Title: MeSH Terms: glaucoma, vision testing, Mechanisms of Perimetric Variability visual loss, perimetry

Summary: Principal Investigator: Disease of the optic nerve, including Neal Weintraub, M.D. glaucoma, is the leading cause of blindness in the United States. Treatment decisions for Project Title: optic nerve diseases are based largely on Inflammatory Mechanisms in Vascular the changes in visual function that occur Disease mostly as a consequence of disease progression. Unfortunately, the decision as to whether change of visual function has occurred is often difficult because of the high Summary: retest variability of conventional visual field testing (perimetry). This variability is so high The overall focus of my research is to that with moderate visual loss, a minimum of understand what causes heart disease six tests are often needed in patients with (atherosclerosis). While a number of risk optic nerve damage to reliably distinguish factors, such as smoking, diabetes, elevated visual field deterioration from random cholesterol, and high blood pressure variation. Our preliminary data show that a contribute to the development of heart substantial portion of the variability of disease, many patients lack these traditional perimetry lies in the type of stimulus used risk factors. My laboratory is attempting to and the testing strategy applied. We learn how inflammation, which is propose the hypothesis that a large portion characteristically present in diseased blood of total perimetric variability in patients with vessels, contributes to the development visual loss is due to a poor signal-to-noise of atherosclerosis. We believe that blood ratio associated with using a small fixed-size vessels respond to factors circulating in the stimulus. To test this hypothesis, we are blood that are perceived as "foreign examining patients with optic nerve diseases invaders", thereby triggering an with conventional automated perimetry (size inflammatory response. This local III) and tests having large-sized and scaled inflammatory response is analagous to what stimuli (size V, multifocal visual evoked happens in other parts of the body, such as response and motion perimetry). Over five the lungs, when invading bacteria are years we are testing 180 patients with optic detected. White blood cells are recruited to neuritis, idiopathic intracranial hypertension, the area in order to help to kill the bacteria. and glaucoma and 60 normals each eight However, recruitment of white blood cells times. We are also studying the associated into blood vessels is harmful and leads to structural-functional correlations. Perimetric atherosclerosis and other types of vascular variability and the reliable identification of disease, such as abdominal aortic

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aneurysms. We believe that many sub- collection of phagocytes and inflammatory clinical infections, such as bronchitis, sinus exudates from experimental animals; in vitro infections, peridontal disease (gingivitis), and ex vivo functional assays of phagocyte and diverticulitis (infections in the colon) (or cell-free protein)-bacteria interactions release substances into the blood such as phagocytosis, bacterial killing and that trigger inflammation of blood vessels. digestion; expression and purification of Also, we think that certain viruses may recombinant proteins including PLA2 contribute to the development of heart variants; and construction and use of disease. We are trying to understand how bacterial mutants. The long-term objectives blood vessels sense and respond of this work concern two fundamental to infections, and how the inflammation questions: 1) What determines the extent of causes the vessels to become diseased. bacterial digestion and disassembly during One area of emphasis is oxidative stress, a host antibacterial action? 2) What regulates condition caused by excess free radical the action of defined phospholipases on the production. Free radicals are abundantly PL of biological membranes? The proposed produced by white blood cells and therefore studies are likely to provide new insights may be a direct consequence of the related to the mechanisms of host defenses inflammatory process in blood vessels. We in infection and of the regulation of are studying the enzymes produce the free phospholipase action in biological radicals and whether treatments that membranes more generally. scavenge free radicals may also attenuate FINDINGS: We have discovered a the disease processes. previously unrecognized antibacterial collaboration between the extracellular MeSH Terms: atherosclerosis, oxidative inflammatory fluid Group IIA PLA2 and the stress, inflammation, free radicals, respiratory burst NADPH oxidase of PMN aneurysm against ingested Staphylococcus aureus. Effects are seen both when otherwise limiting amounts of extracellular PLA2 are present before phagocytosis or even after Principal Investigator: bacteria have already been ingested. These Jerrold Weiss, Ph.D. effects require catalytically active Group IIA PLA2 and are not seen with other closely Project Title: related but non-antibacterial human Neutrophils and Bacterial Phospholipid secretory (low Mr) PLA2. Experiments are Degradation in progress to better define the mechanism of oxidase-induced increased PLA2 activity vs. S. aureus and the means by which extracellular PLA2 gains access to PL of Summary: ingested bacteria. CLINICAL RELEVANCE: S. aureus is an OBJECTIVES: This proposal concerns the important human pathogen responsible for role and regulation of the action of significant acute and chronic infectious mammalian Group IIA phospholipase A2 disease and sequelae. In chronic (PLA2) during inflammation. Our specific granulomatous disease (CGD), these aims are to examine: 1) the molecular bacteria are frequent causes of serious and determinants of the antibacterial actions of repeated infections. These clinical findings mammalian Group IIA PLA2; 2) the originally helped define the clinically regulation and role of PLA2 antibacterial important role of the phagocyte respiratory action at inflammatory sites; and 3) the burst oxidase in host defense against molecular determinants and possible bacterial infection. The discovery of an physiologic role for Group IIA PLA2 action important role of the phagocyte respiratory against host cell-derived membrane burst oxidase in the action of the potently phospholipids (PL). These aims have their antistaphylococcal Group IIA PLA2 in origin in our earlier work and their pursuit will neutrophil-rich inflammatory settings therefore rest heavily on well-tested indicates a previously unappreciated methods used in our laboratory including: integration of cellular and extracellular,

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oxidative and non-oxidative antibacterial protein has been isolated and cloned in our systems in host defense against S. aureus. laboratory and implicated as an important The apparent importance of oxidase intracellular and extracellular cytotoxin activation in PLA2 action may also help against GNB and negative regulator of host explain the chronic inflammation that responses to endotoxin. Our specific aims characterizes this disease (i.e. persistence are to further define the molecular of undegraded bacterial lipids and possibly determinants of interactions of BPI with other envelope components could drive intact GNB, shed outer membrane (OM) protracted inflammatory and immunoligc vesicles (“blebs”) and purified endotoxin, the reactions and, hence, immunopathology). cellular and molecular fates of endotoxin Even in the presence of normal PMN, following these interactions and digestion of ingested S. aureus is limited, if accompanying host cellular responses. PLA2 is limiting, possibly creating METHODS: The pursuit of these aims rests circumstances favorable for persistence of heavily on well-tested methods used in our small numbers of viable bacteria and larger laboratory including: induction and collection amounts of bacterial remnants that may lead of inflammatory exudates from experimental to recurrent infection and chronic animals; purification of native BPI and inflammatory/immunologic disorders. In expression and purification of recombinant settings of dimished cardiovascular function, wild-type and mutant proteins; construction compromised tissue perfusion may also of bacterial mutants to permit high specific create circumstances leading to incomplete metabolic labeling of bacterial endotoxin to elimination of normally incidental bacterial facilitate study of protein:endotoxin:cellular incursions and contribute significantly to interactions. chronic disease. FINDINGS: We have identified a novel role of scavenger receptor ligands in regulating MeSH Terms: gram-negative bacteria, BPI-dependent delivery of purified and gram-positive bacteria, neutrophils, membrane-associated endotoxin to CD14+ inflammation, endotoxin, phospholipids, monocytes. Depending on dose, SR ligands bactericidal, phospholipases, may either markedly enhance or suppress deacylases, membranes, staphylococcus BPI-dependent cellular delivery of endotoxin aureus, antibiotics suggesting that variables in the inflammatory environment, independent of BPI and endotoxin per se, may be important determinants of the efficiency of BPI- Principal Investigator dependent endotoxin clearance. Initial Jerrold Weiss, Ph.D. cellular responses to uptake of BPI:coated endotoxin-rich particles are “silent” (i.e. no Project Title: detetable activation of cellular pro- Microbicidal Activity of Leukocytes: Active inflammatory responses). However, the Factors persistence of structurally intact endotoxin and co-delivery of other potentially bioactive OM components to these cells raises the possibility that clearance of BPI-coated Summary: endotoxin-rich particles could be linked to activation of antigen presenting cells. OBJECTIVES: This project concerns the During most incidental exposures to GNB, further exploration of mechanisms of host this may provide a physiologic link between responses and defense against Gram- innate immune recognition of endotoxin by negative bacteria (GNB) with a focus on the BPI and activation of adaptive immune molecular and functional characterization of responses. However, under more the bactericidal/permeability-increasing exaggerated conditions, this pathway may protein (BPI), an endotoxin-binding lead to antigen presentation of BPI coupled antibacterial protein of neutrophils. This with induction of cellular co-stimulatory protein will be studied both as an isolated responses leading to induction of BPI auto- protein and as a mobilized component of antibodies that are characteristic of PMN-rich inflammatory exudates. This conditions marked by acute or recurrent

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GNB infections and neutrophil mobilization wild-type littermates, while transgenic mice and activation. overexpressing ASIC1 exhibited more fear. CLINICAL RELEVANCE: Insights gained Because ASIC1 levels regulated the degree from studies of BPI-endotoxin interactions of acquired fear, these data suggest that will likely provide a better understanding of ASIC1 may contribute to anxiety disorders. host response to endotoxin and defense Moreover, because disrupting ASIC1 against invading GNB and may contribute to reduced anxiety without causing any gross the design and development of new physical or behavioral abnormalities, ASIC1 therapeutic agents active against invasive antagonists might provide a novel and safe GNB infections and endotoxemia needed approach for anxiolysis in the clinical setting. when endogenous defenses are limiting and Experiments exploring the role of ASIC1 in conventional antibiotics inadequate. A brain function and behavior, and exploring better understanding of factors that regulate the potential therapeutic value of ASIC1 BPI-dependent disposal of endotoxin and antagonists are currently underway. bacterial OM remnants and responses by the cells to which these bacterial products MeSH Terms: ASIC channel, fear, are deposited may also yield a better conditioning, post traumatic stress understanding of how endotoxin-driven disorder inflammation is normally resolved and how acute or persistent GNB infections can lead to inflammatory and/or immunologic sequelae including the generation of auto- Principal Investigator: antibodies to important host defense Mary E. Wilson, M.D. proteins. Project Title: MeSH Terms: lipoplysaccharides, Protective Immunity to Leishmania chagasi neisseria meningitidis, inflammation, endotoxin, endotoxin-binding proteins, capsule, deacylases, outer membrane, lipoproteins Summary:

Leishmania chagasi causes the potentially fatal disease visceral leishmaniasis in South Principal Investigator: America. Cure or protection from all John Wemmie, Ph.D. Leishmania sp. infections in mice is caused by a cellular immune response due to Project Title: certain T cells (CD4+ type 1 cells) that Exploring the Role of ASIC1 in Fear produce the protein interferon-gamma (IFN- gamma). In mice that get progressive infection with L. chagasi, the function of Summary: these “curative” T cells is inhibited by another protein produced in abundance by Anxiety disorders such as posttraumatic immune cells, called transforming growth stress disorder (PTSD) pose a major threat factor-beta (TGF-beta). During this project to veterans' health. Understanding the we are characterizing the immune molecular mechanisms of PTSD may lead to responses that lead to either the production improved treatment and prevention of this of the disease-curing cytokine IFN-gamma disabling illness. We recently found that the or the disease exacerbating cytokine TGF- acid sensing ion channel ASIC1 contributes beta. Our studies address which cells to Pavlovian fear conditioning, an important produce TGF-beta in the whole mouse, and animal model of acquired anxiety and PTSD. the role that the parasite itself plays in We generated knockout mice lacking the inducing the production of TGF-beta. ASIC1 gene and produced transgenic mice Finally, having characterized the curative overexpressing ASIC1. We found that and non-curative immune responses, we will ASIC1 knockout mice had less fear than test a number of vaccine strategies to immunize mice and elicit a curative immune

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response. These strategies involve use of associated with the different outcomes of DNA vaccines, and the use of non-infectious human L. chagasi infection. We are forms of the bacterium Listeria acquiring clinical and family information, and monocytogenes. Overall, these studies are obtaining DNA from individuals living in intended to delineate the roles of different neighborhoods outside of Natal where L. cytokine responses during infection and chagasi infection is highly endemic. Using immunization of mice with L. chagasi, in this information we are taking two preparation for generating a human vaccine. approaches to determining whether there are genetic loci associated with infection. MeSH Terms: parasitology; leishmania; First, we are examining alleles of candidate leishmania infantum; vaccines; DNA genes that we expect to influence the vaccines immune response to the organism. Second, we are performing a genome-wide scan to identify markers that are linked to different outcomes of infection. During future years, Principal Investigator: we hope to extend these results to studies of Mary E. Wilson, M.D. genetically determined antileishmanial immune responses in Brazilians that Project Title: develop different outcomes of disease. Immunogenetics of Leishmania chagasi Ultimately we hope to identify specific alleles Infection that influence human immune responses and thereby modify the course of disease.

MeSH Terms: parasitology; leishmania; Summary: leishmania infantum; human genetics

Visceral leishmaniasis is a potentially fatal tropical disease that affects humans living in Principal Investigator: endemic tropical and sub-tropical climates. Mary E. Wilson, M.D.; John E. Donelson, The disease is caused by the protozoan Ph.D. Leishmania chagasi. L. chagasi infection of humans leads to a variety of outcomes ranging from asymptomatic infection to Project Title: progressive fatal disease. Furthermore, The Major Surface Protease of many people living in endemic regions do Trypanosomatid Protozoa not harbor evidence of L. chagasi infection despite their likely exposure to the parasite. Several studies including our own have Summary: documented familial clustering of L. chagasi infection. This proposal will test the hypothesis that genetic factors predispose Leishmania chagasi is the protozoan individuals to develop the different clinical causing the fatal human disease visceral outcomes after L. chagasi infection. leishmaniasis in South America. Other Numerous cases of visceral leishmaniasis Leishmania species cause cutaneous have occurred in neighborhoods outside the leishmaniasis, which is prevalent in both the city of Natal in northeast Brazil over the past Old and the New World. Trypanosoma 10 years. In collaboration with Dr. Selma brucei sp. cause African trypanosomiasis, or Jeronimo in Natal we have examined >1400 sleeping sickness. These protozoan individuals living in these endemic parasites are cousins, despite the fact that neighborhoods. Using a combination of they cause very different human diseases. historical and clinical information we Both Leishmania species and Trypanosoma determined whether they had asymptomatic brucei subspecies have on their surface a or symptomatic L. chagasi infection, and major protein antigen which has enzymatic their likelihood of exposure. We propose to protease activity. The protein is called MSP use a genetic approach to identify factors or GP63. MSP enables Leishmania to resist killing by complement in human serum, and

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it helps them enter into macrophages where phagocytes during leishmania infection, and the parasite is “safe” from killing by the that parasites can inhibit responses leading immune system. The role of MSP in to oxidant generation. Which of these trypanosomes is not as well established. actually predominates, the macrophage or During this project we are examining the the parasite, may reflect the sum of expression of genes encoding the MSPs of opposing effects. The purpose of this Leishmania and of Trypanosoma brucei. proposal is to determine which macrophage Studies address the way the parasites antimicrobial mechanisms are mobilized to transcribe MSP RNA and translate MSP kill intracellular leishmania, and whether the proteins to enhance their survival in different effects of the parasite on its host cell allow host environments. The studies will or prevent intracellular parasite killing. We examine where the proteins are located will study infections with Leishmania within the parasites, and how they interact chagasi, the protozoan causing fatal visceral with human cells. We will compare the leishmaniasis in Latin America. Our overall functions of MSP proteins in T. brucei and L. hypothesis states that the outcome of chagasi. We hope through these leishmania infection depends on the balance comparisons to learn more about the between macrophage microbicidal events, functions of these similar proteins in each of parasite defense mechanisms, and parasite- these human pathogens. initiated changes in the capacity of macrophages to express microbicidal or MeSH Terms: parasitology; leishmania; immunoregulatory factors. Using a variety leishmania infantum; glycoprotein GP63; of biochemical, microscopic, and molecular trypanosoma brucei biology methods, we are defining the effector molecules that kill leishmania in the mammalian host, and at parasite mechanisms to evade these killing Principal Investigator: molecules. It is our hope that a better Mary E. Wilson, M.D. understanding of the means through which leishmania can be killed by host Project Title: macrophages will lead to new methods for Interactions of Leishmania chagasi with Host treating the disease. Our results will set the Macrophages stage for future investigations of the opposing host and parasite factors that lead to different outcomes of L. chagasi infection in humans residing in a highly endemic Summary: region of Brazil.

MeSH Terms: parasitology; leishmania; Leishmania sp. are obligate intracellular leishmania infantum; respiratory burst; protozoa that reside in mononuclear macrophage phagocytes of their mammalian hosts. They cause a variety of human infections in individuals residing in or traveling to the Principal Investigator: many countries where these infections are Mary E. Wilson, M.D.; John E. Donelson, endemic. Recent military operations have Ph.D. led to leishmania infections in the American veteran population. The final effector cell that eradicates leishmania is the Project Title: macrophage in which the parasite resides. Training in Mechanisms of Parasitism Activated macrophages can kill intracellular NIH/NIAID T32 (Training Grant) microbes by generating toxic oxygen radicals including superoxide, nitric oxide and their toxic byproducts (hydroxyl radical, Summary: peroxynitrite). Published literature contains conflicting reports showing that oxygen This grant funds three predoctoral and three radicals are formed in mononuclear postdoctoral US trainees to work with faculty

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on studies of parasitology, microbial leishmaniasis, both systemically and locally pathogenesis, or microbial immunology in in lesions. DNA will be collected to begin to several departments at the University of study disease susceptibility genes. Project Iowa. 2 will examine the contribution of factors derived from the sand fly vector to the MeSH Terms: parasitology, bacteria, immune response to disease. Project 3 is a immunology, training support family study of visceral leishmaniasis, focusing on the epidemiology and natural history of disease. A family study is being performed to look at the contribution of Principal Investigator: genetic factors to disease. Mary E. Wilson, M.D.; Jeffrey Murray, M.D. MeSH Terms: leishmania, immunology, Project Title: vaccines, parasitology, leishmania braziliensis International Maternal and Child Health Research and Training NIH Fogarty Foundation T32 (Training Grant) Principal Investigator: Patricia L. Winokur, M.D. Summary: Project Title: The purpose of this training grant is to Antibiotic Resistance in Food Animals and provide short-term and long-term training Humans opportunities for Brazilian students, postdoctoral fellows and junior faculty to train in laboratories at the University of Iowa. The focus of the training is on genetics of Summary: human diseases. Salmonella and E. coli are important MeSH Terms: genetics, human genetics, pathogens in humans and animals. Both Brazil, training support organisms have been linked to food-borne transmission where outbreaks have been associated with ingestion of contaminated poultry, meat, milk or dairy products. Our Principal Investigator: laboratory has recently identified Salmonella Mary E. Wilson, M.D.; Edgar Carvalho, and E. coli isolates obtained from porcine, M.D., Ph.D. bovine and human sources that are resistant to a very important human antibiotic, Project Title: ceftriaxone. This antibiotic is one of the main NIH/NIAID Tropical Medicine Research therapeutic agents for many infections in Center P50: Pathogenesis of Leishmaniasis: humans. All of the bacterial strains we Host, Parasite and Vector identified contained a very rare gene and Project 1: “Host and Parasite Factors in this gene was present on a type of DNA that Mucosal Leishmaniasis” can be readily transferred from one bacterium to another. Analysis of these genes and the pieces of DNA that carry Summary: these antibiotic resistance genes demonstrated that there were very strong This program is a Tropical Medicine molecular similarities between the animal Research Center focused on leishmaniasis and human isolates. These data suggest in northeastern Brazil. The purpose of that transfer of this resistance gene has project 1 is to characterize human immune occurred between food animals and man. responses to L. braziliensis in mucosal

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We have developed a rapid molecular Gender 60% 60% male n.s. screening techniques to detect the male ceftriaxone resistance gene in field samples. GAF 68.9 + 78.90 + >.05 We have been testing food products for E. 11.75 10.40 coli and Salmonella that contain this resistance gene. We have also been using Table 2: Psychiatric disorders and this test to detect ceftriaxone resistant E. coli Functional Somatic Syndromes in in the surface waterways of Iowa. There is PGW Veterans with Fibromyalgia significant fecal contamination of the Iowa and Controls surface waterways and it is concerning that poor waste management may be a way for FMS Controls p-value this very concerning antibiotic resistance to (n=40) (n=40) spread from place to place. A better Any 37(92%) 12 (30%) >.01 understanding of antibiotic resistance in Psychiatric organisms associated with food-borne Diagnosis illness will facilitate the development of 36 10 (25%) >.01 control strategies needed to protect animal (90%) health, the food supply and human health. Major Depression MeSH Terms: antibiotic resistance, Anxiety 30 2 (5%) >.01 ceftriaxone, foodborne infections Disorder (75%) Chronic 38(95%) 0 >.01 Fatigue Irritable 25 0 >.01 Principal Investigator: Bowel (62%) Catherine Woodman, M.D. Syndrome Chronic 20(50%) 5(12.5%) >.01 Project Title: Headache Fibromyalgia in Persian Gulf War Veterans: A Family Study Probands were age and gender matched. Fibromyalgia probands met American College of Rheumatology criteria. Subjects completed a Structured Clinical Interview for Summary: DSM-IV, the Functional Somatic Syndrome Interview, the NEO Personality Interview, This study of PGW veterans who were and the MOS-36. The results for probands recruited from the Persian Gulf Registry at are summarized in Table 1 and 2. Probands the Iowa City VAMC. Preliminary data is with FMS had significantly more psychiatric available for 40 probands, 40 controls and illness and comorbid functional somatic their first-degree relatives. The Registry syndromes than control veterans. Veterans includes PGW veterans who receive their with FMS had more than one Axis I health care at the Iowa City VAMC and psychiatric disorders 85% of the time, and those that have not received care there. comorbid anxiety and depression was most The study included 30% PGW veterans who common. They had lower quality of life, as received care at the Iowa City VAMC within measured by the MOS-36, more somatic the year prior to the study and 70% who had preoccupation, and higher scores on the not. NEO. First-degree relatives over 18 were contacted and interviewed. The results for Table 1: Demographics of PGW Veterans relatives are summarized in Table 3 and 4. with Fibromyalgia and Controls Relatives of FMS probands had significantly more fibromyalgia and psychiatric disorders FMS Controls p- than relatives of controls. (n=40) (n=40) value Age 31.35 + 32.82 + n.s. 6.9 8.1

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Table 3: Demographics of First Degree

FMS Controls p-value Principal Investigator: (n=202) (n=226) Chaoqun Yao, M.D., Ph.D.

Any Psychiatric 71(35%) 13 (8%) >.01 Diagnosis Project Title: Major 61 (30%) 12 (8%) >.01 Expression of the Major Surface Protease of Depression Leishmania Chagasi Anxiety 54 (27%) 5 (2%) >.01 Disorder Fibromyalgia 45 (22%) 4 (2%) >.01 Summary: Irritable Bowel 25 (12%) 1 (<1%) >.01 Syndrome Leishmania protozoa shuttle between a Chronic 35 (17%) 8 (4%) >.01 sand-fly vector and a mammalian host. Fatigue They cause human leishmaniasis that Relatives of PGW Veterans with affects 12 million people worldwide. Fibromyalgia and First-Degree Relatives of Leishmaniasis is important to the Control Veterans Department of Veterans Affairs health care system for two reasons. First, military FMS Controls p- personnel deployed in endemic area such (n=202) (n=226) value as Iraq, Kuwait, and Afghanistan may suffer Age 43.2 + 42.9 + n.s. from the disease. Second, Americans are 17.4 17.1 increasingly threatened by the disease even Gender 49% 48% n.s. in homeland, since domestic raised hunting male male dogs throughout 21 states have been found GAF 78.9 + 83.10 + .05 infected with the parasite, and dogs are the 10.34 10.40 most important reservoir hosts of human leishmaniasis. Leishmania spp. are coated Table 4: Psychiatric disorders and with a major surface protease (MSP or Functional Somatic Syndromes in GP63), which plays a vital role in their First Degree Relatives of PGW survival in mammalian hosts. At least 18 Veterans with Fibromyalgia and genes encode MSP in Leishmania chagasi. First-Degree Relatives of Control These fall into three categories (MSPL, Veterans MSPS, and MSPC) according to unique sequences in their 3’ untranslated regions. Expression of the different MSP gene Preliminary data revealed that PGW classes is post-transcriptionally regulated in veterans with FMS a significantly higher rate concert with parasites’ virulence and life of chemical sensitivity, dyspnea, and cycle stages. The reason they possess allergies than control veterans. Relatives in redundant genes encoding the same this study were not assessed for chemical protease is not clear. Most studies of gene sensitivity, dyspnea, or a history of atopy or expression in Leishmania address regulation asthma. of RNA levels, but do not address protein expression because of technical difficulties MeSH Terms: Persian Gulf War, veterans, in working with these protozoa. We propose stress, fibromyalgia, genetics several novel approaches to studying MSP protein expression. The goal of this project is to define which MSP class genes are expressed in different parasite stages, and to develop methods to functionally knockout each gene class separately. Objectives are: (1) To overexpress MSP genes in MSP-low parasite lines, and study cellular localization and growth-related expression. (2) To generate antibodies to detect the products of

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MSPC genes, which encode a unique velocity, thereby suggesting that vascular membrane anchor from MSPS and MSPL. related defects are primarily responsible for (3) To generate fluorescent reporters to the development of diabetic neuropathy in localize MSP gene products, and (4) To Type 2 diabetes. In addition, we propose develop the system for double stranded that hyperglycemia combined with RNA interference in L. chagasi, with the hyperlipidemia via formation of advanced purpose of selectively deleting function of glycation end products and/or an increase in individual gene classes. An understanding oxidative stress contribute to the of the mechanisms regulating MSP development and progression of vascular expression could lead to unique methods for and neural dysfunction in ZDF-obese manipulating parasite virulence factors, and diabetic rats. To test our hypotheses we are pave a way for new treatment modalities for addressing the following two objectives: 1) this fatal parasitic infection. Determine the sequential development and progression of vascular and neural MeSH Terms: Leishmania, major surface dysfunction in ZDF-obese diabetic rats, and protease, parasitic disease, gene 2) Determine whether treatment of ZDF- expression obese diabetic rats with rosiglitazone, aminoguanidine, pyridoxamine, α-lipoic acid or M40403 improves vascular and neural dysfunction. Our previous studies have Principal Investigator: demonstrated that vascular and neural Mark A. Yorek, Ph.D. dysfunction is apparent in ZDF-obese diabetic rats following 4 weeks of Project Title: hyperglycemia. We have not determined Pathology of Type 2 Diabetes whether slowing of motor nerve conduction velocity (neural dysfunction) in Type 2 diabetes is preceded by vascular dysfunction as in streptozotocin-induced Summary: diabetic rats, an animal model for Type 1 diabetes. If vascular dysfunction precedes The goal of this project is to determine the neural deficits in Type 2 diabetes it would cause of vascular and neural dysfunction in suggest that abnormal vascular activity is Type 2 diabetes. Obesity and many years responsible for early diabetic neuropathy. of escalating insulin resistance, chronic Such a result would suggest that hyperinsulinaemia and an ultimate failure of investigators should focus on correcting vascular dysfunction as a means of treating pancreatic islet β-cells to cope with the diabetic neuropathy. In this regard, our progressive demand for insulin characterize studies we will determine the efficacy of the pathology of Type 2 diabetes. The treating ZDF-obese diabetic rats with insidious nature of this process usually rosiglitazone, aminoguanidine, means that Type 2 diabetes often remains undiagnosed until the patient presents with pyridoxamine, α-lipoic acid or M40403, on chronic complications. The natural history of vascular and neural function. Combined, diabetic complications in Type 2 diabetes is these studies will provide us with a better less well known than in Type 1 diabetes in understanding and insight for potential part because of the difficulty in identifying treatment of diabetic vascular and neural the precise onset of the disease. To disease in Type 2 diabetes. address this issue we are examining the sequential development of vascular and MeSH Terms: diabetes mellitus Type 2, neural dysfunction and potential treatment of obesity, oxidative stress, peripheral vascular and neural complications in the nerve, antioxidants male Zucker diabetic obese rat (ZDF/Drt-fa), an animal model for Type 2 diabetes. We hypothesize that vascular dysfunction and reduction in endoneurial blood flow precedes slowing of motor nerve conduction

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and blood flow to peripheral nerves. Given this Principal Investigator: information one project in the laboratory is to test the hypothesis that compromising the function of Mark A. Yorek, Ph.D. CGRP contributes to nerve ischemia and the morphological/progressive changes that occur Project Title: during the second phase of diabetic neuropathy. Vascular Disease in Diabetic Neuropathy The objectives of this study are: 1) Determine whether loss of CGRP-sensory nerve innervation causes diabetes-like changes in Summary: vascular and neural function, 2) Determine the sequential effect of diabetes on CGRP sensory nerve innervation and bioactivity in The long range goal of my laboratory has been to epineurial arterioles in relation to morphologic determine the cause of diabetic vascular and changes in the peripheral nerves, and 3) neural disease. The pathophysiology of diabetic Determine whether the decrease in CGRP- neuropathy is not well understood. In a quality of sensory nerve innervation and CGRP bioactivity life assessment, patients with diabetic neuropathy caused by diabetes can be prevented and/or had significantly higher scores indicating an reversed. impaired quality of life compared to diabetic patients without diabetic neuropathy or non- MeSH Terms: diabetes mellitus, oxidative diabetic controls. This assessment emphasizes stress, peripheral nerve, antioxidants, the need for further research for the CGRP prevention/treatment of diabetic neuropathy. In diabetic animal models the pathophysiology of diabetic neuropathy can be divided into two phases. The first phase is characterized by Principal Investigator: metabolic derangements of the nerve and Mark A. Yorek, Ph.D. vascular tissue. The metabolic derangements are readily reversible and can also be prevented Project Title: by a variety of interventions, which implies that Effect of Angiotensin Converting Enzyme the cause of diabetic neuropathy is multifactorial. Inhibitor and/or Angiotensin 2 Receptor The second phase of diabetic neuropathy is Antagonist on Diabetic Neuropathy characterized by morphological derangement of peripheral nerves thought to be caused by ischemia and/or loss of neurotrophic factors. We have little understanding of the pathogenesis Summary: responsible for the second phase of diabetic neuropathy. To date clinical interventions to An effective treatment for diabetic prevent/treat diabetic neuropathy have been neuropathy is currently not available. disappointing. However, many of the treatments However, several well known classes of were designed to prevent the pathogenesis that drugs that have been proven to be safe and occurs during the first phase of diabetic beneficial for the treatment of hypertension neuropathy and failed when applied to patients are becoming a common form of treatment with clinical symptoms. It is likely that these for renal and cardiovascular disease in patients had already experienced damage to diabetes patients. These drugs known as nerve and vascular tissue, consistent with the angiotensin converting enzyme (ACE) second phase of diabetic neuropathy. In recent inhibitors and angiotensin II receptor studies, we have determined that epineurial antagonists (ARBs) have been shown to arterioles of the sciatic nerve are innervated by have antioxidant and neuroprotective sensory nerves containing the neuropeptide properties. Previously, my laboratory has calcitonin-gene related peptide (CGRP). We demonstrated that diabetes-induced have demonstrated that the innervation of oxidative stress, which can be prevented epineurial arterioles by sensory nerves containing with antioxidants, is a major contributor to CGRP and CGRP-mediated vascular relaxation diabetes vascular and neural disease. are decreased in diabetes. Neuropeptides such Therefore, we believe that ACE inhibitors as CGRP are thought to regulate vascular tone and/or ARBs have great potential for

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preventing the development/progression of experiments we have discovered a number diabetic neuropathy. To test this hypothesis of roles that this protein can do. One of we are treating streptozotocin-induced these roles is that it promotes the secretion diabetic rats, a model for type diabetes, with of the immunological regulator called GM- an ACE inhibitor and/or ARB. For our CSF. The latter can promote the immune studies we are using both prevention and function of the brain to ward off infection intervention protocols. After the desired when the brain is invaded by treatment we are conducting studies to microorganisms. But when the brain suffers determine whether the applied treatments from an autoimmune disease such as prevent and/or improve diabetes-induced multiple sclerosis (MS) in which the immune disorders in vascular or neural function. It is cells spuriously attack the brain cells, GM- our hope that these studies will provide a CSF can aggravate the situation by rationale for investigating in human diabetes enhancing immunity. Since GMF activates the potential of ACE inhibitors and/or ARBs GM-CSF, we have found a new approach to for the treatment of diabetic neuropathy. treat MS by manipulating the amount of GMF in the brain. We are currently testing MeSH Terms: diabetes mellitus, oxidative this hypothesis. If successful we hope to be stress, peripheral nerve, antioxidants, able to contribute to the therapy of such angiotensin-converting-enzyme devastating diseases as MS. inhibitors, angiotensin 2 MeSH Terms: neurology, biochemistry, multiple sclerosis, growth substances, proteins, immunologic factors, Principal Investigator: autoimmunity Asgar Zaheer, Ph.D.

Project Title: Principal Investigator: Glia Maturation Factor as a Signaling Nicholas ZavaZava, M.D., Ph.D. Molecule

Project Title:

Immune-Privilege and Tolerogenicity of Summary: Stem Cells

Proteins are molecules that carry out a great variety of functions in the cell and in the Summary: body as a whole. The kind of proteins that a cell produces is dependent on the genetic code in the DNA and on the expression of Research on stem cells have become very these genes. Each organ in the body has its promising based on published data and the signature proteins that makes that organ unique properties of embryonic stem cells unique. For example, there are specific brain not found in any other cell types found in the proteins that are exclusive to the brain and human body. There are three different types that carries out a host of activities that of stem cells, namely i) those found in bone promote our memory, thinking, and marrow and in peripheral blood, generally influences our mood and behavior. Glia referred to as hematopoietic stem cells; ii) maturation factor (GMF) is one of such adult or mesenchymal stem cells that can be proteins in the brain. We isolated and detected in solid organs such as the heart or sequenced this protein some years ago and liver and; iii) embryonic stem cells that are have produced this protein by derived from embryos or by nuclear transfer. biotechnology. We are able to engineer the Of these three different types of stem cells, brain cells to make it produce an excess only embryonic stem cells are capable of amount of this protein or, alternatively, to differentiating into any desired tissue or make it devoid of this protein. In this manner organ under optimal culture conditions. we can figure out how the protein is actually Therefore these cells hold the promise for working in the brain. In doing these future therapies directed at treating

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degenerative diseases such as Parkinson’s stem cells such as their expression of death Disease, Alzheimer’s Disease, diabetes and receptors will be studied. Ultimately, the others. Our project focuses on establishing a potential of embryonic stem cells will better reliable protocol for the protection of be understood on completion of these transplanted organs from rejection using studies. embryonic stem cells. Results from these studies could lead to improved organ MeSH Terms: embryonic stem cells, transplantation without the need for tolerance, organ transplantation, expensive immunosuppression, which rejection, immunosuppression, generally leads to significant reduction of the transplantation, regenerative medicine quality of life for patients. Further, the results could be beneficial to the treatment of degenerative diseases.

MeSH Terms: stem cells, transplantation, tolerance, rejection, immune privilege

Principal Investigator: Nicholas Zavazava, M.D., Ph.D.

Project Title: Potency of Embryonic Stem cells to Induce Graft Tolerance

Summary:

Organ transplantation remains the only form of treatment for many patients suffering from organ failure, particularly for those suffering from end-stage renal function, liver failure, cardiac failure and diabetes. Accompanied by the chronic shortage of organs available for transplantation, rejection is a difficult problem for those who ultimately get transplanted. Our laboratory recently observed that organ rejection can be prevented by embryonic stem cells. This effect is specific to the organ donor indicating that these cells may be effective in educating the immune system of the recipient not to reject. Our laboratory seeks to achieve a state of mixed chimerism, i.e. a situation where an animal has foreign blood cells circulating in its own blood system without their rejection. Achieving this state of immunological results in protection from rejection and obviates the need for the use of immunosuppression. Here we seek to characterize embryonic stem cells immunologically thereby laying the basis for their possible use in clinical transplantation. The molecular characteristics of embryonic

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INDEX Cell Cycle, 10 Surface receptor, 9 Cerebral blood flow, 37 A Cerebrovascular accident, 13, 22 Cervical Absorption, 33 Cancer, 15 Acute respiratory distress syndrome, 24, 25 Cytology, 31 Adjuvants, 23 Smears, 31 Adrenergic Spine, 16 Alpha-agonists, 26 CGRP, 49 Beta-agonists, 26 Cholesterol, 11 Adverse drug events, 19 Cirrhosis, 5, 32 Aging, 6 Coinfection, 35 AIDS, 35 Compressive optic neuropathy, 20 Aneurysm, 40 Conditioning, 42 Angiotensin 2, 50 Co-receptor, 35 Angiotensin-converting enzyme inhibitors, Coronary, 29 50 Disease, 30 Antibiotic resistance, 46 Vessels, 28 Antibiotics, 41 CpG, 23 Antibody response, 4 Cyclin dependent kinases, 10 Anticoagulants, 13 Cystic fibrosis, 26 Antioxidants, 7, 8, 48, 49, 50 Cytidylyltransferase, 24, 25 Arachidonate 12-lipoxygenase, 28 Cytokines, 5, 8, 11 Arachidonic acid, 28 Atrial Fibrillation, 13 D Occlusive diseases, 10 Atherosclerosis, 6, 7, 22, 40 Deacylases, 41, 42 Artificial disc, 15 Decision support system, 30 ASIC channel, 42 Dehydroepiandrosterone, 9 Autoimmunity, 4, 50 Diabetes, 34 Diabetes Mellitus, 49, 51 Type 2, 29, 48 B Diabetic foot, 12 Diagnostic immunology, 2, 4 B cells, 4 Diskectomy, 16 Bacteria, 17, 45 DNA Vaccines, 43 Bacterial pneumonia, 8 Drug Bactericidal, 41 Monitoring, 13 Biochemistry, 50 Therapy, 12 Blood pressure, 7 Brazil, 45 E

C Elastin, 27 Electric stimulation therapy, 12

Emphysema, 27 Ca-Co-2 cells, 11 Endothelium, 22 Calpain, 24 Endotoxin, 14, 18, 41, 42 Cancer, 17 Binding proteins, 42 Capsule, 42 Endotoxines, 17 Cardiovascular disease(s), 7, 22, 30 Epithelial amiloride sensitive Na+ channel, CD4 cell, 11 38 CD14 cell, 14 Evidence based medicine, 30 Ceftriaxone, 46

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Human Genetics, 43, 45 F Papillomavirus, 15 Hypertension, 7, 9, 20, 35 Fatty acids, 11, 34 Fear, 42 Fibromyalgia, 47 I Fibrosis, 5, 17, 18 Flow cytometry, 2, 3, 4 Immune privilege, 51 Foodborne infections, 46 Immunity, 2 Free radicals, 5, 40 Immunodeficiency, 4 Immunologic factors, 50 Immunology, 2, 3, 4, 11, 45 G Immunosuppression, 51 Immunotherapy, 23 Gastric emptying, 33 Infection, 18, 21, 28 Gastrointestinal transit, 33 Inflammation, 1, 5, 8, 14, 21, 40, 41, 42 GB virus, 35 Injury, 17, 18 Gene, 25 Innate immunity, 21, 28 Expression, 38, 48 Insulin, 34 Transfer, 26 Sensitivity, 34 Genetics, 45, 47 Intestines, 11 Glaucoma, 39 Intradiscal pressure, 15 Globus pallidus, 12 Ion channels, 35 Glucose, 34 Ischemia, 26 Glutamate, 37 Glycoprotein GP63, 44 G-protein, 9, 10 K Gram-negative Bacteria, 40 Kidney, 9, 20 Bacterial infections, 14 Collecting ducts, 38 Gram-positive bacteria, 41 Growth substances, 50 Gynecologic health, 31 L

Laminectomy, 16 H Leishmania, 43, 44, 45, 48 Braziliensis, 45 Head and neck cancer, 15 Infantum, 43, 44 Health services Lipopolysaccharide(s), 14, 42 Research, 30 Binding protein, 14 Utilization, 31 Lipoproteins, 42 Hearing Liver disease, 32 Aids, 36, 37 Lumbar spine, 15 Loss, 36, 37 Lung infection, 17, 18 Heart failure, 7, 9 Lymphomal, 4 Congestive, 39 Hemorrhage, 13 Hepatitis M C, 32 Chronic, 32 Macrophage(s), 5, 17, 18, 21. 44 HIV, 35, 50 Major surface protease, 48 Home care services, 39 Mechanoreceptor, 37 Homocysteine, 22 Medulla, 37 Hospital mortality, 32 Membranes, 41 Host defense, 2 Metabolic syndrome X, 29

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Microcirculation, 28 Satisfaction, 37 Mitochondria, 34 Perimetry, 39 Molecular biology, 4 Peripheral nerve, 48, 49, 50 Motion analysis, 15, 16 Peristalsis, 33 Multiple sclerosis, 50 Persian Gulf War, 47 Muscle, smooth, 10 Phagocytosis, 1, 21 Myocardial Phospholipases, 41 Infarction, 7 Phospholipids, 41 Ischemia, 30 Polypharmacy, 19 Myeloperoxidase, 28 Pons, 37 Post traumatic stress disorder, 42 Potassium channels, 6 N Prasterone, 9 Pressoreceptors, 6, 7 NADPH Prostaglandins, 20 oxidase, 5 Prostate cancer, 23 quinone, 8 Proteins, 50 Natural ligand, 35 Protheses Neisseria meningitidis, 42 Heart valve, 13 Nerves, 9 Pseudomonas Aeruginosa, 8 Neurology, 50 Purkinje fibers, 26 Neurons afferent, 20 Neurosurgery, 15, 16 Neurosurgical procedures, 12 Q Neurotransmitters, 20, 37 Neutrophil(s), 1, 28, 41 Quality Nitric oxide, 9, 37 Of health care, 30 NK cells, 2, 3, 4 Of indicators, 30 Nodose ganglia, 6

R O Rape, 31 Obesity, 34, 48 Reactive oxygen species, 1 Optical coherence tomography, 20 dicumarol, 8 Organ transplantation, 51 Receptor Outcomes Bradykinin, 10 Assessment, 32 G-protein coupled, 10 Research, 32 IL-2, 11 Outer membrane, 42 Regenerative medicine, 51 Oxidase, 28 Rehabilitation of the hearing impaired, 36, Oxidative stress, 5, 29, 40, 48, 49, 50 37 Oxygen Rejection, 51 Derived free radicals, 6 Renin angiotensin system, 20 Free radicals, 7 Respiratory Burst, 44 Diseases, 5 P Retinoids, 27 Rheumatology, 4 Pancreatic cancer, 8 Risk adjustment, 32 Parasitic disease, 48 Parasitology, 43, 44, 45 Parasympathetic, 37 S Parkinson’s disease, 12 Patient Sepsis, 14 Centered care, 30 Sexual Violence, 31

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Signal transduction, 10 Ventricular, 26 Skin, 17 Veterans, 19, 32, 47 Neoplasms, 17 Viral Sodium, 9 Genes, 15 Channels, 6, 35 Proteins, 15 Solitary tract, 37 Virology, 4 Staphylococcus aureus, 21, 41 Virus replication, 15 Stem cells, 51 Vision testing, 39 Embryonic, 51 Visual loss, 20, 39 Stress, 47 Substance P, 20 Subthalamic nucleus, 12 W Sudden cardiac death, 7 Superoxide anion, 5 Warfarin, 13 Surfactant, 24, 25, 26 Women veterans, 31 Sympathetic nerve activity, 6, 7, 34 Wound infection, 12 Wounds and injuries, 17 T Z T cells, 2, 4 Tachycardia, 26 Zucker rats, 29 Telemedicine, 39 Telephone, 36, 39 Thromboembolism, 13 TLR4, 14 Tolerance, 11, 51 Toll-like receptors,14 Training support, 45 Transmembrane signaling, 4 Transplantation, 51 Treatment outcome, 36 Questionnaires, 37 Tropoelastin, 27 Trypanosoma brucei, 44 Tuberculosis, 21 Tumor Immunology, 2 Necrosis Factor, 24, 25 Suppressor proteins, 10

U

Ubiquitin protein ligase, 38 Ulcer, 1 Ultraviolet, 17 Urocanic acid, 17

V

Vaccines, 23, 43, 45 Vascular, 10 Endothelium, 9

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