AimsTo investigate if nuclear NF-κB p65 expression in ex vivo isolated peripheral blood mononuclear cells correlates with urinary MCP-1 or RANTES and the severity of type 2 diabetic nephropathy.MethodsAccording to their urinary albumin-to-creatinine ratio (uACR), 107 patients with type 2 diabetes (eGFR >60 ml/min) were divided into normal albuminuria group (DN0 group, 38 cases), microalbuminuria group (DN1 group, 38 cases), and macroalbuminuria group (DN2 group, 31 cases), compared with matched healthy normal control group (NC group, 30 cases). Nuclear NF-κB p65 protein expression levels in peripheral blood mononuclear cells were detected by western blotting. Real-time quantitative polymerase chain reaction was used to detect NF-κB p65 mRNA expression and ELISA assay was used to detect the levels of urinary MCP-1 and RANTES.ResultsNuclear NF-κB p65 protein and NF-κB p65 mRNA expression levels in peripheral blood mononuclear cells, urinary MCP-1/Cr and RANTES/Cr were all significantly higher in all diabetes groups as compared with NC group. In particular, the increase of nuclear NF-κB p65 protein and NF-κB p65 mRNA expressions, urinary MCP-1/Cr and RANTES/Cr all correlated with the severity of type 2 diabetic nephropathy as indicated by the increase in uACR. Pearson correlation analysis indicated that both urinary MCP-1/Cr and RANTES/Cr were positively correlated with nuclear NF-κB p65 protein or NF-κB p65 mRNA levels. Stepwise multiple regression analysis showed that nuclear NF-κB p65 protein or NF-κB p65 mRNA was an independent variable for urinary MCP-1/Cr, and MCP-1/Cr and RANTES/Cr were two independent variables for uACR.ConclusionOur research demonstrates that nuclear NF-κB p65 protein and mRNA expressions in ex vivo isolated peripheral blood mononuclear cells well correlate with urinary MCP-1/Cr, RANTES/Cr and the severity of type 2 diabetic nephropathy.

Immunohistochemistry staining showed that Prx1 expressed in the cytoplasm of renal tubular epithelial cells, in the kidneys of UUO rats. The reduction was confirmed by both IHC and real-time polymerase chain reaction following a course of renal tubulointerstitial fibrosis in UUO rats and a decrease of Prx1 occurred concomitantly with an elevation of TUNEL-positive cells. Fluorofenidone (AKF-PD), a new anti-tubulointerstitial fibrotic agent, attenuated Prx1 reduction in UUO rats. Furthermore, hydrogen peroxide (H2 O2 )-derived oxidative stress activated p38 MAPK, and induced apoptosis in NRK-52E cells; knockdown of Prx1 sensitized both events in NRK-52E cells, and overexpression of Prx1 diminished the apoptosis and the phosphorylation of p38 CONCLUSION: Downregulation of Prx1 occurred in renal tubular epithelial cells of UUO rats and patients with obstructive nephropathy. Prx1 may alleviate the pathogenesis by inhibiting H2 O2 -induced apoptosis via inhibiting the p38 MAPK pathway. Prx1 may represent a useful target for a protective therapy towards renal tubulointerstitial fibrosis.

Tubular epithelial-mesenchymal transition (EMT) has been widely accepted as the underlying mechanisms of renal interstitial fibrosis (RIF). The production of reactive oxygen species (ROS) plays a vital role in tubular EMT process. The purpose of this study was to investigate the involved molecular mechanisms in TGF-beta-induced EMT and identify the potential role of glutathione S-transferase alpha 3 (GSTA3) in this process. The iTRAQ screening was performed to identify protein alterations of the rats underwent unilateral-ureteral obstruction (UUO). Protein expression of GSTA3 in patients with obstructive nephropathy and UUO rats was detected by immunohistochemistry. Protein and mRNA expression of GSTA3 in UUO rats and NRK-52E cells were determined by Western blot and RT-PCR. siRNA and overexpression plasmid were transfected specifically to assess the role of GSTA3 in RIF. The generation of ROS was measured by dichlorofluorescein fluorescence analysis. GSTA3 protein and mRNA expression was significantly reduced in UUO rats. Immunohistochemical analysis revealed that GSTA3 expression was reduced in renal cortex in UUO rats and patients with obstructive nephropathy. Treating with TGF-β1 down-regulated GSTA3 expression in NRK-52E cells, which have been found to be correlated with the decreased expression in E-cadherin and megalin and increased expression in α-smooth muscle actin. Furthermore, knocking down GSTA3 in NRK-52 cells led to increased production of ROS and tubular EMT, whereas overexpressing GSTA3 ameliorated ROS production and prevented the occurrence of tubular EMT. GSTA3 plays a protective role against tubular EMT in renal fibrosis, suggesting GSTA3 is a potential therapeutic target for RIF.

Abstract. The RhoA/ROCK signaling pathway plays a significant role in transforming growth factor (TGF)-β1-mediated epithelial-mesenchymal transition (EMT). It remains unclear, however, whether the RhoA/ROCK signaling pathway mediates TGF-β1-induced EMT by promoting the dissolution of tight junctions (TJs) in renal proximal tubular epithelial cells. In this study, we aimed to investigate the association between TGF-β1-mediated Rho/ROCK signaling and TJs in a cell line derived from human renal proximal tubular cells (HK-2 cells). HK-2 cells were treated with 5 ng/ml TGF-β1 for 0, 12, 24 and 48 h. Zona occludens protein 1 (also known as tight junction protein 1; ZO-1) and occludin mRNA and protein levels were determined by real-time PCR and western blot analysis, respectively. The HK-2 cells were then divided into three groups: a control group (serum-free culture medium for 24 h); a TGF-β1 group (treated with 5 ng/ml TGF-β1 for 24 h); and a TGF-β1 + Y-27632 (a specific ROCK inhibitor) group (pre-treated with 10 µM Y-27632 for 2 h and subsequently treated with 5 ng/ml TGF-β1 for 24 h). The levels of ZO-1 and occludin were detected by real-time PCR, western blot analysis and immunofluorescence. As shown by our results, the mRNA and protein levels of ZO-1 and occludin were decreased in the HK-2 cells following treatment with TGF-β1 in a time-dependent manner; in addition, ZO-1 and occludin levels in the TGF-β1 + Y-27632 group were significantly increased compared with those of the TGF-β1 group (P<0.05), with no significant changes compared with the control group. Our results indicate that the Rho/ROCK signaling pathway mediated by TGF-β1 plays a role in the dissolution of TJs during EMT.

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Purpose: To investigate whether Niban protein plays a role in renal interstitial fibrosis by regulating renal tubular epithelial cell apoptosis and explore the underlying mechanism. Methods: Unilateral ureteral obstruction (UUO) model was performed in C57B/6J mice, and divided into sham operation group and groups of days 3, days 7, and days 14. Niban expression was detected by immunohistochemistry and Western blot. TUNEL assays were used to detected apoptosis. Niban siRNA and overexpression Niban plasmid were transfected in HK-2 cells respectively to explore apoptosis related mechanisms of Niban during angiotensin II (AngII)and endoplasmic reticulum (ER) stress-induced injury. Results: With the development of obstruction, Niban's expression decreased gradually while apoptosis increased. Silencing of Niban not only increased the AngII-and ER stress-induced apoptosis, but also promoted the expression of caspase 8, caspase 9, Bip, and Chop. Overexpression of Niban reduced AngII-induced apoptosis and the expression of caspase 8 and caspase 9. Conclusions: Niban protein is involved in apoptosis regulation in HK-2 cells, and most likely via caspase-dependent pathway.

Niban decreased in renal tubular cells of patients of obstructive nephropathy, UUO rats and TGF-β1 stimulated HK-2 cells. Suppressing Niban increases apoptosis in HK-2 cells. Niban may be associated with apoptosis of HK-2 cells.

Objective: To investigate the causes of peritonitis in patients with peritoneal dialysis (PD) using continuous quality improvement (CQI) to develop effective interventions and reduce the occurrence of peritonitis. Methods: A quality control team consisting of 10 members, including the department head, four nephrologists and four nurses, all specialized in PD care, and the head nurse, was established at the Peritoneal Dialysis Center of the Third Xiangya Hospital of Central South University. All patients with peritonitis occurring between 1 July 2010 and 31 December 2011 (pre-CQI period) were analyzed and compared with data obtained between January 2012 (implementation of CQI) and March 2013 to investigate possible causes of peritonitis and to develop corresponding interventions. Fishbone analysis, including laboratory parameters, was carried out monthly. Results: Gastrointestinal tract dysfunction, nonstandard procedures and malnutrition were found to be the top three risk factors for peritonitis. Gastrointestinal tract dysfunction was the likely cause of peritonitis in 42.8% of the subjects before CQI and 36.0% after CQI (p50.05). Nonstandard procedures were the cause of peritonitis in 33.3% of the subjects before CQI and 24.0% after CQI (p50.05). The overall incidence of peritonitis reduced from once every 40.1 patient months before the CQI to once every 70.8 patient months after CQI (p50.05). The incidence of Grampositive bacteria peritonitis reduced from once every 96.9 patients per month before CQI to once every 209.1 patient months after CQI (p50.05), whereas the incidence of Gram-negative bacteria peritonitis reduced from once every 234.2 patient months before CQI to once every 292.8 patient months after CQI. Conclusion: CQI can effectively reduce the occurrence of PD-related peritonitis.