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目的:研究汉防己甲素对人多发性骨髓瘤干细胞CD138~- B细胞增殖、凋亡的影响及作用机制。方法:利用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)比色法检测汉防己甲素对CD138~- B细胞增殖的影响;应用流式细胞仪检测汉防己甲素对CD138~- B细胞周期的影响以及药物诱导细胞凋亡的作用;应用蛋白免疫印迹法(Western blot)检测汉防己甲素对CD138~- B细胞中裂解的半胱氨酸天冬氨酸蛋白水解酶(cleaved-Caspase)-3,cleaved-Caspase-9蛋白表达的影响。结果:与空白组及溶剂组比较,10,20,30,40μmol·L~(-1)汉防己甲素处理6,12,24 h对CD138~- B细胞的增殖均具有显著的抑制作用(P<0.01);与空白组及溶剂组比较,汉防己甲素30,40μmol·L~(-1)时,G_0/G_1期细胞显著增多(P<0.01),S期细胞明显减少(P<0.05,P<0.01),汉防己甲素10,20,30,40μmol·L~(-1)时,细胞凋亡率显著增加(P<0.01);与空白组及溶剂组比较,20,30,40μmol·L~(-1)汉防己甲素处理后,cleaved-Caspase-3,cleaved-Caspase-9蛋白表达显著升高(P<0.01),10μmol·L~(-1)汉防己甲素处理时,cleaved-Caspase-3蛋白表达明显增高(P<0.05)。结论:汉防己甲素通过阻滞细胞周期在G_0/G_1期抑制CD138~- B细胞的增殖,通过活化细胞内与凋亡相关蛋白cleaved-Caspase-3,cleaved-Caspase-9促进细胞凋亡。
Objective: To study the effect and mechanism of tetrandrine on proliferation and apoptosis of human multiple myeloma stem cell CD138 ~ - B cells. Methods: The effect of tetrandrine on the proliferation of CD138 ~ - B cells was detected by MTT assay with 3- (4,5-dimethylthiazol-2) -2,5-diphenyltetrazolium bromide The effect of tetrandrine on the cell cycle of CD138 ~ - B and the effect of drug-induced apoptosis were detected by flow cytometry. The effect of tetrandrine on CD138 ~ - B cells was detected by Western blot Effect of cleaved caspase-3 and cleaved-Caspase-9 protein expression. Results: Compared with blank group and solvent group, 10, 20, 30, 40μmol·L -1 tetrandrine prevented the proliferation of CD138 ~ - B cells at 6, 12 and 24 h P <0.01). Compared with the blank group and the solvent group, the cells in G 0 / G 1 phase increased significantly (P <0.01) and the cells in S phase decreased significantly when treated with 30 and 40 μmol·L -1 of tetrandrine (P < 0.05, P <0.01). The apoptotic rate was significantly increased by 10, 20, 30, 40μmol·L -1 tetrandrine (P <0.01). Compared with the blank group and the solvent group, , Cleaved-Caspase-3 and cleaved-Caspase-9 protein expressions were significantly increased after treated with 40 μmol·L -1 tetrandrine (P <0.01), while those of 10 μmol·L -1 tetrandrine At the time of treatment, cleaved-Caspase-3 protein expression was significantly increased (P <0.05). CONCLUSION: Tetrandrine can inhibit the proliferation of CD138 ~ - B cells in G_0 / G1 phase by arresting the cell cycle, and promote the apoptosis by activating the apoptosis related proteins cleaved-Caspase-3 and cleaved-Caspase-9.