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[摘要]
目的 研究人参皂苷Rh2(Rh2)对人白血病细胞KG1α增殖的抑制作用,并从自噬凋亡角度来探讨其机制。方法 采用CCK-8法检测Rh2对KG1α细胞增殖的抑制作用;流式细胞术(FCM)检测细胞凋亡;Hoechest染色观察细胞核染色质的形态;吖啶橙染色观察Rh2对细胞自噬的影响;Western blotting和RT-PCR检测Rh2对白血病细胞自噬重要蛋白和基因表达的影响;运用自噬抑制剂(3-MA)研究自噬对细胞增殖和凋亡的影响。结果 CCK-8显示Rh2在低浓度时能有效抑制KG1α细胞增殖,且其抑制作用具有浓度和时间依赖性;FCM检测和Hoechest染色结果显示Rh2能增加细胞的凋亡,使染色质呈凋亡形态改变;吖啶橙染色发现Rh2组细胞绿色荧光增强,细胞出现大量的酸性自噬小泡;Western blotting和RT-PCR结果发现Rh2上调Beclin-1、LC3A、LC3B、激活型Caspase-3表达和增加Bax/Bcl-2值,并激活MAPK、ATK、ERK信号通路;细胞自噬剂(3-MA)会削弱Rh2对KG1α细胞的增殖抑制和促凋亡作用。结论 Rh2可能通过激活MAPK、ATK、ERK信号通路,诱导细胞自噬途径,从而抑制KG1α细胞增殖和促进其凋亡。
[Key word]
[Abstract]
Objective To investigate the antitumour activity of ginsenoside Rh2 against human leukemia KG1α cells through apoptosis and autophagy pathway. Methods CCK-8 assay was used to screen the most effective ingredient on the proliferations among ginsenoside Rh2 in leukemia KG1α cell line; FCM detected cell apoptosis; Hoechst staining observed the cell morphological changes of apoptosis; Acridine staining detected Rh2 effected on autophagy; Western blotting and RT-PCR detected the expression levels of the proteins closely associated with autophagy and apoptosis. After joining autophagy inhibitors, using CCK-8 to test the proliferation activity of cells, cell apoptosis was measured by FCM. Results CCK-8 indicated that Rh2 could inhibit the proliferation of KG1α cells significantly with dose- and time-dependent manners; FCM indicated that Rh2 induced apoptosis; Hoechest staining showed that KG1α cells had typical apoptotic morphological changes by treated Rh2; Acridine staining revealed that Rh2 cause increase in the number of acidic autophagy vesicles in cells, causing cell autophagy levels increased; Western blotting and RT-PCR results showed that Rh2 increased the expression of Beclin-1, LC3A, and LC3B, activated Caspase-3 and Bax/Bcl-2 rates, and MAPK, ATK, and ERK signaling pathway; After using autophagy inhibitors(3-MA), autophagy crippled that Rh2 inhibited the proliferation and induced apoptosis in KG1α cells. Conclusion Ginsenoside Rh2 could significantly enhance autophagy through activated MAPK, ATK, and ERK signaling pathway, and then inhibit the proliferation and induce apoptosis in KG1α cells.
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[基金项目]
国家自然科学基金资助项目(31271368)