期刊论文

Protective mechanism of NHE-I-siRNA on human renal tubular epithelial cell from ischemic reperfusion injury induced by antimycin A

中文

中华肾脏病杂志

CHINESE JOURNAL OF NEPHROLOGY

1001-7097

2008

24

8

560-565

10.3321/j.issn:1001-7097.2008.08.015

国家重点基础研究发展计划(973计划)(2007CB507403);国家自然科学基金(30670893);解放军总医院苗圃基金(06MP09)

本校为其他机构

目的 通过构建钠-氢交换蛋白1(NHE-1)小干扰RNA(siRNA)抑制肾小管上皮细胞NHE-1的表达,探讨其对细胞缺血再灌注损伤的保护作用机制.方法 根据人NHE-1全长基因序列设计并合成NHE-1-siRNA,转染人肾小管上皮细胞系(HKC),以无关siRNA转染组作为对照.利用10 μmol/L抗霉素A诱导细胞来模拟缺血缺氧环境.用RT-PCR、Western印迹法检测NHE-1的表达.用BCECF/AM、Fluo-3/AM和SBFI-AM标记HKC,通过激光共聚焦显微镜分别检测其细胞内pHi、Ca~(2+)和Na~+浓度的变化.用Hoechst 33342染色技术及AnnexinV/PI染色结合流式细胞仪技术检测细胞凋亡情况.利用荧光探针JC-1检测细胞线粒体膜电位的变化.结果 特异性NHE-1-...

Objective To explore the mechanism of protecting cells from ischemic reperfusion injury by constructing specific small interference RNA （siRNA） to inhibit Na^＋-H^＋ exchanger-1 （NHE-1） expression in human renal tubular epithelial cell （HKC）. Methods The siRNA was designed and synthesized based on human NHE-1 complete sequence, and was transfected into HKC. The irrespective siRNA transfected group was used as control. The cells were treated with 10 μmol/L antimycin A to induce ischemia and anoxyaemia environment. NHE-1 expression was exa...