Vascular Endothelial Cell Injury Research Articles

Lycopene alleviates oxidative stress-induced cell injury in human vascular endothelial cells by encouraging the SIRT1/Nrf2/HO-1 pathway

ABSTRACT Background and objective: Epidemiological research have displayed that dietary intake rich in lycopene, an antioxidant, is negatively correlated with the risk of cardiovascular disease (CVD). This study aimed to investigate whether the intervention with different concentrations of lycopene could attenuate H2O2-induced oxidative stress injury in human vascular endothelial cells (VECs). Methods: The human VECs HMEC-1 and ECV-304 were incubated with a final concentration of 300 µmol/L H2O2, followed by they were incubated with lycopene at doses of 0.5, 1, or 2 µm. Subsequently, cell proliferation, cytotoxicity, cell adhesion, reactive oxygen species (ROS) contents, adhesion molecule expression, oxidative stress levels, pro-inflammatory factor production, the apoptosis protein levels, and the silent information regulator-1 (SIRT1)/nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) pathway protein levels were tested by CCK-8 kit, lactate dehydrogenase (LDH) kit, immunofluorescence labeling, cell surface enzyme immunoassays (EIA), enzyme-linked immunosorbent assay (ELISA), as well as Western blot assays, respectively. Results: Under H2O2 stimulation, HMEC-1 and ECV-304 cell proliferation and the SIRT1/Nrf2/HO-1 pathway protein expression were significantly reduced, whereas cytotoxicity, apoptosis, cell adhesion molecule expression, pro-inflammatory and oxidative stress factors production were apparently encouraged, which were partially countered by lycopene intervention in a dose-dependent manner. Conclusion: Lycopene alleviates H2O2-induced oxidative damage in human VECs by reducing intracellular ROS levels, inflammatory factor production, cell adhesiveness, and apoptosis rate under oxidative stress conditions through activation of the SIRT1/Nrf2/HO-1 pathway.

The role of CNPY2 in endothelial injury and inflammation during the progress of atherosclerosis.

Vascular endothelial cells (VECs) injury is closely related to the occurrence and development of atherosclerosis. Canopy FGF signaling regulator 2 (CNPY2), a novel unfolded protein response promoter, has been reported to activate the PERK-CHOP pathway. This study aimed to explore whether CNPY2 is associated with atherosclerosis mediated by VEC injury. By establishing ApoE-/- mouse atherosclerosis model and oxidized low-density lipoprotein (ox-LDL) cell model, we found that CNPY2 was abnormally highly expressed in ApoE-/- mice and ox-LDL-induced mouse aortic endothelial cells (MAECs). Exogenous CNPY2 can significantly aggravate the activation, inflammation, and apoptosis of MAECs induced by ox-LDL and promote the activation of PERK/eIF2α/CHOP signal. The PERK inhibitor GSK2606414 can inhibit CNPY2-induced MAECs injury and PERK signal activation. In addition, in vivo animal experiments furtherly confirmed that CNPY2 could aggravate the process of atherosclerosis in ApoE-/- mice by activating PERK signaling. In conclusion, this study indicated that high level of CNPY2 induces VECs injury by activating PERK signaling and thus participating in the progress of atherosclerosis.

Circ_0001953 contribute to retinal vascular endothelial cell injury induced by high glucose through regulating miR 186

Purpose: To investigate the effects of circ_0001953/miR-186 on human retinal vascular endothelial cell (HRVEC) injury evoked by high glucose. Methods: A cell model (HG group) was established using high glucose-treated HRVECs, while untreated HRVECs were used as the control group (Con group). The levels of endothelin-1 (ET-1), ICAM-1 and IL-6 were evaluated by ELISA and the content of malondialdehyde (MDA) and superoxide dismutase (SOD) in HRVECs were determined. Apoptosis rate was tested adopting flow cytometry. The interrelationship between circ_0001953 and miR-186 was assessed using dual luciferase reporter assay. Measurement of Bax and Bcl-2 was implemented via western blot. Results: In HG group, circ_0001953 increased while miR-186 was downregulated, ET-1, IL-6, ICAM-1, and apoptosis rate increased and accompanied with up-regulated Bax content and declined Bcl-2 protein level. Furthermore, the content of MDA increased and SOD decreased. MiR-186 was a target of circ_0001953. Conclusion: Inhibition of circ_0001953 can repress inflammation, oxidative stress and apoptosis in HRVECs by up-regulating the expression of miR-186.

PDLIM1 inhibits cell migration and invasion in diabetic retinopathy via negatively regulating Wnt3a

The injury of vascular endothelial cells is a crucial factor in the development of diabetic retinopathy (DR). PDLIM1 (a member of the PDZ and LIM protein family) has been reported to exert an essential function in vascular diseases. This study aimed to elucidate the role of PDLIM1 on retinal vascular endothelial cells in DR. Immunofluorescence staining was used to localize the expression of PDLIM1 in the mouse retina. In some tumor diseases, PDLIM1 has been reported to play a key role in regulating the Wnt pathway. However, no in-depth reports have been found in DR. Retinal capillary endothelial cells (RCECs) were treated with high-glucose and high-lipid (HG/HL) culture medium, and siRNA transfection to investigate the role of PDLIM1 in DR. PDLIM1 and Wnt3a expression was confirmed by qRT-PCR and western blotting. Flow cytometry, Transwell assay, and scratch assay were used to test the ability of cell apoptosis, migration, and invasion. PDLIM1 was mainly expressed in the retinal pigment epithelium (RPE), ganglion cell layer (GCL), inner plexus layer (IPL), and outer plexus layer (OPL). HG/HL increased Wnt3a levels and promoted cell’s ability of apoptosis, migration, and invasion, which were reversed by the knockdown of PDLIM1. PDLIM1 was found to play a protective role in diabetic retinopathy by counter-regulating Wnt3a. PDLIM1 ameliorates cell apoptosis, migration, and invasion by negatively regulating Wnt3a in RCECs of DR, which suggests that PDLIM1 might be a promising therapeutic target for DR treatment.

Development of nanoparticles for delivering recombinant human secretory leukocytes protease inhibitor (rhSLPI) for reducing vascular endothelial cell injury

1.4.1.Myocardial ischemia/reperfusion (I/R) is the pathophysiological cause of ischemic heart disease (IHD), which is a leading cause of death globally. I/R injury impacts and damages vascular endothelial cells as well as cardiomyocytes, leading to endothelial dysfunction and then worsened cardiac injury consequently. Previous studies have shown that secretory leukocyte protease inhibitors (SLPI) could provide both cardio- and vasculo-protective benefits against I/R injury. However, SLPI can be destroyed by circulating enzymes and have a short half-life, meaning there are clinical limitations to using SLPI as a therapeutic agent. To overcome these limitations, studies have been conducted on human serum albumin (HSA) nanoparticles for recombinant human SLPI delivery, but not for vascular endothelial cells subjected to I/R injury. In this study, HSA nanoparticles were fabricated using the nanoprecipitation method. Subsequently, rhSLPI-HSA nanoparticles were generated by adding 150 µl SLPI (0.75 µg/µl) to human serum albumin to produce particles. Then, the size and zeta potential of blank-HSA and rhSLPI nanoparticles were measured by field emission scanning electron microscopy (FE-SEM) and dynamic light scattering (DLS). The results showed that the size of blank-HSA nanoparticles and rhSLPI-nanoparticles was 147 nm and 200–300 nm, respectively. The zeta potential of blank-HSA nanoparticles and rhSLPI-nanoparticles was −16.9 mV and −13 mV, respectively. Afterward, the cytotoxicity of both types of nanoparticles was examined on human vascular endothelial cell lines (EA.hy 926) by an MTT assay. The results showed that there was no toxicity in either type of nanoparticle on the vascular endothelial cells. The optimal concentrations for rhSLPI-HSA-NPs were 1 ug/ml and 10 ug/ml, which could significantly increase cell viability. The current study demonstrated the possibility of being able to generate human serum albumin nanoparticles encapsulated with recombinant human SLPI (rhSLPI-HSA nanoparticles) with no cytotoxicity, which could be used for determining the vasculo-protective effect of rhSLPI-HSA nanoparticles against an in vitro simulated I/R-induced vascular endothelial cell injury.

Single cell RNA-seq resolution revealed CCR1+/SELL+/XAF+ CD14 monocytes mediated vascular endothelial cell injuries in Kawasaki disease and COVID-19

IntroductionThe COVID-19 pandemic provide the opportunities to explore the numerous similarities in clinical symptoms with Kawasaki disease (KD), including severe vasculitis. Despite this, the underlying mechanisms of vascular injury in both KD and COVID-19 remain elusive. To identify these mechanisms, this study employs single-cell RNA sequencing to explore the molecular mechanisms of immune responses in vasculitis, and validate the results through in vitro experiments. MethodThe single-cell RNA sequencing (scRNA-seq) analysis of peripheral blood mononuclear cells (PBMCs) was carried out to investigate the molecular mechanisms of immune responses in vasculitis in KD and COVID-19. The analysis was performed on PBMCs from six children diagnosed with complete KD, three age-matched KD healthy controls (KHC), six COVID-19 patients (COV), three influenza patients (FLU), and four healthy controls (CHC). The results from the scRNA-seq analysis were validated through flow cytometry and immunofluorescence experiments on additional human samples. Subsequently, monocyte adhesion assays, immunofluorescence, and quantitative polymerase chain reaction (qPCR) were used to analyze the damages to endothelial cells post-interaction with monocytes in HUVEC and THP1 cultures. ResultsThe scRNA-seq analysis revealed the potential cellular types involved and the alterations in genetic transcriptions in the inflammatory responses. The findings indicated that while the immune cell compositions had been altered in KD and COV patients, and the ratio of CD14+ monocytes were both elevated in KD and COV. While the CD14+ monocytes share a large scale of same differentiated expressed geens between KD and COV. The differential activation of CD14 and CD16 monocytes was found to respond to both endothelial and epithelial dysfunctions. Furthermore, SELL+/CCR1+/XAF1+ CD14 monocytes were seen to enhance the adhesion and damage to endothelial cells. The results also showed that different types of B cells were involved in both KD and COV, while only the activation of T cells was recorded in KD. ConclusionIn conclusion, our study demonstrated the role of the innate immune response in the regulation of endothelial dysfunction in both KD and COVID-19. Additionally, our findings indicate that the adaptive immunity activation differs between KD and COVID-19. Our results demonstrate that monocytes in COVID-19 exhibit adhesion to both endothelial cells and alveolar epithelial cells, thus providing insight into the mechanisms and shared phenotypes between KD and COVID-19.

Homoplantaginin attenuates high glucose-induced vascular endothelial cell apoptosis through promoting autophagy via the AMPK/TFEB pathway.

Vascular endothelial cell (VEC) injury is a key factor in the development of diabetic vascular complications. Homoplantaginin (Hom), one of the main flavonoids from Salvia plebeia R. Br. has been reported to protect VEC. However, its effects and mechanisms against diabetic vascular endothelium remain unclear. Here, the effect of Hom on VEC was assessed using high glucose (HG)-treated human umbilical vein endothelial cells and db/db mice. In vitro, Hom significantly inhibited apoptosis and promoted autophagosome formation and lysosomal function such as lysosomal membrane permeability and the expression of LAMP1 and cathepsin B. The antiapoptosis effect of Hom was reversed by autophagy inhibitor chloroquine phosphate or bafilomycin A1. Furthermore, Hom promoted gene expression and nuclear translocation of transcription factor EB (TFEB). TFEB gene knockdown attenuated the effect of Hom on upregulating lysosomal function and autophagy. Moreover, Hom activated adenosine monophosphate-dependent protein kinase (AMPK) and inhibited the phosphorylation of mTOR, p70S6K, and TFEB. These effects were attenuated by AMPK inhibitor Compound C. Molecular docking showed a good interaction between Hom and AMPK protein. Animal studies indicated that Hom effectively upregulated the protein expression of p-AMPK and TFEB, enhanced autophagy, reduced apoptosis, and alleviated vascular injury. These findings revealed that Hom ameliorated HG-mediated VEC apoptosis by enhancing autophagy via the AMPK/mTORC1/TFEB pathway.

Activating autophagy and ferroptosis of 3‑Chloropropane‑1,2‑diol induces injury of human umbilical vein endothelial cells via AMPK/mTOR/ULK1.

3‑Chloropropane‑1,2‑diol (3‑MCPD) is an internationally recognized food pollutant. 3‑MCPD has reproductive, renal and neurotoxic properties. However, whether 3‑MCPD induces human umbilical vein endothelial cell (HUVEC) injury has not been previously reported. In the present study, HUVECs were treated using 2µg/ml 3‑MCPD for 24h at 37˚C. The effects of 3‑MCPD on HUVEC proliferation and cell cycle arrest, death and senescence were then assessed using Cell Counting Kit‑8 (CCK‑8), flow cytometry and β‑galactosidase staining, respectively. Whether 3‑MCPD induced ferroptosis was evaluated using JC‑1 and FerroOrange staining and transmission electron microscopy. A small interfering RNA targeting AMPK was used to assess whether 3‑MCPD promoted ferroptosis via AMPK signaling. The results demonstrated that 3‑MCPD inhibited HUVEC proliferation in a dose‑dependent manner and induced cell cycle arrest. Furthermore, 3‑MCPD promoted senescence in HUVECs with elevated DNA damage and cell death. The CCK‑8 results demonstrated that ferroptosis and autophagy inhibitors significantly reversed cell death caused by 3‑MCPD. Moreover, 3‑MCPD increased mitochondrial membrane potential, which indicated that 3‑MCPD contributed to mitochondrial dysfunction. 3‑MCPD also markedly increased intracellular Fe2+ levels and lipid peroxidation in HUVECs. The present study assessed the underlying mechanism by which 3‑MCPD activated autophagy and ferroptosis in HUVECs. The data demonstrated that 3‑MCPD significantly increased phosphorylation levels of AMPK and unc‑51 like autophagy activating kinase (ULK1) but significantly decreased phosphorylation of mTOR in HUVECs. Furthermore, silencing of AMPK significantly reversed the increase in autophagy, lipid peroxidation and Fe2+ induced by 3‑MCPD. In conclusion, 3‑MCPD demonstrated a significant damaging effect on HUVECs via induction of autophagy and ferroptosis; such effects may be mediated by AMPK/mTOR/ULK1 signaling. To the best of our knowledge, the present study was the first to demonstrate the mechanism of 3‑MCPD‑induced vascular endothelial cell injury and lays a molecular foundation for the prevention of 3‑MCPD‑related vascular diseases.

Rosiglitazone reduces diabetes angiopathy by inhibiting mitochondrial dysfunction dependent on regulating HSP22 expression

The effects of rosiglitazone (RSG) in patients with type 2 diabetes mellitus (T2DM) remain controversial. Here, we first used network pharmacology to identify the common targets of RSG in the treatment of diabetes angiopathy (DA). Enrichment analysis found that the common genes were involved in the inflammatory response, leukocyte cell-cell adhesion, mitochondrion organization and oxidative stress. Our previous research confirmed that heat shock protein 22 (HSP22) suppresses diabetes-induced endothelial activation and injury by inhibiting mitochondrial reactive oxygen species (mtROS) formation and dysfunction. We then constructed HSP22 knockout mice with T2DM to investigate whether RSG protected the vascular endothelium by upregulating HSP22. Our study suggested that RSG reduced vascular endothelial cell activation and injury by decreasing monocyte adhesion and cytokine secretion and simultaneously upregulating HSP22 expression. Mechanistically, RSG inhibited mitochondrial oxidative stress and dysfunction by regulating PPAR-γ in a manner partially dependent on expression of HSP22, resulting in reduced DA.

Melatonin attenuates high glucose‑induced endothelial cell pyroptosis by activating the Nrf2 pathway to inhibit NLRP3 inflammasome activation.

Endothelial injury induced by hyperglycemia is the most critical initial step in the development of diabetic vasculopathy. The aim of this present study was to explore the prevention and treatment strategies and elucidate the specific mechanism of diabetes‑induced vascular endothelial injury. Melatonin, a hormone secreted by the pineal gland to regulate biological rhythm, serves an important role in maintaining human physiological function. Pyroptosis is a type of newly discovered inflammatory cell death. The current study first found by western blotting that melatonin could activate nuclear factor erythroid 2‑related factor 2 (Nrf2) pathway in human umbilical vein endothelial cells (HUVECs) under high glucose (HG) condition. Second, it found that pretreatment with Luzindole, a specific inhibitor of melatonin receptor (MT1/MT2), significantly reduced the activation of Nrf2 pathway by melatonin in HUVECs. It also found that pretreatment with melatonin or a specific NOD‑like receptor family, pyrin domain‑containing 3 (NLRP3) inhibitor (MCC950) pretreatment reduced HG‑induced endothelial cell pyroptosis. Finally, it was found that the protective effect of melatonin against reactive oxygen species/NLRP3 inflammasome pathway activation induced by HG in HUVECs was decreased after Nrf2 knockdown. In conclusion, the present study showed that melatonin may serve a protective role in HG‑induced vascular endothelial cell pyroptosis by activating the Nrf2 pathway to inhibit NLRP3 inflammasome activation. In addition, it was further found that melatonin attenuated HG‑induced vascular endothelial cell injury by interacting with its receptors (MT1/MT2) to promote activation of Nrf2 pathway.

Circular RNA circ_0026218 Suppressed Atherosclerosis Progression via miR-338-3p/SIRT6 Axis.

Multiple circular RNAs (circRNAs) are implicated in atherosclerosis (AS) pathogenesis. In fact, how circRNA 0026218 (circ_0026218) functions in AS remains unknown, and thus the functions and mechanisms of circ_0026218 in the injury of vascular endothelial cells are to be investigated. Microarray analysis was employed to screen out differentially expressed circRNAs in AS. A cell model was mimicked by treating Human umbilical vein endothelial cells (HUVECs) with oxidized low-density lipoprotein (ox-LDL). circ_0026218, microRNA-338-3p (miR-338-3p) and silent information regulator 6 (SIRT6) expressions in HUVECs with ox-LDL treatment were probed by qRT-PCR. The cell proliferative capabilities were exposed by CCK-8 assay. The contents of interleukin 6 (IL-6), interleukin 1β (IL-1β), and tumor necrosis factor α (TNF-α) were measured by ELISA. Oxidative stress kits were utilized to detect the levels of reactive oxygen species (ROS), superoxide dismutase (SOD), and malondialdehyde (MDA). Flow cytometry was adopted to analyze the level of apoptosis of HUVECs. Dual-luciferase reporter gene assay and RIP assay were leveraged to expose the interplay between miR-338-3p and circ_0026218 or SIRT6 3'-UTR, respectively. In addition, the impacts of circ_0026216 and miR-338-3p on SIRT6 protein expressions were subjected to Western blot. circ_0026218 was greatly depleted in ox-LDL-stimulated HUVECs. circ_0026218 overexpression promoted viability of HUVECs in vitro and inhibited inflammatory response, oxidative stress, and apoptosis. circ_0026218 could adsorb miR-338-3p and positively modulated SIRT6 expressions via sponging miR-338-3p. Upregulation of this miRNA reversed the influence of circ_0026218 overexpression on ox-LDL-caused injury and apoptosis of HUVECs. Collectively, circ_0026218 upregulates SIRT6 expression through decoying miR-338-3p, thereby inhibiting ox-LDL-initiated injury of HUVECs. circ_0026218 is involved in the pathogenesis of AS.

Downregulation of miR-210-3p Attenuates High Glucose-Induced Angiogenesis of Vascular Endothelial Cells via Targeting FGFRL1

Introduction: Vascular endothelial cell injury and angiogenesis induced by hyperglycemia are the main pathological basis of vascular complications in diabetes mellitus. Our study aimed to investigate the role and mechanism of miR-210-3p in high glucose (HG)-induced angiogenesis. Methods: Human umbilical vein endothelial cells (HUVECs) were treated with HG to mimic the pathological process of hyperglycemia. HUVECs were divided into the control group, HG group, HG+inhibitor-NC group, and HG+miR-210-3p inhibitor group. Proliferation and migration were tested by wound healing assay, tube formation, and Transwell assay. Quantitation real-time PCR and Western blots were performed to determine the expression of miR-210-3p and relative proteins, respectively. Results: The level of miR-210-3p significantly increased in HUVECs treated by HG. The knockdown of miR-210-3p attenuated the tube formation, proliferation, and migration of cultured HUVECs in vitro to inhibit angiogenesis by increasing the expression of fibroblast growth factor receptor-like 1 (FGFRL1) and then attenuating the phosphorylation of signal transducer and activator of transcription 3 (STAT3), extracellular regulated protein kinases, and protein kinase B (Akt). Conclusion: Our study revealed that miR-210-3p might be a promising target for treating diabetic-associated vascular injury.

Simvastatin reduces high uric acid-induced oxidative stress and inflammatory response in vascular endothelial cells via nuclear factor E2-related factor 2 (Nrf2) signaling.

Increased oxidative stress and inflammatory response are risk factors for kidney and cardiovascular diseases in patients with hyperuricemia. Uric acid (UA) has been reported to cause inflammation and oxidative damage in cells by inhibiting the nuclear factor E2-related factor 2 (Nrf2) pathway. Notably, Simvastatin (SIM) can regulate the Nrf2 pathway, but whether SIM can regulate inflammatory response and oxidative stress in vascular endothelial cells induced by high UA via this pathway has not been clarified. To demonstrate this speculation, cell activity, as well as apoptosis, was estimated employing CCK-8 and TUNEL, respectively. Indicators of oxidative stress and inflammation were assessed by related kits and western blotting. Subsequently, the effects of SIM on signaling pathways were examined using western blotting. The result showed that after UA exposure, oxidative stress was activated and inflammation was increased, and SIM could reverse this trend. Meanwhile, SIM could inhibit high UA-induced apoptosis. In addition, western blotting results showed that SIM reversed the down-regulation of the expression of Nrf2 pathway-related proteins caused by high UA. SIM alleviated the inflammatory response as well as inhibiting oxidative stress through the Nrf2 pathway, thereby attenuating high UA-induced vascular endothelial cell injury.

Blocking CCR10 Expression Activates m6A Methylation and Alleviates Vascular Endothelial Cell Injury.

Cardiovascular disease, one of the most common types of disease in clinical practice today, carries a very high risk of disability and death. This research aims to examine genome-wide changes in injured human dermal microvascular endothelial cells (HDMECs) using the Ribonucleic Acid sequencing (RNA-Seq) technique, and to search for key genes influencing N6-methyladenosine (m6A) methylation, thus gaining new insights into future clinical diagnosis and treatment of cardiovascular diseases (CVDs) and laying a foundation for follow-up research. Impaired HDMECs (injury group), established by endotoxin intervention, were analyzed by RNA-Seq for differentially expressed genes (DEGs) relative to normal HDMECs (control group). Then, DEGs that might be associated with m6A methylation were selected for expression blocking to observe m6A methylation alterations. The migration, angiogenesis, and inflammatory response of damaged HDMECs were detected by cell scratch assay, western blotting, and Enzyme-linked Immunosorbent Assay (ELISA) experiments, respectively. In this study, 20 DEGs were screened out from the two groups by RNA-Seq, of which 17 were up-regulated and 3 were down-regulated. The C-C motif chemokine receptor 10 (CCR10) was selected for subsequent analysis. Real-Time Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR) identified elevated CCR10 expression and reduced m6A methylation levels in the injury group (p < 0.05). After blocking CCR10 expression in damaged HDMECs by BI6901 (a CCR10 specific blocker) m6A methylation, cell activity, vascular endothelial growth factor A (VEGFA) and CD31 protein expression, as well as relative length and branches of tube formation were found to be increased compared with the injury group, while the levels of inflammatory factors interleukin-1 (IL-1), interleukin-1 (IL-6) and tumor necrosis factor-α (TNF-α) were decreased (p < 0.05). Blocking CCR10 expression can activate m6A methylation, promote cell activity, inhibit inflammatory reactions and alleviate HDMEC injury, which may become a breakthrough in future diagnosis and treatment of cardiovascular diseases.

G0S2 ameliorates oxidized low-density lipoprotein-induced vascular endothelial cell injury by regulating mitochondrial apoptosis.

Oxidative low-density lipoprotein (ox-LDL)-induced endothelial cell damage is a major risk factor for atherosclerosis and its related cardiovascular diseases. The G0/G1 switch gene 2 (G0S2) is a multifunctional protein which has been poorly studied in atherosclerosis. In this study, ox-LDL was utilized to construct a human aortic endothelial cell (HAEC) injury model. It was found that ox-LDL impaired cell viability, augmented lactate dehydrogenase (LDH) release, and reduced G0S2 levels in HAECs in a dose-dependent manner. Further, G0S2 overexpression improved the viability and restrained apoptosis of HAECs treated by ox-LDL. Conversely, G0S2 depletion decreased the viability and aggravated apoptosis of HAECs treated by ox-LDL. At the molecular level, G0S2 overexpression significantly increased the secretion of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPH-Px), promoted intracellular reactive oxygen species (ROS) production and malondialdehyde (MDA) content in HAECs under either normal or ox-LDL conditions. Meanwhile, the ox-LDL-induced mitochondrial dysfunction, as demonstrated by decreased mitochondrial membrane potential, translocation of mitochondrial cytochrome c (Cyt-c) to the cytoplasm, and activation of caspase-3 and caspase-9, was significantly reversed by G0S2 overexpression. In addition, G0S2 overexpression promoted the activation of AMP-activated protein kinase (AMPK) and increased the expression of nuclear factor erythroid-2-related factor-2 (Nrf2), sirtuin 1 (SIRT1) and heme oxygenase 1 (HO-1) under normal and ox-LDL conditions. This study demonstrated that G0S2 protects against ox-LDL-induced vascular endothelial cell injury by regulating oxidative damage and mitochondrial homeostasis and may be a promising target for the treatment of atherosclerosis.

The KLF4-p62 axis prevents vascular endothelial cell injury via the mTOR/S6K pathway and autophagy in diabetic kidney disease.

Diabetic kidney disease (DKD) is a complication of systemic diabetic microangiopathy, which has a high risk of developing into end-stage renal disease and death. This study explored the mechanism underlying autophagy in DKD vascular endothelial cell injury. DKD and vascular endothelial cell injury models were established using Sprague Dawley rats and human umbilical vein endothelial cells (HUVECs). HUVECs overexpressing Kruppel-like factor 4 (KLF4) were constructed by transient transfection of plasmids. Biochemical determination of urinary protein and blood urea nitrogen (BUN), superoxide dismutase (SOD), and creatinine (Scr) levels was performed. Renal pathology was observed by periodic acid-Schiff (PAS) staining. Cell Counting Kit-8 (CCK8), terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL), and immunocytochemistry (ICC) were used to analyse the growth and apoptosis of HUVECs. Microtubule-associated protein light chain 3 (LC3) expression was observed by immunofluorescence (IF). The reactive oxygen species (ROS) levels were measured using flow cytometry. Monocyte chemoattractant protein-1 (MCP-1), KLF4, and tumour necrosis factor alpha (TNF-α) levels were detected using enzyme-linked immunosorbent assay (ELISA). The expression of KLF4, p62 protein, and LC3 was analysed using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR). S6 kinase (S6K), p70 ribosomal S6 kinase (p-S6K), Beclin1, ATG5, LC3, p62, Caspase-3, mammalian target of rapamycine (mTOR), and phsophorylated mTOR (p-mTOR) expressions were detected by western blotting. PAS-positive substances (polysaccharide and glycogen) and S6K protein levels increased, and LC3 protein expression decreased in DKD rats. The levels of urinary protein, BUN, and Scr increased, and KLF4 decreased in DKD rats. High glucose (HG) levels decreased the proliferation and increased the apoptosis rate of HUVECs. The expression of ROS, TNF-α, MCP-1, and p62 increased, while the expression of SOD, KLF4, Beclin1, ATG5, and LC3 decreased in HG-induced HUVECs. KLF4 overexpression significantly increased Beclin1, ATG5, and LC3 protein expression and decreased p62 protein expression compared to the oe-NC group in HG-induced HUVECs. KLF4 overexpression inhibits the expression of Caspase-3, p-mTOR, and p-S6K in HG-induced HUVECs. KLF4-p62 axis improved vascular endothelial cell injury by regulating inflammation and the mTOR/S6K pathway in DKD.

Extracellular vesicle-mediated transfer of lncRNA CLDN10-AS1 aggravates low-density lipoprotein-induced vascular endothelial injury.

Oxidized low-density lipoprotein (ox-LDL) stimulation impairs the oxidation-reduction equilibrium in vascular endothelial cells (VECs) and contributes to atherosclerosis (AS). This study probed the mechanisms of extracellular vesicle (EV)-mediated transfer of lncRNA CLDN10 antisense RNA 1 (CLDN10-AS1) in ox-LDL-induced VEC injury. Initially, VEC injury models were established by treating human umbilical vein endothelial cells (HUVECs) with ox-LDL. EVs were isolated from HUVECs (HUVECs-EVs) and identified. CLDN10-AS1, microRNA (miR)-186, and Yin Yang 1 (YY1) expressions in ox-LDL-treated HUVECs and EVs derived from these cells (ox-EVs) were measured. HUVECs were incubated with EVs, after which the cell viability, apoptosis, and concentrations of proinflammatory cytokines and oxidative stress markers were measured. We discovered that CLDN10-AS1 and YY1 were upregulated in ox-LDL-treated HUVECs, whereas miR-186 was downregulated. ox-EVs treatment elevated CLDN10-AS1 expression in HUVECs and ox-EVs overexpressing CLDN10-AS1 promoted VEC injury. Besides, CLDN10-AS1 is competitively bound to miR-186 and promoted YY1 expression. Rescue experiments revealed that miR-186 overexpression or YY1 suppression partially reversed the roles of ox-EVs overexpressing CLDN10-AS1 in ox-LDL-induced VEC injury. Lastly, clinical serum samples were collected for verification. Overall, CLDN10-AS1 carried by HUVECs-EVs into HUVECs competitively bound to miR-186 to elevate YY1 expression, thereby aggravating ox-LDL-induced VEC injury.

HDAC inhibitors alleviate uric acid-induced vascular endothelial cell injury by way of the HDAC6/FGF21/PI3K/AKT pathway.

Uric acid (UA) accumulation triggers endothelial dysfunction, oxidative stress and inflammation. Histone deacetylase (HDAC) play a vital role in regulating the pathological processes of various diseases. However, the influence of HDAC inhibitor on UA-induced vascular endothelial cell injury (VECI) remains undefined. Hence, this study aimed to investigate the effect of HDACs inhibition on UA-induced vascular endothelial cell dysfunction and its detailed mechanism. UA was employed to induce human umbilical vein endothelial cell (HUVEC) injury. Meanwhile, potassium oxonate- and hypoxanthine-induced hyperuricemia mouse models were also constructed. A broad-spectrum HDAC inhibitor trichostatin A (TSA) or selective HDAC6 inhibitor Tubsastain A (TubA) was given to HUVECs or mice to determine whether HDACs can affect UA-induced VECI. Results showed pretreatment of HUVECs with TSA or HDAC6 knockdown attenuated UA-induced VECI and increased FGF21 expression and phosphorylation of AKT, eNOS and FoxO3a. These effects could be reversed by FGF21 knockdown. In vivo, both TSA and TubA reduced inflammation and tissue injury while increased FGF21 expression and phosphorylation of AKT, eNOS and FoxO3a in the aortic and renal tissues of hyperuricemia mice. Therefore, HDACs, especially HDAC6 inhibitor, alleviated UA-induced VECI through upregulating FGF21 expression and then activating PI3K/AKT pathway. This suggests that HDAC6 may serve as a novel therapeutic target for treating UA-induced endothelial dysfunction.

Bone marrow stromal cells attenuate oxygen and glucose deprivation followed by re‑oxygenation‑induced brain microvascular endothelial cell injury.

Ischemic stroke is a severe threat to the health of older individuals. Bone marrow mesenchymal stem cells (BMSCs) have been implicated in ischemic stroke. Urokinase‑type plasminogen activator (uPA) and its specific receptor (uPAR) are associated with the pathological process of ischemic stroke. However, the relationship between BMSCs and uPA/uPAR in ischemic stroke remains unclear. For simulating the occurrence of an ischemic stroke in vitro, human cerebral microvascular endothelial cells (HBMECs) were subjected to oxygen and glucose deprivation followed by re‑oxygenation (OGD/R) and were then cocultured with BMSCs. 3,4,5‑dimethylthiazol‑2,5‑diphenyltetrazolium bromide and bromodeoxyuridine staining were used for measuring cell viability and proliferation. Flow cytometry was performed for assessing cell apoptosis. Endothelial cell tube formation was determined using angiogenesis assays. Alterations in the protein and gene expression in HBMECs were evaluated using western blot analysis and quantitative reverse transcription‑polymerase chain reaction, respectively. OGD/R considerably inhibited the viability and proliferation of HBMECs by inducing apoptosis, which was reversed by BMSCs. Consistently, OGD/R‑induced inhibition of angiogenesis was attenuated by BMSCs. In addition, BMSCs could protect HBMECs against OGD/R‑induced injury by positively regulating the uPA/uPAR/stromal cell‑derived factor‑1α (SDF‑1α)/C‑X‑C chemokine receptor type 4 (CXCR4) pathway, and uPA/uPAR could mediate the SDF‑1α/CXCR4 pathway in OGD/R‑treated HBMECs. Therefore, this study provides novel strategies to investigate the specific role of BMSCs in ameliorating OGD/R‑induced vascular endothelial cell injury.

Study on the mechanism of paeonol in improving low-density lipoprotein-induced human vascular endothelial cell injury

To investigate the effects of paeonol on low-density lipoprotein-induced human vascular endothelial cell injury and its molecular mechanisms. Human umbilical vein endothelial cells (HUVECs) were divided into 9 groups, normal control (NC) group, ox-LDL group (100 ng/L ox-LDL), low, medium, and high-dose paeonol groups (60 μmol/L, 120 μmol/L, 240 μmol/L paeonol+100 ng/L ox-LDL), ox-LDL+small interfering RNA negative control (si-NC) group, ox-LDL+circ_0003204 small interfering RNA (si-circ_0003204) group, middle dose group+ox-LDL+circ_0003204 overexpression negative control (pcDNA-NC) group, middle dose group+ox-LDL+circ_0003204 overexpression (pcDNA-circ_0003204) group, three replicate wells in each group. MTT flow cytometry, and Western blot were used to detect cell proliferation, apoptosis and protein (CDK2, Bcl2, p27, Bax) expressions, respectively. Malondialdehyde (MDA) and superoxide dismutase (SOD) kit were used to detect MDA content and SOD activity; real-time quantitative PCR (RT-qPCR) was used to detect the expression of circ_0003204. Compared with the NC group, the proliferation activity, protein expressions of CDK2 and Bcl2, and SOD activity of HUVECs in the ox-LDL group were decreased significantly (P＜0.05), and the apoptosis rate, protein expressions of p27 and Bax, MDA content, and circ_0003204 expression were increased significantly (P＜ 0.05). Compared with the ox-LDL group, the proliferation activity, protein (CDK2, Bcl2) expressions and SOD activity of HUVECs in the low, medium and high dose paeonol groups were increased significantly (P＜0.05), and the apoptosis rate, protein (p27, Bax) expressions, MDA content And circ_0003204 expression were decreased significantly (P＜ 0.05). Compared with ox-LDL+si-NC group, the proliferation activity, protein (CDK2, Bcl2) expressions, SOD activity of HUVECs in ox-LDL+si-circ_0003204 group were increased significantly (P＜0.05), the apoptosis rate, protein (p27, Bax) expressions, and the content of MDA were decreased significantly (P＜0.05). Compared with the middle-dose+ox-LDL+pcDNA-NC group, the HUVECs proliferation activity, protein (CDK2, Bcl2) expressions, and SOD activity in the middle-dose+ox-LDL+pcDNA-circ_0003204 group were decreased significantly (P＜0.05), and the levels of circ_0003204, apoptosis rate, protein (p27, Bax) expressions and MDA content were increased significantly (P＜0.05). Paeonol can inhibit ox-LDL-induced apoptosis and oxidative stress of human umbilical vein endothelial cells, and alleviate human umbilical vein endothelial cell injury. The mechanism of action may be related to the down-regulation of circ_0003204 expression.