This project focuses on the relationship between luminal lipid digestion and cholesterol absorption, with particular attention paid to the role of pancreatic phospholipase A/2 (PLA2) and pancreatic lipase (PL) in this process. The specific hypothesis is that cholesterol absorption efficiency is correlated to the efficiency of luminal lipid digestion, which in turn is dictated by the amount and type of bile acid and the activity of lipolytic enzymes present in the intestinal lumen. We postulate that: (1) bile acids and lipolytic enzymes interact in the re- modeling of luminal lipids prior to their absorption by the enterocytes; (2) variations in the level of pancreatic lipolytic enzyme expression contribute to individual differences in cholesterol absorption efficiency; and (3) ectopic expression of pancreatic lipolytic enzymes in the biliary epithelium is a feasible treatment for fat malabsorption associated with pancreatic insufficiency and cystic fibrosis.
Four specific aims are proposed to address these hypotheses:
Specific Aim 1 will determine the effect of bile acid composition on luminal lipolysis. Experiments will be performed in vitro to determine the effects of bile acid composition on (i) PLA2 and PL activities, (ii) cholesterol distribution among lipid emulsions, phospholipid vesicles, and mixed micelles, and (iii) cholesterol uptake by intestinal cells in culture and mucosal sheets. The in vitro observations will be correlated with in vivo affects of bile acid supplementation on luminal lipolytic enzyme activities, cholesterol solubilization, and cholesterol absorption efficiency in human subjects.
Specific Aim 2 will produce PLA2 knockout mice to test eh importance of PLA2 in cholesterol absorption.
Specific Aim 3 will produce PL knockout mice to test the impact of PL gene deletion on fat and cholesterol absorption. The PL knockout mice produced in this study will be used in Specific Aim 4 to determine if recombinant adenovirus mediated PL gene transfer to the biliary epithelium can restore fat absorption in the absence of pancreatic lipase expression by the pancreas. These studies will provide nutritionists and clinicians with mechanistic information to design effective dietary and/or therapeutic treatment for diseases due to diet- induced hyperlipidemia or to aberrant fat digestion and transport due to pancreatic diseases.
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