The alpha2 and alpha3 integrins are required for morphologic differentiation of an intestinal epithelial cell line.
BACKGROUND
The molecular mechanisms controlling intestinal epithelial cell differentiation are poorly defined because of the difficulty of growing normal intestinal cells. We have taken advantage of the ability of the Caco-2 cell line to acquire a glandular phenotype in 3-dimensional (3-D) culture systems to investigate the role of alpha2 and alpha3 integrins in morphologic differentiation.


METHODS
Caco-2 cells transfected with sense or antisense DNA constructs of alpha2 or alpha3 integrins were grown in 3-D Matrigel or collagen I in the presence or absence of integrin function-blocking antibodies. We used light and confocal microscopy, BrDU incorporation, TUNEL assay, a fluorometric adhesion assay, FACS analysis, and Western blot analysis to study the effect of extracellular matrix (ECM) and integrins on morphology, polarization, proliferation, apoptosis, cell adhesion, and integrin expression.


RESULTS
Compared to collagen I, Caco-2 cells cultured in 3-D Matrigel display cytoskeletal and adherens junction rearrangements and decreased proliferation consistent with cellular differentiation. These changes, which are inhibited by alpha2 and alpha3 blocking monoclonal antibodies and alpha2 and alpha3 antisense DNA transfection, were associated with an increase in alpha3 integrin expression.


CONCLUSIONS
We demonstrated that signaling through both constitutively expressed alpha2 integrin and Matrigel-induced alpha3 integrin expression is required to acquire a differentiated phenotype in Caco-2 cells.
Zhang
Xihong
X
University of Minnesota Cancer Center, Department of Surgery, University of Minnesota, Minneapolis, MN, USA.
Cromwell
John W
JW
Kunjummen
B Daniel
BD
Yee
Douglas
D
Garcia-Aguilar
Julio
J
D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001672:Biocompatible Materials; D018938:Caco-2 Cells; D002454:Cell Differentiation / Q000502:physiology; D003094:Collagen; D024042:Collagen Type I; D016373:DNA, Antisense / Q000494:pharmacology; D004338:Drug Combinations; D006801:Humans; D020287:In Situ Nick-End Labeling; D039421:Integrin alpha2 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D039422:Integrin alpha3 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D019012:Integrin beta1 / Q000276:immunology; D007413:Intestinal Mucosa / Q000166:cytology* / Q000378:metabolism*; D007797:Laminin; D011509:Proteoglycans; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection; D015854:Up-Regulation / Q000502:physiology
D000911:Antibodies, Monoclonal; D001672:Biocompatible Materials; D024042:Collagen Type I; D016373:DNA, Antisense; D004338:Drug Combinations; D039421:Integrin alpha2; D039422:Integrin alpha3; D019012:Integrin beta1; D007797:Laminin; D011509:Proteoglycans; C059485:matrigel; D003094:Collagen