rhMG53作为低温保护剂对小鼠BMSCs生物学特性的影响
Effects of rhMG53 as a special cryoprotectant on biological characteristics of mouse bone marrow mesenchymal stem cells

目的:探讨重组人MG53蛋白(rhMG53)作为干细胞低温保护剂的特殊组分对小鼠骨髓间充质干细胞(BMSCs)冷冻保存后生物学特性的影响。方法:常规培养BMSCs,分为对照组(DMEM/F12培养基、FBS、DMSO的体积比为5:4:1并添加30 mg/L BSA)和rhMG53组(DMEM/F12培养基、FBS、DMSO的体积比为5:4:1并添加30 mg/L rhMG53)冻存。冻存1 a后,复苏细胞,qRT-PCR检测两组细胞冻存复苏后干性相关基因Nanog、OCT4和SOX2 mRNA的表达; CCK-8法检测细胞增殖; 台盼蓝染色检测细胞存活能力; 茜素红S染色和油红O染色分别检测两组干细胞的成骨和成脂分化; 免疫荧光检测两组细胞成神经分化相关因子DCX和NSE的表达。结果:与对照组相比,rhMG53组BMSCs的Nanog、OCT4和SOX2 mRNA表达无明显变化(P>0.05),但细胞增殖能力增强,存活率升高,成骨、成脂和成神经分化效率增加(P均<0.05)。结论:rhMG53作为保护剂的特殊组分对冷冻保存的BMSCs有较好的保护效果。
Aim:To observe the effects of recombinant human MG53 protein(rhMG53),as a special component of the low temperature protective agent of stem cells, on the biological characteristics of mouse bone marrow mesenchymal stem cells(BMSCs).Methods:BMSCs were expanded and defined as control group(VDMEM/F12:VFBS:VDMSO=5:4:1,added 30 mg/L BSA)and rhMG53 group(VDMEM/F12:VFBS:VDMSO=5:4:1, added 30 mg/L rhMG53).After one year of cryopreservation,BMSCs were resuscitated.The expressions of stemness related genes Nanog,OCT4 and SOX2 were detected by qRT-PCR; CCK-8 was used to detect cell proliferation; Trypan blue staining was used to detect cell viability.Alizarin red S staining and oil red O staining were used to detect the osteogenic and adipogenic differentiation of the stem cells.Immunofluorescence was used to detect the expressions of DCX and NSE genes related to neural differentiation.Results: Compared with control group,there were no differences in the expressions of the stemness related genes in the rhMG53 group(P>0.05),while the proliferation ability of BMSCs was enhanced,the cells survival rate was increased,and the efficiency of osteogenesis, adipogenesis and neurogenic differentiation was promoted(P<0.05).Conclusion:rhMG53,as a specific components of low temperature protective agent,does not affect the stemness of BMSCs,and could improve the proliferation and survival abilities,as well as osteogenic,adipogenic,and neural differentiation efficiency of the stem cells after cryopreservation and resuscitation