1,8-桉叶油素调节自噬改善高糖诱导的内皮细胞损伤

目的探讨1,8-桉叶油素(1,8-Cineole)通过调节自噬来改善高糖(HG)诱导人主动脉内皮细胞(HAECs)损伤的保护作用。方法采用HG作用HAECs 60 h,不同剂量的1,8-Cineole进行干预,运用噻唑蓝(MTT)法检测细胞活力,乳酸脱氢酶(LDH)检测细胞毒性,Western blot法检测Beclin1、LC3-Ⅱ/Ⅰ、p62、caspase-3和caspase-9的蛋白表达,及使用自噬抑制剂氯喹(CQ)作用HAECs后,检测Beclin1、LC3-Ⅱ/Ⅰ、p62、caspase-3和caspase-9的表达情况。结果1,8-Cineole提高细胞活力,减少LDH含量,激活自噬和抑制细胞凋亡。与空白组比较,CQ组的Beclin1、LC3-Ⅱ/Ⅰ、p62、caspase-3和caspase-9表达增加,同时观察到CQ+HG+CH组与CQ+HG组比较时,以上蛋白差异均无统计学意义。结论1,8-Cineole对高糖诱导HAECs损伤具有保护作用,其作用与改善自噬流相关。     

Hepatotoxicity and endothelial dysfunction induced by high choline diet and the protective effects of phloretin in mice

The involvement of choline and its metabolite trimethylamine-N-oxide (TMAO) in endothelial dysfunction and atherosclerosis has been repeatedly confirmed. Phloretin, a dihydrochalcone flavonoid usually present in apples, possesses a variety of biological activities including vascular nutrition. This study was designed to investigate whether phloretin could alleviate or prevent high choline-induced vascular endothelial dysfunction and liver injury in mice. Mice were provided with 3% high choline water and given phloretin orally daily for 10 weeks. The high choline-treated mice showed the significant dyslipidemia and hyperglycemia with the impaired liver and vascular endothelium (p < 0.01). Administration of phloretin at 200 and 400 mg/kg bw significantly reduced the choline-induced elevation of serum TC, TG, LDL-C, AST, ALT, ET-1 and TXA2 (p < 0.01), and markedly antagonized the choline-induced decrease of serum PGI2, HDL-C and NO levels. Furthermore, phloretin elevated hepatic SOD and GSH-Px activities and decreased hepatic MDA levels of the mice exposed to high choline water. Moreover, histopathological test with the H&E and Oil Red O staining of liver sections confirmed the high choline diet-caused liver steatosis and the hepatoprotective effect of phloretin. These findings suggest that high choline causes oxidative damage, and phloretin alleviate vascular endothelial dysfunction and liver injury.     

Ameliorative effect of berberine on endothelial dysfunction in diabetic rats induced by high-fat diet and streptozotocin

Berberine can improve insulin resistance, lower blood glucose, and regulate lipid metabolism disorders which cause endothelial dysfunction, leading to vascular complications of type 2 diabetes mellitus. The aim of the present study was to investigate the effects of berberine on endothelial dysfunction of aortas in type 2 diabetes mellitus rats and its mechanism. Wistar rats were randomly divided into four groups: diabetic rats, control rats, diabetic rats treated with berberine (100 mg/kg), and control rats treated with berberine. The serum fasting blood glucose, insulin, total cholesterol, triglyceride and nitric oxide (NO) levels were tested. Acetylcholine-induced endothelium-dependent relaxation and sodium nitroprusside induced endothelium-independent relaxation were measured in aortas for estimating endothelial function. The expression of endothelial nitric oxide synthase (eNOS) mRNA was measured by RT-PCR, and the protein expressions of eNOS and NADPH oxidase (NOX4) were analyzed by western blot. The results showed that berberine significantly decreased fasting blood glucose, and triglyceride levels in diabetic rats. Berberine also improved endothelium-dependent vasorelaxation impaired in aorta. The expressions of eNOS mRNA and protein were significantly increased, while NOX4 protein expression was decreased in aortas from diabetic rats with berberine treatment. Moreover, serum NO levels were elevated after berberine treatment. In conclusion, berberine restores diabetic endothelial dysfunction through enhanced NO bioavailability by up-regulating eNOS expression and down-regulating expression of NADPH oxidase.     

Rosiglitazone attenuates indoxyl sulphate‐induced endothelial dysfunction

Indoxyl sulphate is a protein‐bound uraemic toxin that has deleterious effects on the cardiovascular system. Rosiglitazone (RGZ) is an insulin sensitizer used for glycaemic control in type 2 diabetes. Rosiglitazone has been shown to be beneficial for cardiovascular disease because of its pleiotropic effects. Whether RGZ can improve indoxyl sulphate‐induced endothelial damage has not been investigated. In the present in vitro study, we examined the effects of RGZ on indoxyl sulphate‐induced endothelial injury. Endothelial cells were exposed to RGZ (5 and 10 μmol/L) and then treated with indoxyl sulphate (100 and 1000 μmol/L) for 48 h. Indoxyl sulphate upregulated intracellular cell adhesion molecule‐1, vascular cell adhesion molecule‐1 and monocyte chemotactic protein‐1 expression. Indoxyl sulphate also increased the abundance of NADPH oxidase 4 (NOX4) and nuclear factor (NF)‐κB. Both extracellular signal‐regulated kinase (ERK) 1/2 and p38 mitogen‐activated protein kinase (MAPK) signalling pathways were activated after 48 h treatment with indoxyl sulphate. Pretreatment of cells with both concentrations of RGZ improved indices of endothelial injury. In addition, RGZ attenuated the increase in NOX4 and NF‐κB and prevented the activation of the ERK1/2 and p38 MAPK signalling pathways. We conclude that RGZ ameliorates indoxyl sulphate‐induced endothelial injury.     

ZnS nanoarchitectures induced dysfunction of vascular endothelial cells in vitro and in vivo

ZnS nanoarchitectures have been intensively investigated recently because of their applications in optoelectronics and adsorption capacity. The potential hazard of ZnS nanoarchitectures is not well known. In this study, we investigated the toxicity of ZnS nanoarchitectures on vascular endothelial cell (VEC) in vitro and in vivo. The results showed that ZnS could inhibit human umbilical vein endothelial cell (HUVEC) proliferation at 50 and 200 μg/mL. Endothelial nitric oxide synthase (eNOS) activity, nitric oxide (NO), and reactive oxygen species productions were increased, which was companied with the decrease in caveolin‐1 level. The endothelium of the aortic root was damaged and the NO levels in serum were elevated in the mice treated with 5 or 10 mg/kg ZnS for 3 and 6 days, but the body could repair the damage. The data suggested that the high concentration of ZnS could induce dysfunction of VECs through decreasing caveolin‐1 and elevation of the eNOS activity and thus present toxicity. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 755–768, 2015.     

Lipocalin-2 deficiency prevents endothelial dysfunction associated with dietary obesity: role of cytochrome P450 2C inhibition

BACKGROUND AND PURPOSE

Lipocalin-2 is a pro-inflammatory adipokine up-regulated in obese human subjects and animal models. Its circulating levels are positively correlated with the unfavourable lipid profiles, elevated blood pressure and insulin resistance index. Augmented lipocalin-2 has been found in patients with cardiovascular abnormalities.The present study was designed to investigate the role of lipocalin-2 in regulating endothelial function and vascular reactivity.

EXPERIMENTAL APPROACH

Wild-type and lipocalin-2 knockout (Lcn2-KO) mice were fed with either a standard chow or a high-fat diet. Blood pressures and endothelium-dependent relaxations/contractions were monitored at 2 week intervals.

RESULTS

Systolic blood pressure was elevated by high-fat diet in wild-type mice but not in Lcn2-KO mice. Endothelial dysfunction, reflected by the impaired endothelium-dependent relaxations to insulin and augmented endothelium-dependent contractions to ACh, was induced by high-fat diet in wild-type mice. In contrast, Lcn2-KO mice were largely protected from the deterioration of endothelial function caused by dietary challenges. The eNOS dimer/monomer ratio, NO bioavailability, basal and insulin-stimulated PKB/eNOS phosphorylation responses were higher in aortae of Lcn2-KO mice. Administration of lipocalin-2 attenuated endothelium-dependent relaxations to insulin and promoted endothelium-dependent contractions to ACh. It induced eNOS uncoupling and elevated COX expression in the arteries. Treatment with sulphaphenazole, a selective inhibitor of cytochrome P450 2C9, improved endothelial function in wild-type mice and blocked the effects of lipocalin-2 on both endothelium-dependent relaxations to insulin and endothelium-dependent contractions to ACh, as well as eNOS uncoupling.

CONCLUSIONS

Lipocalin-2, by modulating cytochrome P450 2C9 activity, is critically involved in diet-induced endothelial dysfunction.     

Interleukin-2 protects against endothelial dysfunction induced by high glucose levels in rats

AIMS: Interleukin-2 (IL-2) can modulate cardiovascular functions, but the effect of IL-2 on vascular endothelial function in diabetes is not known. We hypothesized that IL-2 may attenuate endothelial dysfunction induced by high glucose or diabetes. So the aim of this study was to investigate the effect of IL-2 on endothelium-response of aortas incubated with high glucose or from diabetic rats and its underlying mechanism. METHODS: Acetylcholine (ACh)-induced endothelium-dependent relaxation (EDR), sodium nitroprusside (SNP)-induced endothelium-independent relaxation (EIR), superoxide dismutase (SOD) and nitric oxide synthase (NOS) were measured in aortas isolated from non-diabetic rats and exposed to a high glucose concentration and from streptozotocin-induced diabetic rats. RESULTS: Incubation of aortic rings with high glucose (44 mM) for 4 h resulted in a significant inhibition of EDR, but had no effects on EIR. Co-incubation with IL-2 for 40 min prevented the inhibition of EDR caused by high glucose in a concentration-dependent manner. Similarly, high glucose decreased SOD and NOS activity in aortic tissue. IL-2 (1000 U/ml) significantly attenuated the decrease of SOD and NOS activity caused by high glucose. In addition, EDR declined along with the decrease of serum NO level in aortas from STZ-induced diabetic rats. Injection of IL-2 (5000 and 50,000 U kg(-1) d(-1), s.c.) for 5 weeks prevented the inhibition of EDR and the decrease of serum NO levels caused by diabetes. CONCLUSIONS: IL-2 significantly ameliorated the endothelial dysfunction induced by hyperglycemia, in which the activation of the NO pathway and SOD may be involved.     

木犀草素减轻氧化应激引起的血管内皮损伤

目的：探讨木犀草素对叔丁基过氧化氢致血管内皮损伤的保护作用及相关机制。方法：首先通过制备大鼠胸主动脉环,观察木犀草素对叔丁基过氧化氢所致血管张力变化的影响;再采用叔丁基过氧化氢诱导血管内皮细胞氧化损伤模型,观察木犀草素对其细胞形态学变化及细胞活力的影响,并用RT-PCR检测eNOS和COX-1 mRNA的含量变化。结果：木犀草素能够浓度依赖性地对抗叔丁基过氧化氢导致的血管舒张功能损伤及细胞损伤作用,且浓度依赖性地减弱叔丁基过氧化氢对内皮细胞eNOS mRNA表达抑制的影响。结论：木犀草素是一种有效的舒血管物质,它可以起到抗氧化的作用,减轻氧化应激反应,并可能通过维持eNOS活性等血管内皮途径舒张血管。     

Na+/H+ exchanger inhibitor prevented endothelial dysfunction induced by high glucose

The aims of this study were to examine whether cariporide, a selective Na+/H+ exchanger inhibitor, has protective effects against endothelial dysfunction induced by high glucose in vitro and to investigate the potential mechanisms. Exposure of rat aorta rings to high glucose (44 mmol/L) for 6 hours caused an inhibition of acetylcholine-induced endothelium-dependent relaxation but had no effect on sodium nitroprusside-induced endothelium-independent relaxation. Treatment with cariporide (0.01, 0.1, 1 micromol/L) of aortic rings incubated with high-glucose medium attenuated the impaired endothelium-dependent relaxation in a dose-dependent manner. Furthermore, high glucose resulted in an increase of malondialdehyde and a decrease of superoxide dismutase activity in rat aorta rings, and these effects were reversed by cariporide. In addition, cariporide was able to inhibit the activation of Na+/H+ exchanger induced by high glucose in cultured endothelial cells. These findings suggest that the endothelial dysfunction induced by high glucose in vitro is caused by the activation of Na+/H+ exchanger.     

动脉粥样硬化过程中内皮细胞功能障碍及药物治疗

动脉粥样硬化与血管内皮细胞之间关系密切，越来越多的证据表明，内皮细胞功能障碍是动脉粥样硬化形成早期的始动环节。本文就内皮细胞功能障碍与动脉粥样硬化的关系以及各类药物尤其是他汀类药物对内皮细胞功能障碍的治疗作用作一介绍。     

Chronic administration of BMS309403 improves endothelial function in apolipoprotein E-deficient mice and in cultured human endothelial cells

BACKGROUND AND PURPOSE

Adipocyte fatty acid-binding protein (A-FABP) is up-regulated in regenerated endothelial cells and modulates inflammatory responses in macrophages. Endothelial dysfunction accompanying regeneration is accelerated by hyperlipidaemia. Here, we investigate the contribution of A-FABP to the pathogenesis of endothelial dysfunction in the aorta of apolipoprotein E-deficient (ApoE^−/−) mice and in cultured human endothelial cells.

EXPERIMENTAL APPROACH

A-FABP was measured in aortae of ApoE^−/−mice and human endothelial cells by RT-PCR, immunostaining and immunoblotting. Total and phosphorylated forms of endothelial nitric oxide synthase (eNOS) were measured by immunoblotting. Changes in isometric tension were measured in rings of mice aortae

KEY RESULTS

A-FABP was expressed in aortic endothelium of ApoE^−/− mice aged 12 weeks and older, but not at 8 weeks or in C57 wild-type mice. Reduced endothelium-dependent relaxations to acetylcholine, UK14304 (selective α₂-adrenoceptor agonist) and {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187 (calcium ionophore) and decreased protein presence of phosphorylated and total eNOS were observed in aortae of 18 week-old ApoE^−/− mice compared with age-matched controls. A 6 week treatment with the A-FABP inhibitor, BMS309403, started in 12 week-old mice, improved endothelial function, phosphorylated and total eNOS and reduced plasma triglyceride levels but did not affect endothelium-independent relaxations. The beneficial effect of BMS309403 on UK14304-induced relaxations was attenuated by Pertussis toxin. In cultured human microvascular endothelial cells, lipid-induced A-FABP expression was associated with reduced phosphorylated eNOS and NO production and was reversed by BMS309403.

CONCLUSIONS AND IMPLICATIONS

Elevated expression of A-FABP in endothelial cells contributes to their dysfunction both in vivo and in vitro.     

Reversal of SIN-1-induced eNOS dysfunction by the spin trap,DMPO, in bovine aortic endothelial cells via eNOS phosphorylation

Background and Purpose

Nitric oxide (NO) derived from eNOS is mostly responsible for the maintenance of vascular homeostasis and its decreased bioavailability is characteristic of reactive oxygen species (ROS)-induced endothelial dysfunction (ED). Because 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), a commonly used spin trap, can control intracellular nitroso-redox balance by scavenging ROS and donating NO, it was employed as a cardioprotective agent against ED but the mechanism of its protection is still not clear. This study elucidated the mechanism of protection by DMPO against SIN-1-induced oxidative injury to bovine aortic endothelial cells (BAEC).

Experimental Approach

BAEC were treated with SIN-1, as a source of peroxynitrite anion (ONOO⁻), and then incubated with DMPO. Cytotoxicity following SIN-1 alone and cytoprotection by adding DMPO was assessed by MTT assay. Levels of ROS and NO generation from HEK293 cells transfected with wild-type and mutant eNOS cDNAs, tetrahydrobiopterin bioavailability, eNOS activity, eNOS and Akt kinase phosphorylation were measured.

Key Results

Post-treatment of cells with DMPO attenuated SIN-1-mediated cytotoxicity and ROS generation, restoration of NO levels via increased in eNOS activity and phospho-eNOS levels. Treatment with DMPO alone significantly increased NO levels and induced phosphorylation of eNOS Ser¹¹⁷⁹ via Akt kinase. Transfection studies with wild-type and mutant human eNOS confirmed the dual role of eNOS as a producer of superoxide anion (O₂⁻) with SIN-1 treatment, and a producer of NO in the presence of DMPO.

Conclusion and Implications

Post-treatment with DMPO of oxidatively challenged cells reversed eNOS dysfunction and could have pharmacological implications in the treatment of cardiovascular diseases.     

川芎嗪对高糖诱导的人视网膜血管内皮细胞增殖的影响

目的 观察川芎嗪对高糖诱导的人视网膜血管内皮细胞（HRCECs）增殖的影响。方法 将HRCECs细胞分为对照（生理盐水）组、模型（25 mmol/L葡萄糖）组和川芎嗪低、中、高浓度（50、100、200 μmol/L）组,培养48 h后,MTT细胞毒实验检测细胞增殖变化,流式细胞技术检测细胞周期,ELISA法检测血管内皮生长因子（VEGF）表达。结果 与对照组比较,高糖对HRCECs细胞增殖、分裂（M）期比例、上清VEGF浓度均具有显著促进作用（P<0.05、0.01）;与模型组比较,低、中、高浓度川芎嗪对高糖诱导的HRCECs细胞增殖、分裂（M）期比例、上清VEGF浓度均发挥显著抑制作用（P<0.05、0.01）,且呈浓度相关性。结论 川芎嗪可能通过抑制高糖诱导的HRCECs细胞VEGF高表达,阻滞细胞周期,发挥抑制HRCECs细胞增殖的作用。     

Folic acid reverses endothelial dysfunction induced by inhibition of tetrahydrobiopterin biosynthesis

While folic acid has been shown to reverse endothelial dysfunction, the exact underlying mechanism remains elusive. Here, folic acid reversed both the endothelial dysfunction and increased production of superoxide following depletion of rabbit aortic ring tetrahydrobiopterin (BH₄) levels with 2,4-diamino-6-hydroxy-pyrimidine (DAHP) and N-acetyl-5-hydroxy-tryptamine (NAS). Incubation with l-nitroarginine methyl ester also attenuated the production of superoxide. DAHP and NAS reduced BH₄ concentrations in both aorta and cultured porcine aortic endothelial cells. Folic acid had no effect on BH₄ concentrations in either preparation. The superoxide anion scavenger Tiron but not folic acid reversed the endothelial dysfunction produced in aortic rings by inhibition of copper–zinc superoxide dismutase with diethyldithiocarbamic acid. Neither folic acid nor its metabolite 5-methyltetrahydrofolate prevented the in vitro oxidation of BH₄. This study demonstrates that folic acid reverses the endothelial dysfunction induced by BH₄ depletion independently of either the regeneration or stabilization of BH₄ or an antioxidant effect.     

Simvastatin prevents inflammation-induced aortic stiffening and endothelial dysfunction

AIMS

The aim of this study was to determine whether simvastatin would protect against inflammation-induced aortic stiffening and endothelial dysfunction.

METHODS

Aortic pulse wave velocity (aPWV) and flow-mediated dilatation (FMD) were assessed three times, at baseline, after a 14 day administration of simvastatin or placebo and 8 h after Salmonella typhi vaccination in 50 healthy subjects.

RESULTS

Following vaccination there was a significant increase in aPWV in the placebo group (5.80 ± 0.87 vs. 6.21 ± 0.97 m s⁻¹, 95% CI 0.19, 0.62, P = 0.002) but not the simvastatin group (5.68 ± 0.73 vs. 5.72 ± 0.74 m s⁻¹, 95% CI −0.19, 0.27, P = 0.9; P = 0.016 for comparison). Whereas FMD response was reduced in the placebo group (6.77 ± 4.10 vs. 5.27 ± 2.88%, 95% CI −2.49, −0.52, P = 0.02) but not in the simvastatin group (7.07 ± 4.37 vs. 7.17 ± 9.94%, 95% CI −1.1, 1.3. P = 0.9, P < 0.001 for comparison). There was no difference in the systemic inflammatory response between groups following vaccination. However, there was a significant reduction in serum apolipoprotein A-I (Apo A-I) in the placebo, but not in the simvastatin, group.

CONCLUSIONS

Simvastatin prevents vaccination-induced aortic stiffening and endothelial dysfunction. This protective mechanism may be due to preservation of the Apo A-I lipid fraction, rather than pleiotropic anti-inflammatory effects of statins.     

The effect of pitavastatin calcium on endothelial dysfunction induced by hypercholesterolemia

Objective: To investigate whether endothelial function can be improved by the treatment of pitavastatin calcium via its antioxidant properties in hypercholesteremia patients.

Methods: Forty patients with hypercholesteremia were randomized to receive pitavastatin calcium 1 or 2 mg/day for 8 weeks. Among them, four people were lost in the follow-up period. Before and after treatment, clinical and biochemical characteristics, markers of oxidative stress (plasma 8-iso-prostaglandin F_2α and serum gp91phox) were determined and concomitantly endothelium-dependent brachial artery flow-mediated dilation (FMD) was measured by ultrasound examination. Thirty healthy subjects were chosen as controls.

Results: For individuals with hypercholesteremia, total cholesterol, low-density lipoprotein cholesterol (LDL-C) and serum gp91phox were significantly increased (p < 0.001 for all) and plasma 8-iso-prostaglandinF2α (8-iso-PGF2α) was significantly higher (p < 0.05), while FMD was obviously impaired (p < 0.001). Total cholesterol, LDL-C and serum gp91phox were significantly reduced (p < 0.001 for all), plasma 8-iso-PGF2α was lower and FMD was significantly improved after pitavastatin calcium treatment compared with those before treatment in any group (p < 0.05 for both). However, there was no significant difference between the 1-mg and 2-mg pitavastatin calcium groups post-therapy.

Conclusions: Endothelial dysfunction induced by hypercholesteremia can be ameliorated by pitavastatin calcium treatment, which occurs in part through its antioxidative properties.     

Atorvastatin inhibits homocysteine-induced dysfunction and apoptosis in endothelial progenitor cells

Aim:

To investigate the protective effects of atorvastatin on homocysteine (Hcy)-induced dysfunction and apoptosis in endothelial progenitor cells (EPCs) and the possible molecular mechanisms.

Methods:

EPCs were divided into six groups: Hcy treatment groups (0, 50, and 500 μmol/L) and atorvastatin pretreatment groups (0.1, 1, and 10 μmol/L). EPC proliferation, migration, in vitro vasculogenesis activity, and apoptosis rate were assayed by the MTT assay, modified Boyden chamber assay, in vitro vasculogenesis kit, and AnnexinV-FITC apoptosis detection kit, respectively. The level of reactive oxygen species (ROS) in cells was measured using H₂DCF-DA as a fluorescence probe. The activity of NADPH oxidase was evaluated with lucigenin-enhanced chemiluminescence. NO in the supernatant was detected by the nitrate reductase assay. The eNOS mRNA expression and p-eNOS, p-Akt, p-p38MAPK protein expression were measured by RT-PCR and Western blotting analysis, respectively. Caspase-3 activity was determined by colorimetric assay.

Results:

Hcy does-dependently impaired the proliferation, migration and in vitro vasculogenesis capacity of EPCs, induced cell apoptosis, increased ROS accumulation and NADPH oxidase activation, and decreased the secretion of NO compared with the control group (P<0.05 or P<0.01). The detrimental effects of Hcy were attenuated by atorvastatin pretreatment. Furthermore, Hcy caused a significant downregulation of eNOS mRNA, p-eNOS, and p-Akt protein expression as well as an upregulation of p-p38MAPK protein expression and caspase-3 activity. These effects of Hcy on EPCs were reversed by atorvastatin in a does-dependent manner.

Conclusion:

Atorvastatin inhibited homocysteine-induced dysfunction and apoptosis in endothelial progenitor cells, which may be related to its effects on suppressing oxidative stress, up-regulating Akt/eNOS and down-regulating the p38MAPK/caspase-3 signaling pathway.