CAJ | 학술논문

目的利用RNA干扰技术(RNAi)研究体外合成的siRNA分子对高尔基体膜上钙离子ATP酶1(plasma membrane-related Ca~(2+)ATPase-1,PMR1)的基因沉默作用,并探讨沉默后对胰岛细胞分泌胰岛素的影响.方法委托公司合成抗PMR1基因的siPMR1,用脂质体方法介导siPMR1转染胰岛NIT-1细胞,采用RT-PCR法观察siPMR1转染前后胰岛细胞中PMR1 mRNA的表达变化,并用放射免疫分析法测定胰岛细胞分泌的胰岛素浓度.结果 siPMR1转染胰岛NIT-1细胞后能明显抑制PMR1 mRNA表达,siPMR1转染的胰岛NIT-1细胞能显著增加胰岛素分泌量.结论 siPMR1 可明显沉默PMR1基因表达,并促进胰岛细胞分泌胰岛素,为用RNAi技术沉默PMR1靶点治疗糖尿病提供了理论基础.
목적 이용RNA간우기술(RNAi)연구체외합성적siRNA분자대고이기체막상개리자ATP매1(plasma membrane-related Ca~(2+)ATPase-1,PMR1)적기인침묵작용,병탐토침묵후대이도세포분비이도소적영향.방법 위탁공사합성항PMR1기인적siPMR1,용지질체방법개도siPMR1전염이도NIT-1세포,채용RT-PCR법관찰siPMR1전염전후이도세포중PMR1 mRNA적표체변화,병용방사면역분석법측정이도세포분비적이도소농도.결과 siPMR1전염이도NIT-1세포후능명현억제PMR1 mRNA표체,siPMR1전염적이도NIT-1세포능현저증가이도소분비량.결론 siPMR1 가명현침묵PMR1기인표체,병촉진이도세포분비이도소,위용RNAi기술침묵PMR1파점치료당뇨병제공료이론기출.
Objective To assess the effect of RNA interference-mediated gene silencing of plasma membrane-related Ca~(2+)-ATPase-1 (PMR1) gene on the insulin secretion in islet β cells NIT-1 in vitro. Methods A small interfering RNA duplex (siPMR1) corresponding to the nucleotides 337-357 of mouse PMR1 cDNA was introduced into NIT-1 cells via liposomes.The gene silencing effect was assessed by RT-PCR, and the total insulin level in the transfected cells was measured by radioimmunoassay. Results Transfection with siPMRl resulted in obviously reduced PMR1 expression and increased insulin secretion in NIT-1 cells. Conclusion The synthesized siPMRl can significantly silence the expression of PMR1 and promote the secretion of insulin in the islet cells in vitro, which shed light on further studies of RNAi-based therapy of diabetes.