小分子RNA干扰组织因子在新生猪胰岛细胞中的表达  

siRNA-mediated tissue factor knockdown in porcine neonatal islet cell clusters in vitro

作　　者：暨明[1] 易受南[2,3] 于德玲[2] 王维[2] 

机构地区：[1]中南大学基础医学院生理学系,长沙410078 [2]中南大学湘雅三医院细胞移植与基因治疗中心,长沙410013 [3]澳大利亚悉尼大学Westmead医院移植与肾病研究中心

出　　处：《中南大学学报（医学版）》2011年第12期1141-1146,共6页Journal of Central South University ：Medical Science

基　　金：国家自然科学基金(30900359)~~

摘　　要：目的:利用小分子RNA(siRNA)在基因水平对新生猪胰岛细胞进行修饰,抑制细胞组织因子的表达。方法:设计5对siRNA转染新生猪胰岛细胞,用real-time PCR方法筛选组织因子基因沉默效果最好的siRNA或组合,同时流式细胞仪检测siRNA转染对细胞活性的影响,real-time PCR和流式细胞仪分别检测细胞组织因子的基因及蛋白沉默水平。结果:根据real-time PCR结果筛选出组织因子基因沉默效果最好的3对siRNA组合,转染新生猪胰岛细胞后,real-time PCR及流式细胞仪检测新生猪胰岛细胞组织因子的基因沉默效率达60%,蛋白表达水平降低约50%,同时流式细胞仪检测结果提示siRNA转染对新生猪胰岛细胞的活性没有明显的影响。结论:3对siRNA组合在体外特异性抑制新生猪胰岛细胞组织因子的表达。Objective To determine the genetic modification on neonatal porcine islet cell clusters（NICC） by small interfering RNA（siRNA）-mediated tissue factor（TF） knockdown in vitro.Methods Porcine NICC were transfected with 5 pairs of designed siRNA respectively or in different combinations with lipofectamine 2000.Transfected NICC were analyzed for TF gene by real-time PCR to select the siRNA which worked best.Meanwhile,the viability of NICC after the TF siRNA transfection was examined by FACS.The efficiency of TF gene and protein suppression was measured by real-time PCR and and FACS respectively.Results Real-time PCR and FACS showed that a 60% reduction in the TF gene expression and a 50% reduction in the protien level of TF on NICC were achieved by transfecting 3 pairs of selected siRNA.The siRNA transfection had no significant effect on the viability of NICC which was analyzed by FACS.Conclusion The expression of TF on porcine NICC is efficiently suppressed by 3 pairs of designed siRNA in vitro.

关 键 词：SIRNA 组织因子 新生猪胰岛细胞 

分 类 号：R363[医药卫生—病理学]