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首页> 中文期刊> 《中国医师杂志》 >β-谷甾醇诱导人胃癌细胞自噬与凋亡的作用及机制研究

β-谷甾醇诱导人胃癌细胞自噬与凋亡的作用及机制研究

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Objective To study the effect and mechanism of autophagy in gastric cancer cells induced by β-sitosterol in vivo and in vitro,providing experimental evidence for antitumor study of β-sitosterol on gastric cancer.Methods Human gastric cancer SGC-7901 cells were cultured in vitro.The control group was treated with phosphate buffer (PBS) only,and the experimental group was treated with different concentrations of β-sitosterol.Tetrazolium salt colorimetry (MTT) was used to detect the inhibitory effect of β-sitosterol on gastric cancer cells.Flow cytometry was used to detect the apoptotic effect of β-sitosterol on gastric cancer cells.Green fluorescent protein labeling (GFP) was used to detect the induction of autophagy by β-sitosterol.Western blot was used to detect the expression of signal pathway proteins phosphorylated phosphoinositid 3-kinase (p-PI3K),phosphorylated protein kinase B (p-AKT) and phosphorylated mammalian target of rapamycin (p-mTOR).The nude mice model of gastric cancer was established by SGC-7901 cells.β-sitosterol was injected abdominally once every 3 days for 42 days.The weight and volume of transplanted tumors were observed.The expression of LC3-Ⅱ and LC3-Ⅰ in transplanted tumors was detected by immunohistochemistry.Results β-sitosterol could inhibite the viability and promote the apoptosis of SGC-7901 cells,inducing the autophagy of SGC-7901 cells when its concentration reached 20 μmol/L.The autophagy rate of experimental group was (19.32 ±2.51,32.25 ±3.18,57.45 ±4.02,78.93 ±4.21),which was significantly higher than that in the control group (7.28 ± 2.83,P ＜ 0.05).In the experimental group,the expression of p-PI3K,p-AKT and mTOR was down-regulated by β-sitosterol,which was significantly different from that in the control group (P ＜ 0.05).The inhibition rate of the experimental group was (69.8 ± 3.6) ％,and the weight and volume of the transplanted tumors were significantly reduced compared with those in the control group (P ＜ 0.05).β-sitosterol increased the expression of LC3-Ⅱ,decreased expression of LC3-Ⅰ,leading the rise of ratio of LC3-Ⅱ to LC3-Ⅰ in nude mouse tumor (t =7.28,P =0.008).Conclusions β-sitosterol can inhibite proliferation and promote apoptosis of SGC-7901 cells.The antitumor mechanism may related to the autophagy induced by beta-sitosterol through PI3K/AKT/mTOR pathway.%目的 从体内外两个方面研究β-谷甾醇诱导胃癌细胞自噬的作用及机制,为β-谷甾醇抗胃癌作用研究提供实验依据.方法 体外培养人胃癌SGC-7901细胞,设对照组和实验组,对照组仅用磷酸盐缓冲液(PBS)处理,实验组以不同浓度β-谷甾醇处理.四唑盐比色法(MTT)检测β-谷甾醇对胃癌细胞的抑制作用,流式细胞术检测β-谷甾醇对胃癌细胞的促凋亡作用,绿色荧光蛋白标记法(GFP)检测β-谷甾醇对胃癌细胞自噬诱导作用,蛋白印迹法(Western-blot)检测信号通道蛋白磷酸化磷脂酰肌醇3激酶(p-PI3K)、磷酸化蛋白激酶B(p-AKT)及磷酸化哺乳动物雷帕霉素靶蛋白(p-mTOR)的表达.用SGC-7901细胞制造胃癌裸鼠模型,经腹胫注射β-谷甾醇,3d给药1次,共42d,观察移植瘤重量体积变化,免疫组化法检测移植瘤组织LC3-Ⅱ、LC3-Ⅰ表达.结果 实验组β-谷甾醇浓度达20 μmol/L时可明显抑制SGC-7901细胞活力,促进SGC-7901凋亡,与对照组比较差异有统计学意义(P ＜0.05或P＜0.01);β-谷甾醇诱导SGC-7901细胞发生自噬效应,(20、40、80、160μmol/L)实验组细胞自噬率(19.32±2.51、32.25±3.18、57.45±4.02、78.93 ±4.21)与对照组(7.28±2.83)比较,差异有统计学意义(P＜0.05,P＜0.01);实验组β-谷甾醇下调p-PI3K、p-AKT及mTOR表达,与对照组比较差异有统计学意义(P＜0.05);实验组抑瘤率为(69.8±3.6)％,移植瘤的重量和体积明显减轻变小,与对照组比较差异均有统计学意义(P＜0.05);实验组β-谷甾醇上调裸鼠移植瘤细胞LC3-Ⅱ表达,下调LC3-Ⅰ表达,使LC3-Ⅱ/LC3-Ⅰ (7.76±0.35)升高,与对照组(0.32±0.21)比较,差异有统计学意义(t=7.28,P =0.008).结论 β-谷甾醇能抑制胃癌细胞增殖并促进其凋亡,其抗癌机制可能与β-谷甾醇通过PI3 K/AKT/mTOR通路诱导细胞自噬作用有关.

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