CAJ | 학술논문

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.
세포모형시연구세포분화원리이급진행고통량사선적유효공구.위료건립특이성표기적지방세포분화모형,구건료포괄지방세포분화특이성표체기인PPARγ2적계동자재내적재체(pPPARγ2-promoter-EGFP),용전천공방법전염소서3T3-L1 전지방세포,용현미형광관찰화RT-PCR학인PPARγ2기인적내원표체.결과현시,EGFP기인성공전입3T3-L1전지방세포,관찰도세포분화과정중EGFP표체화지방적루,RT-PCR분석표명EGFP대표료은정이진실적PPARγ2기인적내원성표체.건립료유지방조직특이표체기인PPARγ2적표체공제적EGFP표기적소서3T3-L1전지방세포계,목전국내외상미견용동양방법대전지방세포진행특이성표기.해세포계장위지방세포분화궤리연구이급위항비반증화항당뇨병약물사선제공유력공구.
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 pPPARγ2-promoter-EGFP was constructed by inserting a 660bp sequence of mouse PPARγ2 promoter into the AseⅠ and KpnⅠ sites of pEGFP-N3 and transferred into 3T3-L1 preadipocyte cells. The cells were induced to differentiate and the expression of PPARγ2 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 pPPARγ2's, which indicated that the EGFP gene was transferred into the mouse 3T3-L1 preadipocyte cells, and its expression was under the control of pPPARγ2 promoter. RT-PCR assays showed that the EGFP expression authentically represented the stable expression of PPARγ2. In conclusion, a preadipocyte cell line expressing EGFP under the control of the promoter of adipocyte-specific expression gene PPARγ2 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.