Beneficial Effects of Rosuvastatin and L-Arginine on High Glucose-Induced Oxidative Stress in Human Umbilical Vein Endothelial Cells

The aim of the present study was to investigate the mechanism of homocysteine (Hcy) induced oxidative stress in the human umbilical vein endothelial cells (HUVECs). The HUVECs were isolated from umbilical vein vascular wall of 12 patients and treated with Hcy. The malondialdehyde (MDA) level was measured using the thiobarbituric acid (TBA) method. The expressions of superoxide dismutase 2 (SOD2), endothelial nitric oxide synthase(eNOS), and intercellular adhesion molecule 1 (ICAM-1) were detected by Western blot and RT-PCR. The genome-wide DNA methylation assay was performed using the Infinium Human Methylation 450 BeadChip. The specific DNA methylation was determined using bisulfite sequencing analysis. To evaluate the role of sorbin and SH3 domain-containing protein 1 (SORBS1), the HUVECs were transfected with small interfere RNA (siRNA) targeting SORBS1 (SORBS1-siRNA). Hcy induced MDA level in HUVECs, and increased ICAM-1 expression both in protein and mRNA levels. The protein and mRNA levels of SOD2 and eNOS were inhibited by Hcy induction. However, the effects of Hcy on MDA level and expressions of SOD2, eNOS, and ICAM-1 were attenuated by folic acid (Fc) and vitamin B12 (B12) treatment. DNA total methylation level in Hcy treated cells was significantly decreased compared to the control group, while the DNA total methylation levels were increased after treatment with Fc and B12. The methylation level of SORBS1 in Hcy treatment group was higher than that of control group. And the methylation level of SORBS1 induced by Hcy was attenuated by Fc and B12 treatment. SORBS1-siRNA transfection induced the MDA levels and reduced the expressions of SOD2 in HUVECs. We indicated that Hcy induced oxidative stress in HUVECs via regulating methylation of SORBS1. We also found that Fc and B12 treatment attenuated the oxidative stress and inflammation induced by Hcy in HUVECs. The findings indicated that Fc and B12 might be effective agents for the treatment of Hcy induced AS.

L-Threoascorbic acid treatment promotes S. aureus-infected primary human endothelial cells survival and function, as well as intracellular bacterial killing, and immunomodulates the release of IL-1β and soluble ICAM-1

ABSTRACTOxidative stress plays a critical role in the pathogenesis of diabetic vascular complications. Trans-δ-viniferin (TVN), a polyphenolic compound, has recently attracted much attention as an antioxidant exhibiting a hypoglycemic potential. In the present study, we aimed at investigating the protective effect of TVN against high glucose-induced oxidative stress in human umbilical vein endothelial cells (HUVECs) and the potential mechanism involved. We found that TVN attenuated reactive oxygen species (ROS) production, increased catalase (CAT) activity and decreased malondialdehyde (MDA) levels to ameliorate cell survival induced by 35 mM glucose. Meanwhile, it inhibited high glucose-induced apoptosis by maintaining Ca2+ and preserving mitochondrial membrane potential (MMP) levels. The immunoblot analysis indicated that TVN efficiently regulated the cleavage of caspase family, p53, Bax and Bcl-2, all mediated by SIRT1. Furthermore, the increased level of SIRT1 induced by TVN was inhibited by nicotinamide and siRNA-medicated SIRT1 silencing (si-SIRT1), thereby confirming the significant role of SIRT1 in these events. In conclusion, our results indicated that TVN efficiently reduced oxidative stress and maintained mitochondrial function related with activating SIRT1 in high glucose-treated HUVECs. It suggested that TVN is pharmacologically promising for treating diabetic cardiovascular complications.

We investigated whether the fermented soymilk extract (FSE) has protective effects against high glucose-induced oxidative stress in human umbilical vein endothelial cells (HUVECs). FSE was prepared via fermentation of soymilk with Bacillus subtilis followed by methanol extraction. To determine the protective effect of FSE, oxidative stress was induced by exposing of HUVECs to the high glucose (30 mM) for 48 hr. Exposure of HUVECs to high glucose for 48 hr resulted in a significant (p<0.05) decrease in cell viability, catalase, SOD and GSH-px activity and a significant (p<0.05) increase in intracellular ROS level and thiobarbituric acid reactive substances (TBARS) formation in comparison to the cells treated with 5.5 mM glucose. However, at concentration of 0.1 ㎎/㎖, FSE treatment decreased intracellular ROS level and TBARS formation, and increased cell viability and activities of antioxidant enzymes including catalase, SOD and GSH-px in high glucose pretreated HUVEC. These results suggest that FSE may be able to protect HUVECs from high glucose-induced oxidative stress, partially through the antioxidative defense systems.

Atherosclerosis (AS) manifests with arterial intimal injury, lipid deposition and chronic inflammation, which is a key pathogenic cause of cardio-cerebrovascular disorders. LncRNA MIR4697HG was downregulated in human advanced atherosclerotic plaques. This study probed the precise biological functions and downstream regulatory mechanisms of MIR4697HG during AS progression. MIR4697HG levels in atherosclerotic plaque tissues and normal arterial intima were measured by RT-qPCR. An injury model of human umbilical vein endothelial cells (HUVECs) was induced through treating with oxidative low-density lipoprotein (ox-LDL). MIR4697HG overexpression plasmids (pcDNA-MIR4697HG) was transfected into ox-LDL-treated HUVECs, and then cell viability, apoptosis, reactive oxygen species (ROS) level, oxidative stress marker protein malondialdehyde (MDA) level and superoxide dismutase (SOD) activity, and adhesion molecule intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) levels in HUVECs were determined. Moreover, the binding between MIR4697HG and fused in sarcoma (FUS) was checked with RNA pull-down assay. The interaction between FUS and annexin A5 (ANXA5) was gauged with Co-immunoprecipitation. Then MIR4697HG/FUS/ANXA5 axis mediated HUVEC functions were accessed with rescue experiments. Additionally, an AS model was established via feeding a high-fat diet for ApoE-/- mice, and lentivirus MIR4697HG overexpression vector (Lv-MIR4697HG) was injected into AS mice followed by detection of atherosclerotic plaque area in mice. MIR4697HG was downregulated in atherosclerotic plaque tissues and HUVECs stimulated by ox-LDL. MIR4697HG overexpression attenuated ox-LDL-induced HUVEC viability inhibition, apoptosis, oxidative stress and adhesion molecule release. Moreover, MIR4697HG bound with FUS and facilitated FUS expression in HUVECs. FUS knockdown abrogated the functions of lncRNA MIR4697HG overexpression in ox-LDL induced HUVEC injury. Besides, FUS could bind with ANXA5. FUS overexpression inhibited ox-LDL induced HUVEC injury, while ANXA5 knockdown reversed these effects. Additionally, Lv-MIR4697HG reduced atherosclerotic plaque area in ApoE-/- mice. LncRNA MIR4697HG mitigated ox-LDL-induced apoptosis, oxidative stress and adhesion molecule release in HUVECs and alleviated AS progression in mice through the FUS/ANXA5 axis.

Inhibition of YAP by lenvatinib in endothelial cells increases blood pressure through ferroptosis

This study investigated the protective effect of the butanol (BuOH) fraction from Laminaria japonica (BFLJ) extract on high glucose-induced oxidative stress in human umbilical vein endothelial cells (HUVECs). Freeze-dried L. japonica was extracted with distilled water, and the extracted solution was mixed with ethanol then centrifuged. The supernatant was subjected to sequential fractionation with various solvents. The BuOH fraction was used in this study because it possessed the strongest antioxidant activity among the various solvent fractions. To determine the protective effect of the BFLJ, oxidative stress was induced by exposing of HUVECs to the high glucose (30 mM) or normal glucose (5.5 mM) for 48 hr. Cell viability, lipid peroxidation, glutathione (GSH) concentration, and antioxidant enzyme activities such as catalase, superoxide dismutase (SOD), glutathione peroxidase (GSH-px), and glutathion reductase (GSH-re) were measured. Exposure of HUVECs to high glucose for 48 hr resulted in a significant (p＜0.05) decrease in cell viability, SOD, GSH-px and GSH-re and a significant (p＜0.05) increase in thiobarbituric acid reactive substances (TBARS) formation in comparison to the cells treated with 5.5 mM glucose or untreated with glucose. BFLJ treatment decreased TBARS formation and increased cell viability, GSH concentration, and activities of antioxidant enzymes including catalase, SOD, GSH-px, and GSH-re in high glucose pretreated HUVECs. These results suggest that BFLJ may be able to protect HUVECs from high glucose-induced oxidative stress, partially through the antioxidative defence systems.

Protective effect of Cordyceps militaris against high glucose-induced oxidative stress in human umbilical vein endothelial cells

Antioxidants can prevent pathological damage caused by hyperglycaemia-induced oxidative stress associated with diabetes. In the present study, we investigated whether the brown alga Ecklonia cava has protective effects against high glucose-induced oxidative stress in Human umbilical vein endothelial cells (HUVECs). For that purpose, we prepared an enzymatic digest from E. cava (ECC) by using the carbohydrate, Celluclast. High glucose treatment induced HUVECs cell death, but ECC, at a concentration of 10 or 100 microg mL(-1), significantly inhibited the high glucose-induced cytotoxicity. Furthermore, treatment with ECC dose-dependently decreased thiobarbituric acid reactive substances (TBARS), intracellular generation of reactive oxygen species, and the nitric oxide level increased by high glucose. In addition, ECC treatment increased activities of antioxidant enzymes including catalase, superoxide dismutase and glutathione peroxidase in high-glucose pretreated HUVECs. High glucose levels induced the overexpression of inducible nitric oxide synthase, cyclooxygenase-2 and nuclear factor-kappa B proteins in HUVECs, but ECC treatment reduced the overexpression of these proteins. These results suggest that ECC is a potential therapeutic agent that will reduce the damage caused by hyperglycaemia-induced oxidative stress associated with diabetes.

Decreased production and/or impaired action of nitric oxide (NO) play a role in the pathogenesis of atherosclerotic cardiovascular disease and erectile dysfunction (ED) in diabetic patients. Under hyperglycemic conditions, formation and accumulation of advanced glycation end products (AGE) have been known to progress, thus contributing to tissue damage in diabetes. However, effects of inhibitors of phosphodiesterase-5 (PDE-5), an enzyme that catalyzes the degradation of cyclic guanosin-monophosphate (cGMP) and subsequently blocks the actions of NO, on AGE-exposed endothelial cells remain unknown. Therefore, this study investigated whether and how vardenafil, an inhibitor of PDE-5, could block the deleterious effects of AGE on human umbilical vein endothelial cells (HUVEC). Gene and protein expression was analyzed in quantitative real-time reverse transcription polymerase chain reaction (RT-PCR) and western blots, respectively. Intracellular formation of reactive oxygen species (ROS) was evaluated with dihydroethidium staining. AGE increased receptor for AGE (RAGE) mRNA and protein levels in HUVEC, both of which were significantly inhibited by the treatments with 30nM vardenafil or 5μM 8-Br-cGMP, an analogue of cGMP. Further, vardenafil reduced the AGE-induced ROS generation and subsequently inhibited up-regulation of monocyte chemoattractant protein-1 (MCP-1) mRNA levels in HUVEC. We demonstrated here for the first time that vardenafil could block the AGE-induced up-regulation of MCP-1 mRNA levels in HUVEC by suppressing RAGE expression and subsequent ROS generation via elevation of cGMP. Our present results suggest that vardenafil directly acts on endothelial cells and it could work as an anti-inflammatory agent against AGE.

Oxidative stress regulates dysfunction and senescence of vascular endothelial cells. The DNA damage response and its main signaling pathway involving ataxia telangiectasia mutated (ATM) have been implicated in playing a central role in mediating the actions of oxidative stress; however, the role of the ATM signaling pathway in vascular pathogenesis has largely remained unclear. Here, we identify ATM to regulate oxidative stress-induced endothelial cell dysfunction and premature senescence. Oxidative stress induced senescence in endothelial cells through activation/phosphorylation of ATM by way of an Akt/p53/p21-mediated pathway. These actions were abrogated in cells in which ATM was knocked down by RNA interference or inhibited by specific inhibitory compounds. Furthermore, the in vivo significance of this regulatory pathway was confirmed using ATM knock-out mice in which induction of senescent endothelial cells in the aorta in a diabetic mouse model of endothelial dysfunction and senescence was attenuated in contrast to pathological changes seen in wild-type mice. Collectively, our results show that ATM through an ATM/Akt/p53/p21-dependent signaling pathway mediates an instructive role in oxidative stress-induced endothelial dysfunction and premature senescence.

Glucagon-like peptide-1 (GLP-1) inhibits advanced glycation end product (AGE)-induced up-regulation of VCAM-1 mRNA levels in endothelial cells by suppressing AGE receptor (RAGE) expression

BackgroundErectile dysfunction (ED) is a well-known complication of diabetes, affecting up to 75% of diabetic men. Although the etiology of diabetic ED is multifactorial, endothelial dysfunction is considered to be a pillar of its pathophysiology. Endothelial dysfunction is caused by the harmful effects of high glucose levels and increased oxidative stress on the endothelial cells that comprise the vascular endothelium. The aim of this study was to identify the proteomic changes caused by high glucose-induced oxidative stress and explore the role of heme oxygenase 1 (HMOX1) in it.MethodsThe cellular proteomic response to hypoxanthine-induced oxidative stress in human umbilical vein endothelial cells (HUVECs) was analyzed by isobaric tags for relative and absolute quantitation (iTRAQ) combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Differentially expressed proteins (DEPs) were analyzed through Network and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Further validation assays was performed to validate the role of HMOX1.ResultsThe results showed that 66 and 76 DEPs were markedly upregulated and downregulated, respectively, for HUVECs oxidative stress. Among these proteins, we verified eight dysregulated genes by quantitative reverse transcription PCR, including nucleolin (NCL), X-ray repair cross-complementing protein 6 (XRCC6), ubiquinol-cytochrome C reductase binding protein (UQCRB), non-POU domain containing octamer binding (NONO), heme oxygenase 1 (HMOX1), nucleobindin 1 (NUCB1), DEK, and chromatin target of prmt1 (CHTOP). Further, using overexpression and genetic knockdown approaches, we found that HMOX1 was critical for the oxidative stress response in HUVECs.ConclusionsWe found that HMOX1 was closely related to the oxidative stress response induced by hypoxanthine. To the best of our knowledge, this study is the first overview of the responses of HUVECs to oxidative stress. The findings will contribute to analyses of the detailed molecular mechanisms involved in the pathogenesis of endothelial dysfunction and related molecular mechanisms in ED patients.

Background: Rheumatoid arthritis (RA) is a long-term autoimmune condition marked by persistent inflammation of the joints and various systemic complications, including endothelial dysfunction, atherosclerosis, and pulmonary fibrosis. Oxidative stress is a key contributor to the pathogenesis of RA, potentially exacerbating vascular damage and promoting pro-angiogenic and profibrotic processes. Objective: This study aims to investigate the effects of sera from RA patients on human umbilical vein endothelial cells (HUVECs), focusing on the induction of oxidative stress, endothelial cell proliferation, migration, and collagen type I synthesis. Methods: Twenty-eight serum samples were collected from RA patients and healthy donors (HDs). HUVECs were exposed to these sera, and intracellular reactive oxygen species (ROS) levels were fluorescently detected using H2DCF-DA. Cell viability was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell migration was evaluated through a scratch wound assay, and collagen type I synthesis was measured using a lentiviral vector expressing the green fluorescent protein (GFP) under the control of the human COL1A1 gene promoter. Results: Exposure to RA sera resulted in a significant increase in intracellular ROS levels in HUVECs compared to HD sera, indicating an elevated state of oxidative stress. RA sera also promoted endothelial cell proliferation and migration, suggesting a pro-angiogenic stimulus. Additionally, RA sera significantly increased collagen type I synthesis in HUVECs, implicating a potential role in profibrotic processes associated with RA. Conclusion: The results of this study emphasize the importance of circulating factors in RA sera in promoting oxidative stress, endothelial dysfunction, and pro-angiogenic and profibrotic phenotypes in endothelial cells. These processes may contribute to the vascular and fibrotic complications observed in RA, highlighting the necessity for additional research into focused therapeutic approaches to alleviate these effects.

Coenzyme Q 10 prevents high glucose-induced oxidative stress in human umbilical vein endothelial cells