雷公藤内酯醇对多发性骨髓瘤细胞凋亡及组蛋白H3K4甲基化的影响

目的：探讨雷公藤内酯醇（TPL）对多发性骨髓瘤RPMI8226细胞增殖、凋亡和组蛋白H3K4甲基化的影响。方法：以人多发性骨髓瘤细胞株RPMI8226为研究对象，在不同浓度（10、20、40、80、160 nmol/L） TPL中共培养不同时间（24 h、48 h、72 h）后，采用噻唑蓝（MTT）法检测细胞增殖活性；流式细胞术检测细胞凋亡和细胞周期；Western blot法检测组蛋白H3K4me2、H3K4me3的甲基化状态，实时荧光定量RT-PCR分析组蛋白甲基化酶SMYD3和组蛋白去甲基化酶LSD1的表达水平。结果：TPL对RPMI8226细胞有明显的增殖抑制作用，呈剂量和时间依赖性（P<0.05）；TPL对RPMI8226细胞有明显诱导凋亡的作用，并且随着TPL作用浓度的增加，细胞凋亡比例逐渐增加（P<0.05）；同时TPL还可以诱导RPMI8226细胞周期阻滞于G₂/M期；TPL以浓度依赖性降低组蛋白H3K4me2、H3K4me3的甲基化水平（P<0.05，P<0.01），并抑制SMYD3和上调LSD1的表达（P<0.05）。结论：TPL可抑制RPMI8226细胞增殖、引起细胞周期阻滞于G₂/M期，并诱导其凋亡；通过抑制组蛋白甲基化酶SMYD3和增强组蛋白去甲基化酶LSD1的表达，降低组蛋白H3K4me3和H3K4me2的甲基化水平，这可能是TPL诱导多发性骨髓瘤细胞凋亡和抗肿瘤作用的机制之一。

Effects of triptolide on the apoptosis and H3K4 protein methylation in multiple myeloma cells

Objective: To investigate the effects of triptolide (Chinese Traditional Medicine)on the apoptosis and H3K4 protein methylation in multiple myeloma cells.Methods: The RPMI8226 cells were cultured with different concentrations(10,20,40,80 and 160 nmol/L)of triptolide for different incubation time (24 h,48 h and 72 h). The inhibition of triptolide on RPMI8226 cell proliferation was detected by MTT assay. Apoptosis and cell cycle distribution were evaluated by flow cytometry.The expressions of H3K4me2 and trimethylation of histone H3 lysine 4(H3K4me3) in RPMI8226 cells were assayed by Western blot. The changes of expressions of histone methylase SET and MYND domain containing 3(SMYD3) and histone demethylase lysine specific demethylase 1(LSD1) in RPMI8226 cells were verified by qRT-PCR.Results: Triptolide had obvious inhibitive effects on proliferation of RPMI8226 cells and showed a dose-and time-dependent manner(P<0.05). Triptolide induced apoptosis and G₂/M cell cycle arrest in a dose-dependent manner(P<0.05). Triptolide decreased histone H3K4me2 and H3K4me3 expression in a dose-dependent manner(P<0.05, P<0.01). SMYD3 was significantly depressed at protein expression in a dose-dependent manner(P<0.05), but LSD1 was up-regulated (P<0.05).Conclusion: Triptolide could inhibit RPMI8226 cell proliferation,induce the apoptosis and cause G₂/M arrest,meanwhile,significantly inhibit the protein expressions of H3K4me2 and H3K4me2 with alter the expression of SMYD3 and LSD1.The effects is probably related to the antitumor mechanism of MM cells induced by triptolide.