Fig 1: Overexpression of caspases is not sufficient to accelerate islet cell apoptosis.832/13 and primary rat islets were untreated (no virus) or treated with recombinant adenoviruses containing cDNAs corresponding to human Apaf-1, human caspase 3, human caspase 9 or the control gene, ß-galactosidase (ß-gal), as indicated. 72 h post-infection, 832/13 cells (A) and primary rat islets (B) were treated with DMSO (control) or thapsigargin (100 nM or 1 µM, respectively) for 18 h. Clarified lysates were examined by immunoblot analysis. Endogenous rat (r) and overexpressed human (h) proteins are labeled accordingly. Of note, Apaf-1 and caspase 9 antibodies only detect human (overexpressed) proteins. (C, D) 72 h post adenoviral infection, rat islets were treated with DMSO (control) or thapsigargin (1 µM) for up to 72 h. Islet cells were stained for TUNEL, caspase 3, and insulin and counterstained with DAPI. Cells were imaged using a high content imager and analyzed using Cellomics software. (C) Percentages of insulin+ TUNEL+ islet cells are shown. (D) Comparison of the percentages of caspase 3+ (overexpressing), TUNEL+ islet cells vs. caspase 3- (non-overexpressing), TUNEL+ islet cells are shown. (C, D) Data represent the mean +S.E.M. of 3 independent experiments. * p = 0.05 as compared to DMSO treated cells.