X-H Qu1, K Zhang. 1. Department of Kidney Medicine, Affiliated Hospital of Jining Medical University, Jining, Shandong, China. jll3h7zdzr73l@sina.com.
Abstract
OBJECTIVE: Phosphatidylinositol 3-kinase/protein kinase B ((PI3K/AKT) signaling pathway plays a role in regulating cell survival and apoptosis. Phosphatase and tensin homologue deleted on chromosome ten (PTEN) can negatively regulate PI3K/AKT signaling pathway, while DJ-1 (Parkinson gene 7) can negatively regulate expression and function of PTEN. DJ-1-PTEN/PI3K/AKT signaling pathway plays a role in the regulation of ischemic reperfusion (IR) injury. Bioinformatics analysis showed that there was a targeted complementary binding site between microRNA-122 (miR-122) and 3'-UTR of DJ-1 mRNA. This study aimed to investigate the effects of miR-122 in regulating DJ-1-PTEN/PI3K/AKT signaling pathway and acute renal I-R injury. MATERIALS AND METHODS: Rat renal artery was clamped and restored after 30 min to establish renal IR injury model. Renal tissue samples were collected at 10 h and 20 h after operation. miR-122 and DJ-1 mRNA were detected with quantitative Real-time PCR (qRT-PCR). DJ-1 protein was tested by using Western blot. Blood urea nitrogen (BUN) and serum creatinine (SCr) were measured. Rat tubular epithelial cells, RRTEC, were cultured in vitro and divided into transfection (miR-NC group) and treatment group (miR-122 inhibitor group). IR treatment was performed after 72 h of transfection. DJ-1, PTEN, AKT, and phosphorylated AKT (p-AKT) were detected using Western blot. Cell apoptosis and reactive oxygen species (ROS) were measured with flow cytometry. RESULTS: Compared with Sham group, blood BUN and SCr contents significantly increased (p < 0.05), miR-122 level significantly elevated (p < 0.05), while DJ-1 mRNA and protein markedly declined (p < 0.05) in IR model rats. Compared with control group, I-R treatment significantly up-regulated miR-122 and PTEN expressions (p < 0.05), decreased DJ-1 and p-AKT levels (p < 0.05), and induced apoptosis and ROS production (p < 0.05) in RRTEC cells. Transfection with miR-122 inhibitor markedly up-regulated DJ-1 expression (p < 0.05), enhanced PTEN/PI3K/AKT pathway activity (p < 0.05), and reduced apoptosis and ROS production (p < 0.05). CONCLUSIONS: MiR-122 increased significantly, while DJ-1 declined significantly during renal I-R injury. Down-regulation of miR-122 markedly elevated DJ-1, enhanced PTEN/PI3K/AKT pathway activity, and inhibited apoptosis and ROS generation in rat renal tubular epithelial cells to alleviate IR injury.
OBJECTIVE: Phosphatidylinositol 3-kinase/protein kinase B ((PI3K/AKT) signaling pathway plays a role in regulating cell survival and apoptosis. Phosphatase and tensin homologue deleted on chromosome ten (PTEN) can negatively regulate PI3K/AKT signaling pathway, while DJ-1 (Parkinson gene 7) can negatively regulate expression and function of PTEN. DJ-1-PTEN/PI3K/AKT signaling pathway plays a role in the regulation of ischemic reperfusion (IR) injury. Bioinformatics analysis showed that there was a targeted complementary binding site between microRNA-122 (miR-122) and 3'-UTR of DJ-1 mRNA. This study aimed to investigate the effects of miR-122 in regulating DJ-1-PTEN/PI3K/AKT signaling pathway and acute renal I-R injury. MATERIALS AND METHODS:Rat renal artery was clamped and restored after 30 min to establish renal IR injury model. Renal tissue samples were collected at 10 h and 20 h after operation. miR-122 and DJ-1 mRNA were detected with quantitative Real-time PCR (qRT-PCR). DJ-1 protein was tested by using Western blot. Blood ureanitrogen (BUN) and serum creatinine (SCr) were measured. Rat tubular epithelial cells, RRTEC, were cultured in vitro and divided into transfection (miR-NC group) and treatment group (miR-122 inhibitor group). IR treatment was performed after 72 h of transfection. DJ-1, PTEN, AKT, and phosphorylated AKT (p-AKT) were detected using Western blot. Cell apoptosis and reactive oxygen species (ROS) were measured with flow cytometry. RESULTS: Compared with Sham group, blood BUN and SCr contents significantly increased (p < 0.05), miR-122 level significantly elevated (p < 0.05), while DJ-1 mRNA and protein markedly declined (p < 0.05) in IR model rats. Compared with control group, I-R treatment significantly up-regulated miR-122 and PTEN expressions (p < 0.05), decreased DJ-1 and p-AKT levels (p < 0.05), and induced apoptosis and ROS production (p < 0.05) in RRTEC cells. Transfection with miR-122 inhibitor markedly up-regulated DJ-1 expression (p < 0.05), enhanced PTEN/PI3K/AKT pathway activity (p < 0.05), and reduced apoptosis and ROS production (p < 0.05). CONCLUSIONS:MiR-122 increased significantly, while DJ-1 declined significantly during renal I-R injury. Down-regulation of miR-122 markedly elevated DJ-1, enhanced PTEN/PI3K/AKT pathway activity, and inhibited apoptosis and ROS generation in rat renal tubular epithelial cells to alleviate IR injury.