Autophagic flux induced by serum deprivation was enhanced when Bcl-2 was down-regulated and inhibitors of autophagy and apoptosis rescue cell from death.

<p>A: Western blot analysis of LC3-II and p62 expression in SH-SY5Y cells. Seventy-two hours after transfection with indicated oligonucleotides, the cells were subjected to starvation for 12 hrs in the present or absence of 50 nM Baf A1. B: Quantitative analysis of optical densities of the LC3-II/LC3-I protein bands with Sigma Scan Pro 5. Bars represent Mean ± SD (n = 3). Statistical analysis was carried out with unpaired t-test (*p<0.05, Bcl-2 siRNA #6 group vs negative control group). C: Quantitative analysis of cell viability with MTT assay. SH-SY5Y cells were treated with Bcl-2 siRNA #6 for 72 hrs, and then were treated with 10 µg/ml E64D, 50 nM Baf A1 and 50 µM Z-VAD. Cells were either maintained in normal medium or subjected to 12 hrs or 24 hrs of starvation prior to cell viability assay. **p<0.01, ***P<0.001 represent the indicated groups vs Bcl-2 RNAi #6 starvation 12 hrs group; ##p<0.01 represent the indicated groups vs Bcl-2 RNAi #6 starvation 24 hrs group. D: Representative western blot image of LC3 in SH-SY5Y cells subjected to normal or serum-free medium in the presence or absence of 10 µg/ml E64D for 12 hrs. E and F: Quantitative analysis of cell viability with MTT assay. SH-SY5Y cells were pre-treated with HA 14-1 or ABT-737, then the cells were either maintained in normal or starvation medium for 12 hrs or 24 hrs with E64D, Baf A1 or Z-VAD. **p<0.01 represent the indicated groups vs HA 14-1/ABT-737 starvation 12 hrs groups; #p<0.05, ##p<0.01 represent the indicated groups vs HA 14-1/ABT-737 starvation 24 hrs groups. Statistical analysis was either carried out with Student t-test, or carried out with ANOVA followed by Dunnett t-test. Data represent mean ± SD for combined data from three independent experiments, each experiment has six replicate wells.