Prevention of renal ischemia-reperfusion injury by short hairpin RNA of endothelin A receptor in a rat model

Abstract

Endothelin A receptor (ETaR) is a key molecule involved in a variety of biological events such as vessel contraction and inflammatory response in ischemia-reperfusion (I/R) injury. RNA interference using short hairpin RNA (shRNA) is a powerful tool to silence gene expression. Here, the effect of ETaR shRNA on I/R injury in rats was studied. A more effective shRNA sequence out of two constructed into plasmid vectors was selected using the A-10 cell line, and was then applied to a rat model. Twenty-eight male Sprague-Dawley rats were randomized into four groups: Sham, shRNA, vector and phosphate-buffered saline (PBS). Renal I/R injury was induced by clamping the left renal pedicle for one hour followed by reperfusion for 24 h. ETaR shRNA (100 μg) plasmid was administered by renal vein injection 48 h before clamping. The expression of both ETaR mRNA and protein was lowered by ETaR shRNA treatment compared with that in the vector and PBS groups; serum creatinine and blood urea nitrogen were significantly decreased; the semi-quantitative score of renal structural damage was improved; the mRNA level of endothelin 1 (ET-1), tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), macrophage inflammatory protein 2 (MIP-2) and monocyte chemoattractant protein 1 (MCP-1) was reduced, but nitric oxide (NO) production in kidney tissues was increased (P < 0.05). In conclusion, ETaR shRNA partially silenced ETaR expression in I/R injury kidneys, reduced the mRNA level of ET-1, inflammatory mediators including TNF-α, IL-6, MIP-2 and MCP-1, increased NO production, and ultimately improved renal function and structure.