3T3-L1细胞与人骨髓间充质干细胞体外成脂及脂肪细胞功能的比较研究

Comparison of Adipogenesis and Adipocyte Functions of 3T3-L1 Cells and Human Bone Marrow Mesenchymal Stem Cells In Vitro

目的:探讨3T3-L1细胞与人骨髓间充质干细胞(mesenchymal stem cells,MSCs)体外成脂及脂肪细胞功能的差异。方法:用密度梯度离心和贴壁培养的方法分离纯化人骨髓MSC,在倒置显微镜下观察细胞形态,体外向脂肪方向诱导分化,并通过油红O染色和吸光度值(OD值)检测其分化程度;qRT-PCR检测过氧化物酶体增殖物激活受体γ(PPARγ)、脂肪酸结合蛋白(FABP4)、CCAAT增强子结合蛋白(C/EBPα)mRNA表达水平;通过诱导的脂肪细胞与THP-1细胞共培养,加入1μg/ml的阿糖胞苷,在48 h测定其对肿瘤细胞的化疗抵抗作用。结果:通过油红O染色和测定吸光度值发现,3T3-L1细胞组脂质聚集量多于MSC组,且OD值大于MSC组(P<0.05);qRTPCR结果显示,3T3-L1来源的脂肪细胞的成脂基因PPARγ、FABP4和C/EBPαmRNA的相对表达量高于人骨髓MSC的脂肪细胞(P<0.05);共培养实验结果表明,含脂肪细胞组THP-1细胞存活数较对照组多(P<0.05),且3T3-L1细胞组与MSC组之间具有统计学差异(P<0.05)。结论:3T3-L1细胞的体外成脂能力比人骨髓MSC强;脂肪细胞具有促使THP-1细胞耐受化疗的作用,且在3T3-L1细胞组的作用大于人骨髓MSC组。

Objective:To compare the adipogenesis and the adipocyte function between 3T3-L1 cells and human bone marrow mesenchymal stem cells(MSCs) in vitro.Methods:By density gradient centrifugation and adherent culture,the MSCs were isolated from human bone marrow and purified.The cell morphology was observed under an inverted microscope.After the induction of adipogenic differentiation,the differentiation level was detected by oil red 0staining and OD values.The expression of PPARγ,FABP4 and C/EBPα mRNA was detected by quantitative reverse transcriptase-polymerase chain reaction(qRT-PCR).Adipocytes and THP-1 cells were co-cultured by adding 1 μg/ml cytarabine.The ability of chemotherapy resistance was measured after 48 h.Results:The Oil Red 0 staining and measuring the absorbance showed that the lipid content in 3T3-L1 cells group was more than that in MSCs group,and the OD value was higher than that in MSCs group(P<0.05).The results of qRT-PCR showed that the relative expression of PPARγ,FABP4 and C/EBPα mRNA of 3T3-L1-derived adipocytes was higher than that of human bone marrow MSCs-derived adipocytes(P <0.05).Coculture experiments showed that the number of viable THP-1 cells in the group containing adipocytes was more than that in the control group(P <0.05).The difference between 3T3-L1 cell group and MSC group was statistically significant(P<0.05).Conclusion:The ability of adipogenesis of 3T3-L1 cells is higher than that of human bone marrow MSCs in vitro.Adipocytes can protect THP-1 cell line against cytarabine,and the effect of adipocytes from 3T3-L1 cell group is greater than that from the human bone marrow MSC group.