Induction of apoptosis in BPH stromal cells by adenoviral-mediated overexpression of caspase-7

Purpose: We hypothesized that expression/activity of critical components of the apoptotic pathway can be used to induce apoptosis of a human prostate cell line derived from benign prostatic hyperplasia (BPH) tissue. Materials and Methods: We analyzed the apoptotic pathway in BPH cells treated with the powerful inducer of apoptosis, staurosporine (STS), and adenoviruses overexpressing caspase-3, -7, or the control gene lacZ. Results: Twelve hours post-STS, most BPH cells were floating in the culture medium, TUNEL staining was widespread, and DEVDase activity (the catalytic activity of type II caspases) was increased. The pan-caspase inhibitor, Z-VAD-FMK, prevented STS- induced apoptosis. Based on these observations, we performed immunoblot analysis for the three known group II caspases (that is caspase-2, -3 and - 7), but none of them was detected with three commercially available antibodies. Nevertheless, in view of the presence of increased DEVDase activity, we reasoned that a group II caspase must be a critical mediator of apoptosis in this model. If correct, we postulated that overexpression and activation of a type II caspase should cause apoptosis. To test this hypothesis, we coupled the cDNAs encoding caspase-3 and caspase-7 to adenoviral vectors and obtained constructs AvC3 and AvC7. Cells infected with AvC3 or AvC7 overexpressed the protein for caspase-3 or -7 within 24 to 48 hours. Caspase-3 overexpression did not cause apoptosis above that observed in cells receiving the control adenovirus expressing the lacZ cDNA (AvLac-Z). In contrast, caspase-7 overexpression induced massive apoptosis. BPH cells were then infected with increasing multiplicity of infection (MOI) of AvC7 and AvlacZ. A positive correlation was found between the amount of caspase-7 expressed and the level of DEVDase activity measured. AvC7 at MOIs of 25:1 and 50:1 induced apoptosis in about 50% of BPH cells at 72 hours post- infection. This effect was AvC7 specific, because the same MOIs of AvlacZ were not apoptogenic. Conclusions: Adenoviral-mediated overexpression of caspase-7 induces apoptosis of BPH-derived cells.