Involvement of Bcl-2 and Bax in photodynamic therapy-mediated apoptosis - Antisense Bcl-2 oligonucleotide sensitizes RIF 1 cells to photodynamic therapy apoptosis

Photodynamic therapy (PDT), a promising treatment modality, is an oxidative stress that induces apoptosis in many cancer cells in vitro and tumors in vivo. Understanding the mechanism(s) involved in POT-mediated apoptosis may improve its therapeutic efficacy. Although studies suggest the involvement of multiple pathways, the triggering event(s) responsible for PDT-mediated apoptotic response is(are) not clear. To investigate the role of Bcl-2 in POT-mediated apoptosis, we employed Bcl-2-antisense and -overexpression app roaches in two cell types differing in their responses toward PDT apoptosis , In the first approach, we treated radiation-induced fibrosarcoma (RIF 1) cells, which are resistant to silicon phthalocyanine (Pc 4)-PDT apoptosis, with Bcl-2-antisense oligonucleotide. This treatment resulted in sensitizat ion of RIF 1 cells to PDT-mediated apoptosis as demonstrated by i) cleavage of poly(ADP-ribose) polymerase, ii) DNA ladder formation, iii) terminal de oxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) positive cells, and iv) DEVDase activity. This treatment also resulted in oligonucl eotide concentration-dependent decrease in cell viability and down-regulati on of Bcl-2 protein with a concomitant increase in apoptosis, However, the level of Bax, a pro-apoptotic member of Bcl-2 family, remained unaltered. I n the second approach, an overexpression of Bcl-2 in PDT apoptosis-sensitiv e human epidermoid carcinoma (A431) cells resulted in enhanced apoptosis an d up-regulation of Bax following PDT. In both the approaches, the increased Bax/Bcl-2 ratio was associated with an increased apoptotic response of PDT , Our data also demonstrated that PDT results in modulation of other Bcl-2 family members in a way that the overall ratio of pro apoptotic and anti-ap optotic member proteins favors apoptosis.