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. 2014 Mar 13;5(3):e1128. doi: 10.1038/cddis.2014.95

Figure 8.

Figure 8

PUMA mediated ER stress-induced apoptosis in vitro. (a) GES-1 and SGC7901 cells were treated with the ER stress inducer tunicamycin, and the western blotting assay showed that GRP78 and cleaved caspase-4 were significantly induced, and PUMA and cleaved caspase-3 were markedly upregulated. β-Actin was used as the loading control. (b) TUNEL (green) staining showed that tunicamycin remarkably induced apoptosis in GES-1 and SGC7901 cell lines. Cell nuclei (blue) were counterstained by DAPI ( × 200). (c) The apoptotic index was calculated by counting a minimum of 20 randomly selected fields following TUNEL staining. The index was obtained by dividing the TUNEL-positive cells by the total number of cells. (d) Western blotting showed that PUMA-siRNA significantly knocked down PUMA expression in GES-1 and SGC7901 cell lines. (e) PUMA-siRNA did not affect GRP78 expression and caspase-4 activation after tunicamycin treatment, however, PUMA knocked down evidently inhibited ER stress-induced caspase-9 and caspase-3 activation by tunicamycin. (f) The ratio of densitometry units of cleaved caspase-3/β-actin was represented in two cell lines. The values are expressed as the means±S.D., and the values were achieved by three separate experiments. *P<0.01 versus 0 h, #P<0.01 versus 24 h without PUMA-siRNA treatment