加味参芪地黄汤通过PI3K/AKT/mTOR通路抑制糖酵解治疗激素敏感性前列腺癌作用的机制研究

Modified Shenqi Dihuang Decoction acts on hormone⁃sensitive prostate cancer by inhibiting glycolysis through the PI3K/Akt/mTOR signaling pathway

目的:评估加味参芪地黄汤对人激素敏感型LNCaP细胞的影响及作用机制。方法:通过加味参芪地黄汤干预LNCaP细胞,采用MTT法检测细胞增值,流式细胞术检测细胞凋亡情况;采用葡萄糖摄取试剂盒、乳酸比色测定试剂盒测定葡萄糖摄取量、乳酸生成量;采用Western印迹检测细胞凋亡相关蛋白Bcl⁃2、Bax和cleaved⁃caspase⁃3,糖酵解相关蛋白HK2、GLUT1、PKM2和LDHA,以及PI3K/Akt/mTOR通路相关蛋白的表达情况;采用糖酵解诱导剂寡霉素,PI3K激活剂740 Y⁃P干预观察对加味参芪地黄汤作用的影响。结果:加味参芪地黄汤以剂量依赖方式抑制LNCaP细胞增值,并诱导其凋亡,同时上调Bax、cleaved⁃caspase 3水平,降低Bcl⁃2水平;加味参芪地黄汤能显著降低LNCaP细胞葡萄糖摄取量、乳酸生成量,并抑制HK2、GLUT1、PKM2和LDHA表达水平;加味参芪地黄汤能显著抑制AKT、PI3K和mTOR的磷酸化水平;寡霉素和740 Y⁃P能逆转加味参芪地黄汤对LNCaP细胞增值的抑制作用;740 Y⁃P能逆转加味参芪地黄汤对葡萄糖摄取量、乳酸生成量,和对糖酵解相关蛋白HK2、GLUT1、PKM2和LDH⁃A的抑制作用。结论:加味参芪地黄汤通过抑制PI3K/AKT/mTOR信号通路,抑制糖酵解,进而抑制LNCaP细胞增值。

Objective:To evaluate the effect of Modified Shenqi Dihuang Decoction(MSDD)on human hormone⁃sensitive LN⁃CaP prostate cancer cells and its action mechanism.Methods:LNCaP prostate cancer cells were treated with MSDD,followed by de⁃tection of the proliferation and apoptosis of the cells by MTT assay and flow cytometry respectively and measurement of glucose uptake and lactate production by glucose uptake assay and colorimetry respectively.The expressions of the apoptosis⁃related proteins Bcl⁃2,Bax and cleaved⁃caspase⁃3,glycolysis⁃related proteins HK2,GLUT1,PKM2 and LDHA,and PI3K/AKT/mTOR pathway⁃related pro⁃teins in the LNCaP cells were determined by Western blot.The effect of MSDD on the LNCaP cells was observed with the glycolysis in⁃ducer oligomycin and the PI3K activator 740 Y⁃P.Results:MSDD inhibited the proliferation,induced the apoptosis,increased the levels of Bax and cleaved⁃caspase 3 and decreased the level of Bcl⁃2 in the LNCaP cells in a dose⁃dependent manner.After MSDD in⁃tervention,the glucose uptake and lactate production in the LNCaP cells were significantly reduced,the expressions of HK2,GLUT1,PKM2 and LDHA and the phosphorylation levels of Akt,PI3K and mTOR were markedly suppressed.Oligomycin and 740 Y⁃P re⁃versed the inhibitory effect of MSDD on the proliferation of the LNCaP cells,and 740 Y⁃P reversed that on glucose uptake,lactic acid production and the expressions of the glycolysis⁃related proteins HK2,GLUT1,PKM2 and LDHA in the LNCaP cells.Conclusion:Modified Shenqi Dihuang Decoction inhibits the proliferation of LNCaP prostate cancer cells by suppressing glycolysis and the PI3K/Akt/mTOR signaling pathway.