从mTOR信号通路研究风柜斗草醇提物含药血清诱导HepG2细胞凋亡作用及机制

To study the apoptosis effect and mechanism of HepG2 cell induced by the serum containing alcohol extract of Sarcopyramis nepalensis wall from mTOR signaling pathway

关键词:  风柜斗草  含药血清  HepG2  凋亡  mTOR信号通路  

Key words:  Sarcopyramis nepalensis Walll  ? serumcontaining  HepG2  Apoptosis  mTOR signaling pathway.

摘要:

摘 要：目的：研究风柜斗草含药血清对肝癌HepG2细胞mTOR信号通路中相关基因及细胞增殖与凋亡的影响。方法：将肝癌HepG2细胞体外培养，分为对照组、风柜斗草含药血清高、中、低剂量组，采用CCK-8、细胞克隆形成检测HepG2细胞增殖情况，FDA染色观察HepG2细胞活力，Transewell检测HepG2细胞侵袭能力；Annexin V-FITC/PI双染法检测HepG2细胞凋亡情况；Western blot检测各组细胞中Caspase-9、Caspase-3、Bax、Bcl-2、mTOR、p-mTOR蛋白表达水平。结果：风柜斗草各含药血清组均可降低HepG2细胞增殖能力与细胞活力（P<0.01），抑制HepG2细胞克隆形成率（P<0.01或P<0.05）与侵袭能力，诱导HepG2细胞凋亡。风柜斗草含药血清组能上调caspase-9、caspase-3、bax、p-mTOR/mTOR蛋白表达（P<0.01），下调bcl-2蛋白表达（P<0.01），且具有一定的浓度依赖性；风柜斗草中剂量含药血清联合雷帕霉素抑制剂（Rapamycin）可协同增强caspase-9、caspase-3、bax、p-mTOR/mTOR蛋白表达（P<0.01），下调bcl-2蛋白表达（P<0.01）。结论：风柜斗草含药血清可有效抑制HepG2细胞增殖与侵袭，促进细胞凋亡，并能抑制HepG2细胞mTOR信号通路mTOR蛋白磷酸化，上调促凋亡蛋白caspase-9、caspase-3、bax表达，下调抗凋亡蛋白bcl-2表达。 关键词：风柜斗草；含药血清；HepG2；mTOR信号通路；

Abstract:

ABSTRACT: OBJECTIVEThis study investigates the effects of Sarcopyramis nepalensis Wall. medicated serum on the mTOR signaling pathway, cell proliferation, and apoptosis in hepatocellular carcinoma HepG2 cells.METHODS HepG2 cells were cultured in vitro and divided into a control group and high-, medium-, and low-dose Sarcopyramis nepalensis Wall. medicated serum groups. Cell proliferation was assessed using the CCK-8 assay and colony formation assay. Cell viability was observed using FDA staining, and cell invasion ability was evaluated using the Transwell assay. Apoptosis was detected using Annexin V-FITC/PI double staining. The protein expression levels of Caspase-9, Caspase-3, Bax, Bcl-2, mTOR, and p-mTOR were measured by Western blot analysis.RESULTS All Sarcopyramis nepalensis Wall. medicated serum groups significantly reduced the proliferation capacity and viability of HepG2 cells (P < 0.01), inhibited the colony formation rate (P < 0.01 or P < 0.05) and invasion ability of HepG2 cells, and induced apoptosis in HepG2 cells. The Sarcopyramis nepalensis Wall. medicated serum groups upregulated the protein expression of caspase-9, caspase-3, Bax, and p-mTOR/mTOR (P < 0.01) and downregulated the protein expression of Bcl-2 (P < 0.01), showing a certain concentration-dependent effect. Furthermore, the medium-dose Sarcopyramis nepalensis Wall. medicated serum combined with the mTOR inhibitor Rapamycin synergistically enhanced the protein expression of caspase-9, caspase-3, Bax, and p-mTOR/mTOR (P < 0.01) and downregulated the protein expression of Bcl-2 (P < 0.01).CONCLUSION The Sarcopyramis nepalensis Wall. medicated serum effectively inhibited the proliferation and invasion of HepG2 cells, promoted cell apoptosis, and suppressed the phosphorylation of mTOR protein in the mTOR signaling pathway. Additionally, it upregulated the expression of pro-apoptotic proteins caspase-9, caspase-3, and Bax, while downregulating the expression of the anti-apoptotic protein Bcl-2.