Renal Interstitial Fibrosis Research Articles

Fluorofenidone attenuates renal fibrosis by inhibiting lysosomal cathepsin‑mediated NLRP3 inflammasome activation.

Currently, no antifibrotic drug in clinical use can effectively treat renal fibrosis. Fluorofenidone (AKFPD), a novel pyridone agent, significantly reduces renal fibrosis by inhibiting the activation of the NOD-like receptor thermal protein domain associated protein 3 (NLRP3) inflammasome; however, the underlying mechanism of this inhibition is not fully understood. The present study aimed to reveal the molecular mechanism underlying the suppression of NLRP3 inflammasome activation by AKFPD. It investigated the effect of AKFPD on NLRP3 activation and lysosomal cathepsins in a unilateral ureteral obstruction (UUO) rat model, and hypoxia/reoxygenation (H/R)-treated HK-2 cells and murine peritoneal-derived macrophages (PDMs) stimulated with lipopolysaccharide (LPS) and ATP. The results confirmed that AKFPD suppressed renal interstitial fibrosis and inflammation by inhibiting NLRP3 inflammasome activation in UUO rat kidney tissues. In addition, AKFPD reduced the production of activated caspase-1 and maturation of IL-1β by suppressing NLRP3 inflammasome activation in H/R-treated HK-2 cells and murine PDMs stimulated with LPS and ATP. AKFPD also decreased the activities of cathepsins B, L and S both in vivo and in vitro. Notably, AKFPD downregulated cathepsin B expression and NLRP3 colocalization in the cytoplasm after lysosomal disruptions. Overall, the results suggested that AKFPD attenuates renal fibrosis by inhibiting lysosomal cathepsin-mediated activation of the NLRP3 inflammasome.

Shen-Shuai-II-Recipe inhibits tubular inflammation by PPARα-mediated fatty acid oxidation to attenuate fibroblast activation in fibrotic kidneys

BackgroundShen Shuai Ⅱ Recipe (SSR) is clinically used to treat chronic kidney diseases (CKDs) with remarkable efficacy and safety. In earlier research, we found the anti-inflammatory, antioxidant, and mitochondrial protective properties of SSR in hypoxic kidney injury model, which is closely related to its renal protection. Further work is needed to understand the underlying molecular mechanisms. PurposeFurther investigation of the mechanisms of action of SSR against renal interstitial fibrosis (RIF) building on previous research leads. MethodsRats receiving CKD model surgery were given with Fenofibrate or SSR once a day for eight weeks. In vitro, the NRK-52E cells were treated with SSR in the presence or absence of 10 μM Sc75741, 0.5 μM PMA, or 1 μM fenofibrate under 1% O2. The effects of SSR on NF-κB/NLRP3 inflammatory cascade, secretion of pro-inflammatory cytokines, fatty acid oxidation (FAO), and renal tubular injury were determined by immunoblotting, luminex liquid suspension chip assay, transmission electron microscopy, and Oil red O staining. Next, we delivered PPARα-interfering sequences to kidney tissue and NRK-52E cells by adeno-associated virus (AAV) injection and siRNA transfection methods. Finally, we evaluated the effect of renal tubular cells on fibroblast activation by co-culture method. ResultsSSR attenuated the release of IL-18, VEGF, and MCP1 cytokines, inhibited the activation of NF-κB/NLRP3 cascade, increased the PPARα, CPT-1α, CPT-2, ACADL, and MCAD protein expression, and improved the lipid accumulation. Further studies have demonstrated that one of the ways in which SSR suppresses the inflammatory response to protect renal tubular cells is through the restoration of PPARα-mediated FAO. In addition, by means of co-culture ways, the results demonstrated that SSR attenuated secretion of inflammatory mediators in NRK-52E cells by PPARα/NF-κB/NLRP3 pathway, thereby inhibiting renal fibroblast activation. ConclusionSSR inhibits RIF by suppressing inflammatory response of hypoxia-exposed RTECs through PPARα-mediated FAO.

Renal and cardiac effects of the PDE9 inhibitor BAY 73-6691 in 5/6 nephrectomized rats.

It has been suggested that the novel selective phosphodiesterase 9 (PDE9) inhibitor may improve cardiac and renal function by blocking 3',5'-cyclic guanosine monophosphate (cGMP) degradation.5/6 nephrectomized (5/6Nx) rats were used to investigate the effects of the PDE9 inhibitor (BAY 73-6691) on the heart and kidney. Two doses of BAY 73-6691 (1mg/kg/day and 5mg/kg/day) were given for 95days.The 5/6Nx rats developed albuminuria, a decrease in serum creatinine clearance (Ccr), and elevated serum troponin T levels. Echocardiographic data showed that 5/6 nephrectomy resulted in increased fractional shortening (FS), stroke volume (SV), and left ventricular ejection fraction (EF). However, 95days of PDE9 inhibitor treatment did not improve any cardiac and renal functional parameter. Histopathologically, 5/6 nephrectomy resulted in severe kidney and heart damage, such as renal interstitial fibrosis, glomerulosclerosis, and enlarged cardiomyocytes. Telmisartan attenuated renal interstitial fibrosis and glomerulosclerosis as well as improved cardiomyocyte size. However, except for cardiomyocyte size and renal perivascular fibrosis, BAY 73-6691 had no effect on other cardiac and renal histologic parameters. Pathway enrichment analysis using RNA sequencing data of kidney and heart tissue identified chronic kidney disease pathways, such as phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt) signaling pathway, complement and coagulation cascades, and nuclear factor kappa B (NF-κB) signaling pathway. PDE9i did not affect any of these disease-related pathways.Two dosages of the PDE9 inhibitor BAY 73-6691 known to be effective in other rat models have only limited cardio-renal protective effects in 5/6 nephrectomized rats.

A gradient model of renal ischemia reperfusion injury to investigate renal interstitial fibrosis.

Background: The progression from acute kidney injury to chronic kidney disease poses a significant health challenge. Nonetheless, a constraint in existing animal models of renal ischemia/reperfusion (I/R) injury is the necessity for a severe injury, almost reaching a life-threatening level, to trigger the subsequent onset of renal fibrosis. Hence, we explored an adapted gradient approach to induce I/R injury, aiming to promote the progression of renal fibrosis while preserving the overall normal functioning of the kidney. Methods: In each group, 6-8 male C57BL/6 mice were used for model construction, with all undergoing sodium pentobarbital anesthesia and left kidney removal. Subsequently, a silk thread was passed beneath the lower renal branch, elevating the right kidney under a 20-g weight's tension via a pulley system for durations of 30, 40, or 60min. Afterwards, we lowered the kidney, sutured the wound, and administered intraperitoneal saline. Mice in different groups were euthanized following reperfusion for 1, 3, 7, or 28days. Results: We observed a complete cessation of blood flow in the lower pole, while an incomplete cessation in the upper pole in the elevated kidney. Significant renal impairment was evident on day 1 with a 60min ischemic period (187.0 ± 65.3 vs 17.9 ± 4.8μmol/L serum creatinine in normal; p < .001), but not with 30 or 40min. On day 1, tubular necrosis and hyaline cast formation was evident in both lower and upper poles. On day 3, renal function returned to normal and remained normal through day 28. Histologic damage resolved in the upper pole over days 3 to 7, resulting in normal histology on day 28. By contrast, there was sustained tubular damage tubular in the lower pole on days 3 and 7, which failed to resolve and led to significant renal fibrosis by day 28. Conclusion: We created a model demonstrating clinically "silent" renal fibrosis.

The pathogenic role of succinate-SUCNR1: a critical function that induces renal fibrosis via M2 macrophage

BackgroundRenal fibrosis significantly contributes to the progressive loss of kidney function in chronic kidney disease (CKD), with alternatively activated M2 macrophages playing a crucial role in this progression. The serum succinate level is consistently elevated in individuals with diabetes and obesity, both of which are critical factors contributing to CKD. However, it remains unclear whether elevated succinate levels can mediate M2 polarization of macrophages and contribute to renal interstitial fibrosis.MethodsMale C57/BL6 mice were administered water supplemented with 4% succinate for 12 weeks to assess its impact on renal interstitial fibrosis. Additionally, the significance of macrophages was confirmed in vivo by using clodronate liposomes to deplete them. Furthermore, we employed RAW 264.7 and NRK-49F cells to investigate the underlying molecular mechanisms.ResultsSuccinate caused renal interstitial macrophage infiltration, activation of profibrotic M2 phenotype, upregulation of profibrotic factors, and interstitial fibrosis. Treatment of clodronate liposomes markedly depleted macrophages and prevented the succinate-induced increase in profibrotic factors and fibrosis. Mechanically, succinate promoted CTGF transcription via triggering SUCNR1-p-Akt/p-GSK3β/β-catenin signaling, which was inhibited by SUCNR1 siRNA. The knockdown of succinate receptor (SUCNR1) or pretreatment of anti-CTGF(connective tissue growth factor) antibody suppressed the stimulating effects of succinate on RAW 264.7 and NRK-49F cells.ConclusionsThe causative effects of succinate on renal interstitial fibrosis were mediated by the activation of profibrotic M2 macrophages. Succinate-SUCNR1 played a role in activating p-Akt/p-GSK3β/β-catenin, CTGF expression, and facilitating crosstalk between macrophages and fibroblasts. Our findings suggest a promising strategy to prevent the progression of metabolic CKD by promoting the excretion of succinate in urine and/or using selective antagonists for SUCNR1.

Sulforaphane (Sul) reduces renal interstitial fibrosis (RIF) by controlling the inflammation and TGF-β/Smad signaling pathway

All chronic renal disorders eventually lead to renal interstitial fibrosis (RIF). Chronic inflammation and pro-fibrotic substances are familiar companions of the fibrotic process. The Sulforaphane (Sul) molecule is particularly useful in protecting the liver from oxidative damage. To investigate the Sul effects on fibrosis markers and inflammatory proteins in the kidney of NRK52E cell line and rats and clarify the mechanism of TGF-β/Smad signaling pathway in a rat model of RIF were developed in the present study. Sul (50, 100, and 200 ng/ml) remarkably reduced the gene expressions of tumor necrosis factor (TNF-α), interleukin-6 (IL-6), interleukin (IL)-1β, collagen 3 (COL3A1), collagen 1 (COL1A1), and α-smooth muscle actin (α-SMA) in fibrotic NRK52E cells compared with those in cells inspired by transforming growth factor-α (TGF-α). Histopathological investigations showed that Sul administration retained renal tissue structure and decreased kidney tissue fibrosis in rats subjected to unilateral ureteral blockage (UUO). The expression level of TNF-α, IL-6, IL-1β, COL3A1, COL1A1, and α-SMA in the rats’ kidneys exposed to UUO was also suppressed by the treatment of Sul. In the present study, western blot analysis showed that Sul upregulated the expressions of fibrotic NRK52E cells Smad7 and rat model UUO groups while simultaneously decreasing the stimulation of Smad2/3 and the expressions of cyclooxygenase-2, NF-κB, Smad4, activator protein-1, and high-mobility group protein B1. Ultimately, Sul’s ability to inhibit the TGF-β/Smad pathway and the development of inflammation factors may mitigate RIF.

Retraction: QiShenYiQi Attenuates Renal Interstitial Fibrosis by Blocking the Activation of β-Catenin.

Retraction: QiShenYiQi Attenuates Renal Interstitial Fibrosis by Blocking the Activation of β-Catenin.

Exercise alleviates renal interstitial fibrosis by ameliorating the Sirt1-mediated TGF-β1/Smad3 pathway in T2DM mice.

Renal interstitial fibrosis is the pathophysiological basis of type 2 diabetes mellitus (T2DM). Exercise appears to improve kidney interstitial fibrosis in T2DM, in which silent information regulator factor 2-related enzyme 1 (Sirt1) is a critical regulator. However, the role of Sirt1 in mediating exercise on renal tissue as well as its mechanism remains unknown. T2DM mouse models were created using a high-fat diet mixed with streptozotocin, followed by 8 weeks of treadmill exercise and niacinamide (Sirt1 inhibitor) intervention. Kits for detecting biochemical indices of renal function were used. The pathological appearance and severity of renal tissue were examined using hematoxylin and eosin, Masson and immunohistochemical staining. The mRNA and protein expression of relevant signaling pathway factors were determined to use real-time reverse transcriptase-polymerase chain reaction and western blotting. T2DM can promote renal interstitial fibrosis, increase kidney index, serum creatinine, blood urea nitrogen and 24 h urinary total protein and cause pathological changes in renal tissue and affect renal function. After 8 weeks of exercise intervention, the biochemical indicators in the kidney of T2DM mice were decreased, Sirt1 expression was increased, the expression of TGF-β1, Smad3, collagen type I (COL1) and collagen type III (COL3) were decreased, and the renal interstitial fibrosis, renal tissue structural lesions and renal function were improved. However, after the nicotinamide intervention, renal interstitial fibrosis of T2DM mice was aggravated, and the improvement effect of exercise on renal interstitial fibrosis of T2DM mice was abolished. The upregulation of Sirt1 expression by exercise can inhibit the transforming growth factor β1/Smad3 pathway, thereby inhibiting the expression and deposition of COL1 and COL3 in renal interstitium, thereby improving renal interstitial fibrosis in T2DM.

Amide Proton Transfer-Weighted Magnetic Resonance Imaging for Application in Renal Fibrosis: A Radiological-Pathological-Based Analysis

Plain Language SummaryAmide proton transfer (APT) MRI, a novel molecular MRI technique, could indirectly measure the peptide or protein concentration within tissues based on frequent exchange between amide proton and the surrounding water protons. This new technology has been used in tumor diagnosis. Given pathological conditions linked to tissue fibrogenesis are similar to carcinogenesis, we proposed the hypothesis that APT MRI can be applied to assess collagen deposition in RF. To test this, we constructed animal models for RF and enrolled CKD patients, respectively. The results suggested APT values were more sensitive to the early stage of fibrosis than ADC values, and the cortical APT was highly positively correlated with the degree of fibrosis. We further identified that the combination of two MRI protocols, cortical apparent diffusion coefficient and cortical APT, was an excellent indicator for the diagnosis of fibrosis, with much higher accuracy than a single method. To the best of our knowledge, this is the first study to explore the value of APT imaging for assessing renal interstitial fibrosis. Our findings identify APT as a novel and precise imaging diagnostic tool for future noninvasive assessment of RF. It complements with the existing imaging biomarkers and adds value to conventional MRI.

Engineered extracellular vesicle-encapsulated CHIP as novel nanotherapeutics for treatment of renal fibrosis

Renal interstitial fibrosis (RIF) is a fundamental pathological feature of chronic kidney disease (CKD). However, toxicity and poor renal enrichment of fibrosis inhibitors limit their further applications. In this study, a platform for CKD therapy is developed using superparamagnetic iron oxide nanoparticles (SPION) decorated mesenchymal stem cells derived extracellular vesicles with carboxyl terminus of Hsc70-interacting protein (CHIP) high expression (SPION-EVs) to achieve higher renal-targeting antifibrotic therapeutic effect. SPION-EVs selectively accumulate at the injury renal sites under an external magnetic field. Moreover, SPION-EVs deliver CHIP to induce Smad2/3 degradation in renal tubular cells which alleviates Smad2/3 activation-mediated fibrosis-like changes and collagen deposition. The extracellular vesicle engineering technology provides a potential nanoplatform for RIF therapy through CHIP-mediated Smad2/3 degradation.

Diosmin ameliorates renal fibrosis through inhibition of inflammation by regulating SIRT3-mediated NF-κB p65 nuclear translocation

BackgroundRenal fibrosis is considered an irreversible pathological process and the ultimate common pathway for the development of all types of chronic kidney diseases and renal failure. Diosmin is a natural flavonoid glycoside that has antioxidant, anti-inflammatory, and antifibrotic activities. However, whether Diosmin protects kidneys by inhibiting renal fibrosis is unknown. We aimed to investigate the role of Diosmin in renal interstitial fibrosis and to explore the underlying mechanisms.MethodsThe UUO mouse model was established and gavaged with Diosmin (50 mg/kg·d and 100 mg/kg·d) for 14 days. HE staining, Masson staining, immunohistochemistry, western blotting and PCR were used to assess renal tissue injury and fibrosis. Elisa kits were used to detect the expression levels of IL-1β, IL-6, and TNF-α and the activity of SIRT3 in renal tissues. In addition, enrichment maps of RNA sequencing analyzed changes in signaling pathways. In vitro, human renal tubular epithelial cells (HK-2) were stimulated with TGF-β1 and then treated with diosmin (75 μM). The protein and mRNA expression levels of SIRT3 were detected in the cells. In addition, 3-TYP (selective inhibitor of SIRT3) and SIRT3 small interfering RNA (siRNA) were used to reduce SIRT3 levels in HK-2.ResultsDiosmin attenuated UUO-induced renal fibrosis and TGF-β1-induced HK-2 fibrosis. In addition, Diosmin reduced IL-1β, IL-6, and TNF-α levels in kidney tissues and supernatants of HK-2 medium. Interestingly, Diosmin administration increased the enzymatic activity of SIRT3 in UUO kidneys. In addition, Diosmin significantly increased mRNA and protein expression of SIRT3 in vitro and in vivo. Inhibition of SIRT3 expression using 3-TYP or SIRT3 siRNA abolished the anti-inflammatory effects of diosmin in HK-2 cells. Enrichment map analysis by RNA sequencing indicates that the nuclear factor-kappa B (NF-κB) signaling pathway was inhibited in the Diosmin intervention group. Furthermore, we found that TGF-β1 increased the nuclear expression of nuclear NF-κB p65 but had little significant effect on the total intracellular expression of NF-κB p65. Additionally, Diosmin reduced TGF-β1-caused NF-κB p65 nuclear translocation. Knockdown of SIRT3 expression by SIRT3 siRNA increased the nuclear expression of NF-κB p65 and abolished the inhibition effect of Diosmin in NF-κB p65 expression.ConclusionsDiosmin reduces renal inflammation and fibrosis, which is contributed by inhibiting nuclear translocation of NF-κB P65 through activating SIRT3.

Downregulating ANGPTL3 by miR-144-3p promoted TGF-β1-induced renal interstitial fibrosis via activating PI3K/AKT signaling pathway

Despite observations of decreased ANGPTL3 (angiopoietin-like protein 3) levels in tubular atrophy and renal interstitial fibrosis (RIF), its functional implications and regulatory mechanisms in RIF remain unclear. This investigation employed unilateral ureteral obstruction (UUO) mice as in vivo model and human proximal kidney tubuloepithelial HK-2 cells under TGF-β1 treatment as in vitro model to explore RIF. The RIF extent was evaluated using H & E staining and Masson's trichrome staining. There was a significant decrease in ANGPTL3 levels and an increase in miR-144-3p, accompanied by heightened expressions of α-SMA, p-PI3K, p-AKT, Collagen I, and Fibronectin in the UUO mice and HK-2 cells treated with TGF-β1. Enhancing ANGPTL3 expression or suppressing miR-144-3p mitigated TGF-β1-induced cellular apoptosis, inflammation, and PI3K/AKT signaling pathway activation, as evidenced by altered levels of α-SMA, Collagen I, Fibronectin, and associated signaling markers. Using a bioinformatics approach, a miR-144-3p binding site was discovered on the ANGPTL3 mRNA, and this finding was subsequently confirmed through luciferase reporter assay. In HK-2 cells stimulated with TGF-β1, the suppression of ANGPTL3 negated the effects of inhibiting miR-144-3p. Under comparable conditions, the use of LY294002, an inhibitor of the PI3K/AKT pathway, nullified the effects caused by the knockdown of ANGPTL3. Collectively, these findings indicate that miR-144-3p exacerbates RIF through PI3K/AKT pathway activation by targeting ANGPTL3, highlighting a novel potential therapeutic target for RIF management.

NSD2 modulates Drp1-mediated mitochondrial fission in chronic renal allograft interstitial fibrosis by methylating STAT1

Renal interstitial fibrosis/tubular atrophy (IF/TA) is a prominent pathological feature of chronic allograft dysfunction (CAD). Our previous study has demonstrated that epithelial-mesenchymal transition (EMT) plays a significant role in shaping the development of IF/TA. Nuclear SET domain (NSD2), a histone methyltransferase catalyzing methylation at lysine 36 of histone 3, is crucially involved in the development and progression of solid tumors. But its role in the development of renal allograft interstitial fibrosis has yet to be elucidated. Here, we characterize NSD2 as a crucial mediator in the mouse renal transplantation model in vivo and a model of tumor necrosis factor-α (TNF-α) stimulated-human renal tubular epithelial cells (HK-2) in vitro. Functionally, NSD2 knockdown inhibits EMT, dynamin-related protein 1 (Drp1)-mediated mitochondrial fission in mice. Conversely, NSD2 overexpression exacerbates fibrosis-associated phenotypes and mitochondrial fission in tubular cells. Mechanistically, tubular NSD2 aggravated the Drp-1 mediated mitochondrial fission via STAT1/ERK/PI3K/Akt signaling pathway in TNF-α-induced epithelial cell models. Momentously, mass spectrometry (MS) Analysis and site-directed mutagenesis assays revealed that NSD2 interacted with and induced Mono-methylation of STAT1 on K173, leading to its phosphorylation, IMB1-dependent nuclear translocation and subsequent influence on TNF-α-induced EMT and mitochondrial fission in NSD2-dependent manner. Collectively, these findings shed light on the mechanisms and suggest that targeting NSD2 could be a promising therapeutic approach to enhance tubular cell survival and alleviate interstitial fibrosis in renal allografts during CAD.

Thonningianin A ameliorated renal interstitial fibrosis in diabetic nephropathy mice by modulating gut microbiota dysbiosis and repressing inflammation.

This study was conducted to examine the potential health benefits of thonningianin A (TA) on renal injury and interstitial fibrosis in diabetic nephropathy (DN) mice. In this study, a DN mice model was established using male C57BL/6 mice injected with streptozotocin (STZ, 50mg/kg) intraperitoneally and treated with TA for 12weeks. Firstly, the therapeutic and anti-fibrotic effects of TA on DN were evaluated. Secondly, the effect of TA on renal inflammation was evaluated and Western blot was used to detect the changes of NLRP3/ASC/Caspase-1 pathway-related protein expressions in kidney. Furthermore, the effect of TA on impairments in the intestinal mucosa barrier was evaluated and the changes of lipopolysaccharide (LPS) levels in feces and serum were detected by ELISA. Finally, 16S rRNA sequencing was used to detect alteration of gut microbiota diversity and abundance in mice after TA treatment. The results showed that TA markedly mitigated blood glucose (Glu), decreased 24-h urinary total protein (24hUTP), improved renal dysfunction and kidney index (KI) in DN mice. Furthermore, TA significantly alleviated renal injury and interstitial fibrosis, repressing renal inflammation. Western blot results showed that the NLRP3/ASC/Caspase-1 signaling pathway-related proteins decreased after TA treatment. In addition, TA also ameliorated impairments in the intestinal mucosa barrier and restored the expressions of intestinal tight junction proteins (Claudin-1, Occludin and ZO-1). Subsequently, it reduced LPS levels of DN mice in fecal and serum. Furthermore, 16S rRNA high-throughput sequencing showed that TA modulated gut microbiota dysbiosis and decreased the abundance of Gram-negative bacteria (Proteobacteria and Escherichia-Shigella). This study suggested that TA might exert a beneficial effect on renal interstitial fibrosis in DN mice by modulating gut microbiota dysbiosis, ameliorating impairments in the intestinal mucosa barrier, reducing the production and release of LPS, inhibiting the activation of NLRP3/ASC/Caspase-1 signaling pathway, and repressing renal inflammatory.

Vildagliptin Treatment Ameliorates Renal Interstitial Fibrosis in a Murine Model of Unilateral Ureteral Obstruction.

Renal interstitial fibrosis in mice can be modeled using unilateral ureteral obstruction (UUO). Here, we investigated the anti-fibrotic effects of the dipeptidyl peptidase-4 inhibitor vildagliptin in this model. We found that vildagliptin given in the drinking water at 10.6 ± 1.5 mg/kg/d prevented fibrosis. Mechanistically, UUO was associated with extracellular signal-regulated kinase (ERK) phosphorylation and with the accumulation of the toxic lipid peroxidation product expression of 4-hydroxy-2-nonenal (4-HNE). Both were significantly inhibited by vildagliptin. Similarly, UUO caused reductions in heme oxygenase-1 (HO-1) mRNA in the kidney, whereas interleukin-6 (IL-6) and cyclooxygenase-1 (COX-1) mRNA were increased; these effects were also prevented by vildagliptin. Taking these data together, we propose that vildagliptin reduces renal interstitial fibrosis resulting from UUO by means of its effects on ERK phosphorylation and the amounts of 4-HNE, HO-1, IL-6 and COX-1 in the kidney.

Irisin ameliorates UUO-induced renal interstitial fibrosis through TGF-β1/periostin/MMP-2 signaling pathway.

Renal fibrosis is the most common pathway in progressive kidney diseases. The unilateral ureteral obstruction (UUO) model is used to induce progressive renal fibrosis. We evaluated the effects of irisin on renal interstitial fibrosis in UUO mice. The GSE121190, GSE36496, GSE42303, and GSE96101 datasets were downloaded from the Gene Expression Omnibus (GEO) database. In total, 656 differentially expressed genes (DEGs) were identified in normal and UUO mouse renal samples. Periostin and matrix metalloproteinase-2 (MMP-2) were selected to evaluate the effect of irisin on renal fibrosis in UUO mice. In UUO mice, irisin ameliorated renal function, decreased the expression of periostin and MMP-2, and attenuated epithelial-mesenchymal transition and extracellular matrix deposition in renal tissues. In HK-2 cells, irisin treatment markedly attenuated TGF-β1-induced expression of periostin and MMP-2. Irisin treatment also inhibited TGF-β1-induced epithelial-mesenchymal transition, extracellular matrix formation, and inflammatory responses. These protective effects of irisin were abolished by the overexpression of periostin and MMP-2. In summary, irisin treatment can improve UUO-induced renal interstitial fibrosis through the TGF-β1/periostin/MMP-2 signaling pathway, suggesting that irisin may be used for the treatment of renal interstitial fibrosis.

Qigui-Yishen decoction delays renal fibrosis in mice with chronic kidney disease by regulating TM and PAI-1.

To explore the mechanism of Qigui-Yishen decoction in delaying renal fibrosis in mice by regulating thrombin regulatory protein (Thrombomodulin, TM) and plasminogen activator inhibitor-1 (PAI-1) based on network pharmacology. The active ingredients of Qigui Yishen decoction and their target molecules associated with chronic kidney disease (CKD) were retrieved from websites and databases, sorted out, and screened, and the possible targets of Qigui Yishen decoction for reducing CKD renal fibrosis were predicted and analyzed. Forty Institute of Cancer research (ICR) rats were used to establish a unilateral ureteral obstruction (UUO) model, and divided into several groups: sham operation group, model group, high concentration decoction group (1 g/mL), low concentration decoction group (0.46 g/mL), and benazepril group (0.1 g/mL). At the end of the experiment, the levels of serum creatinine (Scr) and blood urea nitrogen (BUN) were detected. Masson staining was used to observe changes in the renal interstitial fibrosis index. Immunohistochemistry and western blot were used to detect the expressions of TM, PAI-1, transforming growth factor-β1 (TGF-β1) and collagen I (Col I) in kidney tissues, and the differences between groups were compared. Qigui Yishen decoction contains 42 effective ingredients such as sitosterol, mannitol, and quercetin, with 662 drug targets and 16154 disease targets. Analysis revealed 570 potential targets, including TM4SF19, PAIP1, TGF-β1, and Col I-AI. Compared to the sham operation group, all treatment groups exhibited increased Scr and BUN levels (P<0.05) and enhanced renal interstitial fibrosis (P<0.05) after UUO model establishment. Moreover, immunohistochemical results showed significant increases in PAI-1, TGF-β1, and Col I (all P<0.05), and a significant decrease in TM expression (P<0.05). Compared to the model group, the high concentration decoction group, low concentration decoction group and benazepril group had no significant difference in Scr and BUN values (P>0.05), but the renal interstitial fibrosis index was lower (P<0.05). Also, the relative expressions of PAI-1, TGF-β1 and Col I in the kidney tissue of mice were decreased, while the relative expression of TM was increased (P<0.05). Qigi Yishen decoction has the characteristics of multiple components and multiple targets, and can play a role in delaying renal fibrosis by regulating the expression of PAI-1, TGF-β1, Col I, and TM.

LncRNA Meg3 Aggravates Renal Fibrosis Caused by Unilateral Ureteral Obstruction in Rats by Activating the Hedgehog Pathway.

The hedgehog signaling pathway exerts vital functions in regulating epithelial-to-mesenchymal transition (EMT) in renal interstitial fibrosis (RIF). It was reported that lncRNA-maternally expressed gene 3 (lncRNA Meg3) can regulate hepatic fibrosis by regulating the expression of smoothened (Smo) in the hedgehog signaling pathway. However, the specific role of lncRNA Meg3 in renal fibrosis resulting from unilateral ureteral obstruction (UUO) by regulating the hedgehog signaling pathway has not been reported. Hence, this research aimed to expound the effects of lncRNA Meg3 on renal fibrosis induced by UUO in rats via the hedgehog pathway. Peripheral blood was collected from patients with chronic kidney disease (CKD, CKD group) and healthy volunteers (Normal group) at the same period. In addition, 6-week-old male Sprague-Dawley (SD) rats were divided to Sham, UUO, UUO+shRNA Negative control (shNC), and UUO+sh-Meg3 groups, and their kidney tissues and serum were gathered. Next, quantitative real-time polymerase chain reaction (qRT-PCR) was employed for detecting the lncRNA Meg3 expression level in the serum of patients and renal tissue of rats; kits for testing levels of blood urea nitrogen (BUN), creatinine (Cr), hydroxyproline (HYP), and 24-hour urine protein (24-up) in rats of each group; hematoxylin and eosin (HE) staining and Masson staining for observing kidney tissue and renal fibrosis level in rats; western blot for measuring levels of collagen type III (Col III), α-Smooth muscle actin (α-SMA), fibronectin, E-cadherin, sonic hedgehog (Shh), patched (Ptch) protein, smoothened (Smo) protein and glioma-associated oncogene homolog 1 (Gli1) protein expression. LncRNA Meg3 was highly expressed in CKD patients and UUO rats (p < 0.01). In contrast to the UUO+shNC group, knocking down lncRNA Meg3 improved renal injury, relieved pathological renal lesions, and reduced kidney fibrosis and related protein levels. It inhibited the hedgehog pathway in kidney tissues of UUO rats (p < 0.05 and p < 0.01). LncRNA Meg3 can aggravate UUO-induced rat renal fibrosis by activating the hedgehog pathway.

Effect of miR-1297 on Kidney Injury in Rats with Diabetic Nephropathy through the PTEN/PI3K/AKT Pathway.

This study aimed to determine the influence of miR-1297 on kidney injury in rats with diabetic nephropathy (DN) and its causal role. A DN rat model was established through right kidney resection and intraperitoneal injection of streptozotocin (STZ). Sham rats did not undergo right kidney resection or STZ injection. The DN rats were divided into the DN model and antagomiR-1297 treatment groups. Kidney morphology was observed using hematoxylin and eosin staining. Renal function indices, including blood urea nitrogen (BUN), serum creatinine (SCr), and urinary protein, were measured using kits. Levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-1β, superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) were determined through enzyme-linked immunosorbent assay (ELISA). Fibrin (FN), collagen type I (Col I), and α-smooth muscle actin (α-SMA) were assessed through western blotting and real-time reverse transcription-polymerase chain reaction. Apoptosis was detected using terminal deoxynucleotidyl transferase dUTP nick end labeling staining. miR-1297 targets were predicted using bioinformatic software and verified through luciferase reporter assay. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN)/phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) pathway expression was analyzed through western blotting. AntagomiR-1297 reduced BUN (p = 0.005), SCr (p = 0.012), and urine protein (p < 0.001) levels and improved kidney tissue morphology. It prevented renal interstitial fibrosis by decreasing FN, Col I, and α-SMA protein levels (all p < 0.001). AntagomiR-1297 increased SOD (p = 0.001) and GSH-Px (p = 0.002) levels. Additionally, it reduced levels of cell inflammatory factors, including TNF-α, IL-6, and IL-1β (all p < 0.001), and alleviated apoptosis (p < 0.001) in rat kidney tissue with DN. miR-1297 was pinpointed as a target for PTEN. AntagomiR-1297 increased PTEN expression and suppressed PI3K and AKT phosphorylation (all p < 0.001). AntagomiR-1297 can mitigate renal fibrosis, renal inflammation, apoptosis, and oxidative stress levels through the PTEN/PI3K/AKT pathway.

Phosphodiesterase inhibitor ameliorates senescent changes of renal interstitial pericytes in aging kidney.

Pericytes are mesenchymal cells that surround endothelial cells, playing a crucial role in angiogenesis and vessel maturation. Additionally, they are associated with interstitial fibrosis as a major contributor to renal myofibroblasts. In this study, we aim to investigate whether the phosphodiesterase inhibitor, pentoxifylline (PTX), can ameliorate aging-related functional and histological deterioration in the kidney. We subjected aging C57BL/6 mice, dividing into young, aging, and PTX-treated aging groups. Renal function, albuminuria, and histological changes were assessed. Interstitial pericytes were assessed by immunohistochemistry analysis. We examined changes in pericytes in elderly patients using human kidney tissue obtained from healthy kidney donors for kidney transplantation. Invitro experiments with human pericytes and endothelial cells were performed. Aging mice exhibited declined renal function, increased albuminuria, and aging-related histological changes including mesangial expansion and tubulointerstitial fibrosis. Notably, number of pericytes declined in aging kidneys, and myofibroblasts increased. PTX treatment ameliorated albuminuria, histological alterations, and microvascular rarefaction, as well as modulated angiopoietin expression. Invitro experiments showed PTX reduced cellular senescence and inflammation. Human kidney analysis confirmed similar pericyte changes in aging kidneys. The phosphodiesterase inhibitor, PTX preserved microvascular density and improved renal interstitial fibrosis and inflammation in aging mice kidneys. These protective effects were suggested to be associated with the amelioration of pericytes reduction and the transition to myofibroblasts. Additionally, the upregulation of angiopoietin-1 expression may exert potential impacts. To the best of our knowledge, this is the first report on the changes in renal interstitial pericytes in aging human kidneys.