RACK1对硼替佐米诱导的多发性骨髓瘤细胞凋亡及MAPK/ERK通路的影响

Effect of RACK1 on Bortezomib-induced Apoptosis and MAPK/ERK Pathway in the Multiple Myeloma Cells

目的:探讨蛋白激酶C受体(Receptor for activated C kinase1,RACK1)对硼替佐米(Bortezomi,Bor)诱导的多发性骨髓瘤(Multiple myeloma,MM)细胞凋亡及MAPK/ERK通路的影响。方法:选取6例岳阳市第一人民医院收治的MM患者及6名正常体检者,用实时荧光定量PCR检测血浆及人MM细胞系中RACK1 m RNA的表达。将MM细胞分为3组:对照组(不干预)、Bor组(75n M的Bor干预12 h)和Bor+siRACK1组(RACK1 si RNA转染24 h后再行Bor干预)。CCK-8法检测各组细胞中的细胞存活率,Hoechest 33342染色检测细胞凋亡,Western Blot检测MAPK/ERK通路相关蛋白表达。结果:与正常体检者相比,MM患者血浆及MM细胞系中RACK1 m RNA表达显著增加(P<0.05)。Bor作用12 h、24 h和48 h可显著降低MM细胞的存活率(P<0.05)。与对照组相比,Bor组和Bor+siRACK1组细胞存活率显著降低,Bor+siRACK1组细胞存活率明显高于Bor组(P<0.05)。Hoechest 33342染色显示对照组细胞核染色均一,未见凋亡小体,Bor组见少量凋亡小体,而Bor+siRACK1组细胞见大量凋亡小体,表现为核固缩或碎块状;与对照组相比,Bor组和Bor+siRACK1组细胞中多发性骨髓瘤细胞凋亡率显著增加(P<0.05),Bor+siRACK1组多发性骨髓瘤细胞凋亡率明显高于Bor组(P<0.05)。三组间多发性骨髓瘤细胞凋亡率对比差异有统计学意义(P<0.05)。与对照组相比,Bor组和Bor+siRACK1组细胞中p-P38和p-ERK的表达显著降低,而Bor+siRACK1组p-P38和p-ERK的表达低于Bor组(P<0.05),3组间P38和ERK的表达对比差异无统计学意义(P>0.05)。结论:RACK1沉默可增强Bor诱导的MM细胞凋亡及生长抑制,其机制可能与MAPK/ERK途径抑制有关。

Objective:To investigate the effect of RACK1 on Bordzomib(Bor)induced apoptosis and MAPK/ERK pathway in multiple myeloma(MM)cells.Methods:6 cases of MM patients admitted to the First People’s Hospital of Yueyang and 6 normal subjects were selected.Real-time quantitative PCR was used to detect the expression of RACK1 m RNA in the plasma and human MM cell lines.MM cells were divided into 3 groups:control group(no intervention),Bor group(75 nM Bor intervention for 12 h)and Bor+siRACK1 group(RACK1 si RNA transfection for 24 h followed by Bor intervention).The cell viability of each group were detected by CCK-8 method,the apoptosis was detected by Hoechest 33342 staining,and the expressions of MAPK/ERK pathway-related proteins were detected by Western Blot.Results:Compared with normal subjects,RACK1 m RNA expression was significantly increased in the plasma of MM patients and human MM cell lines(P<0.05).Bor significantly reduced the viability of MM cells at 12 h,24 h and 48 h(P<0.05).Compared with the control group,the viability in the Bor group and the Bor+siRACK1 group were significantly lower than that of the Bor group(P<0.05).Hoechest 33342 staining showed that the control group had uniform nuclear staining,and no apoptotic bodies was found,while a small number of apoptotic bodies were found in the Bor group,a large number of apoptotic bodies were observed in the Bor+siRACK1 group,which showed nuclear pyknosis or fragmentation.Compared with the control group,the apoptotic rate of MM cells in the Bor group and the Bor+siRACK1 group were significantly increased(P<0.05),and the apoptotic rate of MM cells in the Bor+siRACK1 group was significantly higher than that in the Bor group(P<0.05).Compared with the control group,the expressions of p-P38 and p-ERK were significantly decreased in the Bor group and the Bor+siRACK1 group,and the expression of p-P38 and p-ERK in the Bor+siRACK1 group was lower than the Bor group(P<0.05).There were no significant difference in the expressions of P38 and ERK among the 3 groups(P>0.05).Conclusions:RACK1 silencing enhances Bor-induced apoptosis and growth inhibition of MM cells,and its mechanism may be related to the inhibition of MAPK/ERK pathway.