Intestinal barrier function is compromised in inflammatory, infectious, ischemic, and immune-mediated intestinal disease. In the previous cycle we studied mechanisms and impact of tumor necrosis factor- (TNF-) induced barrier loss using in vitro and in vivo models to: i) define TNF-induced, myosin light chain kinase- (MLCK-) dependent tight junction regulation;ii) show that MLCK is required for TNF-induced diarrhea;iii) identify and characterize the human MLCK promoter;iv) document MLCK activation in human inflammatory bowel disease (IBD);and v) show that chronic epithelial MLCK activation stimulates mucosal immune cells and sensitizes mice to experimental IBD. We have now begun to define the mechanisms by which TNF specifically activates MLCK1, a long MLCK splice variant, to trigger endocytosis of the tight junction protein occludin. This creates tight junction 'leaks'that allow paracellular flux of large macromolecules without affecting ion selectivity. Our preliminary data show that expression of constitutively-active MLCK in vivo increases flux across the 'leak'pathway. Moreover, unlike in vitro MLCK activation, chronic in vivo MLCK activation alters ion selectivity of the paracellular barrier. This is due to increased mucosal IL-13 production, which induces epithelial claudin-2 expression to create small, cation-selective 'pores'that allow Na+, but not large macromolecules, to traverse the paracellular pathway. This in vivo result emphasizes the presence of, as well as interactions between, two distinct pathways across the tight junction;a high capacity 'pore'pathway that allows small uncharged solutes and specific ions to pass and a low capacity 'leak'pathway that is permeable to larger, uncharged macromolecules but is not ion selective. The fact that 'pore'and 'leak'pathways interact through the mucosal immune system, i.e. IL-13 production, suggests that these pathways may make distinct contributions to disease pathogenesis. These and other data have led to the central hypothesis that at least two mechanisms of regulation modulate unique paracellular pathways, i.e. 'pore'and 'leak,'to differentially impact mucosal homeostasis and disease pathogenesis.
The aims of this application are to i) define the mechanisms by which TNF triggers MLCK1 perijunctional trafficking and enzymatic activation to enhance flux across the tight junction 'leak'pathway;ii) identify the mechanisms that regulate claudin-2 expression and 'pore'pathway flux;and iii) determine the impact of MLCK, occludin, and claudin-2 expression on 'pore'and 'leak'pathway function as well as initiation and progression of chronic intestinal disease. As a whole, these studies will define the relative roles of specific epithelial regulatory processes and immune signaling in modulation of 'pore'and 'leak'pathway paracellular flux and disease pathogenesis. The data, which will provide new understanding of the mechanisms by which intestinal barrier function contributes to human health and disease, will lay the foundation necessary for development of novel therapeutic strategies to correct barrier dysfunction.

Public Health Relevance

The intestinal lining (epithelium) must maintain a barrier that keeps the intestinal contents separate from the remainder of the body. This function is frequently compromised in intestinal disease and has been implicated as an early step in disease development. The proposed studies will advance mechanistic understanding of barrier regulation and dysregulation and, therefore, lead to development of novel therapeutic approaches to improve human health.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK068271-07
Application #
8111221
Study Section
Special Emphasis Panel (ZRG1-DKUS-C (03))
Program Officer
Grey, Michael J
Project Start
2004-07-01
Project End
2014-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
7
Fiscal Year
2011
Total Cost
$474,305
Indirect Cost
Name
University of Chicago
Department
Pathology
Type
Schools of Medicine
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
Edelblum, Karen L; Sharon, Gil; Singh, Gurminder et al. (2017) The Microbiome Activates CD4 T-cell-mediated Immunity to Compensate for Increased Intestinal Permeability. Cell Mol Gastroenterol Hepatol 4:285-297
Odenwald, Matthew A; Turner, Jerrold R (2017) The intestinal epithelial barrier: a therapeutic target? Nat Rev Gastroenterol Hepatol 14:9-21
Tsai, Pei-Yun; Zhang, Bingkun; He, Wei-Qi et al. (2017) IL-22 Upregulates Epithelial Claudin-2 to Drive Diarrhea and Enteric Pathogen Clearance. Cell Host Microbe 21:671-681.e4
Wu, Richard L; Vazquez-Roque, Maria I; Carlson, Paula et al. (2017) Gluten-induced symptoms in diarrhea-predominant irritable bowel syndrome are associated with increased myosin light chain kinase activity and claudin-15 expression. Lab Invest 97:14-23
France, Marion M; Turner, Jerrold R (2017) The mucosal barrier at a glance. J Cell Sci 130:307-314
Christensen, Britt; Hanauer, Stephen B; Erlich, Jonathan et al. (2017) Histologic Normalization Occurs in Ulcerative Colitis and Is Associated With Improved Clinical Outcomes. Clin Gastroenterol Hepatol 15:1557-1564.e1
Odenwald, Matthew A; Choi, Wangsun; Buckley, Aaron et al. (2017) ZO-1 interactions with F-actin and occludin direct epithelial polarization and single lumen specification in 3D culture. J Cell Sci 130:243-259
Choi, Wangsun; Yeruva, Sunil; Turner, Jerrold R (2017) Contributions of intestinal epithelial barriers to health and disease. Exp Cell Res 358:71-77
Weber, Christopher R; Turner, Jerrold R (2017) Dynamic modeling of the tight junction pore pathway. Ann N Y Acad Sci 1397:209-218
Krug, S M; Bojarski, C; Fromm, A et al. (2017) Tricellulin is regulated via interleukin-13-receptor ?2, affects macromolecule uptake, and is decreased in ulcerative colitis. Mucosal Immunol :

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