Type 1 diabetes results due to autoimmune destruction of pancreatic beta cells. The pro-apoptotic BH3-only protein Bid is a member of the Bcl-2 family that is activated in both extrinsic (death receptor-induced) and intrinsic (Bcl-2-regulated or mitochondrial) cell death pathways in mouse beta cells. We have previously demonstrated that Bid-deficient mouse islets are protected from Fas ligand-mediated as well as granzyme B-mediated apoptosis in vitro. Silencing bid therefore has therapeutic potential to inhibit both the pathways of cell death in beta cells. However, it is not yet known if Bid plays similar central role in human beta cell death. We therefore aimed to test the role of bid in human islet cell apoptosis using siRNA knockdown. 

Freshly isolated human islet cells were dispersed into single cells, cultured overnight and then transfected with bid siRNA. We also transfected siRNA for FADD (Fas-associated via death domain), a molecule in the extrinsic cell death pathway upstream of bid. Using FITC-labelled non-specific siRNA, we found that the transfection efficiency varied from 40-60%. We observed 60-80% reduction in bid transcript as compared to scramble siRNA transfected controls. Exposure of bid siRNA transfected cells to apoptotic stimuli (cytokines + Fas-ligand) in vitro did not result in significant protection from cell death as determined by DNA fragmentation analysis, whereas FADD knockdown was protective. When cultured human islet cells were transfected with siRNAs, we observed that the transfection efficiency was significantly higher (80%). However, bid knock down was still not able to confer protection against Fas-ligand mediated apoptosis. Our results indicate that Fas-ligand signalling in human islets occurs through direct caspase activation in a Bid independent manner, despite the relatively high level of Bid expression in normal human islets. This leaves open the question of the role of Bid in human islet apoptosis.