Tubulointerstitial Fibrosis In Diabetic Nephropathy Research Articles

Cordyceps cicadae polysaccharides attenuate diabetic nephropathy via the miR-30a-3p/TRIM16 axis.

The molecular mechanism of the protective effect of Cordyceps cicadae polysaccharides (CCPs) on renal tubulointerstitial fibrosis in diabetic nephropathy (DN) is still unclear. This study aims to further understand the molecular mechanisms behind the therapeutic benefits of CCP on diabetic nephropathy. Mice were randomly assigned into six groups (n = 8). Cordyceps cicadae polysaccharide dissolved in 5% dimethyl sulfoxide was administered by gavage for 12 consecutive weeks. The CCP doses were divided into low, medium, and high, 75, 150, and 300 mg/kg/day, respectively. The efficacy of CCP was determined by assessing the renal function and histological alterations in diabetic db/db mice. The degree of glomerular mesangial dilatation and sclerosis was evaluated using semiquantitative markers. Cell viability, apoptosis, epithelial-mesenchymal transition (EMT), inflammation, oxidative stress, and mitochondrial reactive oxygen species (ROS) in high glucose (HG)-cultured MPC5 podocytes were determined. The interaction of miR-30a-3p and tripartite motif-containing protein 16 (TRIM16) was examined by luciferase reporter assay. Western blotting, reverse transcription-polymerase chain reaction, and immunofluorescence were used to analyze gene and protein expressions. The in vivo findings illustrated that CCP may protect mice with type 2 diabetes from inflammation and oxidative damage (P < 0.05). Furthermore, CCP has a therapeutic value in protecting renal function and morphology in diabetic nephropathy by reversing podocyte EMT. The in vitro results indicated that CCP dose-dependently inhibited HG-induced apoptosis, EMT, inflammation, oxidative stress, and mitochondrial ROS levels in MPC5 podocytes (P < 0.05). Luciferase reporter assay confirmed the interaction between miR-30a-3p and TRIM16 in MPC5 podocytes cultured in high glucose (P < 0.05). The protective effect of CCP on HG-induced MPC5 can be achieved by miR-30a-3p/TRIM16 axis.

Satellite cell-derived exosome-mediated delivery of microRNA-23a/27a/26a cluster ameliorates renal tubulointerstitial fibrosis in diabetic nephropathy

Objective: Renal tubulointerstitial fibrosis (TIF) is the final convergent pathway of diabetic nephropathy (DN), yet no effective targeted therapies currently can prevent the progression of TIF. Here, as a novel and more effective gene therapy approach, the microRNA (miRNA) clusters provide a method to tackle this issue. Methods: Differentially expressed miRNAs in the serum of DN patients were identified by miRNA sequencing (miRNA-seq). We engineered a kidney targeting exosomes (Exos) vector that contained Lamp2b, an exosomal membrane protein gene fused with rabies virus glycoprotein (RVG) peptide. We transfected this vector into muscle satellite cells and then transduced these cells with adenovirus that expresses miR-23a/27a/26a cluster to manufacture the resultant Exos denoted as RVG-miR-23a/27a/26a-Exos. Db/db mice were injected by tail vein with RVG-miR-23a/27a/26a-Exos and the biodistribution of Exos in db/db mice was imaged by the IVIS. RNA sequencing (RNA-seq) and bioinformatic analysis were performed to explore downstream targets of miRNAs. Results: MiRNA-seq results showed that a total of 23 differentially expressed miRNAs with fold-change &gt;2.0 and p value &lt;0.05 were identified (six downregulated and seventeen upregulated) in serum samples from patients with DN, among which miR-23a, miR-26a, and miR-27a were significantly decreased. Then, it was confirmed that miR-23a, miR-26a and miR-27a expression in the serum of patients with DN were reduced by 47%, 57% and 38%, respectively. Moreover, Pearson's correlation analysis showed that miR-23a, miR-26a and miR-27a had strong negative correlation with TIF and renal function decline. Then, we combined miR-23a, miR-26a and miR-27a as an artificially engineered cluster and successfully engineered RVG-miR-23a/27a/26a cluster loaded exosomes derived from muscle satellite cells, which not only enhanced the stability of miR-23a/27a/26a cluster, but also efficiently delivered more miR-23a/27a/26a cluster homing to injured kidney. More importantly, treatment with RVG-miR-23a/27a/26a-Exos significantly ameliorated tubular injury and TIF in mice with DN. Mechanistically, we showed that miR-23a/27a/26a-Exos could potentiate antifibrotic effects by regulating miRNA cluster-targeting Lpp simultaneously, as well as miR-27a-targeting Zbtb20 and miR-26a-targeting Klhl42, respectively. Conclusions: We established a novel kidney-targeting Exo-based delivery system by manipulating the miRNA-23a/27a/26a cluster to ameliorate TIF in DN effectively, providing a promising therapeutic strategy for DN. This work was supported by the National Natural Science Foundation of China (Nos. 81970664, 82000648, 82070735); the Natural Science Foundation of Jiangsu Province (Nos. BK20211385, BK20200363); the 789 Outstanding Talent Program of SAHNMU (Nos. 789ZYRC202080119, 789ZYRC202090251) and the Science and Technology Development Foundation of Nanjing Medical University (No. NMUB2020049). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

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.

Semicarbazide-sensitive amine oxidase inhibition ameliorates albuminuria and glomerulosclerosis but does not improve tubulointerstitial fibrosis in diabetic nephropathy.

Semicarbazide-sensitive amine oxidase (SSAO) is an enzyme with a unique dual function in controlling inflammation as well as reactive oxygen species (ROS) generation. We have demonstrated benefit of SSAO inhibition in acute kidney fibrosis. However the function of SSAO in chronic kidney disease (CKD) and diabetic kidney disease (DKD) is yet to be determined. We aimed to assess the effectiveness of a SSAO inhibitor (SSAOi; PXS-4728A) as an antifibrotic agent using a diabetic model of CKD. Diabetic mice were treated with SSAOi for 24 weeks and outcomes compared with untreated diabetic mice and telmisartan treated animals as a standard of care comparator. Extracellular matrix markers, fibronectin and oxidative stress, were downregulated in diabetic mice treated with SSAOi compared with untreated diabetic mice. Expression of the pan-leukocyte marker CD45 was also supressed by SSAOi. SSAO inhibition in diabetic mice resulted in a significant reduction in glomerulosclerosis and associated albuminuria compared to untreated diabetic mice. However, the effect of SSAO inhibition was less obvious in the tubulointerstitial compartment than in the glomeruli. Therefore, SSAO may be a potential target for diabetic glomerulosclerosis.

Cyclin G2 regulates canonical Wnt signalling via interaction with Dapper1 to attenuate tubulointerstitial fibrosis in diabetic nephropathy.

Cyclin G2 (CCNG2) is an atypical cyclin that inhibits cell cycle progression and is often dysregulated in human cancers. Cyclin G2 in the occurrence and development of diabetic nephropathy (DN), one of the most severe diabetic complications, has not been fully identified. In this study, we investigated the function and regulatory mechanism of cyclin G2 in DN. In vivo studies revealed that a deficiency of cyclin G2 significantly increased albuminuria and promoted tubulointerstitial fibrosis in established DN. Cyclin G2 regulated the expression of fibrosis‐related proteins via the canonical Wnt signalling pathway in renal tubular epithelial cells. Moreover, the binding of cyclin G2 to Dapper1 (Dpr1/DACT1), a protein involved in Wnt signalling, decreased the phosphorylation of Dpr1 at Ser762 by casein kinase 1 (CK1) and suppressed the Wnt signalling pathway. These findings reveal that cyclin G2 can protect against renal injury and fibrosis associated with DN and, thus, is a new target for the prevention and treatment of diabetic complications.

MiR-192 prevents renal tubulointerstitial fibrosis in diabetic nephropathy by targeting Egr1.

Diabetic nephropathy (DN), as the most common and serious diabetic microvascular complication, has become the first cause of end-stage renal disease (ESRD) in many countries and regions. However, the pathogenesis of renal fibrosis during the development of DN remains unknown. The expression levels of miR-192 and early growth response factor 1 (Egr1) were determined by quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) and Western blotting in the renal tissues of Otsuka-Long-Evans-Tokushima-Fatty (OLETF) and Long-Evans-Tokushima-Otsuka (LETO) rats. The diabetic kidney environment was simulated by a high-sugar medium. The expression levels of miR-192 and Egr1 were further measured in the HK-2 cell line. Egr1 was verified as a potential target of miR-192 by using bioinformatics analysis and luciferase activity assay. The expression level of Egr1 was determined by overexpressing and knocking down the expression of miR-192. In addition, Western blotting was used to determine changes in Transforming growth factor-beta 1 (TGF-β1) and fibronectin (FN). Compared with the kidney tissue of LETO rats, the expression of miR-192 was decreased in OLETF rats, whereas the expression of Egr1 was increased. We found the same phenomenon in the HK-2 cell line cultured in the high-glucose medium. Next, miR-192 can act on Egr1 through 3'-UTR to reduce the expression of Egr1 verified by luciferase assay. In addition, the expression levels of TGF-β1 and FN changed significantly, as the expression level of Egr1 increased or decreased. MiR-192 causes degradation of TGF-β1 and FN through targeting Egr1 and affects the progression of TIF and even DN.

Notch1 regulates PTEN expression to exacerbate renal tubulointerstitial fibrosis in diabetic nephropathy by inhibiting autophagy via interactions with Hes1

Diabetic nephropathy (DN) is a serious clinical microvascular complication of diabetes mellitus. DN is characterized by the accumulation of extracellular matrix, resulting in progressive fibrosis leading to the loss of renal function. Notch1 and phosphatase and tensin homolog deleted on chromosome ten (PTEN) signaling have been associated with fibrosis. Autophagy serves as an essential regulator of tubular cellular homeostasis. However, how these molecules control the balance between fibrosis and autophagy, the main homeostatic mechanism regulating fibrosis, is not well understood. This association was confirmed using Notch1-siRNA in vitro, which prevented the increase in Hes1 and restored PTEN expression. In contrast, transfection with pHAGE-Hes1 repressed PTEN promoter-driven luciferase activity, implying a direct relationship between Hes1 and PTEN. The expression of Notch1 and Hes1 was increased in diabetic db/db mice by western blotting; in contrast, the expression of PTEN was decreased. Importantly, the dysregulation of these signaling molecules was associated with an increase in extracellular matrix proteins (Collagen-I and III) and the inhibition of autophagy. Similar results were evident in response to high glucose concentrations in vitro in the NRK-52e cells. Therefore, the high glucose concentrations present in diabetes promote fibrosis through the Notch1 pathway via Hes1, while inhibiting the PTEN and autophagy. In conclusion, the inhibition of PTEN by Notch1/Hes1 in response to high glucose concentration inhibits autophagy, which is associated with the progression of fibrosis. Therefore, these signaling molecules may represent novel therapeutic targets in diabetic nephropathy.

Complement C5a inhibition moderates lipid metabolism and reduces tubulointerstitial fibrosis in diabetic nephropathy.

Complement C5 mediates pro-inflammatory responses in many immune-related renal diseases. Given that the C5a level is elevated in diabetes, we investigated whether activation of C5a/C5aR signalling plays a pathogenic role in diabetic nephropathy (DN) and the therapeutic potential of C5a inhibition for renal fibrosis. Human renal biopsies from patients with DN and control subjects were used for immunohistochemical staining of complement C5 components. Renal function and tubulointerstitial injury were compared between db/m mice, vehicle-treated mice and C5a inhibitor-treated db/db mice. A cell culture model of tubule epithelial cells (HK-2) was used to demonstrate the effect of C5a on the renal fibrotic pathway. Increased levels of C5a, but not of its receptor C5aR, were detected in renal tubules from patients with DN. The intensity of C5a staining was positively correlated with the progression of the disease. In db/db mice, administration of a novel C5a inhibitor, NOX-D21, reduced the serum triglyceride level and attenuated the upregulation of diacylglycerolacyltransferase-1 and sterol-regulatory element binding protein-1 expression and lipid accumulation in diabetic kidney. NOX-D21-treated diabetic mice also had reduced serum blood urea nitrogen and creatinine levels with less glomerular and tubulointerstitial damage. Renal transforming growth factor beta 1 (TGF-β1), fibronectin and collagen type I expressions were reduced by NOX-D21. In HK-2 cells, C5a stimulated TGF-β production through the activation of the PI3K/Akt signalling pathway. Blockade of C5a signalling by NOX-D21 moderates altered lipid metabolism in diabetes and improved tubulointerstitial fibrosis by reduction of lipid accumulation and TGF-β-driven fibrosis in diabetic kidney.

O20 Low molecular weight fucoidan inhibits fibrotic processes in diabetic nephropathy

Introduction The aim of this study was to determine whether low-molecular-weight fucoidan (LMWF) can reduce TGF-β-mediated pathogenesis of tubulointerstitial fibrosis in diabetic nephropathy(DN) using in vitro and in vivo models. Methods LMWF was administered to diabetic rats at a daily dose of 100 mg/kg body weight by intragastric administration. Following 12 weeks of treatment, we measured body and kidney weight, blood glucose, blood pressure, creatinine and urea concentration in serum and urine, urine output, urinary osmolality, and urinary protein excretion. α-SMA, E-cadherin, fibronectin, TGF-β1, and CTGF were measured by Western blot to evaluate the level of epithelial-to-mesenchymal transition (EMT). Results 10 ng/ml TGF-β1 induced EMT and activated downstream pathways in HK2 cells. Treatment with LMWF significantly reversed EMT and dose-dependently inhibited accumulation of extracellular matrix proteins, including connective tissue growth factor and fibronectin. In type 1 and type 2 diabetic rat models, it was found that diabetes resulted in TGF-β signalling activation and increased expression of TGF-β downstream signals in the kidneys. LMWF significantly reduced blood urea nitrogen (BUN) and blood creatinine compared with the untreated type 1 diabetic group. LMWF also significantly reduced BUN and blood creatinine compared with the untreated type 2 diabetic group. Conclusion Our results demonstrate that LMWF treatment attenuated renal dysfunction and fibrogenesis in the kidneys of type 1 and type 2 diabetic rats. The present study suggests that LMWF may be developed as a candidate drug to prevent or inhibit fibrotic progression in DN.

PDGF receptor-β uses Akt/mTORC1 signaling node to promote high glucose-induced renal proximal tubular cell collagen I (α2) expression.

Increased expression of PDGF receptor-β (PDGFRβ) has been shown in renal proximal tubules in mice with diabetes. The core molecular network used by high glucose to induce proximal tubular epithelial cell collagen I (α2) expression is poorly understood. We hypothesized that activation of PDGFRβ by high glucose increases collagen I (α2) production via the Akt/mTORC1 signaling pathway in proximal tubular epithelial cells. Using biochemical and molecular biological techniques, we investigated this hypothesis. We show that high glucose increases activating phosphorylation of the PDGFRβ, resulting in phosphorylation of phosphatidylinositol 3-kinase. A specific inhibitor, JNJ-10198409, and small interfering RNAs targeting PDGFRβ blocked this phosphorylation without having any effect on MEK/Erk1/2 activation. We also found that PDGFRβ regulates high glucose-induced Akt activation, its targets tuberin and PRAS40 phosphorylation, and finally, mTORC1 activation. Furthermore, inhibition of PDGFRβ suppressed high glucose-induced expression of collagen I (α2) in proximal tubular cells. Importantly, expression of constitutively active Akt or mTORC1 reversed these processes. As a mechanism, we found that JNJ and PDGFRβ knockdown inhibited high glucose-stimulated Hif1α expression. Furthermore, overexpression of Hif1α restored expression of collagen I (α2) that was inhibited by PDGFRβ knockdown in high glucose-stimulated cells. Finally, we show increased phosphorylation of PDGFRβ and its association with Akt/mTORC1 activation, Hif1α expression, and elevated collagen I (α2) levels in the renal cortex of mice with diabetes. Our results identify PDGFRβ as a driver in activating Akt/mTORC1 nexus for high glucose-mediated expression of collagen I (α2) in proximal tubular epithelial cells, which contributes to tubulointerstitial fibrosis in diabetic nephropathy.

MicroRNA-130b improves renal tubulointerstitial fibrosis via repression of Snail-induced epithelial-mesenchymal transition in diabetic nephropathy.

MicroRNA-130b (miR-130b) downregulation has been identified in diabetes, but the role and mechanisms for miR-130b in mediating renal tubulointerstitial fibrosis in diabetic nephropathy (DN) remain unknown. We demonstrated that plasma miR-130b downregulation exhibited clinical and biological relevance as it was linked to increased serum creatinine, β2-microglobulin and proteinuria, increased Snail expression and tubulointerstitial fibrosis in renal biopsies of DN patients. MiR-130b inhibitor caused Snail upregulation and enhanced molecular features of epithelial-to-mesenchymal transition (EMT) in high glucose (30 mM) cultured NRK-52E cells. In contrast, miR-130b mimic downregulated Snail expression and increased epithelial hallmarks. Notably, Snail was identified as an miR-130b direct target and inversely correlated with E-CADHERIN expression. Furthermore, the miR-130b-dependent effects were due to Snail suppression that in turn deregulated E-CADHERIN, VIMENTIN, COLLAGEN IV and α-smooth muscle actin (α-SMA), key mediators of EMT. These effects were reproduced in streptozotocin-induced diabetic rats. Thus, we propose a novel role of the miR-130b-SNAIL axis in fostering EMT and progression toward increased tubulointerstitial fibrosis in DN. Detection of plasma miR-130b and its association with SNAIL can be extrapolated to quantifying the severity of renal tubulointerstitial fibrosis. Targeting miR-130b could be evaluated as a potential therapeutic approach for DN.

MicroRNA-130b Improves Renal Tubulointerstitial Fibrosis via Repression of Snail-Induced Epithelial-Mesenchymal Transition in Diabetic Nephropathy

MicroRNA-130b (miR-130b) downregulation has been identified in diabetes, but the role and mechanisms for miR-130b in mediating renal tubulointerstitial fibrosis in diabetic nephropathy (DN) remain unknown. We demonstrated that plasma miR-130b downregulation exhibited clinical and biological relevance as it was linked to increased serum creatinine, β2-microglobulin and proteinuria, increased Snail expression and tubulointerstitial fibrosis in renal biopsies of DN patients. MiR-130b inhibitor caused Snail upregulation and enhanced molecular features of epithelial-to-mesenchymal transition (EMT) in high glucose (30 mM) cultured NRK-52E cells. In contrast, miR-130b mimic downregulated Snail expression and increased epithelial hallmarks. Notably, Snail was identified as an miR-130b direct target and inversely correlated with E-CADHERIN expression. Furthermore, the miR-130b-dependent effects were due to Snail suppression that in turn deregulated E-CADHERIN, VIMENTIN, COLLAGEN IV and α-smooth muscle actin (α-SMA), key mediators of EMT. These effects were reproduced in streptozotocin-induced diabetic rats. Thus, we propose a novel role of the miR-130b-SNAIL axis in fostering EMT and progression toward increased tubulointerstitial fibrosis in DN. Detection of plasma miR-130b and its association with SNAIL can be extrapolated to quantifying the severity of renal tubulointerstitial fibrosis. Targeting miR-130b could be evaluated as a potential therapeutic approach for DN.

Cdk5 Regulates ERK1/2/PPARγ-mediated Renal Tubulointerstitial Fibrosis in Diabetic Nephropathy

Cdk5 Regulates ERK1/2/PPARγ-mediated Renal Tubulointerstitial Fibrosis in Diabetic Nephropathy

Thioredoxin-interacting protein mediates dysfunction of tubular autophagy in diabetic kidneys through inhibiting autophagic flux

Thioredoxin-interacting protein (TXNIP) expression is ubiquitous and is induced by a variety of cellular stresses, including high intracellular glucose. TXNIP is associated with activation of oxidative stress and tubulointerstitial fibrosis in diabetic nephropathy. Autophagy is a major pathway that delivers damaged proteins and organelles to lysosomes to maintain cellular homeostasis. This study aimed to investigate the dysregulation of autophagy and the regulation of TXNIP on autophagy in renal proximal tubular cells (PTCs) under diabetic conditions. The formation of autophagosomes was measured using transmission electron microscopy, and LC3-II, and the effectiveness of autophagic clearance was determined by p62 expression in diabetic kidney and in human PTCs exposed to high glucose (HG). The results collectively demonstrated increased expression of TXNIP, LC3/LC3-II and p62 in renal tubular cells of mice with diabetic nephropathy and in cultured human PTCs exposed to HG (30 mM/l) for 48 h compared with control. The formation of autophagic vacuoles was increased in HG-induced cells. Furthermore, silencing of TXNIP by siRNA transfection reduced autophagic vacuoles and the expression of LC3-II and p62 in human PTCs exposed to HG compared with control and partially reversed the accumulation of LC3-II and p62 induced by bafilomycin A1 (50 nM/l), a pharmacological inhibitor of autophagy which blocks the fusion of autophagosomes with lysosomes and impairs the degradation of LC3-II and p62. Collectively, these results suggest that hyperglycemia leads to dysfunction of autophagy in renal tubular cells and decreases autophagic clearance. HG-induced overexpression of TXNIP may contribute to the dysfunction of tubular autophagy in diabetes.

Collagen VIII influences epithelial phenotypic changes in experimental diabetic nephropathy

Epithelial-to-mesenchymal transition (EMT) is an important mechanism of renal tubulo-interstitial fibrosis in diabetic nephropathy (DN). Inducers of EMT, among others, are transforming growth factor-β(1) (TGF-β(1)) as well as extracellular collagens. In renal cells of diabetic mice and in kidneys of patients with DN, the expression of collagen VIII (gene: Col8α1/α2) is enhanced and characteristic features of DN in streptozotocin (STZ)-induced diabetic Col8α1/α2 knockout-(KO) mice are attenuated compared with diabetic wild-type mice. This study aimed to investigate whether collagen type VIII may influence the induction of EMT. DN was induced in wild-type and Col8α1/α2-KO mice using the established and widely accepted low-dose STZ model [treatment for 5 consecutive days (50 mg/kg)]. Healthy and diabetic mice were analyzed for changes in renal function and the expression of EMT-related genes and proteins. Renal morphology, fibrosis, and various EMT markers were studied in kidneys using immunohistological and molecular biological methods. Knockout of Col8α1/α2 attenuated albuminuria, extracellular matrix production, as well as fibrosis. Furthermore, the kidneys of diabetic Col8α1/α2-KO mice showed a marked reduction in interstitial myofibroblasts, and in tubular cells the inhibition of the expression of epithelial markers as well as the expression of typical mesenchymal markers was reduced. The present study demonstrates that in contrast to diabetic wild-type mice EMT-like changes were attenuated in diabetic Col8α1/α2-KO mice, which indicates that either collagen VIII may be one of the major inducers of EMT-like changes in kidneys of diabetic wild-type mice or/possibly the lack of Col8α1/α2 disrupts TGF-β(1)-induced EMT-like changes.

IHG-1 Promotes Mitochondrial Biogenesis by Stabilizing PGC-1α

Increased expression of Induced-by-High-Glucose 1 (IHG-1) associates with tubulointerstitial fibrosis in diabetic nephropathy. IHG-1 amplifies TGF-β1 signaling, but the functions of this highly-conserved protein are not well understood. IHG-1 contains a putative mitochondrial-localization domain, and here we report that IHG-1 is specifically localized to mitochondria. IHG-1 overexpression increased mitochondrial mass and stabilized peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α). Conversely, inhibition of IHG-1 expression decreased mitochondrial mass, downregulated mitochondrial proteins, and PGC-1α-regulated transcription factors, including nuclear respiratory factor 1 and mitochondrial transcription factor A (TFAM), and reduced activity of the TFAM promoter. In the unilateral ureteral obstruction model, we observed higher PGC-1α protein expression and IHG-1 levels with fibrosis. In a gene-expression database, we noted that renal biopsies of human diabetic nephropathy demonstrated higher expression of genes encoding key mitochondrial proteins, including cytochrome c and manganese superoxide dismutase, compared with control biopsies. In summary, these data suggest that IHG-1 increases mitochondrial biogenesis by promoting PGC-1α-dependent processes, potentially contributing to the pathogenesis of renal fibrosis.

Maternal exposure to carbon black nanoparticle increases collagen type VIII expression in the kidney of offspring

The potential health risks of inhaling nanomaterials are of great concern because of their high specific activity and their unique property of translocation. Earlier studies showed that exposure to nanoparticles through the airway affects both respiratory and extrapulmonary organs. When pregnant mice were exposed to nanoparticles, the respiratory system, the central nervous system and the reproductive system of their offspring were affected. The aim of this study was to assess the effect of maternal exposure to nanoparticles on the offspring, particularly on the kidney. Pregnant ICR mice were exposed to a total of 100 µg of carbon black nanoparticle on the fifth and the ninth days of pregnancy. Samples of blood and kidney tissue were collected from 3-week-old and 12-week-old male offspring mice. Collagen expression was examined by quantitative RT-PCR and immunohistochemistry. Serum levels of creatinine and blood urea nitrogen were examined. Exposure of pregnant ICR mice to carbon black resulted in increased expression of Collagen, type VIII, a1 (Col8a1) in the tubular cells in the kidney of 12-week-old offspring mice but not in 3-week-old ones. The levels of serum creatinine and blood urea nitrogen, indices of renal function, were not different between the groups. These observations were similar to those of tubulointerstitial fibrosis in diabetic nephropathy. These results suggest that maternal exposure to carbon black nanoparticle induces renal abnormalities similar to tubulointerstitial fibrosis in diabetic nephropathy are induced in the kidney of offspring.

Renal expression of proto-oncogene Ets-1 on matrix remodeling in experimental diabetic nephropathy

The molecular mechanisms of glomerulosclerosis and tubulointerstitial fibrosis in diabetic nephropathy (DN) have received scant attention. Ets-1 proto-oncogene plays a role in matrix remodeling by regulating matrix-degrading enzymes. We investigated the possible role of Ets-1 in the pathogenesis of DN. 6-week-old male Sprague–Dawley rats were divided into two experimental groups as follows: control group ( n = 30) and a Diabetes mellitus group ( n = 40) induced by injection of streptozotozin (STZ). The rats were investigated at 1, 4, 8, 12 and 16 weeks after STZ-treatment. By means of immunohistochemistry, the expression of Ets-1 in glomeruli was significantly increased in STZ-treated rat kidneys from week 1 ( P < 0.05) and reached the peak at week 4 ( P < 0.05), followed by a downward trend at subsequent time points. Similarly, the expression of Ets-1 in the tubulointerstitium was also markedly increased from week 1 ( P < 0.05) and reached a maximum at week 8 ( P < 0.05). By double immunostaining, Ets-1-positive cells were frequently found to co-express matrix metalloproteinase-2 (MMP-2) in STZ-treated rat kidneys. Increased expression of tissue inhibitor of metalloproteinase-2 (TIMP-2) coincided with increased expression of α-smooth muscle actin (α-SMA) in STZ-induced DN. A positive relationship was observed between the expression of Ets-1 in glomeruli or tubulointerstitium and the expression of MMP-2 ( P < 0.01; P < 0.01, respectively) in STZ-treated rat kidneys. The ratio of MMP-2 and TIMP-2 in glomeruli or tubulointerstitium was negatively correlated with deposition of type IV collagen ( P < 0.01; P < 0.01, respectively). These findings suggest that Ets-1 may play a critical role in fine-tuning matrix remodeling of STZ-induced DN.

Cinnamaldehyde impairs high glucose-induced hypertrophy in renal interstitial fibroblasts

Cinnamaldehyde is a major and a bioactive compound isolated from the leaves of Cinnamomum osmophloeum kaneh. To explore whether cinnamaldehyde was linked to altered high glucose (HG)-mediated renal tubulointerstitial fibrosis in diabetic nephropathy (DN), the molecular mechanisms of cinnamaldehyde responsible for inhibition of HG-induced hypertrophy in renal interstitial fibroblasts were examined. We found that cinnamaldehyde caused inhibition of HG-induced cellular mitogenesis rather than cell death by either necrosis or apoptosis. There were no changes in caspase 3 activity, cleaved poly(ADP-ribose) polymerase (PARP) protein expression, and mitochondrial cytochrome c release in HG or cinnamaldehyde treatments in these cells. HG-induced extracellular signal-regulated kinase (ERK)/c-Jun N-terminal kinase (JNK)/p38 mitogen-activated protein kinase (MAPK) (but not the Janus kinase 2/signal transducers and activators of transcription) activation was markedly blocked by cinnamaldehyde. The ability of cinnamaldehyde to inhibit HG-induced hypertrophy was verified by the observation that it significantly decreased cell size, cellular hypertrophy index, and protein levels of collagen IV, fibronectin, and α-smooth muscle actin (α-SMA). The results obtained in this study suggest that cinnamaldehyde treatment of renal interstitial fibroblasts that have been stimulated by HG reduces their ability to proliferate and hypertrophy through mechanisms that may be dependent on inactivation of the ERK/JNK/p38 MAPK pathway.

Effect of activin A on tubulointerstitial fibrosis in diabetic nephropathy

The effect of activin A on tubulointerstitial fibrosis in diabetic nephropathy (DN) using streptozotocin (STZ)-induced diabetic rats and high glucose-cultured HK-2 cells was investigated. Male Wistar rats were randomized into a normal control group (NC) and diabetes mellitus group (DM). Diabetes was induced by i.p. injection of STZ. Six rats were respectively killed 4, 8, 12 and 16 weeks after model establishment in each group. The changes of kidney weight/bodyweight (KW/BW), urine albumin excretion rate (AER) and creatinine clearance rate (Ccr) were determined. The morphology of tubulointerstitium was observed by light microscopy. Further biochemical analysis was provided using immunohistochemistry and real-time polymerase chain reaction. The different parameters in high glucose-cultured HK-2 cells were monitored by western blotting or enzyme-linked immunosorbent assay (ELISA) and the intervention of rh-follistatin on them was investigated. Compared with the NC group, there was marked enlargement in the levels of KW/BW, AER, Ccr and interstitial fibrosis index, and the production of P-Smad2/3 and fibronectin in the DM group from 8 to 16 weeks. Activin betaA, mainly located in tubular epithelial cells, was significantly higher in the DM group than that in the NC group throughout the study periods. Follistatin was abundant in the NC group, but was diminished gradually in the DM group. High glucose may facilitate the synthesis of activin betaA, transforming growth factor (TGF)-beta, P-Smad2/3 and fibronectin in HK-2 cells while rh-follistatin inhibited them except TGF-beta. Activin A is involved in tubulointerstitial fibrosis in DN by inducing the production of fibronectin through Smad signal pathway.