金水鲜联合放疗对A549细胞血管生成及凋亡相关蛋白表达关系的研究

Study on vascular generation and apoptosis related protein expression of A549 cells using Jinshuixian combined with radiotherapy

目的研究金水鲜联合放疗对肺腺癌A549细胞实验处理后的细胞存活力,以及抗血管生成蛋白表达与凋亡相关蛋白表达的关系变化。方法将生长状态良好的A549细胞按处理因素的不同分为4组,生理盐水(NC)组、金水鲜(JSX)组、放射治疗+金水鲜(RT+JSX)组、放射治疗(RT)组,用CCK8试剂盒处理4 h后酶标仪测OD450值,收集细胞裂解后测定蛋白浓度并蛋白凝胶(WB)电泳,分析血管促进因子蛋白血管生成因子(VEGF)/胎盘源性生长因子(PIGF)和突变型P53以及促凋亡蛋白FAS蛋白的表达变化。结果①CCK8检测各组经过处理后细胞的增殖活力,RT+JSX组细胞的增殖活力最低,与JSX组比较,差异有高度统计学意义(P<0.01);JSX组与NC组比较,差异有统计学意义(P<0.05)。②Western-blot结果显示在第48小时,JSX组VEGF蛋白的表达下降明显;而在RT+JSX组中促凋亡蛋白FAS的表达明显增加,突变型P53蛋白的表达下降,与JSX组比较,差异有高度统计学意义(P<0.01),提示联合使用金水鲜增强了放疗自身的促凋亡作用。结论金水鲜联合放疗可能发挥药物单独具有的血管生成抑制作用,并能增强单用放疗的细胞促凋亡效果,考虑为金水鲜在影响血管生成方面能增加放疗的敏感性,激活细胞凋亡相关因子FAS和下调突变型P53,具体作用机制有待进一步证实。

Objective To study cell viability, anti angiogenic protein expression and apoptosis related protein expression after the experimental treatment of Jinshuixian combined with radiotherapy on lung A549 cells. Methods A549cells in good condition were divided into 4 groups including physiological saline（NC） group, Jinshuixian（JSX）group,radiotherapy ＋ Jinshuixian（RT＋JSX） group, radiation therapy（RT） group, according to the treatment factors according to processing factors, OD450were measured after CCK8 kit for 4 h. Determination of protein concentration and WB electrophoresis were collected after cell lysis. VEGF/PIGF and mutant P53 protein and the expression of pro-apoptotic protein FAS were measured after the process. Results ① The cell proliferation activity of each group after Goldwater fresh and radiotherapy, cell proliferation activity of RT＋JSX group was the lowest, compared with that of JSX group, the differences were statistically significant（P 〈0.01）; cell proliferation activity of JSX group was compared with that of NC group, the difference was statistically significant（P〈0.05）. ② Western-blot experiments showed that in the first48 hours, the expression of VEGF protein was decreased significantly in JSX group; the expression of pro-apoptotic protein FAS in RT＋JSX group was increased, the expression of mutant P53 protein was decreased, compared with JSX group, the differences were statistically significant（P〈0.01）, it prompted Jinshuixian enhanced the pro-apoptotic effects of radiotherapy. Conclusion Jinshuixian combined with radiotherapy may has stronger inhibitory effect on angiogenesis, and can enhance the single cell apoptosis effect of radiotherapy, which is considered as the sensitivity of radiotherapy. This combined therapy can also activate the apoptosis related factor FAS and adjustable mutant P53. But the specific mechanism needs further confirmation.