【摘要】 目的:构建骨髓间充质干细胞(BMMSC)与多发性骨髓瘤(MM)细胞共培养体系,探讨共培养后BMMSC对M M细胞迁移和归巢的影响。方法:采用贴壁筛选法培养绿色荧光蛋白(GFP)小鼠BM M SC,建立M M细胞株XG-7细胞和BMMSC间接及直接共培养体系;用CD138磁珠法分离直接共培养的XG-7细胞及BMMSC,用台盼蓝染色计数法检测XG-7细胞的增殖水平、Annexin V/PI测定细胞凋亡水平、单丹(磺)酰戊二胺(MDC)法检测细胞自噬泡数量、在共聚焦显微镜下动态观察BMMSC与PE-CD138标记的XG-7细胞共培养过程的分子流向和分布。结果:经共培养后,BMMSC促进XG-7细胞增殖;单独培养后凋亡率为(17.90±1.46)%,直接和间接共培养后凋亡率分别降至(6.23±0.12)%和(6.97±0.03)%(P<0.01);XG-7细胞单独培养及与BMMSC共培养后,荧光显微镜下均可出现自噬,共培养后的XG-7细胞自噬现象相对少见;在共聚焦荧光显微镜下XG-7细胞是以极化的胞膜区同BMMSC接触,加入甲基β环糊精后极化现象消失。结论:BMMSC可促进XG-7细胞的生长,且增强XG-7细胞抗凋亡及抗自噬能力,影响骨髓瘤细胞的迁移和归巢。更多还原

【Abstract】 Objective:To construct a co-culture system for bone marrow mesenchymal stem cells(BMMSC) and multiple myeloma(MM) cells,and to investigate the effects of co-cultured BMMSC on the migrating and homing of multiple myeloma cells.Methods:The BMMSC from the transgenic mice with green fluorescent protein(GFP) fetal bone were cultured by adherent screening.A co-culture system of BMMSC and MM cell line XG-7 cells was constracted,the proliferation and apoptosis of cells were determined by trypan blue exclusion and Annexin V/PI,respectively,MDC staining was employed to detect the autophagy.The moving direction distribution of molecule in BMMSC and XG-7 cells labeled with PE-CD138 in co-culture process were observed dinamically by confocal microscopy.Results:After co-culture with GFP-BMMSC,the resistance of XG-7 cells to apoptosis and autophagy were enhanced;at the same time,their proliferation increased.Apoptosis rates of XG-7 cells directly and indirectly co-cultured with BMMSC were(6.23 ±0.12)%and(6.97 ±0.03)%respectively,which were lower than that of XG-7 cells cultured alone(17.90 ± 1.46)%(P<0.01).There was low level of autophagy in XG-7 cells co-cultured with BMMSC.XG-7 cells are highly polarized and contained a specialized membrane domain with specific protein and lipid components to contact with BMMSC under confocal microscope.After methyl- β-cyclodextrin treatment,the molecules were normally enriched in the specialized domain.Conclusion:BMMSC can protect XG-7 cells from apoptosis and autophagy,and obviously promote the proliferation of XG-7 cells,and can influence the migrating and homing of multiple myeloma cells.更多还原