Fibrosis In Diabetic Nephropathy Research Articles

Statement of Retraction: Diminution of microRNA-98 alleviates renal fibrosis in diabetic nephropathy by elevating Nedd4L and inactivating TGF-β/Smad2/3 pathway

This article refers to:RETRACTED ARTICLE: Diminution of microRNA-98 alleviates renal fibrosis in diabetic nephropathy by elevating Nedd4L and inactivating TGF-β/Smad2/3 pathway

Cellular phenotypic transitions in diabetic nephropathy: An update.

Diabetic nephropathy (DN) is a major cause of morbidity and mortality in diabetes and is the most common cause of end stage renal disease (ESRD). Renal fibrosis is the final pathological change in DN. It is widely believed that cellular phenotypic switching is the cause of renal fibrosis in diabetic nephropathy. Several types of kidney cells undergo activation and differentiation and become reprogrammed to express markers of mesenchymal cells or podocyte-like cells. However, the development of targeted therapy for DN has not yet been identified. Here, we discussed the pathophysiologic changes of DN and delineated the possible origins that contribute to myofibroblasts and podocytes through phenotypic transitions. We also highlight the molecular signaling pathways involved in the phenotypic transition, which would provide valuable information for the activation of phenotypic switching and designing effective therapies for DN.

Deciphering the Pharmacological Mechanisms of Qidan Dihuang Decoction in Ameliorating Renal Fibrosis in Diabetic Nephropathy through Experimental Validation In Vitro and In Vivo

Objective QiDan DiHuang decoction (QDD) has been proven to have good efficacy in decreasing albuminuria levels, improving renal function, and inhibiting renal fibrosis in diabetic nephropathy (DN). However, the potential mechanism remains unclear. The purpose of this study was to explore the underlying mechanism of QDD for treating DN in vitro and in vivo. Methods Db/db mice were treated with QDD or saline intragastrically for 12 weeks. Non-diabetic db/m mice were used as controls. Rat renal tubular epithelial cells (NRK-52E) were cultured in high glucose conditions. ATF4 siRNA was transfected into NRK-52E cells. Different indicators were detected via UPLC, RT-PCR, western blotting, cell viability assays and apoptosis, transmission electron microscopy, histology, and immunofluorescence staining. Results Db/db mice experienced severe kidney damage and fibrosis, increased levels of PERK, eIF2α, and ATF4, and suppression of renal autophagy compared with db/m mice. The results showed a significant improvement in glucose intolerance, blood urea nitrogen, urine albumin, serum creatinine, and renal fibrosis in db/db mice with QDD treatment. Meanwhile, the application of QDD resulted in the downregulation of PERK, eIF2α, and ATF4 and the upregulation of autophagy in diabetic kidneys. In vitro, the exposure of NRK-52E cells to high glucose resulted in downregulation of the ratio of LC3-II/LC3-I and upregulation of P62, a reduction in the number of autophagosomes and upregulation of fibronectin (FN), collagen IV and TGF-β1 protein, which was reversed by QDD treatment through inhibiting ATF4 expression. Conclusions Taken together, our results suggest that QDD effectively alleviates diabetic renal injuries and fibrosis by inhibiting the PERK-eIF2α-ATF4 pathway and promoting autophagy in diabetic nephropathy.

Decoy receptor 2 mediates the apoptosis-resistant phenotype of senescent renal tubular cells and accelerates renal fibrosis in diabetic nephropathy

Apoptotic resistance leads to persistent accumulation of senescent cells and sustained expression of a senescence-associated secretory phenotype, playing an essential role in the progression of tissue fibrosis. However, whether senescent renal tubular epithelial cells (RTECs) exhibit an apoptosis-resistant phenotype, and the role of this phenotype in diabetic nephropathy (DN) remain unclear. Our previous study was the first to demonstrate that decoy receptor 2 (DcR2) is associated with apoptotic resistance in senescent RTECs and renal fibrosis. In this study, we aimed to further explore the mechanism of DcR2 in apoptosis-resistant RTECs and renal fibrosis in DN. DcR2 was co-localized with fibrotic markers (α-SMA, collagen IV, fibronectin), senescent marker p16, and antiapoptotic proteins FLIP and Bcl2 but rarely co-localized with caspase 3 or TUNEL. DcR2 overexpression promoted renal fibrosis in mice with streptozotocin (STZ)-induced DN, as evidenced by augmented Masson staining and upregulated expression of fibrotic markers. DcR2 overexpression also enhanced FLIP expression while reducing the expression of pro-apoptotic proteins (caspases 8 and 3) in senescent RTECs, resulting in apoptotic resistance. In contrast, DcR2 knockdown produced the opposite effects in vitro and in vivo. Moreover, quantitative proteomics and co-immunoprecipitation experiments demonstrated that DcR2 interacted with glucose-related protein 78 kDa (GRP78), which has been shown to promote apoptotic resistance in cancer. GRP78 exhibited co-localization with senescent and antiapoptotic markers but was rarely co-expressed with caspase 3 or TUNEL. Additionally, GRP78 knockdown decreased the apoptosis resistance of HG-induced senescent RTECs with upregulated cleaved caspase 3 and increased the percentage of apoptotic RTECs. Mechanistically, DcR2 mediated apoptotic resistance in senescent RTECs by enhancing GRP78–caspase 7 interactions and promoting Akt phosphorylation. Thus, DcR2 mediated the apoptotic resistance of senescent RTECs and renal fibrosis by interacting with GRP78, indicating that targeting the DcR2–GRP78 axis represents a promising therapeutic strategy for DN.

Pyrroloquinoline quinone ameliorates renal fibrosis in diabetic nephropathy by inhibiting the pyroptosis pathway in C57BL/6 mice and human kidney 2 cells.

Diabetic nephropathy (DN), which is characterized by renal fibrosis, is a major complication of diabetes, a disease that afflicted more than 460 million people worldwide in 2019. Pyroptosis is an essential signaling pathway in DN-related injuries, such as renal fibrosis. Pyrroloquinoline quinone (PQQ) is a naturally occurring bioactive compound that protects human kidney 2 (HK-2) cells from oxidative stress-induced damage caused by high glucose concentrations. However, the nature and underlying mechanism of the effect of PQQ on DN-related renal fibrosis remains unclear. In this study, we evaluated whether PQQ has potential protective effects against renal fibrosis due to DN by establishing type 1 diabetes in mice via streptozotocin treatment and then inhibiting their pyroptosis signaling pathway. We found that compared to control mice, the area of renal fibrosis and injury were significantly increased in diabetic mice, and this was accompanied by increased levels of expression of collagen Ⅰ and transforming growth factor-β1; increased concentrations of the inflammatory cytokines, interleukin (IL)-1β, IL-6, and tumor necrosis factor-α; and activation of the pyroptosis pathway components nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3), caspase-1, IL-1β, and IL-18. All of these changes were reversed by PQQ treatment. Analogously, we treated cultured HK-2 cells with a high concentration of glucose (35mmol/L), which caused these cells to exhibit significantly increased concentrations of reactive oxygen species (ROS), phosphorylated (p)-nuclear factor kappa B (NF-κB), p-IkappaB, NLRP3, caspase-1, IL-1β, and IL-18, and the loss of mitochondrial transmembrane potential. However, PQQ treatment significantly blunted these effects. In conclusion, in this study we demonstrated that PQQ attenuates renal fibrosis by alleviating mitochondrial dysfunction, reducing ROS production, and inhibiting the activation of the NF-κB/pyroptosis pathway under conditions of DN and hyperglycemia.

MO405: Leucine-Rich Alpha-2-Glycoprotein as a Potential Marker of Mesangial Cell Proliferation in Immunoglobulin a Nephropathy

Abstract BACKGROUND AND AIMS Leucine-rich alpha-2-glycoprotein (LRG1) is a novel proangiogenic factor involved in the abnormal angiogenesis and renal fibrosis in diabetic nephropathy by potentiating endothelial transforming growth factor-beta/activin receptor-like kinase 1 (TGF-β/ALK1) signalling [1]. TGF-β mediates mesangial matrix accumulation culminating in the development of glomerulosclerosis [2], which also occurs in immunoglobulin A nephropathy (IgAN) patients. Our study aims to evaluate LRG1 as a marker of mesangial cell proliferation in IgAN patients. METHODS: Adults with a morphologically confirmed IgAN by the presence of IgA deposits by immunofluorescence microscopy were included in the study. Diabetes mellitus, oncology, acute inflammation processes were exclusion criteria. Serum LRG1 was detected using ELISA. RESULTS A total of 73 patients with IgAN were included [mean age 41 years, interquartile range (IQR) 35–47; range, 21–65; 65.8% men]. The median estimated glomerular filtration rate (eGFR) was 44 (IQR 24–83, range 4–133) mL/min/1.73m2. Kidney biopsies were analyzed according to the Oxford classification: M0 in 10 (13,7%), M1 in 56 (76.7%) patients. The median serum LRG1 was 49 µg/mL (IQR 40.7–57.7, range 29–133.3). Bivariate analyses showed that plasma LRG1 was negatively associated with eGFR (rho = −0.342, P = 0.003). A linear model showed that GFR decline for 1 mL/min was associated with an increase of LRG1 for 0.194 µg/mL. In IgAN patients with an M1-score LRG1 was higher (P = 0027, Mann–Whitney U test), the area under the ROC curve value for serum LRG1 was 0.739. CONCLUSION Serum LRG1 increases with GFR decline in patients with IgAN and at the same time correlates with mesangial hypercellularity according to Oxford classification. Therefore, LRG1 might be a potential marker of advanced disease in IgAN.

Asiatic acid from Cyclocarya paliurus regulates the autophagy-lysosome system via directly inhibiting TGF-β type I receptor and ameliorates diabetic nephropathy fibrosis.

Diabetic nephropathy (DN) fibrosis is a major cause of end-stage renal disease with unsatisfactory therapy drugs and a low 5-year survival rate. There is a lack of specific and effective treatment drugs. In the present study, we report that asiatic acid (AA), a triterpenic acid found in Cyclocarya paliurus, has good anti-fibrosis activity both in vitro and in vivo. The STZ-induced diabetic model of rats was used to investigate the effects of AA on DN fibrosis. A 15-week AA treatment (10 mg kg-1 or 30 mg kg-1) markedly decreased urine albumin and blood urea nitrogen levels, and ameliorated increased mesangial matrix and glomerular fibrosis. HG + TGF-β1-induced HK-2 cells were applied to evaluate the anti-fibrosis effect of AA. The results revealed AA selectively blocked the interaction of TGF-β type I receptor (TGF-βRI) with Smad3 by binding to TGF-βRI, suppressed the subsequent phosphorylation and nuclear translocation of Smad3, and downregulated the major fibrotic protein expression of collagen I, fibronectin and a-smooth muscle actin (α-SMA), thereby switching the progress of epithelial-mesenchymal transition (EMT). Furthermore, the protein levels of LC3 and LAMP1 were significantly altered by AA administration, implying that the autophagy-lysosome system might be involved in DN fibrosis. However, the anti-fibrosis capacity of AA was partly counteracted by an autophagy-lysosome inhibitor (chloroquine). These findings indicate AA could decrease TGF-β1 secretion and suppress tubulointerstitial fibrosis by directly inhibiting TGF-βR1 and activating the autophagy-lysosome system. Altogether, AA may be a potential candidate drug for preventing DN fibrosis.

Long Noncoding RNA MEG3-205/Let-7a/MyD88 Axis Promotes Renal Inflammation and Fibrosis in Diabetic Nephropathy

Aim: The aim of this study was to investigate the role and mechanism of long noncoding RNA (lncRNA) maternally expressed gene 3 (MEG3)-205 in renal inflammation and fibrosis in diabetic nephropathy (DN). Materials and Methods: lncRNA microarray profiling was used to examine differentially expressed lncRNAs of kidney tissues in db/db mice compared to db/m mice. Mouse mesangial cells (mMCs) were cultured in vitro with advanced glycation end products (AGEs) via transfection with lncRNA MEG3-205 siRNAs or plasmids. The role of lncRNA MEG3-205 in vivo was examined in db/db mice treated with long-acting lncRNA MEG3-205 siRNA. The interaction between lncRNA MEG3-205 and let-7a was investigated using luciferase assay and RNA immunoprecipitation assay. Results: lncRNA MEG3-205 was markedly upregulated in renal tissues of db/db mice, DN patients, and AGEs-treated mesangial cells. Overexpression of lncRNA MEG3-205 promoted the secretion of pro-inflammatory cytokines and synthesis of extracellular matrix proteins in mesangial cells. Both lncRNA MEG3-205 and myeloid differentiation primary-response protein 88 (MyD88) could bind to let-7a, and lncRNA MEG3-205 overexpression can significantly rescue the silencing effect of let-7a on MyD88 protein expression in mMCs. Mechanistically, we identified that lncRNA MEG3-205 could act as a competing endogenous RNA by binding with let-7a and thus regulate MyD88. Knockdown of lncRNA MEG3-205 alleviated albuminuria and attenuated renal inflammation and fibrosis in db/db mice. Conclusion: These findings indicated an important role of the lncRNA MEG3-205/let-7a/MyD88 axis in regulating renal inflammation and fibrosis in DN. Targeting lncRNA MEG3-205 might present a promising therapeutic strategy for DN.

Oridonin Ameliorates Renal Fibrosis in Diabetic Nephropathy by Inhibiting Wnt/β-Catenin Signaling Pathway

Oridonin Ameliorates Renal Fibrosis in Diabetic Nephropathy by Inhibiting Wnt/β-Catenin Signaling Pathway

Exosomal hsa_circ_0125310 promotes cell proliferation and fibrosis in diabetic nephropathy via sponging miR-422a and targeting the IGF1R/p38 axis.

Diabetic nephropathy (DN) is still on the rise worldwide, and millions of patients have to be treated through dialysis or transplant because of kidney failure caused by DN. Recent reports have highlighted circRNAs in the treatment of DN. Herein, we aimed to investigate the mechanism by which high glucose‐induced exo‐circ_0125310 promotes diabetic nephropathy progression. circ_0125310 is highly expressed in diabetic nephropathy and exosomes isolated from high glucose‐induced mesangial cells (MCs). High glucose‐induced exosomes promote the proliferation and fibrosis of MCs. However, results showed that the effects of exosomes on MCs can be reversed by the knockdown of circ_0125310. miR‐422a, which targets IGF1R, was the direct target of circ_0125310. circ_0125310 regulated IGF1R/p38 axis by sponging miR‐422a. Exo‐circ_0125310 increased the luciferase activity of the WT‐IGF1R reporter in the dual‐luciferase reporter gene assays and upregulated the expression level of IGF1R and p38. Finally, in vivo research indicated that the overexpression of circ_0125310 promoted the diabetic nephropathy progression. Above results demonstrated that the high glucose‐induced exo‐circ_0125310 promoted cell proliferation and fibrosis in diabetic nephropathy via sponging miR‐422a and targeting the IGF1R/p38 axis.

Pyridoxamine alleviates high glucose induced fibrosis in renal tubular epithelial cell by inhibiting the activity of TGF-β1/Smad3 signaling pathway

Abstract Background Renal fibrosis is one of the main characteristics of diabetic nephropathy. TGF-β1/Smad3 pathway is expected to reveal the pathogenesis of renal fibrosis in diabetic nephropathy (DN). Pyridoxamine (PM), a natural form of vitamin B6, is a powerful inhibitor of advanced glycation end products (AGEs). PM plays an anti-apoptotic, anti-oxidative stress, and fibrosis role in DN. The purpose of this study was to assess whether PM has a protective effect in renal tubular epithelial and to investigate its possible mechanism. Methods The effects of PM were investigated in HK-2 cells induced by high glucose. HK-2 cells were administered with PM at a dose of 1 mmol/L. Western blot and Realtime PCR were used to detect the expression levels of renal fibrosis related proteins. The possible mechanism of PM was examined by expression of transforming growth factor-β1 (TGF-β1)/Smad3 pathway. Results PM could reduce the expression of Fibronectin (FN) and α-smooth muscle actin (α-SMA) induced by high glucose. PM could also affect the activity of TGF-β1/Smad3 pathway in HK-2 cells. FN and α-SMA were up-regulated by overexpression of Smad3 for 48 h. After adding PM, the levels of FN and α-SMA are significantly decreased. Conclusion Our findings indicate that PM showed a protective effect in HK-2 cells through the inhibition of TGF-β1/Smad3 pathway.

Buyang Huanwu Decoction protects against STZ-induced diabetic nephropathy by inhibiting TGF-\u03b2/Smad3 signaling-mediated renal fibrosis and inflammation

BackgroundBuyang Huanwu Decoction (BHD) is a classical Chinese Medicine formula empirically used for diabetic nephropathy (DN). However, its therapeutic efficacies and the underlying mechanisms remain obscure. In our study, we aim to evaluate the renoprotective effect of BHD on a streptozotocin (STZ)-induced diabetic nephropathy mouse model and explore the potential underlying mechanism in mouse mesangial cells (MCs) treated with high glucose in vitro, followed by screening the active compounds in BHD.MethodsMice were received 50 mg/kg streptozotocin (STZ) or citrate buffer intraperitoneally for 5 consecutive days. BHD was intragastrically administrated for 12 weeks starting from week 4 after the diabetes induction. The quality control and quantitative analysis of BHD were studied by high-performance liquid chromatography (HPLC). Renal function was evaluated by urinary albumin excretion (UAE) using ELISA. The mesangial matrix expansion and renal fibrosis were measured using periodic acid-schiff (PAS) staining and Masson Trichrome staining. Mouse mesangial cells (MCs) were employed to study molecular mechanisms.ResultsWe found that the impaired renal function in diabetic nephropathy was significantly restored by BHD, as indicated by the decreased UAE without affecting the blood glucose level. Consistently, BHD markedly alleviated STZ-induced diabetic glomerulosclerosis and tubulointerstitial injury as shown by PAS staining, accompanied by a reduction of renal inflammation and fibrosis. Mechanistically, BHD inhibited the activation of TGF-β1/Smad3 and NF-κB signaling in diabetic nephropathy while suppressing Arkadia expression and restoring renal Smad7. We further found that calycosin-7-glucoside (CG) was one of the active compounds from BHD, which significantly suppressed high glucose-induced inflammation and fibrosis by inhibiting TGF-β1/Smad3 and NF-κB signaling pathways in mesangial cells.ConclusionBHD could attenuate renal fibrosis and inflammation in STZ-induced diabetic kidneys via inhibiting TGF-β1/Smad3 and NF-κB signaling while suppressing the Arkadia and restoring renal Smad7. CG could be one of the active compounds in BHD to suppress renal inflammation and fibrosis in diabetic nephropathy.

PP2 Ameliorates Renal Fibrosis by Regulating the NF-κB/COX-2 and PPARγ/UCP2 Pathway in Diabetic Mice.

Renal fibrosis is characterized by glomerulosclerosis and tubulointerstitial fibrosis in diabetic nephropathy (DN). We aimed to evaluate the effects of PP2 on renal fibrosis of DN. GSE33744 and GSE86300 were downloaded from the GEO database. Firstly, 839 DEGs were identified between nondiabetic and diabetic mice renal glomerular samples. COX-2 was selected to assess the effects of PP2 on renal glomerulosclerosis. In db/db mice, PP2 decreased the expression of COX-2, phosphorylated p65, and fibrotic proteins, accompanied with attenuated renal glomerulosclerosis. In cultured glomerular mesangial cells, high glucose- (HG-) induced p65 phosphorylation and COX-2 expression were attenuated by PP2 or NF-κB inhibitor PDTC. PP2, PDTC, or COX-2 inhibitor NS-398 ameliorated abnormal proliferation and expression of fibrotic proteins induced by HG. Secondly, 238 DEGs were identified between nondiabetic and diabetic mice renal cortex samples. UCP2 was selected to assess the effects of PP2 on renal tubulointerstitial fibrosis. In db/db mice, PP2 decreased the expression of PPARγ and UCP2, accompanied with attenuated renal tubulointerstitial fibrosis and EMT. In cultured proximal tubular cells, HG-induced PPARγ and UCP2 expression was inhibited by PP2 or PPARγ antagonist GW9662. PP2, GW9662, or UCP2 shRNA ameliorated HG-induced EMT. These results indicated that PP2 ameliorated renal fibrosis in diabetic mice.

MUC1 Promotes Mesangial Cell Proliferation and Kidney Fibrosis in Diabetic Nephropathy Through Activating STAT and β-Catenin Signal Pathway.

Diabetic nephropathy (DN) is a complication of diabetes, which leads to most end-stage kidney diseases and threatens health of patients. Mucin 1 (MUC1) is a heterodimeric oncoprotein, which is abnormally expressed in tumors and hematologic diseases. The aim of this study is to clarify the mechanism and role of MUC1 in DN. The mesangial cells (MCs) suffered from high glucose (HG) treatment to mimic DN in vitro. The cell proliferation was detected by Cell Counting Kit-8 assay and 5-ethynyl-2-deoxyuridine (EdU) staining assay. The expression of MUC1 and fibrosis markers: fibronectin, collagen I, and collagen IV were assessed by western blot. In this study, we demonstrated that HG treatment induced MUC1 expression in MCs. With knockdown of MUC1 or overexpressed MUC1 in MCs, the results indicated that knockdown of MUC1 inhibited MCs proliferation and reduced kidney fibrosis markers expression, including fibronectin, collagen I, and collagen IV, whereas overexpression of MUC1 led to opposite results. Mechanically, MUC1 activated signal transducers and activators of transcription (STAT) and β-catenin signal pathway. After added AG490 (STAT inhibitor) or FH535 (β-catenin inhibitor), blocking STAT3 and β-catenin signal pathway attenuated MUC1-induced cell proliferation and fibronectin production in MCs. Finally, knockdown of MUC1 attenuated DN-induced kidney fibrosis in db/db mice. Therapeutic target for DN. In conclusion, MUC1 promotes MCs proliferation and kidney fibrosis in DN through activating STAT and β-catenin signal pathway, which can help to provide a novel therapeutic target for DN.

Sp1-Induced lncRNA Rmrp Promotes Mesangial Cell Proliferation and Fibrosis in Diabetic Nephropathy by Modulating the miR-1a-3p/JunD Pathway.

Diabetic nephropathy (DN) is a serious complication of diabetes mellitus. Long non-coding RNAs (lncRNAs) are regulators in DN progression. However, the regulatory mechanisms of multiple lncRNAs in DN remain to be determined. Our aim was to investigate the function and molecular mechanism of lncRNA RNA component of mitochondrial RNAase P (Rmrp) in DN. Here, we observed that the expression of Rmrp was up-regulated in the kidney of db/db DN mice and high glucose induced glomerular mesangial cells (MC). More importantly, the abnormal transcription of Rmrp was induced by nuclear transcription factor Sp1, which promotes the proliferation and production of fibrotic markers in MC. Subsequently, we screened the miRNAs related to Rmrp and found that Rmrp and miR-1a-3p are co-localized at the subcellular level of MC, and Rmrp could directly binds to miR-1a-3p. Further mechanism research demonstrated that the elevated miR-1a-3p significantly attenuated the proliferation and fibrosis-promoting effects induced by up-regulation of Rmrp. At the same time, we also investigated that miR-1a-3p can directly bind to Jun D proto-oncogene (JunD), thereby regulating the protein level of JunD. Rmrp-induced proliferation and fibrogenesis were reversed by co-transfection with JunD siRNA. In summary, Sp1 induced lncRNA Rmrp could drive the expression of JunD via sponging miR-1a-3p in DN progression.

Podocyte Glucocorticoid Receptors Are Essential for Glomerular Endothelial Cell Homeostasis in Diabetes Mellitus.

BackgroundProteinuria and glomerular segmental fibrosis are inevitable complications of diabetic nephropathy though their mechanisms are poorly understood. Understanding the clinical characteristics and pathogenesis of proteinuria and glomerular segmental fibrosis in diabetic nephropathy is, therefore, urgently needed for patient management of this severe disease.Methods and ResultsDiabetes mellitus was induced in podocyte‐specific glucocorticoid receptor knockout (GRPKO) mice and control littermates by administration of streptozotocin. Primary podocytes were isolated and subjected to analysis of Wnt signaling and fatty acid metabolism. Conditioned media from primary podocytes was transferred to glomerular endothelial cells. Histologic analysis of kidneys from diabetic GRPKO mice showed worsened fibrosis, increased collagen deposition, and glomerulomegaly indicating severe glomerular fibrosis. Higher expression of transforming growth factor‐βR1 and β‐catenin and suppressed expression of carnitine palmitoyltransferase 1A in nephrin‐positive cells were found in the kidneys of diabetic GRPKO mice. Podocytes isolated from diabetic GRPKO mice demonstrated significantly higher profibrotic gene expression and suppressed fatty acid oxidation compared with controls. Administration of a Wnt inhibitor significantly improved the fibrotic features in GRPKO mice. The glomerular endothelium of diabetic GRPKO mice demonstrated the features of endothelial‐to‐mesenchymal transition. Moreover, endothelial cells treated with conditioned media from podocytes lacking GR showed increased expression of α‐smooth muscle actin, transforming growth factor‐βR1 and β‐catenin levels.ConclusionsThese data demonstrate that loss of podocyte GR leads to upregulation of Wnt signaling and disruption in fatty acid metabolism. Podocyte–endothelial cell crosstalk, mediated through GR, is important for glomerular homeostasis, and its disruption likely contributes to diabetic nephropathy.

LncRNA SOX2OT alleviates mesangial cell proliferation and fibrosis in diabetic nephropathy via Akt/mTOR-mediated autophagy

BackgroundAccumulating evidences have demonstrated that long non-coding RNAs (lncRNAs) are involved in the pathophysiology of diabetic nephropathy (DN). lncRNA SOX2OT plays an essential role in many diseases, including diabetes. Herein, we aim to investigate the underlying mechanism of lncRNA SOX2OT in DN pathogenesis.MethodsStreptozotocin-induced DN mouse models and high glucose-induced mouse mesangial cells were constructed to examine the expression pattern of lncRNA SOX2OT. The activation of autophagy was evaluated using immunohistochemistry, immunofluorescence and western blot analysis, respectively. SOX2OT overexpressing plasmid was applied to further verify the functional role of SOX2OT in DN pathogenesis. CCK-8 and EDU assays were performed to the proliferation of mesangial cells. Additionally, rapamycin, the inhibitor of mTOR signaling, was used to further clarify whether SOX2OT controls DN development through Akt/mTOR pathway.ResultslncRNA SOX2OT was markedly down-regulated both in streptozotocin-induced DN mice and high glucose-induced mouse mesangial cells. Moreover, overexpression of lncRNA SOX2OT was able to diminish the suppression of autophagy and alleviate DN-induced renal injury. Functionally, CCK-8 and EDU assays indicated that lncRNA SOX2OT overexpression significantly suppressed the proliferation and fibrosis of mesangial cells. Additionally, an obvious inhibition of Akt/mTOR was also observed with lncRNA SOX2OT overexpression, which was then further verified in vivo.ConclusionIn summary, we demonstrated that lncRNA SOX2OT alleviates the pathogenesis of DN via regulating Akt/mTOR-mediated autophagy, which may provide a novel target for DN therapy.

Overexpression of mm9_circ_013935 alleviates renal inflammation and fibrosis in diabetic nephropathy via the miR-153-3p/NFIC axis.

Circ-RBM4 (mm9_circ_013935) has been revealed to have low expression in the renal tissues of diabetic nephropathy (DN) mice, and its underlying regulatory mechanism remains unexplored. The high glucose (HG) - treated mouse podocytes were used to establish DN cell models. A cell counting kit-8 assay was used to examine the viability of mouse podocytes. The expression of proteins related to fibrosis (collagen I, collagen III, fibronectin) was detected using Western blot. The concentration of inflammation cytokines (tumor necrosis factor α, interleukin 1β (IL-1β), IL-8) in mouse podocytes was assessed by ELISA. The interaction between genes was explored by luciferase reporter assays. HG treatment decreased the viability and elevated the expression of fibrosis and inflammation factors in mouse podocytes. Circ-RBM4 expression was downregulated in HG-treated mouse podocytes. Circ-RBM4 overexpression reversed HG-induced increase in levels of proteins related to fibrosis and the concentration of inflammation factors. The miR-153-3p was revealed to bind with circ-RBM4 and directly targeted nuclear factor I/C (NFIC) in mouse podocytes. Rescue assays indicated that circ-RBM4 attenuated HG-induced fibrosis and inflammation response in mouse podocytes by inhibiting miR-153-3p expression or upregulating NFIC expression. Circ-RBM4 alleviated the renal inflammation and renal fibrosis in DN by targeting the miR-153-3p/NFIC axis.

385-P: Exosomal MiR-146a-5p from Macrophages Inhibits Diabetic Nephropathy Fibrosis by Targeting Smad4

Diabetic kidney disease(DKD) has become the leading cause of end-stage renal disease and the main cause of death of chronic kidney disease. Renal interstitial fibrosis is an important pathological change of DKD, and chronic microinflammation also plays a key role in the occurrence and development of Diabetic Nephropathy(DN). Macrophages are one of the main regulatory cells of inflammation and infiltrate in the kidney tissue of early DN. In addition to producing injury mediators, activated macrophages can also affect the kidney through other mechanisms. Interestingly, there are a large amount of exosomes generated by macrophage under hyperglycemia condition and it is not clear whether the exosomes has any effect on glomerular mesangial cell fibrosis in patients with diabetes. Here, we use a diabetic nephropathy db/db mouse model to determine the effect of macrophage-produced exosomes by miR-146a-5p overexpression or down-regulation on mesangial cell fibrosis. with high glucose, Macrophage proliferation is promoted in mice and miR-146a-5p expression is downregulated in both macrophages and glomerular mesangial cells. we confirmed that Smad4 is the target gene of miR-146a-5p through Luciferase reporter gene system. Fibrosis-associated col I, α-SMA, and Smad4 are upregulated in glomerular mesangial cells. In exosomes, we observed miR-146a-5p is downregulated in high glucose than low glucose. We use the exosomes from the macrophages that are transfected with miR-146a-5p mimics or inhibitor to culture the glomerular mesangial cells. Exosomes from miR-146a-5p mimics significantly downregulated glomerular Mesangial cell fibrosis-related factor α-SMA, col I protein, and Smad4. Exosomes from miR-146a-5p inhibition upregulated these factors. Finally, we found macrophage exosomes from miR-146a-5p overexpression can inhibit glomerular mesangial cell fibrosis, which could provide a potential treatment strategy for DKD. Disclosure W. Zhang: None. H. Jiang: None. Y. Ma: None. H. Peng: None. J. Liu: None. J. Yuan: None. K. Hu: None. B. Zhao: None. H. Wu: None.

MiR-214 promotes renal fibrosis in diabetic nephropathy via targeting SOCS1

Purpose: To elucidate how miR-214 regulates the pathogenesis of diabetic nephropathy (DN).
 Methods: The extent of fibrosis in DN mice kidneys was examined using Masson’s staining. Quantitative polymerase chain reaction (qPCR) was used to determine the levels of miR-214. Dual luciferase reporter assay was used to identify the target of miR-214. The expression of fibrosis marker proteins of high glucose-stimulated NRK-52E cells transfected with miR-214 was determined using western blotting.
 Results: Fibrosis in renal tissue of DN mice was significantly increased and miR-214 was upregulated (p < 0.001). Suppressor of cytokine signaling 1 protein (SOCS1) was the target gene of miR-214, and overexpression of miR-214 promoted fibrosis (p < 0.05, p < 0.001). On the other hand, overexpression of SOCS1 inhibited this process, indicating that miR-214 promoted fibrosis via targeting SOCS1 (p < 0.001). Finally, inhibition of miR-214 c ameliorated renal fibrosis in DN mice (p < 0.01, p < 0.001).
 Conclusions: MiR-214 is upregulated in db/db DN mice kidney tissue; miR-214 regulates renal fibrosis in DN mice by targeting SOCS1.