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Role of TRPM7 channels in hyperglycemia-mediated injury of vascular endothelial cells.

ABSTRACT: This study investigated the change of transient receptor potential melastatin 7 (TRPM7) expression by high glucose and its role in hyperglycemia induced injury of vascular endothelial cells. Human umbilical vein endothelial cells (HUVECs) were incubated in the presence or absence of high concentrations of D-glucose (HG) for 72 h. RT-PCR, Real-time PCR, Western blotting, Immunofluorescence staining and whole-cell patch-clamp recordings showed that TRPM7 mRNA, TRPM7 protein expression and TRPM7-like currents were increased in HUVECs following exposure to HG. In contrast to D-glucose, exposure of HUVECs to high concentrations of L-glucose had no effect. HG increased reactive oxygen species (ROS) generation, cytotoxicity and decreased endothelial nitric oxide synthase protein expression, which could be attenuated by knockdown of TRPM7 with TRPM7 siRNA. The protective effect of silencing TRPM7 against HG induced endothelial injury was abolished by U0126, an inhibitor of the extracellular signal-regulated kinase signaling pathway. These observations suggest that TRPM7 channels play an important role in hyperglycemia-induced injury of vascular endothelial cells.

SUBMITTER: Sun H

PROVIDER: S-EPMC3815131 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Role of TRPM7 channels in hyperglycemia-mediated injury of vascular endothelial cells.

Sun Huawei H Leng Tiandong T Zeng Zhao Z Gao Xiuren X Inoue Koichi K Xiong Zhi-Gang ZG

PloS one 20131101 11

This study investigated the change of transient receptor potential melastatin 7 (TRPM7) expression by high glucose and its role in hyperglycemia induced injury of vascular endothelial cells. Human umbilical vein endothelial cells (HUVECs) were incubated in the presence or absence of high concentrations of D-glucose (HG) for 72 h. RT-PCR, Real-time PCR, Western blotting, Immunofluorescence staining and whole-cell patch-clamp recordings showed that TRPM7 mRNA, TRPM7 protein expression and TRPM7-li ...[more]

PMID: 24223965

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Project description:BackgroundTo determine whether and how exosomes from human umbilical vein endothelial cells (HUVEC-Exos) regulates vascular smooth muscle cells (VSMCs) calcification/senescence in high glucose condition.MethodsHUVEC-Exos were isolated from normal glucose (NG) and high glucose (HG) stimulated HUVECs (NG/HG-HUVEC-Exos) by super speed centrifugation. HUVEC-Exos were identified by transmission electron microscopy and Western blot of CD63. Protein profile in HUVEC-Exos was examined to screen the candidate molecules that mediate HUVEC-Exos function. VSMCs were incubated with HUVEC-Exos. A series of functional assays in vitro were performed to assess the effects of HUVEC-Exos on the calcification/senescence of VSMCs. The role of the candidate protein in HUVEC-Exos-induced VSMCs dysfunction was assessed.ResultsExosomes isolated from HG-HUVEC-Exos induced calcification/senescence in VSMCs as assessed by Alizarin Red Staining, senescence-associated ?-galactosidase (SA-?-gal) staining, and the expression of ALP and p21. HG-HUVEC-Exos significantly increased LDH activity, as well as the product of lipid peroxidation (MDA content), and decreased oxidative stress marker activity, as compared with NG-HUVEC-Exos. Moreover, mechanism studies showed that mitochondrial membrane potential and the expression levels of mitochondrial function related protein HADHA and Cox-4 were significantly decreased in HG-HUVEC-Exos compared to controls. Proteomic analysis showed that HG-HUVEC-Exos consisted of higher level of versican (VCAN), as compared with NG-HUVEC-Exos. Observation under laser confocal microscopy revealed that most green fluorescence of VCAN could overlap with the red fluorescence came from mitochondria, indicating VCAN is mainly localized to the mitochondria of VSMCs. Knockdown of VCAN with siRNA in HUVECs, inhibited HG-HUVEC-Exos-induced mitochondrial dysfunction and calcification/senescence of VSMCs.ConclusionsOur data indicate an intracellular role for VCAN in VSMCs. VCAN participates in hyperglycemia-induced calcification/senescence via modulation of mitochondrial function in VSMCs.

Project description:Diabetic cardiovascular complications are characterised by oxidative stress-induced endothelial dysfunction. Uncoupling protein 2 (UCP2) is a regulator of mitochondrial reactive oxygen species (ROS) generation and can antagonise oxidative stress, but approaches that enhance the activity of UCP2 to inhibit ROS are scarce. Our previous studies show that activation of transient receptor potential vanilloid 1 (TRPV1) by capsaicin can prevent cardiometabolic disorders. In this study, we conducted experiments in vitro and in vivo to investigate the effect of capsaicin treatment on endothelial UCP2 and oxidative stress. We hypothesised that TRPV1 activation by capsaicin attenuates hyperglycemia-induced endothelial dysfunction through a UCP2-mediated antioxidant effect.TRPV1(-/-), UCP2(-/-) and db/db mice, as well as matched wild type (WT) control mice, were included in this study. Some mice were subjected to dietary capsaicin for 14 weeks. Arteries isolated from mice and endothelial cells were cultured. Endothelial function was examined, and immunohistological and molecular analyses were performed.Under high-glucose conditions, TRPV1 expression and protein kinase A (PKA) phosphorylation were found to be decreased in the cultured endothelial cells, and the effects of high-glucose on these molecules were reversed by the administration of capsaicin. Furthermore, high-glucose exposure increased ROS production and reduced nitric oxide (NO) levels both in endothelial cells and in arteries that were evaluated respectively by dihydroethidium (DHE) and DAF-2 DA fluorescence. Capsaicin administration decreased the production of ROS, restored high-glucose-induced endothelial dysfunction through the activation of TRPV1 and acted in a UCP2-dependent manner in vivo. Administration of dietary capsaicin for 14 weeks increased the levels of PKA phosphorylation and UCP2 expression, ameliorated the vascular oxidative stress and increased NO levels observed in diabetic mice. Prolonged dietary administration of capsaicin promoted endothelium-dependent relaxation in diabetic mice. However, the beneficial effect of capsaicin on vasorelaxation was absent in the aortas of UCP2(-/-) mice exposed to high-glucose levels.TRPV1 activation by capsaicin might protect against hyperglycemia-induced endothelial dysfunction through a mechanism involving the PKA/UCP2 pathway.

Dataset Information

Role of TRPM7 channels in hyperglycemia-mediated injury of vascular endothelial cells.

Publications

Role of TRPM7 channels in hyperglycemia-mediated injury of vascular endothelial cells.

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