同型半胱氨酸诱导内皮细胞凋亡的研究进展

近年的研究发现，血浆同型半胱氨酸(homocysteine，HCY)水平升高是心肌梗死、脑梗死等心脑血管疾病以及外周阻塞性血管疾病的一个独立危险因素。动物实验发现，HCY可以通过损伤血管内皮细胞，启动粥样斑块和血栓形成。在粥样斑块内存在明显的内皮细胞凋亡。体外内皮细胞培养也显示HCY能诱导其凋亡，提示HCY可能通过内皮细胞凋亡而促进动脉粥样硬化。目前关于HCY引起内皮细胞凋亡的机制尚不明确，作者就HCY的代谢、HCY与内皮细胞凋亡的关系以及细胞凋亡的产生机制综述如下。     

半胱天冬氨酸蛋白酶-12小干扰RNA对小鼠肝细胞凋亡的阻抑

目的观察小鼠半胱天冬氨酸蛋白酶-12（caspase-12）基因特异性小干扰RNA（siRNA）构建的表达载体pRNAT-casp12对caspase-12基因的抑制及其对内质网应激介导的小鼠肝细胞凋亡的影响。方法以小鼠肝细胞株Hepa1-6为靶细胞，利用脂质体与重组质粒pRNAT-casp-2共转染，分别在转染24、48和72h后收集细胞，采用实时荧光定量PCR分析和Western印迹检测caspase-12的表达；利用毒胡萝b素（TG）诱导细胞，建立内质网应激介导的细胞凋亡模型，通过DNA梯带凝胶电泳检测，选出适合的诱导时间；TG诱导已转染了pRNAT—casp12的干扰组细胞后，以空质粒转染组为对照，利用Western印迹检测caspase-12蛋白表达水平的变化，通过DNA梯带凝胶电泳、流式细胞仪、Hoechst33258染色等方法检测细胞凋亡，观察caspase-12siRNA对细胞凋亡的影响。结果pRNAT—casp12转染细胞24、48和72h后，caspase-12mRNA的水平分别下降了45．6％、72．5％和59．5％；caspase-12蛋白表达下降了17．1％、37．3％和60．1％；2μmol／L TG处理细胞30h后，成功诱导细胞凋亡；与对照组相比，干扰组细胞经TG诱导后，caspase-12蛋白表达水平下降了54．6％，流式细胞仪检测发现早期调亡率下调了51．4％（P〈0．01）。结论小鼠caspase-12siRNA对Hepa1—6细胞caspase-12基因的表达具有显著的抑制作用，能够明显阻抑内质网应激介导的凋亡，有望发展成为新一代抗凋亡药物。     

福辛普利对慢性心力衰竭大鼠心肌细胞凋亡及半胱氨酸天冬氨酸蛋白酶-3的影响

目的探讨福辛普利对慢性心力衰竭(CHF)大鼠心肌细胞凋亡及半胱氨酸天冬氨酸蛋白酶(Caspase)-3表达水平的影响。方法雄性Wistar大鼠50只随机抽取10只为假手术组,其余大鼠采用肾上腹主动脉缩窄法建立大鼠CHF模型,6 w后将成模大鼠随机分为CHF组、Fos10组、FOS20组(福辛普利10、20 mg·kg~(-1)·d~(-1))。治疗8 w后,观察各组大鼠血流动力学指标和左心室重量指数(LVMI);透射电镜观察左室心肌组织形态的改变;TUNEL法检测大鼠左心室心肌细胞的凋亡指数;SP免疫组织化学染色法检测左心室心肌组织Caspase-3蛋白的表达。结果 CHF组与假手术组相比,大鼠左心室舒张末压(LVEDP)、LVMI、心肌细胞凋亡指数、Caspase-3蛋白的表达均明显升高(P<0.01),左心室内压最大上升和下降速率(±dp/dtmax)显著下降(P<0.01);与CHF组比较,Fos10组和Fos20组大鼠LVEDP、LVMI、心肌细胞凋亡指数、Caspase-3蛋白的表达均明显降低(P<0.01),左心室内压最大上升和下降速率(±dp/dtmax)显著升高(P<0.01),且Fos20组效果明显。电镜下观察:Fos10组和Fos20组心肌损伤程度较CHF组明显减轻。结论福辛普利可通过下调Caspase-3的表达抑制心肌细胞凋亡,改善CHF大鼠心室功能及心肌超微结构。     

重构型天冬氨酸特异性半胱氨酸蛋白酶3表达载体诱导人肺腺癌细胞凋亡的意义

目的构建天冬氨酸特异性半胱氨酸蛋白酶3(Caspase3)表达载体,观察Caspase3表达载体对人肺腺癌A549细胞凋亡的影响。方法应用基因重组方法,建立重构型Caspase3基因的真核表达系统pcDNA3.1revCaspase3质粒和野生型pcDNA3.1Caspase3质粒。将实验细胞分为3组转染pcDNA3.1revCaspase3质粒组,转染pcDNA3.1Caspase3质粒组,转染pcDNA3.1质粒空白对照组。前2组用Caspase3抑制剂DEVDfmk干预。应用Caspase3酶活性分析、流式细胞仪及甲基偶氮唑蓝(MTT)比色法检测A549细胞Caspase3酶活性变化及细胞凋亡和增殖情况。结果(1)pcDNA3.1revCaspase3质粒组、pcDNA3.1Caspase3质粒组的A549细胞Caspase3酶活性分别是(11.87±0.92)%、(5.34±0.38)%(t=16.02,P<0.01);用Caspase3抑制剂DEVDfmk干预后,pcDNA3.1revCaspase3质粒组酶活性为(7.04±0.48)%,仍明显高于野生型pcDNA3.1Caspase3质粒组的(4.51±0.20)%(t=11.86,P<0.01)。(2)流式细胞分析结果显示,pcDNA3.1revCaspase3质粒组、pcDNA3.1Caspase3质粒组和pcDNA3.1质粒组的A549细胞凋亡率分别是(20.1±3.5)%、(7.8±2.8)%、(1.4±0.3)%,3组差异有统计学意义(F=44.01,P<0.01)。(3)MTT比色检测显示pcDNA3.1revCaspase3质粒组、pcDNA3.1Caspase3质粒组和pcDNA3.1质粒组的A549细胞生存率分别是(35.7±1.1)%、(72.8±2.9)%、(85.4±4.8)%,转染pcDNA3.1revCaspase3质粒组生存率明显低于其他两组(F=375.07,P<0.01)。结论pcDNA3.1revCaspase3在A549细胞内有较强的自身活化能力,对Caspase3抑制剂抵抗作用较强,可明显诱导A549细胞凋亡并抑制A549细胞生长。     

雷米普利对慢性心力衰竭大鼠肾脏细胞凋亡、半胱氨酸天冬氨酸蛋白酶3及细胞色素C的影响

目的观察雷米普利对慢性心力衰竭大鼠肾脏细胞凋亡、半胱氨酸天冬氨酸蛋白酶(caspase)-3和细胞色素C的影响。方法 40只雄性Wistar大鼠,按随机数字表法平均分组,正常对照组10只,余均采用肾上腹主动脉缩窄法建立CHF大鼠模型,将建模成功的20只随机均分为模型组和雷米普利组,分别采取苏木素-伊红(HE)染色法观察大鼠肾脏细胞的形态结构,原位末端标记法(TUNEL)测定肾脏细胞凋亡率;逆转录聚合酶链反应(RT-PCR)检测肾脏组织中caspase-3及细胞色素C mRNA的表达;Western印迹法检测肾脏组织caspase-3蛋白及细胞色素C蛋白含量。结果与正常对照组相比,模型组肾脏组织凋亡指数(AI)显著增高,caspase-3和细胞色素C mRNA及蛋白表达明显增加(P0.01);与模型组相比,雷米普利组肾脏组织AI显著减少,caspase-3和细胞色素C mRNA及蛋白表达量均显著降低(P0.01)。结论雷米普利能够降低CHF大鼠肾脏组织凋亡率,减少肾组织中caspase-3和细胞色素C mRNA及蛋白表达,其抑制肾脏细胞凋亡的作用可能是改善肾功能的作用机制。     

曲美他嗪与雷米普利对心力衰竭大鼠心肌细胞凋亡及半胱氨酸天冬氨酸蛋白酶3的影响

目的探讨曲美他嗪与雷米普利单用及联合应用对慢性心力衰竭大鼠左心室心肌细胞凋亡情况以及半胱氨酸天冬氨酸蛋白酶3(Caspase-3)表达水平的影响。方法雄性Wistar大鼠50只随机分为假手术组、模型组、曲美他嗪组、雷米普利组和联合用药组(曲美他嗪+雷米普利),每组10只。采用肾上腹主动脉缩窄法制作慢性心力衰竭模型,观察各组左心室舒张末压、左心室压力最大上升及下降速率(±dp/dtmax);TUNEL法检测心肌细胞凋亡指数;SP免疫组织化学法检测各组大鼠左心室心肌Caspase-3水平的表达,RT-PCR测定Caspase-3mRNA水平。结果与模型组比较,曲美他嗪组、雷米普利组和联合用药组左心室舒张末压、心肌凋亡指数明显降低,±dp/dtmax明显升高,差异有统计学意义(P<0.01),Caspase-3蛋白及mRNA表达水平明显降低,差异有统计学意义[(0.14±0.01、0.14±0.02和0.08±0.02)vs(0.22±0.02),(0.54±0.02、0.54±0.02和0.32±0.02)vs(0.91±0.02),P<0.01]。结论曲美他嗪与雷米普利单用及联合应用均可有效改善心功能、抑制心肌细胞凋亡,联合用药的疗效优于单独用药。     

曲美他嗪对心力衰竭大鼠心肌细胞凋亡及半胱氨酸天冬氨酸蛋白酶3表达的影响

目的探讨曲美他嗪对心力衰竭大鼠心肌细胞凋亡及半胱氨酸天冬氨酸蛋白酶(Caspase)-3表达的影响。方法雄性Wistar大鼠随机分为对照组、模型组和实验组各15只。对照组进行模拟手术,模型组和实验组采用肾上腹主动脉缩窄法建立心力衰竭大鼠模型。实验组给予曲美他嗪10 mg·kg~(-1)·d~(-1)持续灌胃28 d。测量3组左室舒张末压力(LVEDP)、左心室压力最大上升和下降速率(±dp/dtmax);检查3组心肌细胞凋亡比例、Caspase-3及细胞色素C蛋白表达水平。结果实验组LVEDP显著高于对照组(P<0.05),而显著低于模型组(P<0.05);实验组±dp/dtmax显著高于模型组(P<0.05),而显著低于对照组(P<0.05);对照组、实验组和模型组心肌细胞凋亡指数(AI)分别为(1.87±0.32)%,(22.52±5.29)%和(39.71±12.58)%,实验组心肌细胞凋亡比例、Caspase-3、细胞色素C蛋白表达明显低于模型组(P<0.05),而显著高于对照组(P<0.05);结论曲美他嗪可通过抑制心力衰竭大鼠心肌细胞凋亡及Caspase-3、细胞色素C蛋白表达来改善心脏功能。     

同型半胱氨酸硫内酯对缺氧神经母细胞瘤系细胞内质网应激途径相关因子表达的影响

目的探讨同型半胱氨酸硫内酯(HTL)对缺氧神经母细胞瘤系细胞内质网应激途径相关因子表达的影响。方法选取人神经母细胞瘤株SHSY5Y细胞采用氯化钴作用24 h制作缺氧模型后,分为模型组、试验A组、试验B组、试验C组、试验D组,每组5例,后4组分别加入HTL 50、100、200和400μmol/L同时刺激。另取处于对数生长期的SHSY5Y细胞为对照组(n=5)。检测各组细胞活性,免疫组织化学染色观察模型组和试验D组、半胱氨酸天冬氨酸酶(Caspase)-12和Caspase-3及TNF-α蛋白表达,ELISA检测各组上述3种蛋白表达。结果与对照组比较,模型组细胞活力明显降低(0.922±0.039 vs 1.143±0.076,P=0.000)。试验A组、试验B组、试验C组、试验D组细胞活力明显低于模型组(P<0.01)。免疫组织化学染色显示,与模型组比较,试验D组TNF-α蛋白表达明显升高(0.34±0.04 vs 0.26±0.01,P=0.014)。模型组、试验A组~D组的Caspase-3、Caspase-12、TNF-α蛋白表达呈上升趋势。与对照组比较,试验C组Caspase-3和试验D组Caspase-3、Caspase-12、TNF-α蛋白表达明显升高(P<0.01)。结论HTL可能通过Caspase-12凋亡途径及TNF-α激活,抑制细胞生长,促进细胞炎性反应发生及凋亡,加重缺氧后损伤。     

上调SIRT1减少同型半胱氨酸诱导的血管内皮细胞凋亡

目的 研究同型半胱氨酸处理对血管内皮细胞中SIRT1表达的影响,并探讨上调血管内皮细胞中SIRT1表达对同型半胱氨酸诱导的血管内皮细胞凋亡的影响.方法 建立同型半胱氨酸诱导的血管内皮细胞凋亡模型,通过CCK8试剂盒检测细胞活性,通过流式细胞仪检测细胞凋亡,通过qPCR法检测mRNA表达和通过免疫印记法检测蛋白表达.结果...     

自由基清除剂MCI-186对高糖环境中内皮细胞凋亡的干预作用

观察内皮细胞在高糖环境及经肿瘤坏死因子α(TNF-α)诱导后的细胞凋亡情况，并加入自由基清除剂MCI-186予以干扰。检测细胞DNA裂解片段及凋亡信号蛋白(caspase)的表达量，以期探讨MCI-186对高糖环境中血管内皮细胞凋亡的干预作用及凋亡机制。发现高糖环境可增加内皮细胞凋亡率，并上调TNF-α诱导的调亡。MCI-186可抑制高糖诱导的细胞凋亡。但无法干预高糖环境中TNF—a诱导的凋亡。     

Effects of antioxidants on homocysteine thiolactone-induced apoptosis in human umbilical vein endothelial cells

Background and objectives Hyperhomocysteinemia is an independent risk factor for cardiovascular disease. Homocysteine thiolactone (HcyT), one of the homocysteine metabolites in vivo, is toxic both in vivo and in vitro. The aim of this study was to investigate the effect of HcyT on apoptotic damage in human umbilical vein endothelial cells (HUVECs) and the role of antioxidants in the reduction of HcyT-induced apoptosis. Methods HUVECs were cultured in DMEM supplemented with 20% heat inactivated fetal bovine serum cell cultures were maintained in a humidified 5% CO2 atmosphere at 37℃. Cytotoxicity was determined by MTT assay, which consists of hypodiploid cells with propidium iodide labeling and intracellular reactive oxygen species levels using 2',7'-dichlorofluorescein diacetate as the probe by flow cytometry. Results HcyT (250-2000μM) induced HUVECs apoptosis in a time- and concentration-dependent manner. Reactive oxygen species levels rose in response to increasing HcyT concentrations at 24-h incubation. The reduction of cell apoptosis by N-acetylcysteine, vitamin E, or pyrrolidine dithiocarbamate, occurred simultaneously with a significant decrease in intracellular reactive oxygen species levels. Conclusion HcyT exerts its cytotoxic effects on endothelial cells through an apoptotic mechanism involving cellular reactive oxygen species production. The capacity of N-acetylcysteine, vitamin E, and pyrrolidine dithiocarbamate to scavenge HcyT-induced cellular reactive oxygen species correlates well with their efficiency to protect against HcyT-promoted apoptotic damage. The protective effect of pyrrolidine dithiocarbamate on cell apoptosis indicates HcyT-generated hydrogen peroxide may provoke cell apoptosis via activating nuclear factor-kappa binding protein.     

PPARγ对ECV-304细胞ICAM-1与VCAM-1表达的影响

目的 观察过氧化物酶体增殖物激活型受体γ(PPARγ)对糖基化终末产物(AGEs)所诱导的血管内皮细胞表达ICAM-1、VCAM-1的影响,探讨PPARγ在糖尿病血管并发症及动脉粥样硬化中的可能作用.方法 体外培养人内皮细胞系ECV-304,应用PPARγ的反义寡核苷酸及激动剂处理细胞,采用流式细胞仪及Western印迹技术检测细胞中ICAM-1及VCAM-1蛋白质的表达.结果 反义阻断PPARγ在mRNA水平上的表达,ECV-304细胞ICAM-1及VCAM-1的表达上调,而PPARγ激动剂Ciglitizone则降低ICAM-1及VCAM-1的表达.结论 PPARγ激活可负调控AGEs诱导的血管内皮细胞ICAM-1和VCAM-1的表达,这是PPARγ抑制糖尿病血管并发症及动脉粥样硬化发生发展的可能机制之一.     

瘦素对人脐静脉内皮细胞表达白细胞介素-8的影响

目的:探讨瘦素对人脐静脉内皮细胞(ECV-304)表达白细胞介素-8(IL-8)的影响。方法:用不同浓度的瘦素刺激ECV-304,检测ECV-304表达IL-8的情况。结果:在相同作用时间下,随着瘦素浓度的升高,IL-8蛋白及IL-8的mRNA的表达也随之升高;在相同瘦素浓度下,随着作用时间的延长,IL-8蛋白及IL-8的mRNA的表达也随之升高。结论:瘦素可以刺激ECV-304表达IL-8,而且呈时间和剂量相关性。     

偏硅酸钠对ECV-304细胞株与单个核细胞黏附的影响

目的 观察偏硅酸钠(Na2SiO3)对氧化低密度脂蛋白(ox-LDL)诱导的ECV-304细胞与人外周血单个核细胞黏附的影响,探讨其抗动脉粥样硬化(AS)作用的机制.方法 ECV-304细胞分为5组:对照组、ox-LDL组、Na2SiO3高剂量组(硅68.5 mg/L)、中剂量组(硅34.2 mg/L)、低剂量组(硅17.1 mg/L).分离正常健康人外周血单个核细胞,计数与各组ECV-304细胞黏附数,检测细胞培养液中IL-8水平;采用流式细胞术测VCAM-1的蛋白表达量;半定量RT-PCR检测VCAM-1、IL-8mRNA的表达.结果 Na2SiO3减少ECV-304单个核细胞黏附数(P＜0.01),降低IL-8浓度(P＜0.01),降低VCAM-1的蛋白表达量(P＜0.05),下调VCAM-1、IL-8 mRNA的表达(P＜0.01).结论 Na2SiO3能够下调ox-LDL诱导的ECV-304 IL-8和VCAM-1的表达.     

Hypoxia inducible factor-1α mediates protective effects of ischemic preconditioning on ECV-304 endothelial cells

AIM: To investigate whether hypoxia inducible factor-1α (HIF-1α) is linked to the protective effects of ischemic preconditioning (IP) on sinusoidal endothelial cells against ischemia/reperfusion injury. METHODS: Sinusoidal endothelial cell lines ECV-304 were cultured and divided into four groups: control group, cells were cultured in complete DMEM medium; cold anoxia/warm reoxygenation (A/R) group, cells were preserved in a 4℃ UW solution in a mixture of 95% N2 and 5% CO2 for 24 h; anoxia-preconditioning (APC) group, cells were treated with 4 cycles of short anoxia and reoxygenation before prolonged anoxia- preconditioning treatment; and anoxia-preconditioning and hypoxia inducible factor-1α (HIF-1α) inhibitor (I-HIF-1) group, cells were pretreated with 5 μm of HIF-1α inhibitor NS398 in DMEM medium before subjected to the same treatment as group APC. After the anoxia treatment, each group was reoxygenated in a mixture of 95% air and 5% CO2 incubator for 6 h. Cytoprotections were evaluated by cell viabilities from Trypan blue, lactate dehydrogenase (LDH) release rates, and intracellular cell adhesion molecule-1 (ICAM-1) expressions. Expressions of HIF-1α mRNA and HIF-1α protein from each group were determined by the RT-PCR method and Western blotting, respectively. RESULTS: Ischemia preconditioning increased cell viability, and reduced LDH release and ICAM-1 expressions. Ischemia preconditioning also upregulated the HIF-1α mRNA level and HIF-1α protein expression. However, all of these changes were reversed by HIF-1α inhibitor NS398.CONCLUSION: Ischemia preconditioning effectively inhibited cold hypoxia/warm reoxygenation injury to endothelial cells, and the authors showed for the first time HIF-1α is causally linked to the protective effects of ischemic preconditioning on endothelial cells.     

Hypoxia inducible factor-1α mediates protective effects of ischemic preconditioning on ECV-304 endothelial cells

AIM: To investigate whether hypoxia inducible factor-1α (HIF-1α) is linked to the protective effects of ischemic preconditioning (IP) on sinusoidal endothelial cells against ischemia/reperfusion injury.METHODS: Sinusoidal endothelial cell lines ECV-304 were cultured and divided into four groups: control group, cells were cultured in complete DMEM medium; cold anoxia/warm reoxygenation (A/R) group, cells were preserved in a 4℃ UW solution in a mixture of 95% N2 and 5% CO2 for 24 h; anoxia-preconditioning (ARC) group, cells were treated with 4 cycles of short anoxia and reoxygenation before prolonged anoxia-preconditioning treatment; and anoxia-preconditioning and hypoxia inducible factor-1α (HIF-1α) inhibitor (I-HIF-1) group, cells were pretreated with 5 μm of HIF-1α inhibitor NS398 in DMEM medium before subjected to the same treatment as group ARC. After the anoxia treatment, each group was reoxygenated in a mixture of 95% air and 5% CO2 incubator for 6 h. Cytoprotections were evaluated by cell viabilities from Trypan blue, lactate dehydrogenase (LDH) release rates, and intracellular cell adhesion molecule-1 (ICAM-1) expressions. Expressions of HIF-1α mRNA and HIF-1α protein from each group were determined by the RT-PCR method and Western blotting, respectively.RESULTS: Ischemia preconditioning increased cell viability, and reduced LDH release and ICAM-1 expressions. Ischemia preconditioning also upregulated the HIF-1α mRNA level and HIF-1α protein expression. However, all of these changes were reversed by HIF-1α inhibitor NS398.CONCLUSION: Ischemia preconditioning effectively inhibited cold hypoxia/warm reoxygenation injury to endothelial cells, and the authors showed for the first time HIF-1α is causally linked to the protective effects of ischemic preconditioning on endothelial cells.     

Caspase-12 mediates carbon tetrachloride-induced hepatocyte apoptosis in mice

AIM: To investigate the role of caspase-12 and its downstream targets in carbon tetrachloride (CCl₄)-induced hepatocyte apoptosis.METHODS: The role of caspase-12 was determined by using caspase-12 knock-out (^-/-) mice. CCl₄ (300 μL/kg body weight) or vehicle (corn oil) was administered to caspase-12^+/+ or caspase-12^-/- mice as a single intraperitoneal injection. The animals were sacrificed 24 h after the CCl₄ treatment. Blood was collected to evaluate liver function by the measurement of the activity of alanine aminotransferase. Liver samples were used for the measurements of reactive oxygen species using plasma malondialdehyde as biomarker, hepatocyte apoptosis was evaluated via terminal transferase-mediated dUTP nick-end labeling and controlled by morphologic study, and cytochrome C release and caspase activations were measured by Western blotting.RESULTS: Administration of a low dose of CCl₄ resulted in hepatocyte apoptosis and acute liver injury in wild-type mice. CCl₄ also induced the generation of reactive oxygen species and induction of endoplasmic reticulum stress in the liver followed by activations of caspase-12, -9 and -3 as well as release of small amounts of cytochrome C. However, in the CCl₄-treated caspase-12^-/- mice, activation of caspase-9 and -3 were significantly attenuated (P < 0.05); no effect was seen in cytochrome C release. CCl₄-induced apoptosis and liver damage was markedly reduced in caspase-12^-/- mice compared to caspase-12^+/+ mice (P < 0.05). The active form of caspase-8 was not detected in either caspase-12^+/+ or caspase-12^-/- mice. There was no significant different in the formation of reactive oxygen species in the livers of caspase-12^+/+ and caspase-12^-/- mice treated with CCl₄.CONCLUSION: Caspase-12 plays a pivotal role in CCl₄-induced hepatic apoptosis through the activation of the downstream effector caspase-3 directly and/or indirectly via capase-9 activation.     

糖基化终末产物对ECV-304细胞ICAM-1及VCAM-1表达的影响

目的探讨糖基化终产物(AGEs)与其受体(RAGE)对ECV-304细胞上血管细胞黏附分子-(1ICAM-1)和细胞间黏附分子-(1VCAM-1)的影响,为研究AGEs在动脉粥样硬化发生发展中的作用提供实验依据。方法采用体外制备的糖基化终产物——糖基化的牛血清白蛋白(AGE-BSA)和反义RAGE寡核苷酸处理ECV-304细胞,通过流式细胞仪技术、Western印迹技术检测ECV-304细胞上ICAM-1、VCAM-1蛋白质的表达。结果AGEs处理ECV-304细胞后,可诱导ECV-304细胞上VCAM-1和ICAM-1的表达增加,且这种效应呈剂量依赖性。采用反义技术阻断RAGE的mRNA水平的翻译后,ECV-304细胞ICAM-1和VCAM-1的表达呈剂量依赖性降低。结论AGEs可通过与其膜受体RAGE的结合激活血管内皮细胞VCAM-1和ICAM-1的表达,这可能是AGEs促动脉粥样硬化发生发展的机制之一。     

氧化低密度脂蛋白对血管内皮细胞骨架的损伤及其机制

目的:观察氧化低密度脂蛋白(OxLDL)对血管内皮细胞(ECV-304)微丝肌动蛋白的影响及同细胞内钙离子变化间的关系,以探讨OxLDL的致动脉粥样硬化机制。方法:采用ECV 304内皮细胞株体外培养,分为空白对照组,OxLDL (200 μg/ml)组,采用激光扫描共聚显微镜分别观察OxLDL作用后12小时胞内钙离子的变化及OxLDL作用24小时后的细胞骨架微丝(F-actin)的改变。结果:OxLDL作用12小时后细胞内钙离子浓度明显升高(P<0.001),而F-actin也在24小后发生明显的破坏。结论:OxLDL可破坏内皮细胞的骨架结构从而导致血管内皮细胞的屏障功能的损害,这一作用可能与动脉粥样硬化形成有关。     

Fas、FasL及Caspase-3在维甲酸诱导胃癌细胞凋亡中的作用

目的:探讨维甲酸诱导胃癌细胞BGC-803凋亡的作用及其与Fas、FasL、Caspase-3表达的关系.方法:以0.001、0.01、0.1、1、10、20 μmol/L的维甲酸作用BGC-803细胞72h后,MTT法检测维甲酸对BGC-803细胞的生长抑制作用;流式细胞术分析维甲酸对胃癌BGC-803细胞凋亡的诱导作用;Hoechst33342/PI双荧光染色观察细胞凋亡;RT-PCR法检测Fas、FasL、Caspase-3基因的mRNA表达变化.结果:0.1、1、10、20μmol/L的维甲酸作用BGC-803细胞72 h后,较对照组(未加药)能显著抑制细胞增殖(32.61%、44.42%、48.14%、51.15%vs 0.657%,均P＜0.01);20μmol/L维甲酸作用BGC-803细胞12、24和48 h后,G2/M期细胞比例显著增加,出现凋亡特征性的亚G1峰;细胞出现染色质凝集、核膜破裂等凋亡特征;作用48 h后,Fas、FasL、Caspase-3 mRNA表达水平均较对照组显著上调.结论:Fas、FasL、Caspase-3参与了维甲酸诱导胃癌细胞凋亡的调控过程.