Metformin attenuates renal interstitial fibrosis through upregulation of Deptor in unilateral ureteral obstruction in rats.

Renal interstitial fibrosis (RIF) is a common pathological process that accompanies chronic kidney disease (CKD) and that progresses to end-stage renal failure (ESRD). Accumulating evidence has revealed that persistent mammalian target of rapamycin (mTOR) activation in kidneys is closely associated with the occurrence and progression of CKD. The DEP domain-containing mTOR interacting protein (Deptor) is an endogenous negative regulator of mTOR. Metformin can attenuate renal fibrosis in an animal model of diabetic nephropathy. Previous studies demonstrated that metformin can attenuate renal fibrosis in several models of CKD. However, the precise mechanisms of this effect are not well understood. The present study aimed to examine the mechanism of action of metformin on unilateral ureteral obstruction (UUO)-induced RIF in rats in vivo. Sprague-Dawley rats were randomly divided into a sham-operated group, three UUO groups examined at different time points (3, 7 and 14 days after UUO surgery), and three metformin-treated groups, treated with three different concentrations of metformin. The metformin-treated groups were administered metformin orally every day for 14 consecutive days following surgery. The protein expression levels of Deptor, α-smooth muscle actin (α-SMA), phosphorylated (p-)mTOR, p-ribosomal protein S6 kinase (p-p70S6K) and CD68 were assessed. The present results suggested that, following UUO, there was a significant reduction of Deptor expression, and an increase in collagen deposition in the extracellular matrix over time, accompanied by an increased expression of several proteins including CD68, α-SMA, p-mTOR and p-p70S6K. Notably, metformin treatment reversed these effects. In conclusion, the present results suggested that metformin attenuated RIF of UUO rats, and the mechanism of action was found to be associated with the increase in Deptor expression and inhibition of the mTOR/p70S6K pathway in the kidneys of UUO rats.

Role of artesunate in TGF­ß1­induced renal tubular epithelial­mesenchymal transdifferentiation in NRK­52E cells.

The implications of epithelial­mesenchymal transdifferentiation (EMT) have extended beyond the confines of renal fibrosis to renal tubulointerstitial fibrosis. It has been proposed that EMT may be one of the mechanisms involved in the pathogenesis of renal fibrosis. However, the underlying mechanisms remain unknown. Transforming growth factor (TGF)­ß1 is considered to be an important cytokine which regulates the transdifferentiation of tubular epithelial cells into myofibroblasts in renal tubulointerstitial fibrosis. In the present study, normal rat kidney tubular epithelial cells (NRK­52E) were treated for 48 h with TGF­ß1 (5 ng/ml) and different concentrations of artesunate (ART; 0.01, 0.1 and 1 µg/ml). Western blotting, reverse transcription­semi quantitative polymerase chain reaction analysis and immunofluorescence staining were used to evaluate the expression of bone morphogenetic protein (BMP)­7, uterine sensitization­associated gene (USAG)­1, E­cadherin, α­smooth muscle actin (α­SMA) and extracellular matrix collagen type I (Col I) mRNA. ART was able to attenuate renal injury in a unilateral ureteral obstruction model. However, its anti­fibrotic effect remains to be elucidated. In the present study, it was observed that ART was able to ameliorate the TGF­ß1­induced alterations in cellular morphology. In addition, ART inhibited the TGF­ß1­induced USAG­1 increase and the decrease in BMP­7. Treatment with ART markedly attenuated the TGF­ß1­induced upregulation of α­SMA and downregulation of E­cadherin. Additionally, ART was able to significantly attenuate the deposition of interstitial collagens, including Col I. The results of the present study further verified the therapeutic efficacy of ART in TGF­ß1­induced renal interstitial fibrosis. These findings indicated that ART may hold the potential to prevent chronic kidney diseases via the suppression of USAG­1 expression or by increasing BMP­7 expression.

Artesunate attenuates unilateral ureteral obstruction-induced renal fibrosis by regulating the expressions of bone morphogenetic protein-7 and uterine sensitization-associated gene-1 in rats.

PURPOSE: Kidney fibrosis is the most common final stage of progressive renal disease. Bone morphogenetic protein-7 (BMP-7) has been shown to be important in both preservation of kidney function and resistance to injury. Recently, it has been realized that uterine sensitization-associated gene-1 (USAG-1) functions as a kidney-specific BMP antagonist. Because of the reported anti-fibrotic effects of artesunate (Art), this study was designed to investigate the effects of Art on renal fibrosis in unilateral ureteral obstruction (UUO) rats and to explore the underlying mechanisms. METHODS: Thirty male Sprague-Dawley rats were randomly divided into five groups: Sham group, UUO group, low-dose Art-treated (Art-L), middle-dose Art-treated (Art-M), and high-dose Art-treated (Art-H) groups. The UUO rat model was established by ligating the left ureter. Fourteen days later, interstitial collagen deposition, expression of USAG-1, BMP-7, E-cadherin, α-smooth muscle actin (α-SMA), fibronectin (FN), collagen I, as well as the inflammatory infiltration levels in the kidneys were assessed. RESULTS: Art treatment significantly attenuated the deposition of interstitial collagens in the UUO rats' kidneys and exhibited the ability to improve renal function, followed by the up-regulated expression of BMP-7 and E-cadherin and the down-regulated expression of USAG-1 and α-SMA. In addition, increased macrophages infiltration in the kidneys of the UUO rats were also attenuated by the administration of Art. CONCLUSIONS: These results indicate that Art is able to improve the renal function decline and renal fibrosis induced by UUO, which may be associated with the up-regulation of BMP-7 and down-regulation of USAG-1. Accordingly, Art may become a potential preventive or therapeutic agent for chronic kidney diseases.

Febuxostat Prevents Renal Interstitial Fibrosis by the Activation of BMP-7 Signaling and Inhibition of USAG-1 Expression in Rats.

BACKGROUND: Renal interstitial fibrosis (RIF) is a common pathology associated with end-stage renal diseases. The activation of bone morphogenetic protein-7 (BMP-7)-Smad1/5/8 pathway seems to alleviate RIF. Uterine sensitization-associated gene-1 (USAG-1), a kidney-specific BMPs antagonist, is associated with the development and prognosis of several renal diseases. Febuxostat is a xanthine oxidase inhibitor that can attenuate the renal dysfunction of patients. The purpose of this study was to investigate the effects of febuxostat on renal fibrosis and to clarify the mechanisms underlying these effects. METHODS: Rats were randomly divided into 6 groups termed a sham-operated group, a unilateral ureteral obstruction (UUO) group, 3 doses of febuxostat groups (low, intermediate and high doses) and a sham group treated with high-dose febuxostat. After 14 days, renal function, relative kidney weight, accumulation of glycogen and collagens were examined by different methods. Expression of α-SMA, transforming growth factor-ß1 (TGF-ß1), BMP-7 and USAG-1 was detected by western blotting and RT-PCR, respectively. The phosphorylation level of Smad1/5/8 was also quantified by western blotting. RESULTS: The renal function was declined, and large amounts of glycogen and collagens were deposited in the kidneys of UUO rats compared with the rats in the sham group. Besides, expression of α-SMA and USAG-1 in these kidneys was elevated, and the TGF-ß1 was also activated, while the BMP-7-Smad1/5/8 pathway was inhibited. Febuxostat reversed the changes stated earlier, exhibiting protective effects on RIF induced by UUO. CONCLUSION: Febuxostat was able to attenuate RIF caused by UUO, which was associated with the activation of BMP-7-Smad1/5/8 pathway and the inhibition of USAG-1 expression in the kidneys of UUO rats.