Figure 4.

Integrin inhibition-induced cell death in mouse NS cells does not require caspase activity. (a) GL-261 NS cells (left) or SMA-560 NS cells (right) were exposed to RAD, cilengitide (1 or 10 μM), MFL (50 ng/ml), staurosporine (50 nM), zVAD-fmk (10 μM) or to combinations thereof for 6 h. DEVD-amc cleaving activity was determined fluorometrically. (b) Whole-cell protein lysates of GL-261 NS or SMA-560 NS exposed to RAD, cilengitide (1 or 10 μM) or staurosporine (50 nM) for 24 h were assessed for full-length and cleaved caspase 3, using actin as a loading control. (c) GL-261 NS cells or SMA-560 NS cells were exposed to RAD, cilengitide (10 μM), zVAD-fmk (1 μM), staurosporine (50 nM), or to a combination of zVAD-fmk (1 μM) and cilengitide (10 μM) or staurosporine (50 nM). Viability was assessed after 24 h of treatment by annexin V/PI staining. (d) Control or crm-A transfectants were exposed to RAD, cilengitide (10 μM) or MFL (50 ng/ml) for 6 h and analyzed for DEVD-amc-cleaving caspase activity (top). Cell death was assessed by annexin V/PI staining after exposure to RAD or cilengitide (10 μM) for 24 h (bottom)