Abstract:

Objective To investigate the mechanism of Fas-mediated apoptosis of MML-1 cells inhibited in G1 phase by PMA. Methods MML-1 cells were pretreated by 10 nmol/L PMA for 24 h, and the apoptosis rates of MML-1 cells induced by 50 ng/mL anti-Fas antibody were detected by flow cytometry before and after cell cycle arrest by PMA. The expression of active caspase-3/8 in anti-Fas antibody-induced MML-1 cells was detected by PI-Triton X and active caspase-3/8 double staining and Western blotting before and after PMA treatment. The expression of apoptosis related protein of caspase-3/8, Bcl-2, FLIP and Akt in cells of G1 phase was detected by Western blotting. Results Twenty-four hours after treatment of MML-1 cells by PMA, the percentage of cells in G1 phase significantly increased (93.77%). Before PMA treatment, the apoptosis rate of MML-1 cells was 56% after Fas-mediated apoptosis for 6 h. Whereas after PMA treatment, the apoptosis rate of MML-1 cells was 14% after Fas-mediated apoptosis for 6 h. It was revealed by flow cytometry and Western blotting that the expression of active caspase-3/8 was significantly inhibited after PMA treatment. Western blotting demonstrated that the expression of Bcl-2 was up-regulated in cells of G1 phase. Conclusion Fas-mediated apoptosis would be inhibited in MML-1 cells arrested in G1 phase, which might be related to the up-regulation of Bcl-2 in G1 phase.

Key words: apoptosis, Bcl-2, G1 phase, Fas