The effects of dietary fat on pancreatic lipase gene expression. Item Type text; Dissertation-Reproduction (electronic) Authors Ricketts, Jennifer Regan. Publisher The University of Arizona. Rights Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author. Download date 30/09/2021 04:34:56 Link to Item http://hdl.handle.net/10150/187287 INFORMATION TO USERS This manuscript .has been reproduced from the microfilm master. UMI films the text directly from the original or copy submitted. Thus, some thesis and dissertation copies are in typewriter face, while others may be from any type of computer printer. The quality of this reproduction is ~ependent upon the quality of the copy submitted. 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Contact UMI directly to order. UMI A Bell & Howell Information Company 300 North Zeeb Road. Ann Arbor. MI48106-1346 USA 313/761-4700 800:521-0600 THE EFFECTS OF DIETARY FAT ON PANCREATIC LIPASE GENE EXPRESSION by Jennifer Ricketts A Dissertation Submitted to the Faculty of the COMMITTEE ON NUTRITIONAL SCIENCES (GRADUATE) In Partial Fulfillment of the Requirements For the Degree of DOCTOR OF PHILOSOPHY In the Graduate College THE UNIVERSITY OF ARIZONA 1 995 UMI Number: 9604513 OMI Microform 9604513 Copyright 1995, by OMI Company. All rights reserved. This microform edition is protected against unauthorized copying under Title 17, United States Code. UMI 300 North Zeeb Road Ann Arbor, MI 48103 2 THE UNIVERSITY OF ARIZONA GRADUATE COLLEGE As members of the Final Examination Committee, we certify that we have read the dissertation prepared by Jennjfer Rjcketts --~~~~~~~---------------------- entitled ____T~h~e~ef~~~ec~t~s~o~f~d~ie~t~ary~t~a~t~o~n~p~a~n~cr~e~a~ti~c~h~·p~a~se~g~en~e~ex~p~r~e~ss~io~n~ ___________ and recommend that it be accepted as fulfilling the dissertation ~nt for the Degree of Doctor of Philosophy M.Q~ Date I 1/9.« /16- Date .zf~,)97S Darrel E. Goll Date Date Final approval and acceptance of this dissertation is contingent upon the candidate's submission of the final copy of the dissertation to the Graduate College. I hereby certify that I have read this dissertation prepared under my direction and recommend that it be accepted as fulfilling the dissertation requirement. DisserQ~oP· ~- Date' I 3 STATEMENT BY AUTHOR This dissertation has been submitted in partial fulfillment of requirements for an advanced degree at The University of Arizona and is deposited in the University Library to be made available to borrowers under rules of the Library. Brief quotations from this dissertation are allowable without special permIssIon, provided that accurate acknowledgment of source is made. Requests for permission for extended quotation from or reproduction of this manuscript in whole or in part may be granted by the head of the major department or the Dean of the Graduate College when in his or her judgment the proposed use of the material is in the interests of scholarship. In all other instances, however, permission must be obtained from the author. SIGNED:~;;~ 4 ACKNOWLEDGEMENTS I would like to thank Dr. Patsy Brannon for her support, effort, and time during the development of this dissertation project. I also would like to thank Dr. David K. Y. Lei, Dr. Charles W. Weber, Dr. Darrel E. Goll and Dr. Michael Wells for their participation in this project. I am grateful to Deb Scott, Suzie Kunz and An Tsai for their help and friendship both inside and outside of the laboratory. Special thanks is extended to Phyllis Reid for her help in final preparation of this dissertation. 5 DEDICATION This dissertation is dedicated to the memory of Dr. B.L. Reid. He was generous in his support and encouragement and he will be missed by all. 6 TABLE OF CONTENTS Page LIST OF TABLES . .. 9 LIST OF FIGURES ........................................ 10 ABSTRACT . .. 11 1. INTRODUCTION ......................................... 12 2. LITERATURE REVIEW. .. 15 LIPID METABOLISM ................................... 15 Dietary Fats . .. 16 Physical properties . .. 16 Health and disease interactions ........................ 17 Digestion . .. 20 An overview ................................... 20 Preduodenal Iipases ............................... 23 Duodenal Iipases . .. 23 Bile salts . .. 25 Absorption . .. 26 Enterocyte . .. 27 Transport ..................................... 29 Tissue Utilization ................................... 30 Triglycerides (TG) . .. 30 Ketones. .. 31 Lipoproteins . .. 31 PANCREAS . .. 33 Endocrine Pancreas .................................. 33 Hormones ..................................... 33 Insulin .................................... 34 Glucagon .................................. 35 Somatostatin ................................ 35 Exocrine Pancreas ................................... 35 Ultrastructure . .. 36 Digestive enzymes . .. 37 Proteases . .. 37 Glycosidase . .. 38 Lipases . .. 38 Nucleases .................................. 39 Synthesis and secretion . .. 39 Dietary and hormonal regulation . .. 42 7 TABLE OF CONTENTS--continued page Dietary regulation . .. 44 Hormonal regulation ........................... 47 PANCREATIC LIPASE .................................. 50 Gene Structure ..................................... 52 Function ......................................... 56 Dietary Regulation . .. 59 Hormonal Regulation ................................. 61 Regulation of Colipase ................................ 63 3. MULTIPLE MECHANISMS OF REGULATION OF PANCREATIC LIPASE IN RATS BY AMOUNT AND TYPE OF DIETARY FAT. .. 65 INTRODUCTION ............................ .. 65 METHODS... .. ... ... .. .. ... ... ... .. ... .. 66 Enzyme Analysis. .. 66 Isolation of Total RNA . .. 68 Quantitation of mRNA ................................ 69 Statistical Analysis . .. 70 RESULTS. .. 70 Enzyme Activities and Plasma Ketones . .. 70 mRNA Levels. .. 74 DISCUSSION . .. 74 4. DIETARY FAT SATURATION HAS DIFFERENTIAL EFFECTS ON PANCREATIC LIPASE ACTIVITY, mRNA LEVELS, AND SYNTHESIS. .. 77 INTRODUCTION ...................................... 77 METHODS. .. ..... .. ... .. ..... .... ..... .. .. ... .. .. 78 Experimental Protocol . .. 78 Enzyme Analysis. .. 79 Isolation of Total RNA . .. 79 Quantitation of mRNA ................................ 80 Pancreatic Acini Isolation .............................. 80 Protein Synthesis . .. 81 2-D Gels. .. 81 Isolation of tRNA ................................ 85 Total tRNA-phenylalanine . .. 86 Radiolabeled tRNA-phenylalanine ...................... 86 Statistical Analysis . .. 86 RESULTS. .. 87 Experiment 1 ...................................... 87 Food consumption . .. 87 8 TABLE OF CONTENTS--continued page Enzyme activities and plasma ketones . .. 87 Pancreatic mRNA Levels. .. 91 Experiment 2 ...................................... 91 Body weights ................................... 91 Relative synthesis ................................ 91 Specific activity of tRNA-phenylalanine .................. 95 DISCUSSION . .. 95 5. THE EFFECTS OF DIETARY FAT ON THE REGULATION OF RAT PANCREATIC LIPASE FOLLOWING INHIBITION OF CHYLOMICRON TRANSPORT FROM THE ENTEROCYTE . .. 99 INTRODUCTION ............ .. 99 METHODS ......................................... 101 Experimental Protocol . .. 101 Enzyme and mRNA analysis ........................... 105 Statistical Analysis . .. 105 RESULTS .......................................... 105 Food Consumption and Body Weight ...................... 105 Enzyme Activities and mRNA Levels . .. 107 Protease activity ................................... 107 Amylase activity and mRNA levels ....................... 107 Lipase activity and mRNA levels . .. 107 DISCUSSION . .. 112 6. SUMMARY AND CONCLUSIONS . .. 116 APPENDIX A . .. 118 REFERENCES .................................... -;. .. 120 9 LIST OF TABLES Table Page 3-1. Dietary Composition 67 3-2. The Effects of Dietary Fat on Enzyme Activities and Ketone Levels ........ 73 4-1. The Effects of Dietary Fat on Body and Pancreatic Weights and Food Intake. .. 88 4-2. The Effects of Dietary Fat on Enzyme Activities and Plasma Ketone Levels ... 89 4-3. The Effects of Dietary Fat on 28s RNA and Amylase mRNA Levels. .. 92 4-4. The Effects of Dietary Fat on Total tRNA Phenylalanine, tRNA-Phenylalanine Specific Activity, and Relative Synthesis of Amylase . .. 96 5-1. Dietary Composition . .. 104 5-2. The Effects of Treatment and Diet on Total Food Consumption, Weight