Renal Fibrosis In Obstructed Kidney Research Articles

Notch Blockade Specifically in Bone Marrow-Derived FSP-1-Positive Cells Ameliorates Renal Fibrosis

The infiltration of inflammatory cells during a kidney injury stimulates myofibroblast activation leading to kidney fibrosis. Fibroblast-specific protein 1 (FSP-1) positive cells have been reported as either myofibroblasts or monocytes during tissue fibrosis. The functions of FSP-1+ cells that are associated with the development of renal fibrosis and the signaling pathways that regulate FSP-1+ cell activation have not been well defined. In mice with unilateral ureteral obstruction (UUO), we characterized FSP-1+ cells and determined the role of the Notch signaling pathway in the activation of bone marrow-derived FSP-1+ cells during kidney fibrosis. In kidneys from mice with UUO, the FSP-1+ cells accumulated significantly in the tubulointerstitial area. By using immunostaining and FSP-1 reporter mice, we found that FSP-1 was co-stained with inflammatory cell markers, but not myofibroblast markers. Results from mice with bone marrow transplantations showed that FSP-1+ cells in obstructed kidneys represent a bone marrow-derived population of inflammatory cells. In cultured FSP-1+ cells, the inhibition of Notch signaling suppressed the activation and cytokine secretion of FSP-1+ cells that were induced by LPS but not by IL-4. The specific KO or blockade of Notch signaling in bone marrow-derived FSP-1+ cells suppressed UUO-induced ECM deposition, the infiltration of FSP-1+ inflammatory cells, and cytokine production. These responses ameliorated myofibroblast accumulation and renal fibrosis in obstructed kidneys. Our study reveals that most FSP-1+ cells in obstructed kidneys are activated macrophages that are derived from bone marrow and that Notch signaling activates the production of M1 cytokines in FSP-1+ monocytes/macrophages, which is important for renal inflammation and fibrosis.

Remdesivir Inhibits Tubulointerstitial Fibrosis in Obstructed Kidneys.

Aim: Kidney impairment is observed in patients with COVID-19. The effect of anti-COVID-19 agent remdesivir on kidneys is currently unknown. We aimed to determine the effect of remdesivir on renal fibrosis and its downstream mechanisms. Methods: Remdesivir and its active nucleoside metabolite GS-441524 were used to treat TGF-β stimulated renal fibroblasts (NRK-49F) and human renal epithelial (HK2) cells. Vehicle or remdesivir were given by intraperitoneal injection or renal injection through the left ureter in unilateral ureteral obstruction (UUO) mice. Serum and kidneys were harvested. The concentrations of remdesivir and GS-441524 were measured using LC-MS/MS. Renal and liver function were assessed. Renal fibrosis was evaluated by Masson’s trichrome staining and Western blotting. Results: Remdesivir and GS-441524 inhibited the expression of fibrotic markers (fibronectin and aSMA) in NRK-49F and HK2 cells. Intraperitoneal injection or renal injection of remdesivir attenuated renal fibrosis in UUO kidneys. Renal and liver function were unchanged in remdesivir treated UUO mice. Two remdesivir metabolites were detected after injection. Phosphorylation of Smad3 that was enhanced in cell and animal models for renal fibrosis was attenuated by remdesivir. In addition, the expression of Smad7, an anti-fibrotic factor, was increased after remdesivir treatment in vitro and in vivo. Moreover, knockdown of Smad7 blocked the antifibrotic effect of GS and RDV on renal cells. Conclusion: Remdesivir inhibits renal fibrosis in obstructed kidneys.

Exosomal miR-21 from tubular cells contributes to renal fibrosis by activating fibroblasts via targeting PTEN in obstructed kidneys.

Rationale: Ureteral obstruction-induced hydronephrosis is associated with renal fibrosis and progressive chronic kidney disease (CKD). Exosome-mediated cell-cell communication has been suggested to be involved in various diseases, including renal fibrosis. However, little is known regarding how exosomes regulate renal fibrosis in obstructed kidneys.Methods: We first examined the secretion of exosomes in UUO (unilateral ureteral obstruction) mouse kidneys and TGF-β1-stimulated tubular epithelial cells (NRK-52E). Exosomes from NRK-52E cells were subsequently harvested and incubated with fibroblasts (NRK-49F) or injected into UUO mice via the tail vein. We next constructed Rab27a knockout mice to further confirm the role of exosome-mediated epithelial-fibroblast communication relevant to renal fibrosis in UUO mice. High-throughput miRNA sequencing was performed to detect the miRNA profiles of TGFβ1-Exos. The roles of candidate miRNAs, their target genes and relevant pathways were predicted and assessed in vitro and in vivo by setting specific miRNA mimic, miRNA inhibitor, siRNA or miRNA LNA groups.Results: Increased renal fibrosis was associated with prolonged UUO days, and the secretion of exosomes was markedly increased in UUO kidneys and TGF-β1-stimulated NRK-52E cells. Purified exosomes from TGF-β1-stimulated NRK-52E cells could activate fibroblasts and aggravate renal fibrosis in vitro and in vivo. In addition, the inhibition of exosome secretion by Rab27a knockout or GW4869 treatment abolished fibroblast activation and ameliorated renal fibrosis. Exosomal miR-21 was significantly increased in TGFβ1-Exos compared with Ctrl-Exos, and PTEN is a certain target of miR-21. The promotion or inhibition of epithelial exosomal miR-21 correspondingly accelerated or abolished fibroblast activation in vitro, and renal fibrosis after UUO was alleviated by miR-21-deficient exosomes in vivo through the PTEN/Akt pathway.Conclusion: Our findings reveal that exosomal miR-21 from tubular epithelial cells may accelerate the development of renal fibrosis by activating fibroblasts via the miR-21/PTEN/Akt pathway in obstructed kidneys.

Trichostatin A Alleviates Renal Interstitial Fibrosis Through Modulation of the M2 Macrophage Subpopulation.

Mounting evidence indicates that an increase in histone deacetylation contributes to renal fibrosis. Although inhibition of histone deacetylase (HDAC) can reduce the extent of fibrosis, whether HDAC inhibitors exert the antifibrotic effect through modulating the phenotypes of macrophages, the key regulator of renal fibrosis, remains unknown. Moreover, the functional roles of the M2 macrophage subpopulation in fibrotic kidney diseases remain incompletely understood. Herein, we investigated the role of HDAC inhibitors on renal fibrogenesis and macrophage plasticity. We found that HDAC inhibition by trichostatin A (TSA) reduced the accumulation of interstitial macrophages, suppressed the activation of myofibroblasts and attenuated the extent of fibrosis in obstructive nephropathy. Moreover, TSA inhibited M1 macrophages and augmented M2 macrophage infiltration in fibrotic kidney tissue. Interestingly, TSA preferentially upregulated M2c macrophages and suppressed M2a macrophages in the obstructed kidneys, which was correlated with a reduction of interstitial fibrosis. TSA also repressed the expression of proinflammatory and profibrotic molecules in cultured M2a macrophages and inhibited the activation of renal myofibroblasts. In conclusion, our study was the first to show that HDAC inhibition by TSA alleviates renal fibrosis in obstructed kidneys through facilitating an M1 to M2c macrophage transition.

Exogenous pancreatic kininogenase protects against renal fibrosis in rat model of unilateral ureteral obstruction.

Tissue kallikrein has protective function against various types of injury. In this study, we investigated whether exogenous pancreatic kininogenase (PK) conferred renoprotection in a rat model of unilateral ureteral obstruction (UUO) and H2O2-treated HK-2 cells in vitro. SD rats were subjected to UUO surgery, then PK (7.2 U/g per day, ip) was administered for 7 or 14 days. After the treatment, rats were euthanized; the obstructed kidneys were harvested for further examination. We found that PK administration significantly attenuated interstitial inflammation and fibrosis, and downregulated the expression of proinflammatory (MCP-1, TLR-2, and OPN) and profibrotic (TGF-β1 and CTGF) cytokines in obstructed kidney. UUO-induced oxidative stress, closely associated with excessive apoptotic cell death and autophagy via PI3K/AKT/FoxO1a signaling, which were abolished by PK administration. We further showed that PK administration increased the expression of bradykinin receptors 1 and 2 (B1R and B2R) mRNA and the production of NO and cAMP in kidney tissues. Coadministration with either B1R antagonist (des-Arg9-[Leu8]-bradykinin) or B2R antagonist (icatibant) abrogated the renoprotective effects of PK, and reduced the levels of NO and cAMP in obstructed kidney. In H2O2-treated HK-2 cells, addition of PK (6 pg/mL) significantly decreased ROS production, regulated the expression of oxidant and antioxidant enzymes, suppressed the expression of TGF-β1 and MCP-1, and inhibited cell apoptosis. Our data demonstrate that PK treatment protects against the progression of renal fibrosis in obstructed kidneys.

Reduced asymmetric dimethylarginine accumulation through inhibition of the type I protein arginine methyltransferases promotes renal fibrosis in obstructed kidneys.

The role of asymmetric dimethylarginine (ADMA) in chronic kidney disease (CKD) is unclear. Through inhibition of type I protein arginine methyltransferases (PRMTs), a novel strategy, we aimed to determine the effect of ADMA on renal fibrosis and explore its underlying working mechanisms. After sham or unilateral ureter ligation (UUO) operation, 20-25 g male c57 mice were treated with vehicle or PT1001B, an inhibitor of type I PRMTs, for 13 d. Moreover, human kidney 2 (HK2) and normal rat kidney 49F (NRK-49F) cells were treated with various concentrations of PT1001B or ADMA in the presence of 2.5 ng/ml TGF-β. We found that treatment with PT1001B increased the deposition of extracellular matrix proteins, the expression of α smooth muscle actin, and connective tissue growth factor in UUO-induced fibrotic kidneys, which is correlated with reduced expression of PRMT1, reduced the production of ADMA, and increased expression of uromodulin. In TGF-β-stimulated HK2 and NRK-49F cells, PT1001B dose-dependently inhibited ADMA production, increased NO concentrations, and enhanced the expression of profibrotic proteins. Exogenous addition of ADMA inhibited the expression of profibrotic proteins dose-dependently and attenuated the profibrotic effect of PT1001B. Moreover, ADMA reduced the NO concentration in PT1001B-treated HK2 cells. Finally, we conclude that ADMA has an antifibrotic effect in obstructed kidneys, and future application of type I PRMT inhibitor should be done cautiously for patients with CKD.-Wu, M., Lin, P., Li, L., Chen, D., Yang, X., Xu, L., Zhou, B., Wang, C., Zhang, Y., Luo, C., Ye, C. Reduced asymmetric dimethylarginine accumulation through inhibition of the type I protein arginine methyltransferases promotes renal fibrosis in obstructed kidneys.

Altered Regulation of Renin-Angiotensin, Endothelin and Natriuretic Peptide Systems in Rat Kidney with Chronic Unilateral Ureteral Obstruction

Background: The present study was designed to investigate the role of the local renin-angiotensin-aldosterone system (RAAS), endothelin (ET) and the natriuretic peptide system (NPS) for the development of renal fibrosis and progressive renal disease in experimental unilateral ureteral obstructed (UUO) rats. Methods: Male Sprague-Dawley rats (180–200 g) were unilaterally obstructed by ligation of the proximal ureters for 14 days. Control rats were treated in the same way, except that no ligature was made. The mRNA expressions of local renin-angiotensin system, aldosterone synthase (CYP11B2), ET-1 and NPS was determined in the cortex by real-time polymerase chain reaction. Results: Following the unilateral ureteral obstruction, the mRNA expressions of angiotensin-converting enzyme 1, ET-1 and transforming growth factor-β₁ (TGF-β₁) were increased, while angiotensin-converting enzyme 2 was decreased in the obstructed kidney compared with the controls. Angiotensin II type 1 receptor was decreased and TGF-β₁ was not changed in contralateral kidney compared with the controls. Atrial natriuretic peptide, brain natriuretic peptide, and C-type natri- uretic peptide expressions were increased in UUO kidneys compared with the controls, while natriuretic peptide receptor-A was decreased in UUO kidneys. Conclusion: The local RAAS and ET-1 was upregulated which may play a role in the progressive renal fibrosis in obstructed kidneys in rats with UUO. The enhanced activity of NPS in UUO kidney may play a role to compensate against progressive renal fibrosis in chronic obstructive uropathy.

Twist Relates to Tubular Epithelial-Mesenchymal Transition and Interstitial Fibrogenesis in the Obstructed Kidney

Epithelial-mesenchymal transition (EMT) is a critical step in renal fibrosis. It has been recently reported that a transcription factor, Twist, plays a pivotal role in metastasis of breast tumors by inducing EMT. In this study, we examined whether Twist relates to renal fibrogenesis including EMT of tubular epithelia, evaluating Twist expression level in the unilateral ureteral obstruction (UUO) model. Kidneys of mice subjected to UUO were harvested 1, 3, 7, and 10 days after obstruction. Compared with control kidneys, Twist mRNA-level significantly increased 3 days after UUO (UUO day 3 kidney) and further augmented until 10 days after UUO. Twist expression increased in tubular epithelia of the dilated tubules and the expanded interstitial areas of UUO kidneys, where cell-proliferating appearances were frequently found in a time-dependent manner. Although a part of tubular cells in whole nephron segment were immunopositive for Twist in UUO day 7 kidneys, tubular epithelia downstream of nephron more frequently expressed Twist than upstream of nephron. In UUO day 7 kidneys, some tubular epithelia were confirmed to coexpress Twist and fibroblast-specific protein-1, a marker for EMT, indicating that Twist is involved in tubular EMT under pathological state. Twist was expressed also in a number of alpha-smooth muscle actin-positive myofibroblasts located in the expanded interstitial area of UUO kidneys. From these findings, the present investigation suggests that Twist is associated with tubular EMT, proliferation of myofibroblasts, and subsequent renal fibrosis in obstructed kidneys.