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目的探讨左旋棉酚诱导淋巴瘤Daudi细胞发生自噬可能机制及对细胞存活率的影响。方法采用CCK-8法检测左旋棉酚在体外对Daudi细胞增殖抑制作用的影响;采用台盼蓝排斥实验检测不同处理对细胞存活率的影响;Western blot检测细胞中自噬相关蛋白LC3、ERK和磷酸化ERK的表达情况;AO染色观察经左旋棉酚处理后Daudi细胞酸性小体的变化情况。结果左旋棉酚能剂量依赖性地抑制Daudi细胞的增殖和促进细胞死亡;AO染色后经左旋棉酚处理的细胞内可观察到大量的酸性小体形成;Western blot显示左旋棉酚能显著上调自噬相关蛋白LC3Ⅱ的表达及增强磷酸化ERK的水平,抑制ERK的磷酸化能下调LC3Ⅱ的表达;ERK抑制剂U0126及自噬抑制剂CQ和3-MA均能显著增强左旋棉酚的杀细胞效力。结论左旋棉酚可能通过ERK通路诱导Daudi细胞发生自噬,抑制ERK介导的自噬能够显著增强左旋棉酚的抗瘤效应。
Objective To investigate the possible mechanism of autophagy induced by L-Gossypol in Daudi cells and its effect on cell viability. Methods CCK-8 was used to detect the effect of L-gossypol on the proliferation inhibition of Daudi cells in vitro. The effects of different treatments on cell viability were detected by trypan blue exclusion assay. The expressions of LC3, ERK and Phosphorylation of ERK expression; AO staining Daudi cells treated with L-gossypol acidic body changes. Results L-gossypol could inhibit the proliferation of Daudi cells and promote cell death in a dose-dependent manner. A large number of acidic bodies were observed in the cells treated with L-gossypol after AO staining. Western blot showed that L-gossypol could significantly up-regulated from The expression of LC3Ⅱand the phosphorylation of ERK increased, and the phosphorylation of ERK inhibited the expression of LC3Ⅱ. The ERK inhibitor U0126 and the autophagy inhibitors CQ and 3-MA significantly enhanced the cytotoxicity of L-gossypol . Conclusion L-gossypol may induce autophagy in Daudi cells via ERK pathway. Inhibition of ERK-mediated autophagy can significantly enhance the anti-tumor effects of L-gossypol.