刺梨多糖抑制PI3K/Akt/mTOR通路和抗氧化作用诱导前列腺癌DU145细胞凋亡

Rose roxburghii polysaccharide-induced apoptosis of prostate cancer DU145 cells by inhibiting PI3K/Akt/mTOR pathway and antioxidant effects

基于磷脂酰肌醇3-激酶(phosphoinositide 3-kinase,PI3K)/蛋白激酶B(protein kinase B,Akt)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)信号通路,和通路相关分子磷酸酯酶与张力蛋白同源物(phosphatase and tensin homolog,PTEN)、B细胞淋巴瘤/白血病-2基因(B-cell lymphoma-2,Bcl-2)、与Bcl-2关联X蛋白(Bcl-2-associated X protein,BAX),探究刺梨多糖抑制前列腺癌DU145细胞增殖,诱导细胞凋亡的作用机制,并探讨刺梨多糖的抗氧化活性。使用不同浓度的刺梨多糖处理前列腺癌DU145细胞,采用CCK-8法检测刺梨多糖对DU145细胞增殖的影响;细胞划痕实验检测刺梨多糖对DU145细胞迁移的影响;Transwell实验检测刺梨多糖对DU145细胞侵袭能力的影响;流式细胞术检测刺梨多糖诱导DU145细胞的凋亡情况;real-time PCR技术和Western blot技术检测刺梨多糖对DU145细胞PI3K、Akt、mTOR、PTEN、BAX、Bcl-2的mRNA及蛋白表达水平的影响。二苯基苦基苯肼(DPPH)和2,2-联氮-二(3-乙基-苯并噻唑-6-磺酸)二铵盐(ABTS)实验检测刺梨多糖的体外抗氧化活性。实验结果表明,刺梨多糖对DU145细胞的增殖、迁移和侵袭均有抑制作用。流式细胞术结果显示,与对照组相比,经刺梨多糖处理后的各组DU145细胞凋亡率随着多糖浓度的增加而升高;抑制性与多糖浓度呈正相关。刺梨多糖对PI3K/Akt/mTOR通路有抑制作用,通过上调PTEN、BAX基因和蛋白的表达,下调Akt、PI3K、mTOR和Bcl-2的表达,诱导DU145细胞凋亡。刺梨多糖对ABTS和DPPH自由基均有一定的清除作用,提示刺梨多糖可能通过清除细胞内活性氧(ROS)诱导DU145细胞凋亡。刺梨多糖可能通过抑制PI3K/Akt/mTOR通路和清除细胞内ROS抑制DU145细胞增殖,诱导细胞凋亡。

Based on the signaling pathway of phosphoinositide 3-kinase(PI3K)/protein kinase B(Akt)/mammalian target of rapa-mycin(mTOR),pathway-related phosphatase and tensin homolog(PTEN),B-cell lymphoma-2(Bcl-2),and Bcl-2-associated X pro-tein(BAX),the mechanism of Rose roxburghii polysaccharides in inhibiting proliferation and inducing apoptosis of prostate cancer DU145 cells was explored,as well as the antioxidant activity of R.roxburghii polysaccharides.Prostate cancer DU145 cells were trea-ted with different concentrations of R.roxburghii polysaccharides.The effect of R.roxburghii polysaccharides on the proliferation of DU145 cells was detected by the CCK-8 method,and the effect of R.roxburghii polysaccharides on the migration of DU145 cells was detected by cell scratch test.In addition,a Transwell assay was conducted to detect the effect of R.roxburghii polysaccharides on the invasion ability of DU145 cells.The apoptosis of DU145 cells induced by R.roxburghii polysaccharides was detected by flow cytome-try.Real-time PCR and Western blot were used to detect the effects of R.roxburghii polysaccharides on the mRNA and protein expres-sion levels of PI3K,Akt,mTOR,PTEN,BAX,and Bcl-2 in DU145 cells.DPPH and ABTS assays were used to determine the an-tioxidant activity of R.roxburghii polysaccharides in vitro.The results showed that R.roxburghii polysaccharides inhibited the prolife-ration,migration,and invasion of DU145 cells.Flow cytometry analysis showed that compared with that of the control group,the apop-tosis rate of DU145 cells in groups treated with R.roxburghii polysaccharides was increased with the increase in the concentration of R.roxburghii polysaccharides,and its inhibition was positively correlated with the concentration of R.roxburghii polysaccharides.R.roxburghii polysaccharides inhibited the PI3K/Akt/mTOR pathway and induced apoptosis of DU145 cells by up-regulating the pro-tein and gene expression of PTEN and BAX and down-regulating the expression of Akt,PI3K,mTOR,and Bcl-2.R.roxburghii poly-saccharides could scavenge ABTS and DPPH radicals to a certain extent,suggesting that R.roxburghii polysaccharides may induce the apoptosis of DU145 cells by scavenging intracellular ROS.R.roxburghii polysaccharides may inhibit the proliferation of DU145 cells and induce its apoptosis by inhibiting the PI3K/Akt/mTOR pathway and clearing intracellular ROS.