Gastrointestinal mucosal wound healing requires the migration of sheets of intestinal epithelial cells across a mucosal defect, together with modulation of the differentiated phenotype of these cells. This process is critical to the response of the gut mucosa to surgical or traumatic injury, inflammatory bowel disease, and peptic or infectious mucosal ulceration. Preliminary investigations have validated a cell culture model for the study of intestinal epithelial cell sheet migration and have demonstrated modulation of the differentiation state of these cells during migration. We have hypothesized that enterocytic differentiation and epithelial sheet migration reflect opposing and mutually exclusive cell specializations. Preliminary data and analogy to studies of tyrosine kinase and phosphoprotein modulation during integrin engagement in other cell types suggest that migrating intestinal epithelial cells adapt to migration rather than nutrient absorption via a signal transduction cascade which begins with engagement of reorganized integrin receptors by matrix proteins and modulation of integrin-associated tyrosine phosphoproteins and has as one result the downregulation of digestive brush border expression. We therefore propose to investigate the mechanisms which regulate phenotype (using digestive brush border expression as a marker) in intestinal epithelial cells migrating over laminin, a physiologically relevant substrate, focusing specifically on initial integrin engagement and tyrosine phosphorylation events in order to begin to trace this signal transduction cascade. In particular, we will elucidate the manner in which phenotype is modulated by engagement of specific integrin receptors for laminin via modulation of integrin- associated tyrosine phosphoproteins. The relevance of these findings will be verified by comparison with protein tyrosine phosphorylation states during inhibition of migration by tyrosine kinase inhibitors, stimulation of migration and modulation of integrin expression and organization using EGF, and induction of differentiation by sodium butyrate, each of which we have previously characterized in this system. Specific antibodies and mRNA probes for proteins of interest will then be developed in order to confirm the biological relevance of these findings in vivo in healing mucosa. Such studies should not only illuminate the regulation of a process of fundamental biological significance (modulation of intestinal epithelial cell phenotype during mucosal wound healing) but may also provide a firm scientific basis for future investigation into and intervention in iatrogenic or pathological disturbances of gastrointestinal healing.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29DK047051-04
Application #
2856768
Study Section
Surgery and Bioengineering Study Section (SB)
Program Officer
Hamilton, Frank A
Project Start
1995-07-01
Project End
2000-12-31
Budget Start
1999-01-01
Budget End
1999-12-31
Support Year
4
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Yale University
Department
Surgery
Type
Schools of Medicine
DUNS #
082359691
City
New Haven
State
CT
Country
United States
Zip Code
06520
Basson, M D (2001) In vitro evidence for matrix regulation of intestinal epithelial biology during mucosal healing. Life Sci 69:3005-18
Sanders, M A; Basson, M D (2000) Collagen IV-dependent ERK activation in human Caco-2 intestinal epithelial cells requires focal adhesion kinase. J Biol Chem 275:38040-7
Basson, M D; Emenaker, N J; Sanders, M A (2000) Alpha integrin subunits regulate human (Caco-2) intestinal epithelial proliferation and phenotype. Cell Physiol Biochem 10:27-36
Yu, C F; Basson, M D (2000) Matrix-specific FAK and MAPK reorganization during Caco-2 cell motility. Microsc Res Tech 51:191-203
Yu, C F; Sanders, M A; Basson, M D (2000) Human caco-2 motility redistributes FAK and paxillin and activates p38 MAPK in a matrix-dependent manner. Am J Physiol Gastrointest Liver Physiol 278:G952-66
Liu, Y W; Sanders, M A; Basson, M D (1999) Loss of matrix-dependent cytoskeletal tyrosine kinase signals may regulate intestinal epithelial differentiation during mucosal healing. J Gastrointest Surg 3:82-94
Basson, M D; Emenaker, N J; Rashid, Z (1998) Effects of modulation of tyrosine phosphorylation on brush border enzyme activity in human Caco-2 intestinal epithelial cells. Cell Tissue Res 292:553-62
Basson, M D (1998) Role of integrins in enterocyte migration. Clin Exp Pharmacol Physiol 25:280-5
Liu, Y W; Sanders, M A; Basson, M D (1998) Human Caco-2 intestinal epithelial motility is associated with tyrosine kinase and cytoskeletal focal adhesion kinase signals. J Surg Res 77:112-8
Perdikis, D A; Davies, R; Zhuravkov, A et al. (1998) Differential effects of mucosal pH on human (Caco-2) intestinal epithelial cell motility, proliferation, and differentiation. Dig Dis Sci 43:1537-46

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