心肌再灌注损伤保护的机制和策略

局部缺血部位快速再灌注虽然保护了心肌,但也引起再灌注损伤。目前还没有减轻再灌注损伤的特效疗法,但近年来研究显示,G蛋白耦联受体（Gprotein-coupledreceptor,GPCR）的激动剂、胰岛素和缺血后处理可以在各种实验条件和各类动物模型中有效抵抗再灌注损伤。这些干预手段启动的心脏保护机制可能包括激活再灌注损伤补救激酶（reperfus ioninjury salvage kinase,RISK）途径、抑制糖原合酶激酶-3β（glycogen synthase kinase3β,GSK-3β）以及抑制线粒体膜通透性转换孔（mitochondrial permeabili tytransition pore,mPTP）开放等。这些研究成果有利于开发治疗急性心肌梗死的有效临床手段。     

缺血后处理降低高胆固醇血症大鼠心肌对缺血/再灌注损伤的易感性及其机制

目的:探讨缺血后处理对高胆固醇血症基础上发生的心肌缺血/再灌注损伤的影响及其可能的机制。方法:建立食源性高胆固醇血症大鼠模型,运用TTC染色、酶活性检测等方法测定缺血/再灌注所致的心肌损伤,用实时定量RT-PCR方法检测心肌组织中低氧诱导因子-1α(HIF-1α)mRNA水平,用Western blot方法检测HIF-1α蛋白水平。结果:高胆固醇血症加重了缺血/再灌注造成的心肌损伤,而缺血后处理显著缩小了高胆固醇血症大鼠缺血/再灌注所致的心梗面积,降低了血清肌酸激酶(CK)的活性,减少了心肌细胞凋亡。同时,缺血后处理提高了高胆固醇血症大鼠缺血心肌组织中HIF-1α的蛋白水平。结论:缺血后处理可以降低高胆固醇血症大鼠心肌对缺血/再灌注损伤的敏感性,其效应与心肌组织中HIF-1α的蛋白水平存在着相关性。     

线粒体通透性转换孔在缺血后处理中的作用和机制

心肌细胞急性缺血后,及时再灌注能够挽救缺血心肌细胞的活力、减少梗死面积、促进心肌细胞功能恢复。但是再灌注是一把“双刃剑”,它产生大量活性氧类（reactive oxygen species,ROS）和Ca2＋超载,开放线粒体通透性转换孔（mitochondrial permeability transition pore,mPTP）,使线粒体肿胀,外膜破裂导致心肌细胞坏死。mPTP是线粒体非特异性的转换孔,由电压依赖性阴离子通道（voltage-dependent anion channel,VDAC）、腺苷酸转位蛋白（adeninenucleotide translocator,ANT）和亲环蛋白D（cyclophilin D,CYPD）组成。mPTP关闭维持线粒体结构完整,是缺血心肌细胞功能恢复的先决条件。缺血后处理通过减少再灌注早期ROS大量释放和拮抗Ca^2＋超载、释放内源性介质、激活再灌注损伤补救激酶（reperfusion injury salvage kinase,RISK）、抑制mPTP开放,从而保护心肌细胞。     

缺血后处理兔心肌蛋白质组学双向凝胶电泳分析的初步研究

目的:研究缺血后处理对缺血再灌注心肌保护的相关蛋白的变化。方法:将6只新西兰大白兔随机分为两组(每组3只):心肌缺血再灌注对照组(I/R组)和缺血后处理组(P组)。两组均接受左冠状动脉前降支阻断30min,开放再灌注180min。缺血后处理组,结扎LAD30min,然后灌注30s,阻断30s,重复4次,继而再灌注直至180min,分别取各组缺血区心肌进行二维凝胶电泳,利用ImageMaster2D软件分析实验结果。结果:P组和I/R组对比,有11个蛋白表达发生了显著变化,其中表达增强的有7个蛋白,表达降低的有4个蛋白。结论:这些差异表达的蛋白可能在缺血后处理对心肌缺血再灌注损伤的保护中发挥了作用。     

缺血后处理内源性心脏保护的研究进展

再灌注疗法是临床治疗心肌缺血最有效的措施,但会引起再灌注损伤,调动机体内源性保护机制可以减轻再灌注损伤,保护缺血心肌。缺血预处理（ischemic preconditioning,IPC）和后处理（ischemic postconditioning,I-postC）是缺血心脏有效的内源性保护现象,可以减轻缺血再灌注（ischemia／reperfusion,I／R）后心肌坏死与心肌功能障碍,减少恶性心律失常的发生。内源性心脏保护的机制主要是通过诱导触发因子释放,经多条细胞内信号转导途径的介导,作用于多种效应器,影响氧自由基产生、钙超载等I／R损伤的关键环节而发挥心肌细胞保护作用。特别是可以在缺血后实施的I-postC具有良好的临床应用前景。本文以I-postC为重点综述内源性心脏保护作用、机制及其临床应用现状。     

骨骼肌和心肌缺血后处理对兔缺血/再灌注心肌的保护作用

目的:观察骨骼肌缺血后处理(RPostC)、心肌的缺血后处理(MPostC)对缺血/再灌注心肌保护作用是否存在差异以及两者联合后作用是否叠加。方法:健康新西兰大白兔3O只,随机分为5组(n=6):缺血对照组(Con)、假手术组(sham)、心肌缺血后处理组(MPostC)和肢体缺血后处理组(RPostC)及心肌缺血后处理联合肢体缺血后处理组(MPostC+RPostC)。采用开胸结扎冠状动脉左室支45 min,再灌注120min方法制作缺血/再灌注模型,采用短暂结扎双侧髂外动脉固定部位5 min造成骨骼肌短暂缺血。以Evans蓝标记心肌缺血区范围,以TTC法检测梗死心肌范围,并分别于缺血前、后及再灌注1、2 h测定血浆磷酸肌酸激酶(CPK)活性和乳酸脱氢酶(LDH)含量。结果:和Con组相比,MPostC和RPostC组心肌梗死范围均明显降低(P<0.05);MPostC联合RPostC组心肌梗死范围与MPostC或RPostC组相比,均进一步降低(均P<0.05)。但MPostC组及RPostC组之间心肌坏死范围未见统计学差异。再灌注120 min末血浆CPK活性及LDH含量也显示相似趋势。结论:骨骼肌缺血后处理及心肌后处理对缺血/再灌注心肌均具有明显保护作用;且两者作用可以叠加;但骨骼肌和心肌后处理之间保护作用未显示统计学差异。     

缺血后处理、ATP后处理减轻兔缺血再灌注损伤:与腺苷受体激活有关

目的:近期实验研究显示,在再灌注的早期给予短暂、重复的缺血再灌(缺血后处理Postconditioning)能够减轻心肌再灌注损伤。本实验旨在探明三磷酸腺苷(ATP)用于缺血后处理是否产生上述保护效应,以及了解腺苷受体在此保护作用机制中的地位。方法:家兔开胸后左前降支均给予40min结扎和180min的再灌注,并随机分为5组:(1)对照组;(2)缺血后处理组;(3)ATP后处理组;(4)缺血后处理 SPT(硫苯茶碱)组;(5)SPT对照组。于实验终点测定心肌梗死面积(TTC染色),血浆CK-MB、SOD、MDA含量。结果:和时照组相比,缺血后处理组与ATP后处理组心梗面积减少(p<0．05),CK-MB也显著降低(p相似文献   

腺苷和乙酰胆碱后适应诱导的心肌保护作用

近年来缺血后适应的提出成为抗再灌注损伤的里程碑,其良好的临床可控性和可靠的保护效应引起人们广泛关注。缺血后适应即在心肌长时间缺血后再灌注之前,进行数次短暂的再灌注,缺血的循环处理,诱导产生心肌保护效应,其循环次数和间隔时间存在种属差异。研究证实后适应不仅限制急性期梗死面积,还可以减轻长期损伤,其是否与保护血管内皮、抑制中性粒细胞介导的氧化损伤相关还存在争议。上调再灌注损伤补救激酶（reperfusion injury salvageHnase,RISK）通路是后适应保护的重要机制之一,即激活磷脂酰肌醇一3激酶（phosphatidy linositol3-kinase,P13K）-Akt途径和,或细胞外信号调节激酶（extracellular signal-regulatedkinase,ERK）途径,抑制线粒体通透性转换孔的开放,减少细胞凋亡和坏死。但是这两条途径的地位和关系还有待于进一步研究。为了更加适用于临床,研究者将机械调控转变为药物干预,观察药物能否模拟缺血后适应发挥保护作用,即药物后适应。腺苷是研究最广泛,也是最有希望成为临床正式用药的一种药物。我们实验室首先提出了乙酰胆碱可以模拟缺血后适应,通过线粒体ATP敏感钾通道发挥心肌保护效应。本文着重阐述缺血后适应保护缺血,再灌注损伤的效应和信号转导通路,尤其是腺苷和乙酰胆碱模拟药物后适应的可能机制和临床应用。     

缺血预处理对缺血/再灌注离体心脏的保护作用

目的：探讨连续多次短暂缺血预处理对缺血/再灌注损伤心肌的保护作用及机制。方法：采用大鼠离体心脏Lan-gendorff灌流模型,观察缺血预处理对心肌缺血/再灌注后不同时间点冠脉流出液中AST、CPK、UDH及冠脉流量,心肌组织中SOD、LPO以及再灌注性心律失常的影响。结果：缺血预处理可以减少缺血/再灌注损伤的心肌冠脉流出液中AST、CPK、LDH的含量,提高心肌SOD活性,降低LPO水平,并且抑制再灌注性心律失常的发生,提高再灌注期间的冠脉流量。结论：缺血预处理对心肌缺血/再灌注损伤具有一定保护作用。     

中药抗心肌缺血再灌注损伤的信号通路研究进展

冠心病发生率、致死率高,严重危害人类健康。心肌缺血再灌注损伤是加重心肌损伤的主要病理机制,干预再灌注损伤挽救激酶、 单磷酸腺苷激酶、蛋白激酶 C 等信号传导通路保护心肌,成为减轻心肌损伤的重要途径之一。综述近 3 年国际期刊收录的中药有效成分、 提取物及复方制剂调节相关信号传导通路, 减轻心肌再灌注损伤的研究进展, 以期为阐释中药的作用特点, 有效防治心血管疾病提供参考。     

HO-1在缺血后处理抗肺缺血再灌注损伤中的作用及其对STAT-3表达的影响

目的研究血红素加氧酶-1(hemeoxygenase-1,HO-1)在缺血后处理(ischemic postconditioning,IPO)抗肺缺血再灌注损伤中的作用机制及其对STAT-3蛋白表达的影响。方法 40只SD雄性大鼠(250-280 g)随机分为假手术组(S)、缺血再灌注组(IR)、缺血后处理组(IPO)及缺血后处理+HO-1抑制剂组(IPO+ZnPP)。称重法计算缺血肺组织干/湿比(W/D),试剂盒检测缺血肺组织MDA水平及MPO与HO-1活性,Western Blot检测HO-1,p-STAT-3蛋白表达水平。结果与S组比较,IR组大鼠W/D、MDA、MPO、HO-1活性及蛋白表达水平均显著增加,而p-STAT-3蛋白表达水平显著降低,IPO可以逆转上述变化,而HO-1特异性抑制剂可以消除IPO对上述指标的影响。结论 IPO可以通过促进HO-1活性及蛋白表达的增加从而激活STAT-3信号通路而发挥抗肺缺血再灌注损伤作用。     

RISK pathway is involved in oxytocin postconditioning in isolated rat heart

The reperfusion injury salvage kinase (RISK) pathway is a fundamental signal transduction cascade in the cardioprotective mechanism of ischemic postconditioning. In the present study, we examined the cardioprotective role of oxytocin as a postconditioning agent via activation of the RISK pathway (PI3K/Akt and ERK1/2).Animals were randomly divided into 6 groups. The hearts were subjected under 30 minutes (min) ischemia and 100 min reperfusion. OT was perfused 15 min at the early phase of reperfusion. RISK pathway inhibitors (Wortmannin; an Akt inhibitor, PD98059; an ERK1/2 inhibitor) and Atosiban (an OT receptor antagonist) were applied either alone 10 min before the onset of the ischemia or in the combination with OT during early reperfusion phase. Myocardial infarct size, hemodynamic factors, ventricular arrhythmia, coronary flow and cardiac biochemical marker were measured at the end of reperfusion.OT postconditioning (OTpost), significantly decreased the infarct size, arrhythmia score, incidence of ventricular fibrillation, Lactate dehydrogenase and it increased coronary flow. The cardioprotective effect of OTpos was abrogated by PI3K/Akt, ERK1/2 inhibitors and Atosiban.Our data have shown that OTpost can activate RISK pathway mostly via the PI3K/Akt and ERK1/2 signaling cascades during the early phase of reperfusion.     

缺血后处理对肺缺血／再灌注损伤的保护作用及其机制

目的：探讨缺血后处理（聃）是否通过抑制P38丝裂原活化蛋白激酶（P38MAPK）活化来减轻再灌注损伤肺细胞的凋亡。方法：雄性SD大鼠40只,随机分成5组（n=8）,即对照组（C组）、肺缺血／再灌注组（I／R组）、肺缺血／再灌注＋缺血后处理组（IPO组）、缺血后处理＋溶剂对照组（D组）、缺血后处理＋SB203580组（SB组）。各组分别于再灌注2h留取左肺组织,检测肺组织湿／干重比（W／D）和总肺含水量（TLW）;光镜观察肺组织形态学结构改变并进行肺组织损伤定量评估（IQA）;原住末端标记法（TUNEL）检测肺细胞凋亡情况并计算凋亡指数（AI）;RT-PCR和免疫组化法测定Bax、Bcl-2基因和蛋白的表达。结果：与C组相比,I／R组W／D、TLW、IQA和AI均显著升高（P〈0．05,P〈0．01）,肺组织结构发生明显损伤;Bcl-2、Bcl-2／Bax基因及蛋白表达明显降低,Bax基因及蛋白表达明显升高（P〈0．05,P〈0．01）;IPO组、D组、SB组与I／R组相比,w／D、TLW、IQA和AI均显著降低（P〈0．05,P〈0．01）,肺组织结构损伤情况有所改善;Bcl-2、Bcl-2／Bax基因及蛋白表达明显升高,Bax基因及蛋白表达明显降低（P〈0．05,P〈0．01）;D组与IPO组比较各项指标均无明显差异（均P〉0．05）;SB组与IPO组相比,肺组织W／D、TLW、IQA和AI均显著降低（P〈0．05,P〈0．01）,肺组织结构未见明显损伤;Bcl-2、Bcl-2／Bax基因及蛋白表达明显升高,Bax基因及蛋白表达明显降低（P〈0．05,P〈0．01）。结论：I／R通过激活P38MAPK导致大鼠肺泡结构严重破坏,肺内细胞大量凋亡;IPO可能是通过抑制P38MAPK通路的激活而减轻L／R损伤。     

TNF-α与G—CSF在脑缺血-再灌注损伤中的研究

再灌注损伤是由多种原因引起的复杂的病理生理过程,而级联的炎症反应是导致脑细胞损伤的重要病理环节。脑缺血再灌注后,浸润的炎性细胞产生的大量炎性介质,在再灌注损伤中占有重要地位。肿瘤坏死因子α（TNF-α）是一种具有广泛生物学功能的细胞因子,参与机体免疫应答和炎症反应TNF-α是细胞间粘附分子-1（ICAM-1）表达的强诱导剂,抑制细胞粘附分子（ICAM-1）表达可显著减低再灌注时白细胞粘附活化,减少损伤脑面积起保护作用。粒细胞集落刺激因子（G-CSF）能通过STAT途径减少缺血区肿瘤坏死因子-α等的释放,引起人们对其在脑缺血-再灌注损伤中的作用的极大关注。     

姜黄素对自发性高血压大鼠海马缺血/再灌注损伤神经元凋亡核通路的影响

目的:探讨自发性高血压大鼠(SHR)脑缺血/再灌注损伤海马神经元凋亡c-Jun氨基末端激酶(JNK)核通路的变化特点,以及姜黄素对其保护作用可能机制。方法:雄性Wistar-Kyoto大鼠(WKY)和SHR,随机分为5组:WKY假手术组(W-Sham组)、缺血/再灌注组(W-I/R组)和SHR假手术组(S-Sham组)、缺血/再灌注组(S-I/R组)、姜黄素100mg/kg预处理组(S-Cur组),上述5个实验组按再灌注时间又分为再灌注2h、6h、1d、3d、7d5个亚组(n=6)。采用四动脉结扎法制备脑缺血/再灌注模型,以TUNEL法检测海马CA1区的细胞凋亡,免疫组化法分析海马CA1区c-jun、c-fos的动态变化。结果:S-Sham组大鼠海马CA1区TUNEL细胞数量和c-jun、c-fos表达高于W-Sham组(P0.05),S-I/R组TUNEL细胞数量和c-jun、c-fos表达高于S-Sham组及W-I/R组(P0.05);S-Cur组TUNEL细胞数量和c-jun、c-fos表达较S-I/R组明显降低(P0.05)。结论:缺血/再灌注更易导致SHR海马神经元凋亡。姜黄素可抑制SHR脑缺血/再灌注损伤海马神经元凋亡,其作用机制可能与抑制c-jun、c-fos蛋白的表达有关。     

Reduction of SIRT1 blunts the protective effects of ischemic post-conditioning in diabetic mice by impairing the Akt signaling pathway

Ischemic post-conditioning (IPO) activates Akt signaling to confer cardioprotection. The responsiveness of diabetic hearts to IPO is impaired. We hypothesized that decreased cardiac SIRT1, a positive regulator of Akt, may be responsible for the impaired responsiveness of diabetic hearts to IPO-mediated cardioprotection. High-fat diet and streptozotocin-induced diabetic mice were subjected to myocardial ischemia/reperfusion (MI/R, 30 min ischemia and 180 min reperfusion) or IPO (three cycles of 10 s of reperfusion and ischemia at the onset of reperfusion). Adenoviral vectors encoding GFP or SIRT1 (Ad-SIRT1) were administered by direct injection into the left ventricular. Our results showed that IPO activated the Akt signaling pathway and reduced MI/R injury in non-diabetic hearts but not in diabetic hearts, in which reduced expression of SIRT1 and increased Akt acetylation were observed. Delivery of Ad-SIRT1 into the diabetic hearts reduced Akt acetylation and restored the cardioprotective effects of IPO by modulating Akt signaling pathway. In contrast, cardiac-specific SIRT1 knockout increased Akt acetylation and blunted the cardioprotective effects of IPO. In in vitro study, transfection with wild-type SIRT1 but not inactive mutant SIRT1 reduced the expression of Akt acetylation and restored the protective effects of hypoxic post-conditioning in high glucose-incubated cardiomyocytes. Moreover, the cardiomyocytes transfected with constitutive Akt acetylation showed repressed Akt phosphorylation and blunted protective effects against hypoxia/reoxygenation injury. These findings demonstrate that the reduction of SIRT1 blunts the protective effects of IPO by impairing Akt signaling pathway and that SIRT1 up-regulation restores IPO-mediated cardioprotection in diabetic mice via deacetylation-dependent activation of Akt signaling pathway.     

线粒体通透性转换孔在缺血预处理脑保护中的作用及机制

目的：线粒体通透性转换孔通透性改变是导致缺血再灌注损伤的原因,线粒体功能的致命性改变最终引起细胞凋亡,本研究旨在观察线粒体通透性转换孔（mitochondrial permeability transition pore,MPTP）在缺血再灌注及缺血预处理脑保护中的作用;方法：将体外培养8天的海马神经元细胞分为五组,正常对照组（A组）,缺血再灌注组（B组）,缺血预处理＋缺血再灌注组（C组）,苍术苷＋缺血再灌注组（D组）,缺血预处理＋苍术苷＋缺血再灌注组（E组）。使用流式细胞术检测各组细胞凋亡率,罗丹明123染色流式细胞术检测线粒体膜电位,Western-blot检测Bcl-2,Bax的表达。结果：与A组比较,其余四组线粒体膜电位均降低,神经元凋亡率升高（P〈0．05）;与B组比较,c组线粒体膜电位升高,神经元凋亡率升高,Bcl-2表达上调,Bax表达下调（P〈0．05）;与c组比较,E组粒体膜电位降低,神经元凋亡率升高,Bcl．2表达下调,Bax表达上调（P〈0．05）。结论：我们在细胞及分子生物学水平对MPTP及缺血预处理的研究后发现,缺血预处理能有效减轻海马神经元缺血再灌注损伤,抑制缺血再灌注后神经细胞凋亡,其机制与抑制MPTP的开放有关。     

Rho kinase activation plays a major role as a mediator of irreversible injury in reperfused myocardium

Intracellular signal transduction events in reperfusion following ischemia influence myocardial infarct development. Here we investigate the role of Rho kinase (ROCK) activation as a specific injury signal during reperfusion via attenuation of the reperfusion injury salvage kinase (RISK) pathway phosphatidylinositol 3-kinase (PI3K)/Akt/endothelial nitric oxide (NO) synthase (eNOS). Rat isolated hearts underwent 35 min of left coronary artery occlusion and 120 min of reperfusion. Phosphorylation of the ROCK substrate protein complex ezrin-radixin-moesin, assessed by immunoblotting and immunofluorescence, was used as a marker of ROCK activation. Infarct size was determined by tetrazolium staining, and terminal dUTP nick-end labeling (TUNEL) positivity was used as an index of apoptosis. The ROCK inhibitors fasudil or Y-27632 given 10 min before ischemia until 10 min after reperfusion reduced infarct size (control, 34.1 +/- 3.8%; 5 microM fasudil, 18.2 +/- 3.1%; 0.3 microM Y-27632, 19.4 +/- 4.4%; 5 microM Y-27632, 9.2 +/- 2.9%). When 5 microM Y-27632 was targeted specifically during early reperfusion, robust infarct limitation was observed (14.2 +/- 2.6% vs. control 33.4 +/- 4.4%, P<0.01). The protective action of Y-27632 given at reperfusion was attenuated by wortmannin (29.2 +/- 6.1%) and N(omega)-nitro-L-arginine methyl ester (30.4 +/- 5.7%), confirming a protective mechanism involving PI3K/Akt/NO. Ezrin-radixin-moesin phosphorylation in risk zone myocardium confirmed early and sustained ROCK activation during reperfusion and its inhibition by Y-27632. Inhibition of ROCK activation at reperfusion reduced the proportion of TUNEL-positive nuclei in the infarcted region. In conclusion, ROCK activation occurs specifically during early reperfusion. Inhibition of ROCK at reperfusion onset limits infarct size through an Akt/eNOS-dependent mechanism, suggesting that ROCK activation at reperfusion may be deleterious through suppression of the RISK pathway.