GUT NUTRIENT METABOLISM: MOLECULAR CELLULAR APPROACHES

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One of the fundamental challenges in intestinal physiology is to elucidate the mechanisms by which micronutrients are absorbed, metabolized and routed by the polarized small intestinal absorptive cell (enterocyte). The long-term goal of this laboratory is to use molecular and cell biological techniques to study micronutrient metabolism and trafficking in the normal and diseased mammalian small intestine. The mechanisms by which the enterocyte acquires, compartmentalizes and metabolizes fatty acids and retinoids are poorly characterized. It is thought that these processes involve interactions with cellular binding proteins. Two remarkably abundant fatty acid binding proteins (intestinal and liver fatty acid binding proteins, I- and L-FABP) are present in human and rodent enterocytes. Two retinol binding proteins (cellular retinol binding protein, CRBP and cellular retinol binding protein II, CRBP II) are also present in fetal (and possibly adult) rat enterocytes. These proteins are members of a family of small, cytosolic proteins that bind hydrophobic ligands. The physiological functions of these proteins are unknown. A potentially useful system for determining the functions of these binding proteins is provided by the human intestinal cell line, Caco-2. This cell line differentiates into a polarized monolayer which has well developed lipid, metabolic capabilities. This proposal has two specific aims: (1) determination of the physiological role(s) of the fatty acid binding proteins in intestinal metabolism of exogenous fatty acids. Specifically, this will be accomplished by using recombinant DNA techniques to systematically alter the content of fatty acid binding proteins in Caco-2 cells. Changes in the metabolic processing of long chain fatty acids and other lipids will be correlated with the fatty acid binding protein phenotype of the cells. (2) determination of the physiological role(s) of CRBP II and CRBP in intestinal vitamin A metabolism. This will be approached by comparing vitamin A uptake and metabolism in Caco-2 cells before and after the introduction of plasmids that direct the synthesis of CRBP II and/or CRBP mRNA.