Collagen gel overlay induces two phases of apoptosis in MDCK cells.

摘要

We previously demonstrated that collagen gel overlay induced cell remodeling to form lumen and apoptosis in Madin-Darby canine kidney cells. In the present study, we established that collagen gel overlay-induced apoptosis was initiated at areas exclusive of cell remodeling within 24 h (first phase) and extended into areas of cell remodeling within 48 h (second phase). Collagen gel overlay-induced apoptosis was accompanied by selective proteolysis of focal adhesion kinase (FAK), talin, p130(cas), and c-src. Upon collagen gel overlay, FAK was initially degraded into a 90-kDa product during the first phase and subsequently into a 80-kDa product during the second phase. Collagen gel overlay-induced apoptosis of focal adhesion complex proteins and apoptosis of the first phase could be blocked only by a protease inhibitor cocktail. In addition, we found that both DEVD-fmk and ZVAD-fmk inhibited secondary proteolysis of FAK, but only ZVAD-fmk blocked collagen gel overlay-induced apoptosis of the second phase.Finally, collagen gel overlay-induced apoptosis and proteolysis of focal adhesion complex proteins were completely inhibited by the combination of protease inhibitor cocktail and ZVAD-fmk. Taken together, collagen gel overlay induces two phases of apoptosis; the first phase is dependent on proteolysis of focal adhesion complex proteins, and the second phase on activation of caspases.