二甲双胍激活Sirt3信号抑制高糖诱导内皮细胞衰老的实验研究

目的: 研究二甲双胍抑制高糖诱导人脐静脉内皮细胞(human umbilical vein endothelial cells, HUVECs)衰老的作用与机制。方法: 传代培养HUVECs,高糖刺激HUVECs 48 h建立血管内皮细胞衰老模型,采用噻唑蓝法(MTT)检测HUVECs活力,2',7'-二氢二氯荧光素二乙酸酯(DCFH-DA)为荧光探针检测细胞内活性氧(ROS)水平,硫代巴比妥酸法检测丙二醛(MDA)含量,酶联免疫吸附实验(ELISA)检测细胞培养上清液中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)的分泌量,β-半乳糖苷酶染色检测细胞衰老水平。使用Sirt3抑制剂3-TYP作用HUVECs后,蛋白免疫印迹法(Western blot)检测p53、PAI-1及Sirt3的蛋白表达水平。结果: 与正常对照组相比,高糖组中HUVECs的活力降低,细胞内ROS的水平和MDA含量增加,IL-6、TNF-α的分泌量增加,衰老阳性细胞的百分比增加,二甲双胍干预后能明显改善高糖诱导HUVECs的上述变化。3-TYP能显著逆转二甲双胍对p53、PAI-1、Sirt3蛋白表达的调控作用。结论: 二甲双胍对高糖诱导的HUVECs衰老具有明显改善作用,其作用机制与激活Sirt3密切相关。     

磷酸二酯酶3型抑制剂西洛他唑对TNF-α诱导人脐静脉内皮细胞释放可溶性粘附分子的影响

目的研究新型磷酸二酯酶3型抑制剂西洛他唑(cilostazol)对TNF-α诱导人脐静脉内皮细胞(HUVECs)释放可溶性细胞粘附分子(sCAMs)的影响,并探讨其作用机制.方法体外培养第4～6代HUVECs,以TNF-α (10 μg*L-1)刺激细胞,并与西洛他唑(1～10 μmol*L-1)共培养 24 h,取培养上清,通过ELISA法测定可溶性血管细胞粘附分子-1(sVCAM-1)、细胞间粘附分子-1(sICAM-1)以及E-选择素(sELAM-1, sE-selectin),并以四唑蓝(MTT)法考察细胞生长状态.结果1～10 μmol*L-1的西洛他唑对TNF-α诱导的HUVECs释放sICAM-1和sE-selectin无明显影响,但显著抑制sVCAM-1的生成,并且该作用被一种非选择性一氧化氮合成酶(NOS)抑制剂Lω-NAME(0.1 μmol*L-1)所阻断.MTT法测定结果显示,西洛他唑作用于HUVECs 24 h,低浓度(1 μmol*L-1)可显著改善细胞生长状态;高浓度(30 μmol*L-1)表现为抑制,而中浓度(10 μmol*L-1)对细胞生长状态几乎无影响.结论西洛他唑显著抑制由TNF-α诱导的HUVECs释放sVCAM-1,该作用可能与激活NOS并通过NO依赖性通路介导有关,提示该药可在一定程度上对抗由细胞因子所引起的部分粘附反应,因此在动脉粥样硬化以及其它心血管疾病的治疗中具有潜在的应用价值.     

白藜芦醇抑制人牙龈成纤维细胞炎症和氧化应激的机制研究

目的 探讨白藜芦醇（RSV）对牙龈卟啉单胞菌脂多糖（LPS）诱导的人牙龈成纤维细胞（HGF）的炎症和氧 化应激的调节作用。方法 原代培养HGF,将细胞分为实验1和实验2：实验1细胞分为对照组、LPS组、RSV 20 μmol/L 组、RSV 40 μmol/L组、RSV 80 μmol/L组、LPS+RSV 20 μmol/L组、LPS+RSV 40 μmol/L组和LPS+RSV 80 μmol/L组;实 验2细胞分为对照组、LPS组、LPS+RSV 40 μmol/L 组、LPS+RSV 40 μmol/L+E5564组和LPS+E5564组。通过CCK-8 法评估细胞活力。利用酶联免疫吸附试验测定白细胞介素（IL）-1β、IL-6、IL-8、肿瘤坏死因子（TNF）-α、超氧化物歧 化酶（SOD）、丙二醛（MDA）和谷胱甘肽过氧化物酶（GSH-Px）水平。通过Western blot分析测量蛋白的表达水平。结 果 20、40和80 μmol/L RSV对HGF均没有明显的细胞毒性作用。在LPS诱导的HGF细胞中,40和80 μmol/L RSV 通过下调IL-1β、IL-6、IL-8和TNF-α的表达减弱炎症反应,降低了MDA的含量,并显著提高了SOD的水平,80 μmol/L RSV显著提高了GSH-Px水平（P＜0.05）。此外,20、40和80 μmol/L RSV可诱导Toll样受体4（TLR4）/MyD88/NF-κB 信号通路的失活,其均可降低TLR4、MyD88和p-p65蛋白的表达水平（P＜0.05）。TLR4抑制剂（E5564）通过下调IL- 1β、IL-6、IL-8和TNF-α的产生以及上调GSH-Px水平进一步增强了RSV对炎症和氧化应激损伤的缓解作用（P＜ 0.05）。结论 RSV可通过诱导TLR4/MyD88/NF-κB信号通路失活,减轻LPS导致的HGF炎症和氧化应激损伤。     

黄芪甲苷在高糖诱导的内皮细胞损伤中的作用研究

目的探讨黄芪甲苷(astragaloside Ⅳ,AS-IV)对高糖诱导的内皮细胞损伤的保护作用及机制。方法体外培养人脐静脉血管内皮细胞(human umbilical vein endothelial cells,HUVECs),使用高浓度葡萄糖(30 mmol/L)诱导细胞损伤,采用实时聚合酶链式反应(real-time polymerase chain reaction,RT-PCR)检测炎症因子肿瘤坏死因子(tumor necrosis factor,TNF)-α、白介素(interleukin,IL)-6和IL-1β的表达。使用不同浓度黄芪甲苷(10、20、50及100 μmol/L)处理高糖损伤的HUVECs,RT-PCR检测炎症因子TNF-α、IL-6和IL-1β的表达水平。黄芪甲苷干预后,酶联免疫吸附法(enzyme linked immunosorbent assay,ELISA)检测TNF-α、IL-6和IL-1β含量,蛋白质印迹法(western blot,WB)检测自噬相关蛋白LC3及Beclin1的表达。进一步将实验分为高糖组、黄芪甲苷组及3-甲基腺嘌呤(3-methyladenine,3-MA)组,RT-PCR检测TNF-α、IL-6和IL-1β的信使RNA(messenger RNA,mRNA)表达水平、WB法检测自噬相关蛋白LC3和Beclin1表达。结果高糖可显著诱导炎症因子TNF-α、IL-6和IL-1β的表达,50 μmol/L黄芪甲苷为最佳抑制炎症反应的浓度。黄芪甲苷组自噬相关蛋白LC3及Beclin1的蛋白水平显著上调,差异具有统计学意义(P<0.05)。与黄芪甲苷组相比,3-MA组自噬相关蛋白LC3和Beclin1的蛋白水平显著降低,TNF-α、IL-6、IL-1β的mRNA表达水平显著上调,差异具有统计学意义(P<0.05)。结论黄芪甲苷可保护内皮细胞免于炎症损伤,其作用机制可能与细胞自噬有关。     

Protective effects of prostaglandin E1 on human umbilical vein endothelial cell injury induced by hydrogen peroxide

Aim:

To investigate the protective effects of prostaglandin E₁ (PGE₁) against H₂O₂-induced oxidative damage on human umbilical vein endothelial cells (HUVECs).

Methods:

HUVECs were pretreated with PGE₁ (0.25, 0.50, and 1.00 μmol/L) for 24 h and exposed to H₂O₂ (200 μmol/L) for 12 h, and cell viability was measured by the MTT assay. LDH, NO, SOD, GSH-Px, MDA, ROS, and apoptotic percentage were determined. eNOS expression was measured by Western blotting and real-time PCR.

Results:

PGE₁ (0.25−1.00 μmol/L) was able to markedly restore the viability of HUVECs under oxidative stress, and scavenged intracellular reactive oxygen species induced by H₂O₂. PGE₁ also suppressed the production of lipid peroxides, such as MDA, restored the activities of endogenous antioxidants including SOD and GSH-Px, and inhibited cell apoptosis. In addition, PGE₁ significantly increased NO content, eNOS protein, and mRNA expression.

Conclusion:

PGE₁ effectively protected endothelial cells against oxidative stress induced by H₂O₂, an activity that might depend on the up-regulation of NO expression.     

杏仁超临界CO2萃取物抗UVA对成纤维细胞的损伤作用

目的探讨杏仁超临界CO2萃取物对UVA诱导的人皮肤成纤维细胞(ESF-1)损伤的保护作用及其作用机制。方法用20 J.cm 2的UVA照射ESF-1细胞建立光老化细胞模型,以不同浓度杏仁超临界CO2萃取物处理光老化细胞,MTT法检测细胞活力,羟胺法检测细胞SOD活性,比色法检测GSH-Px活性,TBA法检测MDA含量,酶联免疫法检测TNF-α、IL-1β分泌量。结果 UVA照射ESF-1细胞后,细胞活力、SOD、GSH-Px活性降低,MDA含量升高,TNF-α、IL-1β分泌量增加(P<0.01),40μg.mL 1和80μg.mL 1杏仁超临界CO2萃取物能显著提高成纤维细胞活力、SOD、GSH-Px活性,降低MDA含量,减少TNF-α、IL-1β分泌(P<0.05或P<0.01)。结论杏仁超临界CO2萃取物能减轻UVA对人皮肤成纤维细胞的损伤,其机制可能与其清除氧自由基,抗脂质过氧化,抑制氧化损伤,减少炎症细胞因子分泌有关。     

Oxymatrine improves palmitic acid-induced vascular endothelial cell injury through Akt-eNOS-NO signaling pathway北大核心CSCD

目的探讨氧化苦参碱(oxymatrine,OMT)对棕榈酸(palmitic acid,PA)致血管内皮细胞损伤的改善作用及其机制。方法MTT法检测人脐静脉血管内皮细胞(human umbilical vein endothelial cells,HUVECs)存活率;流式细胞仪检测细胞凋亡;ELISA法检测细胞内活性氧(ROS)水平以及细胞培养液中乳酸脱氢酶(LDH)、丙二醛(MDA)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)及一氧化氮(NO)水平;Western blot法检测bcl-2、bax、caspase-3、Akt和内皮型一氧化氮合酶(endothelial nitric oxide synthase,eNOS)的蛋白表达。结果OMT对PA诱导的HUVECs细胞存活率下降和LDH水平升高有明显的抑制作用;降低细胞凋亡;上调bcl-2/bax蛋白比值及下调caspase-3蛋白表达;在细胞培养液中降低ROS和MDA水平而提高SOD、GSH-PX和NO水平;上调Akt和eNOS的磷酸化蛋白表达。结论OMT通过Akt-eNOS-NO信号通路改善PA诱导的血管内皮细胞损伤。     

栀子苷对氧化应激损伤血管内皮细胞的保护作用

目的研究栀子苷对体外培养的人脐静脉内皮细胞(HUVEC)氧化损伤的保护作用。方法采用过氧化氢(Hy-drogen peroxide,H2O2)建立体外培养的HUVEC细胞氧化应激损伤模型。将细胞分为正常对照组、H2O2氧化损伤组、H2O2加栀子苷低、中、高剂量组,其中后3组给予栀子苷预培养24h后加入400μmol.L-1H2O2,然后继续培养12h。MTT法检测细胞存活率;检测各组细胞内超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、一氧化氮合酶(NOS)水平和培养液中一氧化氮(NO)水平;检测细胞内活性氧簇(ROS)水平;用流式细胞仪检测细胞凋亡及周期改变。结果栀子苷能明显提高H2O2损伤的内皮细胞的存活率,提高细胞内SOD、GSH-Px、NOS活性,使培养液中NO含量增加,降低细胞内ROS水平,减少H2O2诱导的细胞凋亡率,恢复血管内皮细胞增殖。结论栀子苷具有较强的抗氧化能力及内皮细胞保护作用,可减轻血管内皮细胞的氧化损伤。     

Statins protect human endothelial cells from TNF-induced inflammation via ERK5 activation

3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) exert pleiotropic effects on the cardiovascular system, in part through a decrease in reactive oxygen species (ROS) formation and reduction of vascular inflammation. To elucidate the molecular mechanisms involved in these effects, we investigated the effect of statins on TNF-α-induced ROS production, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) expression in human aortic endothelial cells (HAECs). Exposure of HAECs to TNF-α caused production of ROS via Rac-1 membrane translocation and activation. The Rac-1 activation and ROS liberation mediated TNF-stimulated NF-κB activation and the subsequent VCAM-1 and ICAM-1 expression. Extracellular-signal-regulated kinase 5 (ERK5) plays a central role in inhibiting endothelial inflammation. Immune complex kinase assay of protein extracts from HAECs treated with atorvastatin revealed increased ERK5 activity in a time- and dose-dependent manner. In addition, pretreatment with atorvastatin inhibited TNF-α-induced ROS production and VCAM-1 and ICAM-1 expression. Chemical or genetic inhibition of ERK5 ablated the statins inhibition of Rac-1 activation, ROS formation, NF-κB, VCAM-1 and ICAM-1 expression induced by TNF-α. Taken together, statins, via ERK5 activation, suppress TNF-stimulated Rac-1 activation, ROS generation, NF-κB activation and VCAM-1 and ICAM-1 expression in human ECs, which provides a novel explanation for the pleiotropic effects of statins that benefit the cardiovascular system.     

Protective effects of ligustrazine on TNF-α-induced endothelial dysfunction

To investigate the effects of Ligustrazine, a compound derived from chuanxiong, on tumor necrosis factor-α (TNF-α) stimulated endothelial cells. Human umbilical vein endothelial cells (HUVECs) were stimulated with TNF-α in vitro. Nitric oxide (NO) was measured as a standard of endothelial dysfunction. Two important indicators of autoimmunity, intracellular adhesion molecular-1 (ICAM-1) and heat shock protein 60 (HSP60), were selected to evaluate the influence of Ligustrazine on HUVECs. Ligustrazine (40 μg/ml) significantly reversed the decrease in NO production induced by TNF-α (5 ng/ml) in HUVECs. The expressions of ICAM-1 and HSP60 were increased by TNF-α treatment, but dramatically inhibited by treatment with ligustrazine in TNF-α-stimulated cells. Ligustrazine increased the production of NO in HUVECs and had an immunomodulatory effect on HUVECs stimulated with TNF-α by down-regulating the expression of ICAM-1 and HSP60. These results suggest that ligustrazine protects the endothelium via inhibition of immunological reactions, preventing atherosclerosis.     

Ginsenoside Rg1 reduces toxicity of PM2.5 on human umbilical vein endothelial cells by upregulating intracellular antioxidative state

Ambient airborne particulate matter (PM) is an important environmental pollutant responsible for many human diseases. Oxidative stress is suggested to be involved in PM-induced cell injury. The present study is designed to study unsalutary effects of the organic extracts of PM with an aerodynamic diameter of less than 2.5 μm (PM_2.5) and protective effect of Ginsenoside Rg1 (Rg1) against PM_2.5 on human umbilical vein endothelial cells (HUVECs) in vitro. Cytotoxic effects of the organic extract PM_2.5 on HUVECs were measured by means of HUVEC cell viability and the generation of intracellular reactive oxygen species (ROS). Expression of heme oxygenase-1(HO-1) and Nuclear factor-erythroid 2-related factor 2 (Nrf2) and Nrf2 cytoplasm–nucleus location were assayed. The present results showed that PM_2.5 (50–800 μg/ml) decreased HUVEC viability and increased intracellular generation of ROS and malondialdehyde (MDA) in a concentration dependent manner, but increased HO-1 expression without concentration dependence. Rg1 (10 and 40 μg/ml) diminished PM_2.5-induced HUVEC viability, decrease ROS and MDA generation, increased HO-1 and Nrf2 expression and promoted Nrf2 translocation to nucleus in a concentration dependent manner. These results suggested that organic extracts of PM_2.5 increase oxidative stress and decrease cell viability; Rg1 antagonize PM_2.5-induced excess oxidative stress; HO-1 expression increase and Nrf2 translocation to nucleus may be involved in the effects of both PM_2.5 and Rg1 on HUVECs.     

Sildenafil attenuates oxidative stress and endothelial dysfunction in lead-induced hypertension

Lead (Pb) reduces NO bioavailability, impairs the antioxidant system, and increases the generation of reactive oxygen species (ROS). Pb-induced oxidative stress may be responsible for the associated endothelial dysfunction. Sildenafil has shown nitric oxide (NO)-independent action, including antioxidant effects. Therefore, we examined the effects of sildenafil on oxidative stress, reductions of NO and endothelial dysfunction in Pb-induced hypertension. Wistar rats were distributed into three groups: Pb, Pb + sildenafil and Sham. Blood pressure and endothelium-dependent vascular function were recorded. We also examined biochemical determinants of lipid peroxidation and antioxidant function. ROS levels, NO metabolites and NO levels in human umbilical vein endothelial cells (HUVECs) were also evaluated. Sildenafil prevents impairment of endothelium-dependent NO-mediated vasodilation and attenuates Pb-induced hypertension, reduces ROS formation, enhances superoxide dismutase (SOD) activity and antioxidant capacity in plasma and increases NO metabolites in plasma and HUVECs culture supernatants, while no changes were found on measurement of NO released from HUVECs incubated with plasma of the Pb and Pb + sildenafil groups compared with the sham group. In conclusion, sildenafil protects against ROS-mediated inactivation of NO, thus preventing endothelial dysfunction and attenuating Pb-induced hypertension, possibly through antioxidant effects.     

川芎嗪对血管内皮细胞损伤的保护作用

用 H2 O2 (75μmol· L-1,4h)诱导人胎儿脐静脉内皮细胞 (HUVECs)损伤 ,研究川芎嗪对血管内皮细胞损伤的保护作用 .噻唑蓝 (MTT)比色法检测细胞活性 ,放射免疫法测定细胞培养液中的内皮素样免疫反应物 (ir- ET)水平及前列环素代谢物6-酮前列腺素 1α(6- keto- PGF1α)含量 ,硫代巴比妥酸法测定细胞内脂质过氧化产物丙二醛 (MDA)含量 ,生化自动分析仪测定培养液中乳酸脱氢酶(LDH)活性 .结果显示川芎嗪 0 .3- 1 .2 mmol·L-1对正常 HUVECs具有促进增殖作用 ,并浓度依赖性拮抗 H2 O2 抑制增殖作用 .川芎嗪 0 .3和 0 .6mmol· L-1使细胞培养液中 6- keto- PGF1α含量升高 ,ir- ET含量降低 ,说明增加前列环素释放 ,减少内皮素基础释放量 ,并部分拮抗 H2 O2 损伤HUVECs所致 LDH,内皮素释放增加 ,前列环素释放减少和 MDA生成增加的作用 .提示川芎嗪具有保护 H2 O2 致血管内皮细胞损伤的作用 ,其机理可能与其抗脂质过氧化作用有关     

降糖复方含药血清对氧化损伤内皮细胞的保护作用

目的：研究降糖复方含药血清对H2O2诱导的血管内皮细胞氧化损伤的保护作用。方法：采用H2O2诱导人脐静脉血管内皮细胞（ECV304）氧化损伤模型,观察细胞形态学变化并测定细胞活力、丙二醛（MDA）、超氧化物歧化酶（SOD）的含量,NO含量以及内皮素（ET-1）含量。结果：降糖复方含药血清可明显改善氧化损伤的ECV304细胞形态学变化,提高细胞存活率,降低MDA含量、提高SOD活力、增加NO释放、减少ET-1释放。结论：降糖复方含药血清能对抗H2O2对血管内皮细胞的氧化损伤,起到保护血管内皮细胞的作用。     

Melatonin prevents endothelial dysfunction in SLE by activating the nuclear receptor retinoic acid-related orphan receptor-α

Atherosclerotic cardiovascular disease confers significant morbidity and mortality in patients with systemic lupus erythematosus (SLE). A substantial proportion of patients with SLE display accelerated endothelial dysfunction, which precedes cardiovascular disease. Melatonin and its nuclear receptor retinoid-related orphan receptor alpha (RORα) have been reported to have some protective effects on the development of atherosclerosis. However, the function of melatonin in SLE-induced endothelial dysfunction and the role that RORα plays are still unknown. In this study, we found that RORα protein expression was decreased in aortas of lupus-prone mice and in human umbilical vein endothelial cells (HUVECs) cultured with medium containing sera of patients with SLE. Melatonin-treated HUVECs showed a decrease of pro-inflammatory mRNAs [interleukin-1beta (IL-1β), interferon-gamma (IFN-γ), and tumor necrosis factor-alpha (TNF-α)] under the stimulation of SLE medium. Melatonin increased nitric oxide and antioxidant mRNAs (SOD1, GPX1, and CAT) and downregulated reactive oxygen species (ROS) level in HUVECs, which may subsequently delay endothelial senescence and promote HUVEC proliferation and repair after injury. Melatonin inhibited SLE medium-induced RAW264.7 macrophage migration. HUVECs pretreated with melatonin expressed less adhesion-related proteins (ICAM-1 and VCAM-1); as a result, these cells adhered to fewer peripheral blood monocytes. In addition, we also showed that the protective effects of melatonin on endothelial cells were largely diminished when RORα was knockdown in HUVECs. In conclusion, by targeting the nuclear receptor RORα, melatonin preserves normal functions of endothelium in SLE by its anti-inflammatory, antioxidant, and anti-senescence effects. RORα may have the potential to become a prophylactic or therapeutic target in preventing endothelial dysfunction and atherosclerotic cardiovascular disease in patients with SLE.     

阿托伐他汀与氨氯地平对ox-LDL损伤的人脐静脉内皮细胞LOX-1和eNOS表达的影响

目的以人脐静脉内皮细胞(HUVECs)为载体,探讨阿托伐他汀与氨氯地平对氧化低密度脂蛋白(ox-LDL)损伤的HUVECs内血凝素样氧化型低密度脂蛋白受体(LOX-1)和内皮型一氧化氮合酶(eNOS)表达的影响。方法体外培养HUVECs,采用倒置显微镜观察内皮细胞形态变化,待细胞生长至融合状态时加入ox-LDL、阿托伐他汀、氨氯地平进行干预。随机分为:①对照组(培养基);②ox-LDL组(50mg/L);③阿托伐他汀组(10μmol/L);④氨氯地平组(10μmol/L);⑤阿托伐他汀+氨氯地平组。应用实时荧光定量聚合酶链反应(RT-PCR)检测各组LOX-1mRNA与eNOSmRNA表达的水平。结果与正常对照组比较,ox-LDL使LOX-1mRNA表达增加,eNOSmRNA表达减少;阿托伐他汀、氨氯地平均可使ox-LDL损伤的HUVECs内LOX-1mRNA表达下调,eNOSmRNA表达上调,且二者具有协同作用;混合刺激组与对照组比较,LOX-1mRNA、eNOSmRNA表达差异均无统计学意义(P>0.05)。结论阿托伐他汀和氨氯地平均可下调ox-LDL损伤的HUVECs内LOX-1mRNA的表达,上调eNOSmRNA的表达,且二者具有协同作用。     

阿托伐他汀对内皮细胞微粒诱导人脐静脉内皮细胞VCAM-1和ICAM-1表达的影响

目的:探讨阿托伐他汀对内皮细胞微粒(EMPs)诱导的人脐静脉内皮细胞(HUVECs)表达血管细胞粘附分子(VCAM)-1和细胞间粘附分子(ICAM)-1的影响。方法:取生长良好的第4,5代人脐静脉内皮细胞,将细胞分为3大组:对照组、EMPs组、EMPs+阿托伐他汀组。对照组加入培养基,EMPs组以不同浓度的EMPs(0/mL,1×102/mL,1×103/mL,1×104/mL,1×105/mL)与HUVECs共同孵育24 h,EMPs+阿托伐他汀组以不同浓度的阿托伐他汀(0.05,0.1,1.0,10μmol.L-1)与HUVECs作用1 h后,加入105/mL EMPs共同孵育24 h。分别采用实时荧光定量聚合酶链反应和蛋白免疫印迹方法检测VCAM-1和ICAM-1 mRNA和蛋白的表达。结果:HUVECs受EMPs刺激后,VCAM-1和ICAM-1 mRNA及蛋白表达呈浓度依赖性增加,阿托伐他汀可不同程度上抑制EMPs的作用。结论:阿托伐他汀抗动脉粥样硬化作用可能部分与抑制EMPs诱导的内皮细胞VCAM-1和ICAM-1的表达有关。     

Improving effect and mechanism of fisetin on cell model of diabetic cerebral microangiopathy

OBJECTIVE To explore the effect and mechanism of fisetin on prevention and treatment of diabetic cerebral microangiopathy at the cellular and molecular level. METHODS In vitro experiments, high glucose(HG, 25 mmol·L~(-1)) and palmitic acid(PA, 200 μmol·L~(-1))were used to intervene in the mouse brain microvascular endothelial cells to establish the model of diabetic brain microangiopathy. Groups are divided into control, model(25 mmol·L~(-1) HG+200 μmol·L~(-1) PA, HG+PA) and fisetin(1, 5, 10 and 20 μmol·L~(-1)) groups. The effect of fisetin on hyperglycemic and hyperlipid-induced cell damage was detected by cell counting kit-(CCK-) 8 assay, cytotoxicity was detected by LDH assay. After treatment, the changes of oxidative stress related factors(ROS, NO, i NOS, MMP-2, MMP-9, TIMP-1, MDA, SOD, CAT) were detected by ELISA. RESULTS Compared with the control group, the cell viability of model group was significantly reduced(P<0.01), indicating that the model was successful y prepared.LDH content, the expression levels of ROS, NO, i NOS,MMP-2, MMP-9, TIMP-1 and MDA in the model group significantly increased(P< 0.01), while the expression of SOD and CAT in the model group decreased significantly(P<0.01). Compared with the model group, the activity of the cells, in the fisetin group was significantly increased(P<0.01). LDH content, the expression levels of ROS,NO, i NOS, MMP-2, MMP-9, TIMP-1 and MDA were decreased in the fisetin group(P<0.01), the expression of SOD and CAT, were significantly increased(P<0.01).The expression levels of antioxidant proteins HO-1 and NQO1 were significantly increased in the fisetin group(P<0.05). CONCLUSION Fisetin can improve the cell injury induced by HG and PA, which mimic the diabetic cerebral microvascular lesions, and its mechanism might be related to inhibiting the production of endogenous ROS in cells, activating the expression of antioxidant proteins and inhibiting the production of related oxidative factors.