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


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.
Marcelli
M
M
Departments of Medicine, Baylor College of Medicine and VA Medical Center, Houston, Texas, USA.
Shao
T C
TC
Li
X
X
Yin
H
H
Marani
M
M
Denner
L
L
Teng
B
B
Cunningham
G R
GR
D000256:Adenoviridae / Q000235:genetics*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D053143:Caspase 2; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases / Q000032:analysis*; D002478:Cells, Cultured; D018076:DNA, Complementary; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D020287:In Situ Nick-End Labeling; D007763:Lac Operon; D008297:Male; D011470:Prostatic Hyperplasia / Q000473:pathology*; D019311:Staurosporine / Q000494:pharmacology
D018076:DNA, Complementary; D004791:Enzyme Inhibitors; C505407:CASP3 protein, human; C505520:CASP7 protein, human; D053143:Caspase 2; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases; D019311:Staurosporine