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目的研究雷公藤内酯醇对多发性骨髓瘤U266细胞组蛋白去甲基化酶的影响,探讨其表观遗传调控作用。方法采用MTT法检测细胞增殖活性;Annexin V-FITC/PI双标法流式细胞术检测细胞凋亡;RT-PCR法检测U266细胞组蛋白赖氨酸去甲基化酶1(LSD1)mRNA、组蛋白去甲基化酶(JMJD2B)mRNA表达的变化;激光共聚焦显微技术观察LSD1亚细胞定位情况及蛋白量的变化;Western blotting法检测在雷公藤内酯醇干预下LSD1和JMJD2B蛋白表达的变化。结果雷公藤内酯醇以剂量、时间相关方式抑制U266细胞增殖,与细胞培养24 h的IC50为153.2 nmol/L;以剂量相关方式诱导U266细胞凋亡,雷公藤内酯醇80 nmol/L作用U266细胞24 h后,细胞出现典型的凋亡形态学改变,总凋亡率达32.9%。;以剂量相关方式抑制JMJD2B蛋白的表达,同时促进LSD1蛋白表达。结论雷公藤内酯醇在抑制U266细胞增殖、诱导其凋亡的同时,明显改变LSD1、JMJD2B的表达,其诱导U266细胞凋亡和抗肿瘤效应可能与其调节LSD1、JMJD2B表达有关。
Objective To study the effect of triptolide on histone demethylase of U266 cells in multiple myeloma and to explore its epigenetic regulation. Methods Cell proliferation was detected by MTT assay. Apoptosis was detected by flow cytometry with Annexin V-FITC / PI double staining method. The expression of histone lysine demethylase 1 (LSD1) mRNA in U266 cells was detected by RT- Histone demethylase (JMJD2B) mRNA expression changes; laser confocal microscopy LSD1 subcellular localization and protein content changes; Western blotting assay triptolide alcohol intervention LSD1 and JMJD2B protein expression Variety. Results Triptolide inhibited the proliferation of U266 cells in a dose-and time-dependent manner, with an IC50 of 153.2 nmol / L at 24 h. U266 cells were treated with 80 nmol / L triptolide in dose-dependent manner. U266 cells After 24 h, the cells showed typical morphological changes of apoptosis, with a total apoptosis rate of 32.9%. ; Dose-dependent inhibition of JMJD2B protein expression, while promoting LSD1 protein expression. Conclusion Triptolide can inhibit the proliferation and induce the apoptosis of U266 cells, and significantly change the expression of LSD1 and JMJD2B. The apoptosis induced by triptolide and its anti-tumor effect may be related to the regulation of LSD1 and JMJD2B expression.