The renoprotective effects of berberine via the EP4-Gαs-cAMP signaling pathway in different stages of diabetes in rats

Abstract

Aims: To investigate the renoprotective roles of berberine (BBR) in different stages of diabetic nephropathy (DN) in streptozotocin (STZ)-induced diabetic rats fed a high-sugar and high-fat diet. Methods: Diabetes was induced in mice by intraperitoneal injection of STZ, and the mice were then randomly divided into groups: normal, diabetes, high-sugar and high-fat and BBR (high, median and low dose) groups. The body weight (BW), kidney weight to body weight (KW/BW), blood urea nitrogen, urine total protein to urine creatinine ratio and serum creatinine were measured on different weeks throughout the study. The protein levels of E prostanoid receptor 4 (EP4), Gαs and content of cAMP in the kidney were, respectively, detected by western blot analysis and RIA analysis. Results: In the DN rats, there was remarkable renal damage. BBR restored renal functional parameters, suppressed alterations in histological and ultrastructural changes in the kidney tissues and increased EP4, Gαs and cAMP levels compared with those of the DN model group. In addition, BBR has different therapeutic effects during the different stages of the development of DN, and it works best in the sixth week. Conclusion: These studies demonstrate, for the first time, that BBR exerts renoprotective effects in different stages of DN via EP4- Gαs- AC-cAMP signaling pathway in STZ-induced DN rats fed a high-sugar and high-fat diet.     

Berberine regulates the expression of E-prostanoid receptors in diabetic rats with nephropathy

Diabetic nephropathy (DN) is a major cause of morbidity and mortality in diabetic patients. Effective therapies to prevent the development of this disease and to improve advanced kidney injury are required. Berberine (BBR) has preventive effects on diabetes and its complications. This study is to investigate the effects of BBR on the expression of E-prostanoid receptors (EPs) in rats with high-fat diet and streptozotocin (STZ)-induced DN and underlying molecular mechanisms of BBR on DN rats. DN model was induced in male Sprague–Dawley rats with high-fat diet and low dose of STZ injection. BBR (50, 100, 200 mg/kg/d) were orally administered to rats after STZ injection and conducted for 8 weeks. The levels of interleukin-6 (IL-6) and prostaglandin E₂ (PGE₂) in renal cortex were measured by enzyme-linked immunosorbent assay. Expression of EPs receptors (EP1–EP4) were determined by western blotting. Remarkable renal damage, hyperglycemia and hyperlipidemia were observed in DN rats. BBR could restore renal functional parameters, suppress alterations in histological and ultrastructural changes in the kidney tissues, improve glucose and lipid metabolism disorders, and increase cAMP levels compared with those of DN model group (Wang et al. in Mol Biol Rep 40:2405–2418, 2013). The level of IL-6 and PGE₂ were significantly increased in DN model group compared with normal group, BBR could apparently reduced the level of IL-6 and PGE₂. Furthermore, the expression of EP1 and EP3 were both increased and EP4 was lessened in the DN model group compared with normal group, BBR could down-regulate total protein expression of EP1 and EP3 of renal cortex in DN rats and up-regulate the expression of EP4, and there is no significant difference on the expression of EP2 among all groups. These studies demonstrate, for the first time, that BBR exerts renoprotection in high-fat diet and STZ-induced DN rats by modulating the proteins expression of EPs in EP–G protein–cAMP signaling pathway.     

Renoprotective effects of berberine and its possible molecular mechanisms in combination of high-fat diet and low-dose streptozotocin-induced diabetic rats

Berberine (BBR), an effective compound of Chinese traditional herbal medicine, has preventive effects on diabetes and its complications. In this study, we investigated the therapeutic effects and underlying molecular mechanisms of BBR in rats with high-fat diet and streptozotocin (STZ)-induced diabetic nephropathy model. BBR (50, 100, 200 mg/kg/d) were orally administered to male Sprague–Dawley rats after STZ injection and conducted for 8 weeks. Renal damage was evaluated by kidney weight to body weight ratio (KW/BW), urine microalbumin (UMAlb), urine protein for 24 h (UP24 h), urine creatinine (UCr), and histological examination. Type IV collagen and transforming growth factor-beta1 (TGF-β₁) were detected by immunohistochemistry and ultrastructure of glomeruli was observed. Fasting blood glucose (FBG),serum creatinine (SCr), blood urea nitrogen (BUN), total cholesterol (TC), triglyceride (TG), high-density lipoprotein-cholesterol (HDL-c), low-density lipoprotein-cholesterol (LDL-c) in serum and G protein-coupled receptor kinases (GRKs), cAMP in kidney were measured. Remarkable renal damage, hyperglycemia and hyperlipidemia were observed in DN rats. BBR could restore renal functional parameters, suppress alterations in histological and ultrastructural changes in the kidney tissues, improve glucose and lipid metabolism disorders, and increase cAMP levels compared with those of DN model group. Furthermore, BBR down-regulated total protein expression of GRK2, GRK3 and up-regulated expression of GRK6 of renal cortex in DN rats, but had a slight effects on GRK4 and GRK5. These studies demonstrate, for the first time, that BBR exerts renoprotection in high-fat diet and STZ-induced DN rats by modulating the proteins expression of GRKs in G protein- AC-cAMP signaling pathway.     

Wnt5a promotes renal tubular inflammation in diabetic nephropathy by binding to CD146 through noncanonical Wnt signaling

Immune and inflammatory factors have emerged as key pathophysiological mechanisms in the progression of diabetic renal injury. Noncanonical Wnt5a signaling plays an essential role in obesity- or diabetes-induced metabolic dysfunction and inflammation, but its explicit molecular mechanisms and biological function in diabetic nephropathy (DN) remain unknown. In this study, we found that the expression of Wnt5a and CD146 in the kidney and the level of soluble form of CD146 (sCD146) in serum and urine samples were upregulated in DN patients compared to controls, and this alteration was correlated with the inflammatory process and progression of renal impairment. Blocking the activation of Wnt5a signaling with the Wnt5a antagonist Box5 prevented JNK phosphorylation and high glucose-induced inflammatory responses in db/db mice and high glucose-treated HK-2 cells. Similar effects were observed by silencing Wnt5a with small-interfering RNA (siRNA) in cultured HK-2 cells. Knockdown of CD146 blocked Wnt5a-induced expression of proinflammatory cytokines and activation of JNK, which suggests that CD146 is essential for the activation of the Wnt5a pathway. Finally, we confirmed that Wnt5a directly interacted with CD146 to activate noncanonical Wnt signaling in HK-2 cells. Taken together, our findings suggest that by directly binding to CD146, Wnt5a-induced noncanonical signaling is a contributing mechanism for renal tubular inflammation in diabetic nephropathy. The concentration of sCD146 in serum and urine could be a potential biomarker to predict renal outcomes in DN patients.Subject terms: Kidney diseases, Inflammation     

AANG Prevents Smad3-dependent Diabetic Nephropathy by Restoring Pancreatic β-Cell Development in db/db Mice

Diabetic nephropathy (DN) is a major cause of end-stage kidney disease, where TGF-β1/Smad signaling plays an important role in the disease progression. Our previous studies demonstrated a combination of Traditional Chinese Medicine derived Smad7 agonist Asiatic Acid (AA) and Smad3 inhibitor Naringenin (NG), AANG, effectively suppressed the progression of renal fibrosis in vivo. However, its implication in type-2 diabetic nephropathy (T2DN) is still unexplored. Here, we detected progressive activation of Smad3 but reduction of Smad7 in db/db mice during T2DN development. Therefore, we optimized the dosage and the combination ratio of AANG to achieve a better rebalancing Smad3/Smad7 signaling for treatment of T2DN. Unexpectedly, preventive treatment with combined AANG from week 4 before the development of diabetes and T2DN effectively protected against the onset of T2DN. In contract, these inhibitory effects were lost when db/db mice received the late AANG treatment from 12-24 weeks. Surprisingly, preventive treatment with AANG ameliorated not only T2DN but also the primary disease type-2 diabetes (T2D) with relative normal levels of fasting blood glucose and HbA1c, and largely improving metabolic abnormalities especially on insulin insensitivity and glucose tolerance in db/db mice. Mechanistically, AANG effectively prevented both Smad3-mediated renal fibrosis and NF-κB-driven renal inflammation in the diabetic kidney in vivo and advanced glycation end-products (AGE) stimulated tubular epithelial mTEC cells in vitro. More importantly, we uncovered that preventive treatment with AANG effectively protected against diabetic-associated islet injury via restoring the β cell development in db/db mice. Taken together, we discovered that the early treatment with combined AANG can effectively protect against the development of T2D and T2DN via mechanism associated with protection against Smad3-depenedent islet injury.     

Berberine ameliorates renal injury by regulating G proteins-AC- cAMP signaling in diabetic rats with nephropathy

Diabetic nephropathy (DN) is a progressive kidney disease that is caused by injury to glomerulus and glomerular mesangial cells (MCs) proliferation play a critical role in the pathogenesis of DN. The current studies were undertaken to investigate the protective effects and the possible molecular mechanism of berberine on streptozotocin (STZ)-induced DN rats. Male Wistar rats were randomly assigned to normal control and DN groups of comparable age. Three DN groups received 50, 100 and 200 mg/kg of berberine for 8 weeks via daily intragastrically, respectively. The G proteins-adenylyl cyclase (AC)-cAMP signaling pathway and glomerular MCs proliferation were examined in STZ-induced diabetic rat kidney. Enhanced MCs proliferation and remarkable renal injury were concomitant with activation of Gαi and inhibition of Gαs and cAMP in DN model group. Berberine treatment for 8 weeks abolished the above changes by upregulating the expression of Gαs protein and downregulating the expression of Gαi protein, increasing cAMP level, and inhibiting MCs proliferation compared with model group. Taken together, for the first time, these results demonstrated that berberine can relieve renal injury in DN rats through mediating G proteins-AC-cAMP signaling pathway and inhibiting the abnormal proliferation of MCs by increasing cAMP level, suggesting that berberine could be a potential therapeutic agent for the treatment of DN.     

MAD2B promotes podocyte injury through regulating Numb-dependent Notch 1 pathway in diabetic nephropathy

Rationale: Recent studies have demonstrated that the loss of podocyte is a critical event in diabetic nephropathy (DN). Previously, our group have found that the mitotic arrest deficient protein MAD2B was involved in high glucose (HG)-induced podocyte injury by regulating APC/C activity. However, the exact mechanism of MAD2B implicated in podocyte injury is still lacking.Methods: The experiments were conducted by using kidney tissues from streptozotocin (STZ) induced diabetic mice with or without podocyte-specific deletion of MAD2B and the cultured podocytes exposed to different treatments. Glomerular pathological injury was evaluated by periodic acid-Schiff staining and transmission electron microscopy. The endogenous interaction between MAD2B and Numb was discovered by yeast two-hybrid analysis and co-immunoprecipitation assay. The expressions of MAD2B, Numb and related pathway were detected by western blot, immunochemistry and immunofluorescence.Results: The present study revealed that MAD2B was upregulated in diabetic glomeruli and cultured podocytes under hyperglycemic conditions. Podocyte-specific deletion of MAD2B alleviated podocyte injury and renal function deterioration in mice of diabetic nephropathy. Afterwards, MAD2B was found to interact with Numb, which was downregulated in diabetic glomeruli and HG-stimulated cultured podocytes. Interestingly, MAD2B genetic deletion could partly reverse the decline of Numb in podocytes exposed to HG and in diabetic mice, and the expressions of Numb downstream molecules such as NICD and Hes-1 were decreased accordingly. In addition, overexpression of Numb ameliorated HG-induced podocyte injury.Conclusions: The present findings suggest that upregulated MAD2B expression contributes to Numb depletion and activation of Notch 1 signaling pathway, which ultimately leads to podocyte injury during DN progression.     

Lysophosphatidic acid receptor 1 inhibitor,AM095, attenuates diabetic nephropathy in mice by downregulation of TLR4/NF-κB signaling and NADPH oxidase

Diabetic nephropathy (DN) is one of the major long-term complications of diabetes. Lysophosphatidic acid (LPA) signaling has been implicated in renal fibrosis. In our previous study, we found that the LPA receptor 1/3 (LPAR1/3) antagonist, ki16425, protected against DN in diabetic db/db mice. Here, we investigated the effects of a specific pharmacological inhibitor of LPA receptor 1 (LPA1), AM095, on DN in streptozotocin (STZ)-induced diabetic mice to exclude a possible contribution of LPAR3 inhibition. AM095 treatment significantly reduced albuminuria and the albumin to creatinine ratio and significantly decreased the glomerular volume and tuft area in the treated group compared with the STZ-vehicle group. In the kidney of STZ-induced diabetic mice, the expression of LPAR1 mRNA and protein was positively correlated with oxidative stress. AM095 treatment inhibited LPA-induced reactive oxygen species production and NADPH oxidase expression as well as LPA-induced toll like receptor 4 (TLR4) expression in mesangial cells and in the kidney of STZ-induced diabetic mice. In addition, AM095 treatment suppressed LPA-induced pro-inflammatory cytokines and fibrotic factors expression through downregulation of phosphorylated NFκBp65 and c-Jun N-terminal kinases (JNK) in vitro and in the kidney of STZ-induced diabetic mice. Pharmacological or siRNA inhibition of TLR4 and NADPH oxidase mimicked the effects of AM095 in vitro. In conclusion, AM095 is effective in preventing the pathogenesis of DN by inhibiting TLR4/NF-κB and the NADPH oxidase system, consequently inhibiting the inflammatory signaling cascade in renal tissue of diabetic mice, suggesting that LPAR1 antagonism might provide a potential therapeutic target for DN.     

Berberine regulates mesangial cell proliferation and cell cycle to attenuate diabetic nephropathy through the PI3K/Akt/AS160/GLUT1 signalling pathway

High glucose (HG) is one of the basic factors of diabetic nephropathy (DN), which leads to high morbidity and disability. During DN, the expression of glomerular glucose transporter 1 (GLUT1) increases, but the relationship between HG and GLUT1 is unclear. Glomerular mesangial cells (GMCs) have multiple roles in HG‐induced DN. Here, we report prominent glomerular dysfunction, especially GMC abnormalities, in DN mice, which is closely related to GLUT1 alteration. In vivo studies have shown that BBR can alleviate pathological changes and abnormal renal function indicators of DN mice. In vitro, BBR (30, 60 and 90 μmol/L) not only increased the proportion of G1 phase cells but also reduced the proportion of S phase cells under HG conditions at different times. BBR (60 μmol/L) significantly reduced the expression of PI3K‐p85, p‐Akt, p‐AS160, membrane‐bound GLUT1 and cyclin D1, but had almost no effect on total protein. Furthermore, BBR significantly declined the glucose uptake and retarded cyclin D1‐mediated GMC cell cycle arrest in the G1 phase. This study demonstrated that BBR can inhibit the development of DN, which may be due to BBR inhibiting the PI3K/Akt/AS160/GLUT1 signalling pathway to regulate HG‐induced abnormal GMC proliferation and the cell cycle, supporting BBR as a potential therapeutic drug for DN.     

Berberine Reduces Fibronectin Expression by Suppressing the S1P-S1P2 Receptor Pathway in Experimental Diabetic Nephropathy Models

The accumulation of glomerular extracellular matrix (ECM) is one of the critical pathological characteristics of diabetic renal fibrosis. Fibronectin (FN) is an important constituent of ECM. Our previous studies indicate that the activation of the sphingosine kinase 1 (SphK1)-sphingosine 1- phosphate (S1P) signaling pathway plays a key regulatory role in FN production in glomerular mesangial cells (GMCs) under diabetic condition. Among the five S1P receptors, the activation of S1P2 receptor is the most abundant. Berberine (BBR) treatment also effectively inhibits SphK1 activity and S1P production in the kidneys of diabetic models, thus improving renal injury. Based on these data, we further explored whether BBR could prevent FN production in GMCs under diabetic condition via the S1P2 receptor. Here, we showed that BBR significantly down-regulated the expression of S1P2 receptor in diabetic rat kidneys and GMCs exposed to high glucose (HG) and simultaneously inhibited S1P2 receptor-mediated FN overproduction. Further, BBR also obviously suppressed the activation of NF-κB induced by HG, which was accompanied by reduced S1P2 receptor and FN expression. Taken together, our findings suggest that BBR reduces FN expression by acting on the S1P2 receptor in the mesangium under diabetic condition. The role of BBR in S1P2 receptor expression regulation could closely associate with its inhibitory effect on NF-κB activation.     

Berberine inhibits aldose reductase and oxidative stress in rat mesangial cells cultured under high glucose

Diabetic nephropathy (DN), one of the most serious microvascular complications of diabetes mellitus, is a major cause of end-stage renal disease. Berberine is one of the main constituents of Coptidis rhizoma and Cortex phellodendri. In the present study, we examined effects of berberine (BBR) on renal injury in streptozotocin-induced diabetic rats, and on the changes of aldose reductase (AR) and oxidative stress in cultured rat mesangial cells exposed to high glucose. Fasting blood glucose, blood urea nitrogen, creatinine, and urine protein over 24 h were detected by using the commercially available kits. Cell proliferation, collagen synthesis, aldose reductase (AR), superoxide anion, superoxide dismutase (SOD), and malondialdehyde (MDA) were detected, respectively, by different methods. In streptozotocin-induced diabetic rats, fasting blood glucose, blood urea nitrogen, creatinine, and urine protein over 24 h were significantly decreased in rats treated with 200 mg/kg berberine for 12 weeks compared with diabetic control rats (P < 0.05). This was accompanied by a reduced AR activity and gene expression at both mRNA and protein levels. In cultured rat mesangial cells exposed to high glucose, incubation of BBR significantly decreased cell proliferation, collagen synthesis and AR activity as well as its mRNA and protein levels compared with control cells (P < 0.05). In vitro, BBR also significantly increased SOD activity and decreased superoxide anion and MDA compared with control cells (P < 0.05). These results suggested that BBR could ameliorate renal dysfunction in DN rats, which may be ascribed to inhibition of AR in mesangium, reduction of oxidative stress, and amelioration of extracellular matrix synthesis and cell proliferation. Further studies are warranted to explore the role of AR in DN and the therapeutic implications by AR inhibitors such as BBR.     

DUSP1 recuses diabetic nephropathy via repressing JNK-Mff-mitochondrial fission pathways

Excessive mitochondrial fission has been identified as the pathogenesis of diabetic nephropathy (DN), although the upstream regulatory signal for mitochondrial fission activation in the setting of DN remains unknown. In the current study, we found that dual-specificity protein phosphatase-1 (DUSP1) was actually downregulated by chronic hyperglycemia stimulus. Lower DUSP1 expression was associated with glucose metabolism disorder, renal dysfunction, kidney hypertrophy, renal fibrosis, and glomerular apoptosis. At the molecular level, defective DUSP1 expression activated JNK pathway, and the latter selectively opened mitochondrial fission by modulating mitochondrial fission factor (Mff) phosphorylation. Excessive Mff-related mitochondrial fission evoked mitochondrial oxidative stress, promoted mPTP opening, exacerbated proapoptotic protein leakage into the cytoplasm, and finally initiated mitochondria-dependent cellular apoptosis in the setting of diabetes. However, overexpression of DUSP1 interrupted Mff-related mitochondrial fission, reducing hyperglycemia-mediated mitochondrial damage and thus improving renal function. Overall, we have shown that DUSP1 functions as a novel malefactor in diabetic renal damage that mediates via modifying Mff-related mitochondrial fission. Thus, finding strategies to regulate the balance of the DUSP1-JNK-Mff signaling pathway and mitochondrial homeostasis may be a therapeutic target for treating diabetic nephropathy in clinical practice.     

A bioinformatics and network pharmacology approach to the mechanisms of action of Shenxiao decoction for the treatment of diabetic nephropathy

BackgroundThe epithelial-to-mesenchymal transition (EMT) of renal tubular epithelial cells is the main pathological alteration in diabetic nephropathy (DN). Traditional Chinese medicine (TCM) has been used for the treatment of DN in clinical practice and has been proven to be effective.PurposeThis aim of this study was to shed light on the efficacy of Shenxiao decoction (SXD) on the EMT of renal tubular epithelial cells and the molecular mechanisms of SXD in mice with DN, as well as on the high glucose (HG)- and TGF-β1-induced EMT of NRK-52E and HK-2 cells.Study design and methodsA bioinformatics and network pharmacology method were utilized to construct the active ingredient-target networks of SXD that were responsible for the beneficial effects against DN. The effects of RUNX3 were validated in HG- and TGF-β1-induced EMT processes in NRK-52E and HK-2 cells.ResultsBioinformatics analysis revealed that 122 matching targets were closely associated with the regulation of cell migration and the AGE-RAGE signaling pathway in diabetic complications. The results also revealed that, relative to the mice with DN, the mice in the treatment group had an improved general state and reduced blood glucose levels. The degradation of renal function was ameliorated by SXD. Moreover, the protective effects of SXD were also observed on renal structural changes. Furthermore, SXD suppressed the activation of the transforming growth factor (TGF)-β1/Smad pathway and upregulated the RUNX3 and E-cadherin levels and downregulated the extracellular matrix (ECM) protein levels in mice with DN. SXD was further found to prevent the HG- and TGF-β1-induced EMT processes in NRK-52E and HK-2 cells. Additionally, the overexpression of RUNX3 markedly inhibited the EMT and TGF-β1/Smad pathway induced by HG and TGF-β1 in NRK-52E and HK-2 cells.ConclusionTaken together, these results suggest that SXD maybe alleviate EMT in DN via the inhibition of the TGF-β1/Smad/RUNX3 signaling pathway under hyperglycemic conditions.     

Klotho attenuates diabetic nephropathy in db/db mice and ameliorates high glucose-induced injury of human renal glomerular endothelial cells

Glomerular endothelial cell injury plays an important role in the development and progression of diabetic nephropathy (DN). The expression and function of klotho in glomerular endothelial cells remain unclear. Thus, this study aimed to investigate the expression and the functional role of klotho in DN progression in mice and in high glucose (HG)-induced cell injury of human renal glomerular endothelial cells (HRGECs) and the underlying mechanism. In this study, HRGECs were cultured with media containing HG to induce endothelial cell injury and db/db mice were used as DN model mice. Klotho was overexpressed or knocked down in HRECs to evaluate its role in HG-induced HRGECs injury. klotho-overexpressing adenovirus (rAAV-klotho) was injected into db/db mice via the tail vein to further validate the protective effect of klotho in DN. Decreased klotho expression was observed in DN patients, DN mice, and HG-exposed HRGECs. Furthermore, klotho overexpression significantly abolished the HG-induced HRGECs injury and activation of Wnt/β-catenin pathway and RAAS. In contrast, klotho knockdown exerted the opposite effects. Moreover, klotho attenuated diabetic nephropathy in db/db mice, which was also associated with inhibition of the Wnt/β-catenin pathway and RAAS. In conclusion, klotho attenuates DN in db/db mice and ameliorates HG-induced injury of HRGECs.     

Acetylshikonin from Zicao ameliorates renal dysfunction and fibrosis in diabetic mice by inhibiting TGF-β1/Smad pathway

Diabetic nephropathy (DN) is the major cause of end-stage renal disease in diabetic patients. Zicao, a well-known Chinese traditional medicine, has attracted much attention due to its beneficial effects in various medical fields. In this study, we attempted to investigate the effects and mechanisms of action of acetylshikonin, the main ingredient of Zicao, on renal dysfunction in DN. Our results showed that administration with acetylshikonin not only decreased blood urea nitrogen, urine creatinine and the mean kidney-to-body weight ratio in streptozotocin-induced diabetic mice, but also restored the loss of body weight, whereas the blood glucose was not changed. Masson’s trichrome staining showed that acetylshikonin treatment resulted in a marked decrease in kidney fibrosis from diabetic mice. The increased expression of fibrosis proteins, such as plasminogen activator inhibitor type 1 (PAI-1), connective tissue growth factor, and collagen III and IV, were reduced after acetylshikonin administration. In addition, the expressions of interleukin-1β, interleukin-6, monocyte chemoattractant protein-1, intercellular adhesion molecule 1 and infiltration of macrophages in kidney tissues were decreased in acetylshikonin-treated diabetic mice. Acetylshikonin led to a reduction of transforming growth factor-β1 (TGF-β1) expression and Smad-2/3 phosphorylation, as accompanied by increased Smad7 expression. Furthermore, in vitro treatment with acetylshikonin markedly attenuated TGF-β1-induced the PAI-1, collagen III and IV, and Smad-2/3 phosphorylation in HK2 immortalized human proximal tubule epithelial cells. Acetylshikonin also prevented epithelial-to-mesenchymal transition induced by TGF-β1. Collectively, our study provides evidences that acetylshikonin attenuates renal fibrosis though inhibiting TGF-β1/Smad signaling pathway, suggesting that acetylshikonin may be a novel therapeutic agent for the treatment of DN.     

The role of activation of KАTP channels on hydrogen sulfide induced renoprotective effect on diabetic nephropathy

This work aims to investigate the renal effect of hydrogen sulfide (H₂S), in the experimentally induced diabetic nephropathy, besides the role of activation of АТP-sensitive potassium (K_АTP) channel in that effect. Thirty-two adult male albino rats randomly divided into four groups: Control, streptozotocin-induced diabetic (diabetic nephropathy [DN]), DN+NaHS (the H₂S inducer), and DN+NaHS+Glibenclamide (a selective K_АTP channel blocker) groups. Results showed that kidney functions in the diabetic group improved by NaHS proved by the significant decrease in the measured renal injury markers when compared with the diabetic group with an obvious role of inflammation and oxidative stress. However, the improved kidney functions produced by NaHS was reduced by the combination with Glibenclamide. Glibenclamide combination led also to a significant increase in renal total antioxidant capacity, in addition to a significant decrease in renal total nitric oxide (NO) level. Аccordingly, the results from the present work revealed that the renoprotective effects of H₂S in the case of DN through its effects on renal tissue antioxidants and NO can be partially dependent on activation of K_АTP channels, while its effect on renal tissue proinflammatory cytokines is independent of it.     

Sarsasapogenin alleviates diabetic nephropathy through suppression of chronic inflammation by down-regulating PAR-1: In vivo and in vitro study

BackgroundSarsasapogenin (Sar) shows good effects on diabetic nephropathy (DN) through inhibition of the NLRP3 inflammasome, yet the potential mechanism is not well known.PurposeThis study was designed to explore the regulation of thrombin and/or its receptor protease-activated receptor 1 (PAR-1) on the NLRP3 inflammasome and NF-κB signaling in DN condition, and further expounded the molecular mechanism of Sar on DN.MethodsStreptozotocin-induced diabetic rats were treated by gavage with Sar (0, 20 and 60 mg/kg) for consecutive 10 weeks. Then urine and serum were collected for protein excretion, creatinine, urea nitrogen, and uric acid assay reflecting renal functions, renal tissue sections for periodic acid-Schiff staining and ki67 expression reflecting cell proliferation, and renal cortex for the NLRP3 inflammasome and NF-κB signaling as well as thrombin/PAR-1 signaling. High glucose-cultured human mesangial cells (HMCs) were used to further investigate the effects and mechanisms of Sar.ResultsSar markedly ameliorated the renal functions and mesangial cell proliferation in diabetic rats, and suppressed activation of the NLRP3 inflammasome and NF-κB in renal cortex. Moreover, Sar remarkably down-regulated PAR-1 in protein and mRNA levels but didn't affect thrombin activity in kidney, although thrombin activity was significantly decreased in the renal cortex of diabetic rats. Meanwhile, high glucose induced activation of the NLRP3 inflammasome and NF-κB, and increased PAR-1 expression while didn't change thrombin activity in HMCs; however, Sar co-treatment ameliorated all the above indices. Further studies demonstrated that PAR-1 knockdown attenuated activation of the NLRP3 inflammasome and NF-κB, and Sar addition strengthened these effects in high glucose-cultured HMCs.ConclusionSar relieved DN in rat through inhibition of the NLRP3 inflammasome and NF-κB by down-regulating PAR-1 in kidney.     

丝胶对糖尿病肾病大鼠肾脏TGF-β_1/Smad3信号通路的作用

目的：观察彩色蚕茧提取物—丝胶对糖尿病肾病大鼠肾脏转化生长因子-β1（TGF-β1）和Smad3蛋白表达的影响。方法：60只雄性SD大鼠随机分为5组（n=12）：正常对照组、糖尿病肾病模型组、丝胶治疗组、二甲双胍组和丝胶预防组。模型组、丝胶治疗组、丝胶预防组和二甲双胍组大鼠均建立链脲佐菌素（STZ）致动物模型,以血糖≥16.7 mmol/L作为成模标准;待模型成功建立后,丝胶治疗组大鼠给予丝胶灌胃（2.4 g/（kg.d）,35 d）、二甲双胍组大鼠给予二甲双胍灌胃（55.33 mg/（kg.d）,35 d）,丝胶预防组大鼠于注射STZ前给予同等剂量丝胶灌胃35天。分别检测各组大鼠血糖和肾重/体重;免疫组化染色观察肾脏TGF-β1蛋白的表达;Western blot法观察肾脏Smad3蛋白的表达。结果：与正常对照组大鼠相比,模型组大鼠血糖、肾重/体重和肾脏TGF-β1、Smad3蛋白的表达均明显升高（P〈0.01）。丝胶治疗组、丝胶预防组和二甲双胍组大鼠的血糖和肾脏TGF-β1、Smad3蛋白的表达明显低于模型组（P〈0.01）,且丝胶治疗组、丝胶预防组与二甲双胍组比较无明显差别（P〉0.05）;丝胶治疗组、丝胶预防组和二甲双胍组大鼠的肾重/体重明显低于模型组（P〈0.01）,且丝胶治疗组、丝胶预防组大鼠的肾重/体重明显低于二甲双胍组（P〈0.05）。结论：丝胶可抑制糖尿病肾病大鼠肾脏TGF-β1/Smda3信号通路的激活,减轻肾小球硬化和肾间质纤维化,发挥对糖尿病肾病肾脏损伤的保护和预防保护作用,且作用与二甲双胍相当。     

P53/NRF2 mediates SIRT1's protective effect on diabetic nephropathy

Diabetic nephropathy (DN) is the leading cause of end stage renal disease, posing a severe threat to public health. Previous studies reported the protective role of sirtuin 1 (SIRT1) in DN, encouraging the investigation of more potent and specific SIRT1 activators. SRT2104 is a novel, first-in-class, highly selective small-molecule activator of SIRT1, with its effect and mechanism unknown on DN. To this end, streptozotocin-induced C57BL/6 wild-type (WT) diabetic mice were treated with SRT2104, for 24 weeks. To determine whether SRT2104 acted through inhibition of P53 – a substrate of SIRT1, the P53 activator nutlin3a was administered to the WT diabetic mice in the presence of SRT2104. In order to test whether nuclear factor erythroid 2-related factor 2 (NRF2) – the master of cellular antioxidants – mediated SIRT1 and P53's actions, WT and Nrf2 gene knockout (KO) diabetic mice were treated with SRT2104 or the P53 inhibitor pifithrin-α (PFT-α). In the WT mice, SRT2104 enhanced renal SIRT1 expression and activity, deacetylated P53, and activated NRF2 antioxidant signaling, providing remarkable protection against the DM-induced renal oxidative stress, inflammation, fibrosis, glomerular remodeling and albuminuria. These effects were completely abolished in the presence of nutlin3a. Deletion of the Nrf2 gene completely abrogated the efficacies of SRT2104 and PFT-α in elevating antioxidants and ameliorating DN, despite their abilities to activate SIRT1 and inhibit P53 in the Nrf2 KO mice. The present study reports the beneficial effects of SRT2104 on DN, uncovering a SIRT1/P53/NRF2 pathway that modulates the pathogenesis of DN.