Inhibition of PKC-α enhances the toxicity of TNF-α to H22 cell Lines

Objective To investigate the effect of tumor necrosis factor alpha ( TNF-α) in combination with Go6976 on murine liver cancer cells (H22). Method H22 cells were divided into two groups. Each group was further divided into four subgroups. Group 1 was treated with TNF-α 0, 20, 40, 60ng/ml. Group 2 was treated with TNF-α 0, 20, 40, 60 ng/ml and Go6976 (4.6 nmol/ml). The apoptotic rate, protein kinase C alpha (PKC-α) expression and phosphorylation-PKC-α (p-PKC-α) were detected by flow cytometer and Western blotting respectively in 4 h, 8 h and 16 h. Result Treated with TNF-α (0, 20, 40, 60 ng/ml) for 4 h, the apoptotic rates of H22 cells were 2. 44% ± 0. 31 % , 1. 80% ± 0. 32% , 2. 73% ±0. 14% and 3. 05% ±0. 78% respectively, with no change on the expression of PKC-α and p-PKC-α; For 8 h, the expressions of PKC-α and p-PKC-α in H22 cells were up-regulated with increasing concentration of TNF-α; When PKC-α was inhibited with Go6976 at the same time, the apoptotic rates of cells increased significantly, being 2. 90% ±0.39%, 7.76% ±0.35%, 11.43% ±1.05% and 12.96% ±2.44% , respectively. Moreover, PKC-α and p-PKC-α were down-regulated accordingly. When H22 cells were treated with TNF-α only or combined with Go6976 for 16 h, the result was similar to that of 8 h; The apoptotic rate dropped in the group in which PKC-α was inhibited by Go6976 with TNF-α at 60 ng/ml, but the proportion of necrotic cells increased. Conclusion TNF-α up-regulates the expression of PKC-α and p-PKC-α in H22 cells. Inhibiting the activity of PKC-α significantly enhances the toxicity of TNF-α to H22 cells.