小鼠3T3-L1前脂肪细胞系的增强绿色荧光蛋白标记

细胞模型是研究细胞分化原理以及进行高通量筛选的有效工具。为了建立特异性标记的脂肪细胞分化模型，构建了包括脂肪细胞分化特异性表达基因PPARγ2的启动子在内的载体（pPPARγ2-promoter-EGFP），用电穿孔方法转染小鼠3T3L1 前脂肪细胞，用显微荧光观察和RT-PCR确认PPARγ2基因的内源表达。结果显示，EGFP基因成功转入3T3-L1前脂肪细胞，观察到细胞分化过程中EGFP表达和脂肪积累，RTPCR分析表明EGFP代表了稳定而真实的PPARγ2基因的内源性表达。建立了由脂肪组织特异表达基因PPARγ2的表达控制的EGFP标记的小鼠3T3-L1前脂肪细胞系，目前国内外尚未见用同样方法对前脂肪细胞进行特异性标记。该细胞系将为脂肪细胞分化机理研究以及为抗肥胖症和抗糖尿病药物筛选提供有力工具。

The Labeling of 3T3-L1 Preadipocyte Cells with Enhanced Green Fluorescent Protein

A cell model is desired for adipocyte differentiation investigation and for high-throughput screening of anti-obesity and anti-diabetes molecules from chemical resources due to the world wide epidemic of obesity and diabetes. In order to establish such a cell model, a plasmid of pPPARgamma2-promoter-EGFP was constructed by inserting a 660bp sequence of mouse PPARgamma2 promoter into the Ase I and Kpn I sites of pEGFP-N3 and transferred into 3T3-L1 preadipocyte cells. The cells were induced to differentiate and the expression of PPARgamma2 was detected by the microscopic observation of EGFP and by RT-PCR assays. The results showed that the EGFP gene expression patterns were similar to that of pPPARgamma2's, which indicated that the EGFP gene was transferred into the mouse 3T3-L1 preadipocyte cells, and its expression was under the control of pPPARgamma2 promoter. RT-PCR assays showed that the EGFP expression authentically represented the stable expression of PPARgamma2. In conclusion, a preadipocyte cell line expressing EGFP under the control of the promoter of adipocyte-specific expression gene PPARgamma2 was generated. The cell line provides a powerful approach for the research of adipocyte differentiation and for the high-throughput screening of anti-obesity and anti-diabetes chemicals.