白花蛇舌草对人肾癌780-0细胞体外增殖及凋亡的影响

Effect of Hedyotis Diffusa on Proliferation and Apoptosis for Human Renal Cell Carcinoma 780-0 Cells in Vitro

目的:探讨白花蛇舌草(Hedyotis diffusa)提取物对人肾癌780-0细胞在体外增殖及凋亡的影响。方法:选择体外培养的人肾癌780-0细胞,分别给予不同浓度的白花蛇舌草提取物(0、25、50、100、200、400、600和800μg/ml)处理细胞,采用CCK8法检测细胞的增殖情况,流式细胞技术检测细胞周期和细胞凋亡情况,荧光定量PCR法检测细胞周期蛋白的表达,Western blot检测凋亡相关蛋白及PI3K/Akt信号通路的表达水平,并应用PI3K/Akt激动剂观察细胞增殖及凋亡变化。结果:不同浓度的白花蛇舌草提取物均能抑制肾癌细胞的增殖,其抑制作用随着药物浓度的升高而增强。相较于对照组,白花蛇舌草干预后G0/G1期细胞百分比显著增加,S期细胞百分比显著减少(P<0.05),细胞周期蛋白(Cyclin A、Cyclin B1、CDK2蛋白)的mRNA表达水平下降。白花蛇舌草处理后细胞凋亡率较对照组明显增加(P<0.05),其凋亡蛋白Caspase-3表达增加、抗凋亡蛋白Bcl-2表达减少。白花蛇舌草抑制PI3K/Akt信号通路的激活,而PI3K/Akt激动剂作用可减弱白花蛇舌草对细胞增殖的抑制及促凋亡作用。结论:白花蛇舌草能显著抑制肾癌780-0细胞的增殖及促进其凋亡,这种作用是通过调节PI3K/Akt信号通路所实现的。

Objective:To investigate the effect of Hedyotis diffusa on the proliferation and apoptosis for human renal cell carcinoma 780-0 cells in vitro.Methods:780-0 cells was selected and by Hedyotis diffusa with various concentrations of active components(0,25,50,100,200,400,600 and 800μg/ml).CCK8 assay was used to detect cell proliferation and flow cytometry was used to detect cell cycle and apoptosis rate.mRNA expression of cyclin was detected by fluorescence quantitative PCR.Western blot was applied to detect the expression of apoptosis-related proteins and PI3 K/Akt signaling pathways.Moreover,PI3 K/Akt agonist was used to observe cell proliferation and apoptosis.Results:Various concentrations of Hedyotis diffusa inhibit the proliferation of renal cell carcinoma cells,and its inhibitory effect increased with increasing drug concentration.As compared to the controlled group,the percentage of G0/G1 phase increased significantly while the percentage of S phase decreased in the Hedyotis diffusa treated group(P<0.05),and the mRNA expression of cyclin(cyclin A,cyclin B1 and CDK2 protein)decreased.Apoptosis rate of 780-0 cells increased significantly in the Hedyotis diffusa treated group comparing to the controlled group(P<0.05).The expression of apoptotic protein caspase-3 was upregulated while anti-apoptotic protein Bcl-2 was downregulated.Hedyotis diffusa suppressed the activation of PI3 K/Akt signaling and PI3 K/Akt agonist attenuated the effects of cell proliferation inhibition and apoptosis promotion.Conclusion:Hedyotis diffusa can significantly inhibit the proliferation and promote the apoptosis of renal cell carcinoma 780-0 cells,which is regulated via PI3 K/Akt signaling pathway.