漆黄素通过缝隙连接影响顺铂细胞毒性

Fisetin increases the cytotoxicity of cisplatin by enhancing gap junctional intercellular communication

目的体外观察漆黄素对顺铂细胞毒性作用影响及其机制。方法 CCK-8法观察不同浓度漆黄素对人胶质瘤U87细胞的毒性;用细胞接种荧光示踪法测定不同浓度漆黄素对U87细胞缝隙连接(GJ)功能的影响;标准细胞集落形成分析法观察顺铂的毒性及漆黄素对顺铂毒性的影响;用Western blot法研究漆黄素在影响GJIC(GJ intercellular communication)功能浓度范围内对Cx43表达的影响。结果 CCK-8法显示漆黄素在小于1μmol·L-1的浓度范围内无细胞毒性;细胞接种荧光示踪法显示漆黄素浓度越高,U87细胞GJ通讯的荧光传递功能越强;标准细胞集落形成分析法显示,20μmol·L-1顺铂能够抑制U87细胞的集落形成,而且在有GJ形成的细胞顺铂对细胞集落形成的抑制作用显著高于无GJ形成的细胞;Western blot结果显示漆黄素对Cx43蛋白表达量无明显影响。结论漆黄素可以增强顺铂的细胞毒性,该作用可能与漆黄素增强U87细胞的GJ通讯功能有关,与Cx43蛋白表达水平变化无关。

Aim To investigate the effect of fisetin on the cytotoxicity of cisplatin and its mechanisms. Meth-ods CCK-8 was used to assay the toxicity of fisetin.“Parachute” dye-coupling assay was used to examine dye spread through gap junctions in U87 cells. “Stand-ard colony-forming assay” was used to examine cell survivals of U87 treated with cisplatin with fisetin. Western blot was performed to detect the expression level of Cx43 . Results Fisetin had no cytotoxic effect on U87 cells at the concentration of 0~1μmol · L-1 . Parachute assay demonstrated that fisetin enhanced dye-coupling through gap juntions in U87 cells. Stand-ard Colony forming assay showed that cisplatin inhibi-ted the clonogenic survivals of U87 cells and fisetin could enhance cisplatin-induced inhibitory effect of clo- nogenic survivals on cells with gap junctional formation more than on cells without gap junctional formation. Conclusion These rusults demonstrate that fisetin could increase the sensitivity of glioma ceils to cisplatin through the enhancement of gap junctional intercellular communication but this may not be related to the ex- pression of Cx43.