Cystein, Intestinal Thiols and Goblet Cell Development

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DESCRIPTION (provided by applicant): The proposal originates from our observations of a three to four fold increase in the number of acidomucin-positive goblet cells accompanied by glutathione (GSH) depletion in the small intestinal mucosa of neonatal pigs being nourished parenterally. We hypothesize that the lack of luminal cysteine is the primary insult perceived by the parenterally nourished epithelium, and that the functional endpoint of this nutritional deficiency is a more oxidized intracellular redox potential The identification of this amino acid as a key nutritional factor is based on (a) the high rate of first-pass consumption of dietary cysteine and the low rate of arterial cysteine utilization by the intestine; (b) evidence that cellular redox status is a key determinant of cellular differentiation, and (c) the fact that cysteine is structurally and functionally involved in both intracellular redox status and mucosal defense via its inherent contribution to the thiol buffers GSH and thioredoxin (Trx) on one hand, and the goblet cell-specific secretory mucins and intestinal trefoil factor (ITF) on the other. We hypothesize that parenteral nutrition results in a preferential activation of the mucous cell lineage as well as selective sparing of goblet cells leading to the repopulation (at a reduced rate) of the villus with goblet cells relative to absorptive enterocytes. The alteration in goblet cell development is initiated by hyperoxia that stems from compromised cysteine status. In other words, we hypothesize that goblet cells are the guardians of mucosal redox homeostasis. Our hypothesis will be tested in three specific aims: 1) Determine if a causal relationship exists between epithelial redox status and goblet cell expansion, independent of TPN-associated inflammation. 2) Quantify, in a goblet cell culture model system, the hierarchy of cysteine usage for the synthesis of GSH and Trx versus ITF and secretory mucins, as well as the role of the transsulfuration pathway in the formation of cysteine and sulfate from methionine, as affected by apical versus basal provision of cysteine, and in response to graded levels of hyperoxia. Simultaneously, determine if the source (inorganic vs. cysteine-derived) and flux of sulfate used for sulfomucin biosynthesis is affected by cysteine availability and hyperoxic stress. 3) Test the hypothesis that Cysl31 and/or Cys165, found within the activation domain of Cdx2, are redox sensitive and upon post-translational modification alter the specificity of Cdx2 transcriptional regulation.