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目的:以人肺癌A549细胞为研究对象,探讨玉竹中提取的高异黄酮对A549细胞增殖的抑制作用及其作用机制。方法:从玉竹中提取高异黄酮,作用于肺癌细胞A549;噻唑蓝(MTT)比色法观察高异黄酮12.5,25,50,100 mg·L~(-1)作用于A549细胞6,12,24 h对细胞存活率的影响。通过流式细胞仪检测细胞凋亡及周期变化,并通过蛋白免疫印迹法(Western blot)分析与细胞凋亡相关半胱氨酸天冬氨酸蛋白酶-3(Caspase-3),B细胞淋巴瘤(白血病)-2(Bcl-2),Bcl-2对抗杀伤性蛋白(Bak)和与细胞周期相关的磷酸化周期素依赖激酶2(p-Cdc 2),细胞周期蛋白依赖性激酶2(Cdc 2)及p38蛋白表达。结果:与空白组比较,高异黄酮呈剂量和时间依赖性抑制A549细胞的增殖(P<0.05,P<0.01),高异黄酮对A549细胞处理12 h后,细胞周期阻滞于G2/M期(P<0.05);与空白组比较,25,50,100 mg·L~(-1)高异黄酮可显著促进凋亡蛋白Caspase-3和Bak表达,抑制Bcl-2表达(P<0.01),可显著促进细胞周期蛋白p-Cdc 2和p38表达,抑制Cdc 2表达(P<0.01)。结论:玉竹中提取的高异黄酮对A549细胞增殖具有抑制作用,能使A549细胞阻滞于细胞周期G2/M期,其机制与线粒体介导的细胞凋亡和p38 MAPK通路相关。
OBJECTIVE: To investigate the inhibitory effect of isoflavones extracted from Yuzhu on the proliferation of A549 cells and its possible mechanism. METHODS: High isoflavone was extracted from Polygonatum soongorhizobium for lung cancer cell line A549. MTT assay was used to observe the effect of high isoflavone (12.5, 25, 50, 100 mg · L -1) on A549 cell line 6,12,24 h on cell viability. Flow cytometry was used to detect apoptosis and cell cycle changes. Western blot analysis was used to detect the expressions of caspase-3, B-cell lymphoma, (Leukemia) -2 (Bcl-2), Bcl-2 on the antiproliferative protein (Bak) and the cell cycle associated phosphorylated cyclin dependent kinase 2 (p-Cdc 2), cyclin dependent kinase 2 2) and p38 protein expression. Results: Compared with the blank group, high isoflavone inhibited the proliferation of A549 cells in a dose and time dependent manner (P <0.05, P <0.01). After isoflavones treated A549 cells for 12 h, cell cycle arrest at G2 / M (P <0.05). Compared with the blank group, 25, 50 and 100 mg · L -1 isoflavones could significantly promote the expression of Caspase-3 and Bak and inhibit the expression of Bcl-2 (P <0.01) The expressions of p-Cdc 2 and p38 and Cdc 2 were significantly increased (P <0.01). CONCLUSION: The high isoflavone extracted from Polygonatum can inhibit the proliferation of A549 cells and arrest the cell cycle G2 / M in A549 cells. The mechanism is related to mitochondria-mediated apoptosis and p38 MAPK pathway.