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Clinical and Molecular Analyses of Intestinal Goblet Cell Acidomucins and Cysteine Metabolism

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Title: Clinical and Molecular Analyses of Intestinal Goblet Cell Acidomucins and Cysteine Metabolism
Author(s): Croix, Jennifer A.
Director of Research: Gaskins, H. Rex
Doctoral Committee Chair(s): Donovan, Sharon M.
Doctoral Committee Member(s): Gaskins, H. Rex; Tappenden, Kelly A.; Tapping, Richard I.
Department / Program: Nutritional Sciences
Discipline: Nutritional Sciences
Degree Granting Institution: University of Illinois at Urbana-Champaign
Degree: Ph.D.
Genre: Dissertation
Subject(s): goblet cell mucin taurine cysteine intestine colon
Abstract: Goblet cells are key contributors to intestinal mucosal barrier function through their role in production of mucins and other important secretory products, which include trefoil factor 3 (TFF3) and resistin like molecule β (RETNLB). Goblet cell mucins show great structural heterogeneity but can be broadly classified into neutral and acidic subtypes (chemotypes), which can be further classified into sulfomucins or sialomucins based on the presence of terminal sulfate or sialic acid groups on the oligosaccharide chains. Sulfomucins and sialomucins vary regionally throughout the gastrointestinal tract and changes in their expression have been observed in diseases such as inflammatory bowel disease and colorectal cancer. It is likely that there is variation in these chemotypes among individuals and that both regional and interindividual variation in sulfo- and sialomucins may influence microbial colonization. However, quantitative data and a description of interindividual variation in sulfo- and sialomucin content in the human colon are lacking. There is an absence of data describing the relationship between microbial species and these mucin chemotypes in the human colon. In Chapter 2, we begin to fill these gaps in the literature via a pilot study examining sulfo- and sialomucins in the healthy human colon and their relationship with sulfate reducing bacteria (SRB). Quantitative differences in mucin abundance among specific regions of the colon and individuals are described. In addition, we observe that the relationship between these mucin chemotypes and SRB is not influenced by location in the colon, but rather by the host. In Chapter 3, we begin to identify factors that may contribute to changes in sulfomucin in disease and possibly interindividual variation, using human adenocarcinoma-derived LS174T cells, which have a goblet cell-like phenotype and produce both sulfo- and sialomucins. We specifically focused on the effects of bacterial flagellin, IL-13, and TNFα on the expression of genes encoding the major secretory mucin, MUC2, and Golgi sulfotransfereases, CHST5 and GAL3ST2. In addition, expression of sulfomucin and Sulfo Lea antigen, which is synthesized in part by GAL3ST2, was examined. Overall, the results indicated that both host and microbial factors influence sulfomucin expression, possibly via modulation of CHST5 and GAL3ST2 expression. Because goblet cells synthesize and secrete cysteine-rich products, including mucins, TFF3, and RETNLB, they may have a high cysteine requirement. Furthermore, they may require additional cysteine, which can be catabolized to sulfate, for mucin sulfation. Cysteine is also used in the production of other proteins as well as essential molecules including glutathione, taurine, and pyruvate. However, high cysteine levels are cytotoxic. Mammals regulate cysteine metabolism to maintain levels within a range that is sufficient for synthesis of essential molecules but below the level of cytotoxicity through the use of cysteine dioxygenase (CDO), which catalyzes the first step in cysteine catabolism. In Chapter 4, CDO expression and localization in mouse small and large intestine is examined using immunohistochemical and immunofluorescence staining techniques. We observed that CDO is expressed in goblet cells as well as Paneth and enteroendocrine cells, which are also members of the secretory lineage, while expression was absent in absorptive enterocytes. We postulate that this striking difference in CDO expression between the absorptive and secretory cell lineages is due to higher cysteine catabolism requirements in goblet, Paneth, and enteroendocrine cells relative to absorptive epithelial cells, either to synthesize additional taurine and sulfate or to metabolize excess cysteine when cells are not actively synthesizing cysteine-rich secretory products. In Chapter 5, we examine the possibility that goblet cells are using CDO to synthesize taurine and determine whether inflammatory stimuli alter taurine biosynthesis, using the LS174T cell line as a model. There are two pathways for taurine synthesis. One pathway involves the oxidation of cysteine via CDO to cysteinesulfinate, which is decarboxylated by cysteinesulfinic acid decarboxylase (CSAD) to hypotaurine. The other pathway involves the conversion of cysteine to Coenzyme A, which releases cysteamine during turnover. Cysteamine is then oxidized to hypotaurine via cysteamine dioxygenase (ADO). In both pathways, hypotaurine is then oxidized to taurine. We confirm that LS174T cells synthesize taurine via the ADO pathway and possibly the CDO/CSAD pathway. We also demonstrate that certain inflammatory triggers can enhance taurine biosynthesis and/or export, which may provide taurine for other cell types in an in vivo setting. The anti-inflammatory effects of taurine in the intestine have been described, so these results may represent a novel mechanism by which goblet cells attenuate inflammation. Overall, the findings described in this dissertation provide novel information regarding the role of intestinal goblet cells in mucosal barrier function.
Issue Date: 2011-01-21
URI: http://hdl.handle.net/2142/18592
Rights Information: Copyright 2010 Jennifer A. Croix
Date Available in IDEALS: 2011-01-21
2013-01-22
Date Deposited: 2010-12
 

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