鱼藤素对人骨髓瘤细胞RPMI-8226增殖和凋亡的影响及对核受体共激活因子-3的调控作用

Effect on proliferation and apoptosis in RPMI-8226 human myeloma cells and regulation on steroid receptor coactivator-3 by deguelin

目的观察鱼藤素对人多发性骨髓瘤细胞株RPMI-8226细胞增殖和凋亡的影响及其对核受体共激活因子-3(SRC-3)的调控作用,研究鱼藤素诱导凋亡作用与其调节SRC-3的相互关系。方法采用MTT法检测细胞增殖活性,Annexin-VFITC/PI双标法及Hoechst33258染色法分析细胞凋亡的改变,RT-PCR法检测鱼藤素对RPMI-8226细胞内SRC-3基因表达的影响;免疫组化法检测鱼藤素对RPMI-8226细胞SRC-3蛋白的影响和分布情况。结果鱼藤素能明显抑制RPMI-8226细胞的增殖,其抑制作用呈时间、剂量依赖性,其作用24h的IC50为(54.55±0.40)nmol/L。此外,鱼藤素具有较强的诱导RPMI-8226细胞凋亡的效应,25nmol/L即能诱导(17.04±0.73)%的RPMI-8226细胞发生凋亡;当药物浓度达到100nmol/L时,总凋亡率达(63.57±1.36)%。在RPMI-8226细胞中SRC-3高表达,且主要分布于细胞核,经鱼藤素干预后,SRC-3的mRNA和蛋白表达水平明显下降。结论鱼藤素能明显抑制RPMI-8226细胞的增殖,并诱导其凋亡。而鱼藤素诱导的SRC-3表达量的下调可能参与了其诱导凋亡作用。

Objective To investigate the effects of deguelin on the proliferation inhibition and apoptosis induction in RPMI-8226 cells in vitro, as well as the regulation of steroid receptor coactivator-3 （SRC-3） to explore the relationship between them. Methods The effect of deguelin on the growth of RPMI-8226 cells was studied by MTT assay. Apoptosis was detected through Hoechst 33258 staining and Annexin-V FITC/PI double-labeled cytometry. RT-PCR Technology was applied to assessment of the mRNA expres- sion of SRC-3, whereas, SRC-3 protein expression and localization were determined by using immunohisto- chemistry method. Results Deguelin presented striking proliferation inhibition potency on RPMI-8226 cells in vitro and apoptosis induction activity in a time- and dose-dependent manner. The IC50 value for 24 h was （54.55±0.40） nmol/L, （17.04±0.73）% RPMI-8226 cells went apoptotic when treated with 25 nmol/L deguelin for 24 h, with the dose of deguelin increasing to 100 nmol/L, （63.57±1.36）o/6o cells were apoptotic. Over-expression of SRC-3 was found in RPMI-8226 cells, whereas the mRNA and protein ex- pression levels of SRC-3 were significantly downregulated in RPMI-8226 cells induced by deguelin in a dose-dependent manner. The disposition of SRC-3 was situated mainly at the nuclear, occasionally in the cytoplasm. Conclusion Deguelin exhibited potent proliferation inhibition to the human myeloma cell line RPMI-8226 cells, furthermore, deguelin might induce RPMI-8226 cells apoptosis in a dose-dependent man- ner, which might correspond to the regulation of the expression of SRC-3.