中文摘要：

目的 研究与糖尿病大血管病变有关的新蛋白FAM172A对人胚胎肾细胞（HEK293细胞）凋亡及增殖的影响.方法 以前期研究工作中所构建的pDrive-FAM172A质粒为模板,通过PCR方法扩增出人FAM172A基因编码框,将扩增产物亚克隆到PDC315真核表达载体,经酶切、测序鉴定,选择插入序列正确的PDC315-FAM172A质粒用脂质体2000转染HEK293细胞,同时用PDC315空质粒作为对照转染HEK293细胞.用噻唑蓝（XTT）法、生长曲线法观察过表达FAM172A基因对HEK293细胞增殖的影响.用碘化丙啶（PI）单染色法、膜联蛋白V/PI双染色法观察过表达FAM172A基因对HEK293细胞凋亡及细胞周期的影响.结果 成功构建PDC315-FAM172A真核表达载体,经酶切、测序证实插入序列正确.与PDC315质粒转染相比,XTT显示PDC315-FAM 172A质粒转染使细胞增殖增加约52%（A值为0.21±0.07比0.32±0.06,P〈0.01）;生长曲线显示PDC315-FAM172A质粒转染使HEK293细胞生长速度增快;PI单染色法显示PDC315-FAM172A质粒转染使细胞凋亡减少约38.5%（分别23.79%±1.36%比14.64%±0.95%,P〈0.01）,同时G0～G1期细胞明显减少（分别66.79%±1.73%比58.16%±0.75%,P〈0.01）而S期细胞明显增加（分别22.62%±1.16%比33.56%±0.94%,P〈0.01）;膜联蛋白V/PI双染色法显示PDC315-FAM172A质粒转染使早期凋亡细胞减少约28%（13.63%±0.56%比9.79%±0.39%,P〈0.01）,晚期凋亡细胞减少约29%（7.70%±0.29%比5.43%±0.29%,P〈0.01）.结论 FAM172A蛋白促进HEK293细胞增殖、抑制凋亡、促使细胞进入S期,提示FAM172A蛋白的生理功能之一是参与细胞生长的调控,这为深入研究FAM172A蛋白的生理功能及在糖尿病大血管并发症中的作用提供了线索.

英文摘要：

Objective To study the effect of FAM172A protein related to diabetic macroangiopathy on apoptosis and proliferation in HEK293 cells.Methods The pDrive-FAM172A plasmid constructed in our previous study was used as a template to amplify human FAM172A open reading frame by a polymerase chain reaction.The resulting PCR products were subcloned into the eukaryotic expression vector PDC315 to construct recombinant PDC315-FAM172A plasmid.PDC315-FAM172A plasmid was identified by enzyme cleavage and sequencing analysis.HEK293 cells were transiently transfected respectively with appropriate PDC315 or PDC315-FAM172A plasmid by Lipofectamine 2000 according to the manufacturer＇s instruction.XTT assay and growth curve were used to observe the effect of over-expression of FAM172A gene on HEK293cell proliferation.PI and Annexin V/PI staining method were used to assess the effect of FAM172A gene on apoptosis and cell cycle of HEK293 cell.Results Eukaryotic expression vector PDC315-FAM172A was successfully constructed and identified by enzyme cleavage and sequencing analysis.Compared with PDC315plasmid transfection,the XTT assay showed that optical density （A） value increased by 52% when transfected with PDC315-FAM172A plasmid （0.21±0.07 vs 0.32±0.06,P 〈 0.01）.Growth curve revealed that HEK293 cells transfected with PDC315-FAM172A plasmid proliferated faster than those transfected with PDC315 plasmid.PI staining showed that,as compared with PDC315 plasmid transfection,the apoptotic rate of HEK293 cells transfected with PDC315-FAM172A plasmid decreased by 38.5%（23.79±1.36 vs 14.64±0.95,P 〈 0.01）,cell percentage of G0-G1 phases significantly decreased （66.79±1.73 vs 58.16 ±0.75,P 〈0.01） and cell percentage of S phases significantly increased （22.62 ±1.16 vs33.56 ±0.94,P 〈0.01）.Annexin V/PI staining revealed that,as compared with PDC315 plasmid transfection,the percentage of early and advanced apoptotic cells decreased by 28% （13.63 ±0.56 vs 9.79 ±0.39,P 〈0.01） and 29% （7.70±0.29 vs 5.4