Slfn1通过下调Cyclin D1抑制内皮祖细胞的黏附功能

Slfn1 Reduces the Adhension of Endothelial Progenitor Cells by Decreasing Cyclin D1

目的探讨睡眠因子1(Slfn1)对内皮祖细胞(EPC)黏附功能的影响。方法实验分为空白对照组(未给予任何处理的EPC,N-control组)、腺病毒阴性对照组(Ad-control组)、Slfn1腺病毒载体组(Ad-Slfn1组)、发夹状RNA阴性对照组(Sh RNA-control组)、发夹状RNA Slfn1干扰组(Sh RNA-Slfn1组)。分离培养大鼠骨髓源性EPC,用Sh RNA-control质粒、Ad-Slfn1及相应的对照质粒分别转染EPC,采用Western blot检测细胞Slfn1及Cyclin D1表达,将EPC接种在预先铺好纤维连接蛋白的培养板中,观察其黏附功能。用流式细胞仪检测细胞周期。结果Sh RNA-Slfn1组Slfn1蛋白的表达明显低于Sh RNA-control组,Ad-Slfn1组Slfn1蛋白的表达明显高于Ad-control组,说明转染有效。用Sh RNA-Slfn1减弱EPC Slfn1基因的表达明显增强EPC的黏附能力,过表达Slfn1基因则明显抑制EPC的黏附能力。细胞周期检查结果显示,降低EPC Slfn1基因的表达,EPC的细胞周期进入到S期细胞明显增多,过表达Slfn1基因使EPC的细胞周期停止在G1期。Western blot检测结果发现,Sh RNA-Slfn1组Cyclin D1的蛋白表达明显高于Sh RNA-control组,而Ad-Slfn1组Cyclin D1的蛋白表达则明显低于Ad-control组。结论 Slfn1通过下游靶点Cyclin D1负性调控EPC的黏附功能。

Ahn To evaluate the effect of Schlafen 1 （Slfnl） on the adhension of endothelial progenitor cells （EPC）. Methods Ad-Slfnl, ShRNA-Slfnl, ShRNA-control and Ad-control were transfected into EPC respectively. Then EPC were plated on fibronectin-coated culture dishes, adherent cells were counted. The protein of Slfnl and Cyclin D1 were examined by Western blot. EPC cell cycle was examined by flow cytometry analysis. Results On 48 h after transfected ShRNA-Slfnl, the expression of Slfnl protein was decreased significantly compared to that in ShRNA-control group （P 〈 0. 05）, the transfection of Ad-Slfnl reversed these responses. Overexpression of Slfnl suppressed the adhens- ion of EPC ; conversely, the silencing of Slfnl using shRNA-Slfnl increased the adhension of EPC. In addition, cell cycle was arrested G1 phase in Ad-Slfnl group. Whereas the transfection of shRNA-Slfnl reversed these responses. The expression of Cyclin D1 protein after transfection of shRNA-Slfnl was increased clearly compared to that in ShRNA-control group （P 〈 0. 05 ）, in contrast, overexpression of Slfnl reversed these results. Cyclin D1 was involved in Slfnl-mediated EPC adhension. Conclusion Slfnl reduced the adhension of EPC through Cyclin D1.