脂肪因子chemerin对3T3-L1前脂肪细胞增殖的影响  

Effects of chemerin on the proliferation of 3T3-L1 adipocytes

作　　者：李雪梅[1] 翟丽东[2] 王鹏华[1] 丁敏[1] 徐俊[1] 冯书红[1] 

机构地区：[1]天津医科大学代谢病医院内分泌研究所卫生部激素与发育重点实验室,300070 [2]天津医科大学基础医学院人体解剖和组织胚胎系

出　　处：《中华医学杂志》2015年第26期2099-2103,共5页National Medical Journal of China

摘　　要：目的 探讨脂肪因子chemerin对3T3-L1前脂肪细胞增殖的影响及可能机制.方法 制备小鼠chemerin过表达及RNA干扰重组慢病毒,感染3T3-L1细胞.实验设chemerin基因沉默组、沉默空载体对照组、过表达组、过表达空载体对照组.另在细胞增殖实验中增设细胞外信号调节激酶（ERK）阻断剂组.应用四甲基偶氮唑蓝（MTT）法联合5-溴脱氧尿嘧啶核苷（BrdU）掺入法观察chemerin对3T3-L1细胞增殖的影响;实时定量PCR检测chemerin、ERK1/2及细胞周期蛋白（cyclin）D1基因mRNA表达;Western印迹法检测chemerin、ERK1/2及磷酸化ERK1/2（p-ERK1/2）蛋白水平.结果 MTT及BrdU掺入实验结果显示：与过表达对照组相比chemerin过表达后吸光度（A）值升高（1.09±0.08比0.90±0.09,P＜0.05）,BrdU掺入量增加[（126.5±14.6）％比（100.0±7.0）％,P＜0.05],此效应可被ERK1/2信号通路阻滞剂PD98059抑制,A值（0.91±0.13）及BrdU掺入量（104.3±10.7）％降低,与过表达组相比差异均有统计学意义（均P ＜0.05）;与沉默对照组相比,chemerin基因沉默后A值降低（0.72 ±0.11比0.90 ±0.09,P＜0.05）,BrdU掺入量减少[（77.6±11.8）％比（99.7±6.3）％,P＜0.05];实时定量PCR结果显示：chemerin过表达后与其空载对照组相比chemerin、ERK1/2、cyclinD1表达上调（2.77 ±0.31比1.01 ±0.12,1.39±0.19比0.76±0.30,1.46±0.14比0.88±0.14,1.44±0.17比1.03±0.15,均P＜0.05）;chemerin基因沉默后均表达下调（0.35±0.12比1.25±0.31,0.64±0.15比1.03±0.14,0.56±0.10比1.06±0.29,0.66 ±0.13比1.09±0.19,均P＜0.05）;Western印迹结果显示：chemerin过表达后与其空载对照组相比chemerin、ERK1/2及p-ERK1/2蛋白水平增加（2.18±0.32比1.18±0.14,1.37±0.05比0.97±0.12,1.06±0.09比0.56 ±0.08,均P＜0.05）,chemerin基因沉默后与其对照组相比chemerin、ERK1/2及p-ERK1/2蛋白水平降低（1.00±0.07比1.40±0.17,0.87±0.15比1.20 ±0.12,0.49±0.07比0.91±0.11,均P＜0.05）.结论 chemerin可Objective To explore the effects of chemerin on the proliferation of 3T3-L1 preadipocytes and elucidate its possible mechanism.Methods Recombinant lentivirus-mediated silencing or over-expression of chemerin gene were constructed in 3T3-L1 cells.The proliferation of 3T3-L1 cells was examined by the assays of methyl thiazolyl tetrazolium （MTT） and 5-bromo-2-deoxyuridine （BrdU） incorporation.The expressions of chemerin,ERK1,ERK2 and cyclinD1 in 3T3-L1 cells were detected by real-time reverse transcriptase-polymerase chain reaction （RT-PCR）.The protein levels of chemerin,ERK1/2 and p-ERK1/2 were detected by Western blot.Results MTT and BrdU results showed that absorbance （A） value and BrdU incorporation increased in chemerin over-expression group compared with over-expression control group [1.09 ± 0.08 vs 0.90 ± 0.09,（126.5 ± 14.6） % vs （100.0 ± 7.0） %,both P 〈 0.05].The increase of A value and BrdU incorporation in chemerin over-expression group could be inhibited by PD98059,a blocker of ERK1/2 signal pathway.A value （0.91 ± 0.13） and BrdU incorporation [（104.3 ± 10.7） %] decreased compared with over-expression group （P 〈 0.05）.They also decreased in chemerin gene silencing group compared to silencing control group [0.72 ± 0.11 vs 0.90 ± 0.09,（77.6 ± 11.8） % vs （99.7 ± 6.3） %,both P 〈 0.05].Quantitative real-time PCR revealed that over-expression group had a higher expression in chemerin,ERK1,ERK2 and cyclinD1 than over-expression control group （2.77 ±0.31 vs 1.01 ±0.12,1.39 ±0.19 vs 0.76 ±0.30,1.46 ±0.14 vs 0.88 ±0.14,1.44 ± 0.17 vs 1.03 ± 0.15,all P 〈 0.05）.Chemerin gene silencing group had lower expressions of chemerin,ERK1,ERK2 and cyclinD1 than silencing control group （0.35 ±0.12 vs 1.25 ±0.31,0.64 ± 0.15 vs 1.03±0.14,0.56±0.10vs 1.06±0.29,0.66±0.13 vs 1.09±0.19,allP〈0.05）.Western blot showed that the expressiond of chemerin,ERK1/2 and p-ERK1/2 increased in chemerin over-expression group versus over-expression control group

关 键 词：脂肪因子类 3T3-L1细胞 细胞增殖 慢病毒属 丝裂原活化蛋白激酶1 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]