三七总皂苷对H22细胞的抑制作用及对H22荷瘤小鼠PI3K-AKT-mTOR信号通路的影响

Inhibitory Effects of Panax notoginseng Saponins on H Cells and its Effects on PI3K-AKT-mTOR Signaling Pathway of H Tumor-Bearing Mice

目的:探讨三七总皂苷体内外抑制H22细胞增殖、促进H22细胞凋亡及对PI3K-AKT-mTOR信号通路的影响。方法:不同浓度三七总皂苷体外干预H22细胞24~48 h后,CCK8比色法检测三七总皂苷对H22细胞增殖的抑制作用;Annexin V/PI双荧光染色法检测细胞凋亡率;构建40只雌雄各半C57BL/6小鼠肿瘤模型,随机分为模型组及三七总皂苷125、250、500 mg/kg组,每组10只,灌胃给药,模型组给予等体积生理盐水,2次/d,连续14 d,间隔2 d记录肿瘤大小;给药结束后分离瘤体,称取瘤体质量计算抑瘤率;HE染色观察瘤体细胞形态;Western blot检测AKT、p-AKT、mTOR、p-mTOR、Bax、PARP、Caspase-3蛋白表达。结果:三七总皂苷能有效诱导H22细胞凋亡(P<0.05)。小鼠体内实验结果显示,三七总皂苷高剂量组具有显著的肿瘤抑制作用(P<0.05),并能显著降低肿瘤组织中AKT、mTOR、PARP、Caspase-3及磷酸化蛋白p-AKT、p-mTOR表达,显著升高Bax蛋白表达(P<0.05)。结论:三七总皂苷具有体内外抑制H22细胞增殖、促进H22细胞凋亡的作用,其机制可能与负调控PI3K-AKT-mTOR信号通路有关。

Objective: To study the effect of Panax notoginseng saponins(PNS)on the proliferation and apoptosis of H22 cells through PI3 K-AKT-mTOR signaling pathway in vitro and in vivo.Methods: H22 cells were treated with different concentrations of PNS for 24 to 48 hours, CCK8 colorimetric method was used to detect the inhibitory effect of PNS on the proliferation of H22 cells;Annexin V/PI double fluorescence staining was used to detect the apoptosis rate;Forty half-male and half-female C57 BL/6 tumor models mice were constructed, the mice were randomly divided into mice, PNS 125,250 and 500 mg/kg groups, with 10 animals in each group.The mice were given intragastric administration, and the model group was given equal volume normal saline, twice/d, continuously for 14 days, and the tumor size was recorded at an interval of 2 days.After the administration, the tumor was separated and the tumor mass was measured to calculate the tumor inhibition rate.The morphology of tumor cells was observed by HE staining, the expressions of AKT,p-AKT,mTOR,p-mTOR,Bax, PARP and Caspase 3 proteins were determined by Western blot.Results: PNS could effectively induce apoptosis of H22 cells(P<0.05).In vivo experimental results in mice showed that PNS with the high-dose group had significant tumor inhibition(P<0.05),could significantly decrease the expressions of AKT,mTOR,PARP,Caspase-3 and phosphorylated protein p-AKT and p-mTOR in the tumor tissue, significantly increase the expression of Bax protein(P<0.05).Conclusion: Panax notoginseng saponins can inhibit H22 cells proliferation and promote H22 apoptosis in vitro and in vivo.The mechanism may be related to the negative regulation of PI3 K-AKT-mTOR signaling pathway.