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目的比较羧基荧光素乙酰乙酸(CFSE)与四甲基偶氮唑盐(MTT)比色法检测人脐血间充质干细胞(CB-MSCs)和神经分化的CB-MSCs对大鼠脾淋巴细胞(LCs)刺激作用的敏感程度。方法分别制备刺激细胞和LCs,将刺激细胞分为4组:1.CB-MSCs组;2.Dif-CB-HSCs组:脐血间充质干细胞培养7d后维甲酸(RA)处理4d诱导其神经分化细胞;3.SH-SY5Y组:人源SH-SY5Y细胞(人神经母细胞瘤细胞系)作为阳性对照组;4.Auto-LC组:自体大鼠LCs作为阴性对照组。分别将上述各组刺激细胞与LCs进行混合培养,分别通过酶标仪(MTT法),流式细胞术(CFSE法)检测LCs增殖情况(n=3)。结果CFSE法和MTT法检测结果均显示,CB-MSCs组和Dif-CB-MSCs组刺激LCs增殖明显弱于阳性对照组(SH-SY5Y细胞组),但CFSE法检测结果显示,CB-MSCs组和Dif-CB-MSCs组细胞刺激LCs增殖百分率高于阴性对照Auto-LC组,MTT法检测结果显示,CB-MSCs组和Dif-CB-MSCs组的吸光度(A)值稍低于阴性对照Auto-LC组。结论CFSE法比MTT法能更加客观地反映淋巴细胞增殖情况,在实际应用中更具有优势。
Objective To compare the effects of CB-MSCs and CB-MSCs differentiated by CFSE and MTT on the proliferation of rat splenic lymphocytes (LCs) to stimulate the degree of sensitivity. Methods Stimulating cells and LCs were prepared and divided into 4 groups: 1.CB-MSCs group; 2.Dif-CB-HSCs group: After treated with retinoic acid (RA) for 4 days, Neural differentiation cells; 3.SH-SY5Y group: human SH-SY5Y cells (human neuroblastoma cell line) as a positive control group; 4. Auto-LC group: autologous rat LCs as a negative control group. The stimulating cells in each group were mixed with LCs respectively, and the proliferation of LCs was detected by MTT assay and flow cytometry (CFSE) respectively (n = 3). Results The results of CFSE assay and MTT assay showed that the proliferation of LCs in CB-MSCs group and Dif-CB-MSCs group was significantly weaker than that in positive control group (SH-SY5Y cell group). However, the results of CFSE assay showed that CB- LC-MSCs group and Dif-CB-MSCs group were higher than the negative control Auto-LC group. MTT assay showed that the absorbance (A) value of CB-MSCs group and Dif- -LC group. Conclusion The CFSE method can reflect the lymphocyte proliferation more objectively than the MTT method and has more advantages in practical application.