HG-induced Cell Research Articles

Schisandrin B ameliorates high-glucose-induced vascular endothelial cells injury by regulating the Noxa/Hsp27/NF-κB signaling pathway.

To address the molecular mechanism of the anti-inflammation effects of schisandrin B (Sch B) in atherosclerosis, we examined injured HMEC-1, HBMEC, and HUVEC-12 cells induced by high glucose (HG). Western blot was performed to detect the levels of the proteins Hsp27, Noxa, TLR5, p-IκBα, and p-p65 in HG-induced cells, while ELISA was used to analyze the inflammatory cytokines TNF-α, IL-6, MCP-1, and IL-1β in cells with Hsp27 or Noxa stable expression. Overexpression of Hsp27 upregulated the inflammatory cytokines and the release of IκBα, promoted transportation of p65 into the nucleus, and lastly, affected the inflammation process, while Sch B counteracted the upregulation. In addition, the effect of Noxa overexpression, which is different from Hsp27 overexpression, was consistent with that of Sch B treatment. Sch B may inhibit the inflammatory cascade and alleviate the injury to HMEC-1, HBMEC, and HUEVC-12 cells caused by HG by regulating the Noxa/Hsp27/NF-κB signaling pathway.

Exosomes derived from monocytes and from endothelial cells mediate monocyte and endothelial cell activation under high d-glucose conditions

Diabetes mellitus type 2 (DMT2) is characterized by hyperglycemia and associated with low grade inflammation affecting both endothelial cells and monocytes. Exosomes are nanovesicles, allow communication between endothelial cells and monocytes and have been associated with diabetic complications. In this study we evaluated whether high glucose can activate monocytes and endothelial cells and whether exosomes play a role in this activation. Moreover, we studied whether endothelial cells and monocytes communicate with each other via exosomes under high and basal glncubation. In the first experiment, monomac 6 cells (MM6) were exposed to high glucose (HG; 25 mmol/L) or to exosomes from MM6 exposed to HG (exoMM6-HG) or basal glucose (5.5 mmol/L) (exoMM6-BG). In the second experiment, MM6 were exposed to exosomes from human umbilical vein endothelial cells (HUVECs) and HUVECs to exosomes from MM6. In the third experiment, MM6 and HUVECs were exposed to a mixture of exosomes from MM6 and HUVECs (exoMix). Cell activation was evaluated by measuring the protein surface expression of intracellular adhesion molecule-1 (ICAM-1) by flow cytometry. HG increased ICAM-1 expression in MM6 and monocytic exosomes from HG or BG shown similar effect in HG and BG MM6 cells. Exosomes from HUVECs increased ICAM-1 expression in MM6 cells, incubated under HG or BG, while also exosomes from MM6 increased ICAM-1 expression in HUVECs incubated under HG or BG. The combination of exosomes from both cell types (exoMixHG or exoMixBG) also increased ICAM-1 expression in both type cells in most conditions. However, the exoMixBG reversed the effect of HG in both MM6 and HUVECs. Our results show that HG activated monocytes and endothelial cells and that exosomes play a role in this HG-induced cell ICAM-1 expression. We hypothesize that during DMT2, exosomes may act as a communication mechanism between monocytes and endothelial cells, inducing and maintaining activating of both cell types in the presence of high glucose.

RETRACTED: Baicalin protects human retinal pigment epithelial cell lines against high glucose-induced cell injury by up-regulation of microRNA-145

Diabetic retinopathy (DR) is a common complication of diabetes mellitus, which is a major reason of blindness. Baicalin (BAI) is a flavonoid extracted from Scutellaria baicalensis, whose pharmacological characterizes have been widely reported in various diseases. However, it remains unclear the effect of BAI on DR. The study aimed to confirm the protective effect of BAI on DR. ARPE-19 cells and HRMECs were exposed to the high glucose (HG) environment to construct a cell injury model. After treatment with HG and BAI, cell viability, apoptosis, inflammatory cytokines and ROS generations were determined in ARPE-19 cells and HRMECs. Subsequently, microRNA-145 (miR-145) inhibitor and its negative control were transfected into ARPE-19 cells, and the regulatory effects on HG-and BAI-co-treated cells were detected. NF-κB and p38MAPK signaling pathways were finally examined to state the underling mechanisms. HG treatment significantly induced ARPE-19 cells and HRMECs injury in vitro. BAI significantly promoted cell proliferation, reduced apoptosis, as well as inhibited the release of IL-1β, IL-6, IL-8 and ROS level in HG-injured ARPE-19 cells and HRMECs. Additionally, the expression level of miR-145 was up-regulated in HG-and BAI-co-treated cells. More importantly, miR-145 inhibition reversed the protective effect of BAI on HG-injured ARPE-19 cells. Besides, we observed that BAI inhibited the activations of NF-κB and p38MAPK pathways by up-regulating miR-145. Results demonstrated that BAI exhibited the protective effect against HG-induced cell injury by up-regulation of miR-145.

High glucose-induced circHIPK3 downregulation mediates endothelial cell injury.

High glucose (HG) induces vascular endothelial cell injury. However, the underlying mechanisms are poorly understood. Circular RNA HIPK3 (circHIPK3) is a highly conserved non-coding RNA. Here we show that circHIPK3 is downregulated in HG-treated human umbilical vein endothelial cells (HUVECs) and in primary aortic endothelial cells (HAECs) from diabetic patients. In both HUVECs and HAECs, lentivirus-mediated circHIPK3 overexpression inhibited HG-induced cell death and apoptosis. Contrarily, circHIPK3 silencing by targeted siRNA exacerbated HG-induced endothelial cell death and apoptosis. Further, circHIPK3 downregulation by HG caused microRNA-124 (miR-124) accumulation in HUVECs and HAECs. On the contrary, miR-124 inhibition by the adeno-associated virus (AAV)-packed miR-124 inhibitor protected endothelial cells from HG. Together, circHIPK3 downregulation mediates HG-induced endothelial cell injury. Targeting circHIPK3-miR-124 pathway could potentially be a novel approach for the treatment of diabetic-associated vascular injury.

Astilbin Inhibits High Glucose-Induced Inflammation and Extracellular Matrix Accumulation by Suppressing the TLR4/MyD88/NF-κB Pathway in Rat Glomerular Mesangial Cells.

Diabetic nephropathy (DN) is characterized by inflammatory responses and extracellular matrix (ECM) accumulation. Astilbin is an active natural compound and possesses anti-inflammatory activity. The aim of this study was to evaluate the anti-inflammatory effect of astilbin on high glucose (HG)-induced glomerular mesangial cells and the potential mechanisms. The results showed that HG induced cell proliferation of HBZY-1 cells in a time-dependent manner, and astilbin inhibited HG-induced cell proliferation. The expression and secretion of inflammatory cytokines, including interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α), and ECM components, including collagen IV (Col IV) and fibronectin (FN), were induced by HG. Moreover, TGF-β1 and CTGF were also induced by HG. The induction by HG on inflammatory response and ECM accumulation was inhibited after astilbin treatment. Astilbin treatment also attenuated HG-induced decrease in expression of matrix metalloproteinase (MMP)-2 and MMP-9. The TLR4/MyD88/NF-κB pathway was activated by HG, and the inhibitor of TLR4 exhibited the same effect to astilbin on reversing the induction of HG. TLR4 overexpression attenuated the effect of astilbin on HG-induced inflammatory cytokine production and ECM accumulation. The results suggested that astilbin attenuated inflammation and ECM accumulation in HG-induced rat glomerular mesangial cells via inhibiting the TLR4/MyD88/NF-κB pathway. This work provided evidence that astilbin can be considered as a potential candidate for DN therapy.

Astilbin inhibits high glucose-induced autophagy and apoptosis through the PI3K/Akt pathway in human proximal tubular epithelial cells

Diabetic nephropathy (DN) is the leading cause of end-stage renal disease. It has been found that astilbin, a flavonoid compound, exerts a protective effect on DN. However, the role of astilbin in autophagy during DN is unknown. The human proximal tubular epithelial cells (HK-2 cells) were treated with high glucose (HG, 30 mM) in the presence or absence of astilbin. Cell viability was measured by MTT assay. The autophagy was determined by detecting the expression of LC3-II and p62 using western blot. The cell apoptosis was evaluated by detecting the apoptosis rate, caspase-3 activity, and the expression of Bcl-2 and Bax. The expression levels of protein kinase B (Akt) and p-Akt were detected by western blot. To determine whether the phosphatidylinositol-3-kinase (PI3K)/Akt pathway was involved in the effect of astilbin, cells were treated with the inhibitor of Akt, LY294002. We found that astilbin (10 and 20 μM) did not affect the viability of HK-2 cells, but attenuated HG-induced cell viability. Astilbin attenuated HG-induced autophagy and apoptosis in HK-2 cells. The expression of p-Akt was inhibited by HG treatment, while the inhibitory effect of HG was attenuated by astilbin. Inhibition of the PI3K/Akt signaling resisted the effect of astilbin on HG-induced apoptosis and autophagy. In conclusion, astilbin attenuated HG-induced autophagy and apoptosis in HK-2 cells through the PI3K/Akt pathway. The results indicated that astilbin might be a new therapeutic agent and be useful for improving clinical management of DN.

Exendin-4 Ameliorates High-Glucose-Induced Fibrosis by Inhibiting the Secretion of miR-192 from Injured Renal Tubular Epithelial Cells

Extracellular vesicles (EVs), which contain microRNA (miRNA), constitute a novel means of cell communication that may contribute to the inevitable expansion of renal fibrosis during diabetic kidney disease (DKD). Exendin-4 is effective for treating DKD through its action on GLP-1R. However, the effect of exendin-4 on EVs miRNA expression and renal cell communication during the development of DKD remains unknown. In this study, we found that EVs derived from exendin-4 and high-glucose (Ex-HG)-pretreated HK-2 cells, which was taken up by normal HK-2 cells, resulted in decreased levels of FN and Col-I compared with those treated with HG pretreated HK-2 cells derived EVs. Furthermore, we found this may contribute to a decrease in miR-192 in both HK-2 cells and EVs after exendin-4 treatment in a p53-dependent manner. Besides, we found that the amelioration of renal fibrosis by exendin-4 occurs through a miR-192-GLP-1R pathway, indicating a new pathway by which exendin-4 regulates GLP-1R. The results of this study suggest that exendin-4 can inhibit the transfer of EVs miR-192 from HG-induced renal tubular epithelial cells to normal cells, thus inhibiting GLP-1R down-regulation and protecting renal cells. This study reports a new mechanism by which exendin-4 exerts a protective effect against DKD. Disclosure Y. Jia: None. Y. Xue: None.

Rapeseed protein-derived antioxidant peptide RAP alleviates renal fibrosis through MAPK/NF-κB signaling pathways in diabetic nephropathy

IntroductionKidney fibrosis is the main pathologic change in diabetic nephropathy (DN), which is the major cause of end-stage renal disease. Current therapeutic strategies slow down but cannot reverse the progression of renal dysfunction in DN. Plant-derived bioactive peptides in foodstuffs are widely used in many fields because of their potential pharmaceutical and nutraceutical benefits. However, this type of peptide has not yet been studied in renal fibrosis of DN. Previous studies have indicated that the peptide YWDHNNPQIR (named RAP), a natural peptide derived from rapeseed protein, has an antioxidative stress effect. The oxidative stress is believed to be associated with DN. The aim of this study was to evaluate the pharmacologic effects of RAP against renal fibrosis of DN and high glucose (HG)-induced mesangial dysfunction.Materials and methodsDiabetes was induced by streptozotocin and high-fat diet in C57BL/6 mice and these mice were treated by subcutaneous injection of different doses of RAP (0.1 mg/kg and 0.5 mg/kg, every other day) or PBS for 12 weeks. Later, functional and histopathologic analyses were performed. Parallel experiments verifying the molecular mechanism by which RAP alleviates DN were carried out in HG-induced mesangial cells (MCs).ResultsRAP improved the renal function indices, including 24-h albuminuria, triglyceride, serum creatinine, and blood urea nitrogen levels, but did not lower blood glucose levels in DN mice. RAP also simultaneously attenuated extracellular matrix accumulation in DN mice and HG-induced MCs. Furthermore, RAP reduced HG-induced cell proliferation, but it showed no toxicity in MCs. Additionally, RAP inhibited the mitogen-activated protein kinase (MAPK) and nuclear factor κB (NF-κB) signaling pathways.ConclusionRAP can attenuate fibrosis in vivo and in vitro by antagonizing the MAPK and NF-κB pathways.

Tangeretin inhibits high glucose-induced extracellular matrix accumulation in human glomerular mesangial cells

Tangeretin (5, 6, 7, 8, 4′-pentamethoxyflavone), a natural compound extracted from citrus plants, has been shown to possess a variety of pharmacological activities, including anti-oxidant, anti-tumor, cytostatic and anti-diabetic properties. However, the role of tangeretin in diabetic nephropathy (DN) has not yet been investigated. This study was undertaken to elucidate the effects of tangeretin on high glucose (HG)-induced oxidative stress and extracellular matrix (ECM) accumulation in human glomerular mesangial cells (MCs) and explore the underlying mechanisms. Our results showed that tangeretin significantly inhibited HG-induced the proliferation of MCs. In addition, tangeretin dramatically reduced the levels of reactive oxygen species (ROS) and malondialdhyde (MDA), and induced SOD activity, as well as inhibited the expression of fibronectin (FN) and collagen IV in HG-stimulated MCs. Furthermore, tangeretin efficiently prevented the activation of ERK signaling pathway in HG-stimulated MCs. Taken together, these data indicated that tangeretin inhibits HG-induced cell proliferation, oxidative stress and ECM expression in glomerular MCs, at least in part, through the inactivation of ERK signaling pathway. Therefore, tangeretin may be a potential agent in the treatment of DN.

Overexpression of Sirtuin2 prevents high glucose-induced vascular endothelial cell injury by regulating the p53 and NF-κB signaling pathways.

To investigate the potential role and underlying mechanism of Sirtuin2 (SIRT2) in regulating high glucose (HG)-induced vascular endothelial cell injury by using human umbilical vein endothelial cells (HUVECs). SIRT2 mRNA and protein expression levels were decreased in HG-treated HUVECs. SIRT2 overexpression increased viability, decreased apoptosis and reduced levels of reactive oxygen species in HG-treated HUVECs. SIRT2 overexpression decreased TNF-α expression (146.5±22.8pg TNF-α ml-1) relative to that in the empty vector group (263.5±18.5pg TNF-α ml-1) and decreased MCP-1 expression (63.8±9.85pg MCP-1ml-1) relative to that in the empty vector group (105.8±8.5pg MCP-1ml-1). SIRT2 overexpression decreased the acetylation of p53 by 33% and decreased the acetylation of NF-κB p65 by 58% in HG-treated HUVECs. SIRT2 prevents HG-induced vascular endothelial cell injury through suppressing the p53 and NF-κB signaling pathways.

Salidroside alleviates high glucose-induced oxidative stress and extracellular matrix accumulation in rat glomerular mesangial cells by the TXNIP-NLRP3 inflammasome pathway

Diabetic nephropathy (DN) is a metabolic disease characterized by mesangial cell proliferation and extracellular matrix (ECM) accumulation. Salidroside (SAL) is the major ingredient in Rhodiola rosea and possesses beneficial effects on DN. This study aimed to evaluate the effect of SAL on high glucose (HG)-induced oxidative stress and ECM accumulation and the underlying mechanism. Rat glomerular mesangial cells HBZY-1 were induced by high glucose (HG) in the presence or absence of SAL. Cell proliferation was measured by CCK-8 assay. The reactive oxygen species (ROS) level, malondialdehyde (MDA) level and superoxide dismutase (SOD) activity were detected to evaluate oxidative stress. The expression levels of ECM proteins including fibronectin (FN) and type IV collagen (Coll IV) were detected by qRT-PCR and western blot analysis. The expressions of thioredoxin-interacting protein (TXNIP), nod-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing CARD (ASC), and caspase-1 were assessed by western blot. Si-TXNIP or si-NC was transfected into HBZY-1 cells to inhibit TXNIP-NLRP3 inflammasome pathway. The results showed that SAL treatment alleviated HG-induced cell proliferation. SAL reduced the levels of ROS and MDA, and induced the SOD activity. Besides, the mRNA and protein expressions of FN and Coll IV were decreased by SAL. The expression levels of TXNIP, NLRP3, ASC, and caspase-1 were reduced in the SAL treated cells. In addition, TXNIP knockdown inhibited TXNIP-NLRP3 inflammasome activation and suppressed HG-induced cell proliferation, oxidative stress, and ECM accumulation. In conclusion, SAL alleviated HG-induced oxidative stress and ECM accumulation in rat glomerular mesangial cells by the TXNIP-NLRP3 inflammasome pathway.

MicroRNA-25 inhibits high glucose-induced apoptosis in renal tubular epithelial cells via PTEN/AKT pathway

Diabetic nephropathy (DN) has become the major cause of end-stage renal disease (ESRD). It has been demonstrated that apoptosis of renal tubular epithelial cells induced by hyperglycemia contributes to the pathogenesis of DN. Recent researches have corroborated the critical roles of microRNAs (miRNAs) in the apoptosis of various types of cells including renal tubular epithelial cells. However, the eff ; ;ect of miRNAs on the hyperglycemia-induced apoptosis of renal tubular epithelial cells remains unclear. The aim of this study is to explore the eff ; ;ect of miRNAs on the hyperglycemia-induced apoptosis of renal tubular epithelial cells and its molecular mechanism. Using a miRNA microarray, miRNAs putatively associated with DN were examined in renal biopsy tissue samples from DN patients and healthy controls. Validation analysis of miR-25 level in serum samples and renal biopsy tissue samples was performed using quantitative reverse transcription PCR (qRT-PCR). Then, gain- and loss- of function experiments were performed to determine the protective roles of miR-25 in high glucose-induced damage to renal tubular epithelial cells. Furthermore, the target gene of miR-25 and the downstream signaling pathway were also investigated. Microarray analysis and qRT-PCR revealed that miR-25 was significantly downregulated in renal biopsy tissue and serum samples from DN patients. We also observed that an inverse relationship between serum miR-25 level and proteinuria in DN patients. Meanwhile, miR-25 was decreased in human kidney (HK-2) cells subjected to HG treatment in a time dependent manner. Its overexpression reduced production of reactive oxygen species (ROS), suppressed cell apoptosis in HG-induced cell damage model, which was coupled with the decreased expression of cleaved caspase-3 and activity of caspase-3. Subsequent analyses demonstrated that phosphatase and tensin homolog deleted on chromosome ten (PTEN) was a direct and functional target of miR-25, which was validated by the dual luciferase reporter assay. Most importantly, the overexpression of PTEN effectively reversed the protective effects of miR-25 mimics on renal tubular epithelial cell injury. We also found that the anti-apoptotic effects of miR-25 are dependent on the activation of PTEN/Akt pathway. In addition, we observed that PTEN was upregulated in renal biopsy tissue samples from patients with DN, and an inverse relationship was found between PTEN and miR-25 expression, suggesting that miR-25 may exert its function through regulation of PTEN in DN. Taken together, our study proved that overexpression of miR-25 could ameliorate HG-induced oxidative stress and apoptosis in renal tubular epithelial cells through activation of PTEN/AKT pathway, suggesting that overexpression of miR-25 might provide a potential therapeutic approach for DN.

Rosuvastatin inhibits high glucose-stimulated upregulation of VCAM-1 via the MAPK-signalling pathway in endothelial cells

Objective: The aim of this study is to investigate the molecular mechanisms and effect of rosuvastatin on adhesion molecule induction in human endothelial cells under high-glucose conditions (HG).Methods and results: The effects of rosuvastatin on vascular cell adhesion molecule (VCAM)-1 production and pERK phosphorylation were measured in HG-induced human umbilical vein endothelial cells (HUVECs) with inhibitors targeting the mitogen-activated protein kinase (MAPK) signal pathway. HG increased levels of VCAM-1. Treatment with rosuvastatin inhibited VCAM-1 expression in a concentration- and time-dependent manner. In addition, we investigated the effects of rosuvastatin on the extracellular signal-regulated kinase (ERK) 1/2 signal pathway. Rosuvastatin completely inhibited HG-induced phosphorylation of ERK. ERK/MAPK inhibitors completely prevented the VCAM-1 inhibition effect of rosuvastatin under HG condition.Conclusions: This study demonstrated that rosuvastatin suppresses HG-induced VCAM-1 production via the MAPK signalling pathway, playing a role in the suppression of atherosclerosis.

Tiron ameliorates high glucose-induced cardiac myocyte apoptosis by PKCδ-dependent inhibition of osteopontin.

Tiron functions as an effective antioxidant alleviating the intracellular reactive oxygen species (ROS) or the acute toxic metal overload. Previous studies have shown that cardiac myocyte apoptosis can be effectively inhibited by tiron administration in streptozotocin (STZ)-induced diabetic rats, primary neonatal rat cardiomyocytes (NRVMs), and H9c2 embryonic rat cardiomyocytes. However, the underlying signalling mechanism is ill-defined. In the present study, we found that tiron supplementation significantly inhibited apoptosis of high glucose (HG)-treated NRVMs and the left ventricular cardiomyocytes from STZ-diabetic rat, accompanied with a reduction of osteopontin (OPN) levels as well as an inhibition of PKCδ phosphorylation. OPN knockdown protected NRVMs against HG-induced cell apoptosis. In addition, genetic inhibition of PKCδ mitigated HG-stimulated enhancement of intracellular OPN levels in NRVMs. These findings indicate that ROS-mediated activation of PKCδ upregulated OPN expression, leading to cardiac myocyte apoptosis. Interfering with ROS/PKCδ pathway by antioxidants such as tiron provides an optional therapeutic strategy for treatment and prevention of apoptosis-related cardiovascular diseases including diabetic cardiomyopathy.

Regulation of Na+-K+-ATPase effected high glucose-induced myocardial cell injury through c-Src dependent NADPH oxidase/ROS pathway

Depressed Na+/K+-ATPase activity has long been reported to be involved in diabetic-related cardiomyocyte death and cardiac dysfunction. However, the nature of directly regulating Na+-K+-ATPase in diabetic-related myocardial diseases remains unknown. Hyperglycemia is believed as one of major factors responsible for diabetic-related myocardial apoptosis and dysfunction. In this study, whether inhibiting Na+-K+-ATPase by ouabain or activating Na+-K+-ATPase by DRm217 has functions on high glucose (HG) -induced myocardial injury was investigated. Here we found that addition of DRm217 or ouabain to HG-treated cells had opposite effects. DRm217 decreased but ouabain increased HG-induced cell injury and apoptosis. This was mediated by changing Na+-K+-ATPase activity and Na+-K+-ATPase cell surface expression. The inhibition of Na+-K+-ATPase endocytosis alleviated HG-induced ROS accumulation. Na+-K+-ATPase·c-Src dependent NADPH oxidase/ROS pathway was also involved in the effects of ouabain and DRm217 on HG-induced cell injury. These novel results may help us to understand the important role of the Na+-K+-ATPase in diabetic cardiovascular diseases.

Heat shock protein 90/Akt pathway participates in the cardioprotective effect of exogenous hydrogen sulfide against high glucose-induced injury to H9c2 cells.

It has been reported that exogenous hydrogen sulfide(H2S) protects against high glucose(HG)-induced cardiac injury and has a modulatory effect on heat shock protein(HSP) and Akt, which play a cardioprotective role. In this study, we examined whether the HSP90/Akt pathway contributes to the protective effects of exogenous H2S against HG-induced injury to H9c2 cardiac cells. Our results revealed that the exposure of H9c2 cardiac cells to 35mM glucose(HG) for 1to 24h decreased the expression of HSP90 and markedly reduced the expression level of phosphorylated (p)-Akt in a time-dependent manner. Co-exposure of the cells to HG and geldanamycin(GA; an inhibitor of HSP90) aggravated the inhibition of the p-Akt expression level by HG. Of note, treatment of the cells with 400µM NaHS(a donor of H2S) for 30min prior to exposure to HG significantly attenuated the HG-induced decrease in the expression levels of both HSP90 and p-Akt, along with inhibition of HG-induced cell injury, as indicated by the increase in cell viability and superoxide dismutase(SOD) activity, and by a decrease in the number of apoptotic cells, reactive oxygen species(ROS) generation, as well as by the decreased dissipation of mitochondrial membrance potential(MMP). Importantly, treatment of the cells with GA or LY294002(an inhibitor of Akt) prior to exposure to NaHS and HG considerably blocked the cardioprotective effects of NaHS against the HG-induced injury mentioned above. On the whole, the findings of this study demonstrate that the inhibition of the HSP90/Akt pathway may be an important mechanism responsible for HG-induced cardiomyocyte injury. We also provide novel evidence that exogenous H2S protects H9c2 cells against HG-induced injury by activating the HSP90/Akt pathway.

Repression of miR-217 protects against high glucose-induced podocyte injury and insulin resistance by restoring PTEN-mediated autophagy pathway

Podocyte damage is the initial hallmark of diabetic nephropathy (DN), leading to the increasing morbidity and mortality in diabetic patients. Recent researches have corroborated the critical roles of miRNAs in the pathological progression of DN. Here, elevation of miR-217 was verified in high glucose (HG)-stimulated podocytes. Moreover, blocking miR-217 expression antagonized HG-induced cell injury by attenuating the adverse role of HG on cell viability and inhibiting ROS levels and cell apoptosis. Simultaneously, miR-217 repression restored HG-disrupted insulin resistance by elevating glucose uptake and nephrin expression, an essential component for insulin-induced glucose uptake. Mechanism assay substantiated the defective autophagy in HG-treated podocytes, which was resumed by miR-217 cessation. Importantly, suppressing autophagy pathway with 3-MA alleviated the protective roles of miR-217 down-regulation in podocyte injury and insulin resistance. Luciferase reporter analysis confirmed that PTEN was a target of miR-217 in podocytes. Additionally, blocking PTEN expression restrained autophagy restoration in miR-217-decreased cells. Furthermore, PTEN down-regulation attenuated the beneficial role of miR-217 suppression in HG-induced injury and insulin resistance. Together, this study manifests that miR-217inhibition can protectively antagonize HG-induced podocyte damage and insulin resistance by restoring the defective autophagy pathway via targeting PTEN, representing a novel and promising therapeutic target against diabetic nephropathy.

Helix B surface peptide attenuates diabetic cardiomyopathy via AMPK-dependent autophagy

BackgroundErythropoietin (EPO) has been reported to exert protective effects on a host of damaged tissues. However, the erythropoietic effect of this hormone can result in high risks of thrombosis, stroke, and hypertension, remarkably limiting the clinical use of EPO. Helix B surface peptide (HBSP) is a small peptide derived from the helix-B domain of EPO. Surprisingly, HBSP retains the tissue protective properties of EPO without altering the hematocrit. Thus, we evaluated the possible role of HBSP on diabetic cardiomyopathy. MethodsDiabetes was induced in mice by intraperitoneal injections of streptozocin (STZ). Mice were randomly treated with normal saline or HBSP. Cardiac function, fibrosis, apoptosis, and myocardial mitochondrial morphology were examined. For in vitro experiments, H9C2 myoblast cells were randomly grouped as normal glucose (NG, 5 mM), NG+HBSP (100 nM), high glucose (HG, 33 mM), HG+HBSP (100 nM), HG+HBSP+3-methyladenine (3-MA, 10 mM), HG+rapamycin (Rapa, 100 nM), and HG+HBSP+Compound C (CC, 10 mM). Autophagosomes, LC3 dots, apoptosis and mitochondria membrane potential (MMP) of H9C2 cells were examined.The expressions of LC3, p62, p-AMPK (Thr172) and p-mTOR (Ser2448) were examined by Western blot. ResultsHBSP markedly improved cardiac function, attenuated cardiac interstitial fibrosis, inhibited myocardial apoptosis, and ameliorated mitochondrial ultrastructure in mice with diabetic cardiomyopathy. HG reduced autophagy in H9C2 cells. HBSP enhanced autophagy in HG-treated H9C2 cells. HBSP reduced the apoptosis index of HG-treated H9C2 cells. HBSP increased the MMP of HG-treated H9C2 cells. HBSP increased the levels of p-AMPK (Thr172), and reduced p-mTOR (Ser2448) in HG-treated H9C2 cells, and the increase of p-AMPK (Thr172) was accompanied by the stimulation of autophagy. Autophagy inhibitor 3-MA and AMPK inhibitor CC mitigated HBSP-induced beneficial effect, whereas autophagy inducer Rapa alleviated the HG-induced cell apoptosis. ConclusionsHBSP attenuates diabetic cardiomyopathy via autophagy mediated by AMPK-dependent pathway. HBSP may be a potential therapeutic intervention for diabetic cardiomyopathy.

Ghrelin gene products rescue cultured adult rat hippocampal neural stem cells from high glucose insult.

Adult hippocampal neurogenesis is decreased in type 2 diabetes, and this impairment appears to be important in cognitive dysfunction. Previous studies suggest that ghrelin gene products (acylated ghrelin (AG), unacylated ghrelin (UAG) and obestatin (OB)) promote neurogenesis. Therefore, we hypothesize that ghrelin gene products may reduce the harmful effects of high glucose (HG) on hippocampal neural stem cells (NSCs). The aim of this study was to investigate the role of these peptides on the survival of cultured hippocampal NSCs exposed to HG insult. Treatment of hippocampal NSCs with AG, UAG or OB inhibited HG-induced cell death and apoptosis. Exposure of cells to the growth hormone secretagogue receptor 1a antagonist abolished the protective effects of AG against HG toxicity, whereas those of UAG or OB were preserved. All three peptides attenuated HG-induced decrease in BrdU-labeled and phosphohistone-H3-labeled cells. We also investigated the effects of ghrelin gene products on the regulation of apoptosis at the mitochondrial level. AG, UAG or OB rescued hippocampal NSCs from HG insult by inhibiting intracellular and mitochondrial reactive oxygen species generation and stabilizing mitochondrial transmembrane potential. In addition, cells treated with ghrelin gene products showed an increased Bcl-2 and decreased Bax levels, thereby increasing the Bcl-2/Bax ratio, inhibiting cytochrome c release and preventing caspase-3 activation. Finally, AG-, UAG- or OB-mediated protection was dependent on the activities of adenosine monophosphate-activated protein kinase/uncoupling protein 2 pathway. Our data indicate that ghrelin gene products may act as survival factors that preserve mitochondrial function and inhibit oxidative stress-induced apoptosis.

Histone methyltransferase Suv39h1 attenuates high glucose-induced fibronectin and p21WAF1 in mesangial cells

Suppressor of variegation 3⿿9 homolog 1 (Suv39h1) is a histone methyltransferase that trimethylates lysine 9 of histone H3 (H3K9me3), which results in gene silencing. A previous study found that H3K9me3 and Suv39h1 were decreased in diabetic mouse vascular smooth muscle cells whereas Suv39h1 overexpression attenuated ischemic myocardial injury. Moreover, high glucose (HG) decreased H3K9me3 and Suv39h1 levels in some cells. Thus, we studied the roles of Suv39h1 in HG-induced effects in MES13 (mouse mesangial) cells.We found that HG, chaetocin (a Suv39h1 inhibitor) and Suv39h1 siRNA decreased Suv39h1 while increasing fibronectin and p21WAF1 protein levels. HG increased mRNA while chaetocin increased transcription of fibronectin and p21WAF1genes. Both HG and chaetocin decreased histone H3K9me3 levels at the promoters of fibronectin and p21WAF1 genes. Additionally, Suv39h1 overexpression attenuated HG-induced fibronectin and p21WAF1 mRNA and protein expressions while attenuating HG-induced cell hypertrophy. Suv39h1 overexpression also attenuated HG-suppressed histone H3K9me3 levels at the promoters of fibronectin and p21WAF1 genes. Moreover, LY294002 or the dominant-negative phosphoinositide 3-kinase (PI3K) mutant (οp85) attenuated HG-decreased Suv39h1 and HG-induced fibronectin and p21WAF1 protein expressions.We concluded that HG decreased Suv39h1 via the PI3K pathway in mesangial cells. Inhibition of Suv39h1 increased fibronectin and p21WAF1 expressions. Moreover, Suv39h1 overexpression attenuated HG-induced fibronectin and p21WAF1 expressions and cell hypertrophy while attenuating HG-suppressed histone H3K9me3 levels at the promoters of fibronectin and p21WAF1 genes.