Ezrin在多发性骨髓瘤侧群细胞的表达及意义

摘要

Objective To determine the expression level of Ezrin protein in multiple myeloma side population cells ( SP cells) , and to detect the Ezrin protein expression and evaluate its impli-cations in multiple myeloma. Methods Bone marrow mononuclear cells of multiple myeloma (MM) and RPMI 8226 cell line were extracted, and incubated through the Hoechst 33342 dye. CD138 ( +) cells were selected and collected from NSP cells. Cells negative for Hoechst 33342 were included in the SP gate while cells positive for Hoechst 33342 were collected in the NSP gate by flow cytometry. Cells were pre-incubated with anti-human CD138 antibodies and then CD138 ( +) cells were isolated from NSP cell fraction. Differential proteins of SP and NSP cells were ana-lyzed by LC-MS /MS, and were identified by Sequest software. The expression of Ezrin protein was detected, and then the expression of Ezrin protein was detected by Western blotting. Results Pro-tein expression ratios were believed to be different if the differences were more than 1. 5 folds be-tween the two groups. LC-MS/MS identification exhibited that Ezrin protein expression ratio were o-ver 1. 5 folds and Western blotting also showed that Ezrin expression was higher in the multiple mye-loma SP groups. Conclusion Side population cells with stem cell properties, might be associated with tumor drug resistance and relapse in tumor patients. Ezrin protein is highly expressed in SP of multiple myeloma RPMI 8226 cell lines and primary cells, and such over-expression might be of significance in the development of drug resistance and relapse of multiple myeloma.%目的 检测Ezrin在多发性骨髓瘤侧群中的表达水平, 探讨Ezrin在多发性骨髓瘤中的表达情况及其意义.方法 从RPMI 8226 细胞系及多发性骨髓瘤 ( multiple myeloma, MM) 骨髓提取单个核细胞,通过Hoechst 33342染料进行孵育, 应用流式细胞术得到的Hoechst 33342阴性的细胞为侧群 ( side popula-tion, SP) 细胞, 阳性的细胞为非侧群 ( non-side population NSP) 细胞.从NSP细胞中分选CD138阳性细胞. SP和NSP细胞的, 检测Ezrin蛋白的表达情况, 并用Western印迹验证Ezrin蛋白表达.结果 蛋白质谱鉴定筛选的蛋白中, 以两组表达变化超过1. 5倍作为差异存在. Ezrin蛋白在RPMI 8226细胞系及多发性骨髓瘤原代SP与NSP细胞的比值为4, Western印迹实验结果也表明zrin在SP组较NSP细胞组表达升高.结论 SP细胞群为富含具有干细胞特征的肿瘤起始细胞, 可能与临床上肿瘤患者的耐药和复发相关. Ezrin蛋白在多发性骨髓瘤RPMI 8226细胞系及原代细胞的SP中均具有高表达, 初步探讨其在多发性骨髓瘤复发、耐药中的意义.