胰岛素在缺血心肌保护治疗中的研究进展

自Sodi-Pallares(1962年)首次提出将极化液(GIK)用于治疗缺血性心脏病以来,胰岛素作为GIK的主要成分,主要被当作细胞能量代谢调节剂运用于临床,但其疗效一直存在争议,其争论的焦点主要体现在GIK是否能真正给缺血心肌带来益处。随着对以往胰岛素临床资料回顾性分析以及近年来该领域大量基础临床研究结果,人们对胰岛素作用机制有了新的认识,对其在缺血性心脏病治疗中的价值也给予了认可。本文就近年来胰岛素在心肌保护治疗领域中的研究进展进行综述。1胰岛素/GIK对心肌电稳定和能量代谢的作用Goulston(1912年)首次提出用蔗糖治疗心力衰…     

Endothelin-1-dependent signaling pathways in the myocardium

Endothelin-1 (ET-1) is a locally acting vasoactive peptide that also has profound effects on the contractile properties and growth of the cardiac myocyte. Binding of ET-1 to its transmembrane heptahelical receptors activates G proteins of the G(q) and G(i) classes. Activation of G(q) stimulates hydrolysis of phosphatidylinositol-4,5-bisphosphate, and the diacylglycerol thus formed stimulates protein kinase C. Subsequently, the protein kinase Raf is activated and this leads to activation of the extracellular signal-regulated protein kinase (ERK) subfamily of mitogen-activated protein kinases. Activation of G(i) counteracts β-adrenoceptor-mediated increases in cAMP concentrations. We have attempted to rationalize the established physiological consequences of ET-1 agonism in the cardiac myocyte (that is, on contraction and growth) in terms of activation of these signaling pathways.     

Stunning: Damaging or protective to the myocardium?

Summary There are several potential outcomes of myocardial ischemia. When ischemia is severe and prolonged, irreversible damage occurs and there is no recovery of contractile function. Interventions aimed at reducing mechanical activity and oxygen demand, either before ischemia or during reperfusion, have been shown to delay the onset of ischemic damage and to improve recovery on reperfusion.When myocardial ischemia is less severe but still prolonged, myocytes may remain viable but exhibit depressed contractile function. Under these conditions, reperfusion restores complete contractile performance. This type of ischemia, leading to a reversible, chronic left ventricular dysfunction, has been termed hibernating myocardium. Depression of mechanical activity is, actually, a protective mechanism whereby the hibernating cells reduce their oxygen demands in the setting of reduced oxygen supply.A third possible outcome after a short period of myocardial ischemia is a transient postischemic ventricular dysfunction, a situation termed stunned myocardium. As in the case of hibernating myocardium, the depressed contractile function occurring during stunning could be a protective mechanism, allowing the reperfused cells to gradually recover their metabolism and function.     

Ancestral Notch-mediated segmentation revealed in the cockroach Periplaneta americana

Through division into segments, animal bodies can reach higher degrees of complexity and functionality during development and evolution. The segmentation mechanisms of insects and vertebrates have been seen as fundamentally different at the anatomical and molecular levels, and consequently, independently evolved. However, this conclusion was mostly based on observations of derived insects such as Drosophila. We have cloned the Delta, Notch, and hairy genes in the cockroach Periplaneta americana, a basal insect with short germ-band development, and carried out functional assays of Notch activity during its segmentation. Our results show that, in more basal insects, segmentation involves a similar developmental mechanism to that in vertebrates, including induction of segment formation by cyclic segmental stripes of hairy and Delta expression. This result indicates that Notch-mediated segmentation is the ancestral segmentation mechanism of insects, and together with previous results in the literature [Stollewerk A, Schoppmeier M, Damen WGM (2003) Nature 423:863-865], of arthropods as well. The similarity with vertebrate segmentation might suggest that Notch-mediated segmentation is an ancient developmental mechanism inherited from a common ancestor of insects and vertebrates.     

The protective effect of prazosin on the ischaemic and reperfused myocardium

Experiments were undertaken to establish whether prazosin prevents isolated hearts from gaining excess Ca2+ during post-ischaemic reperfusion, to determine whether this effect is dose-dependent, if it is accompanied by a change in the energy-rich phosphate reserves, and whether prazosin is effective when added only upon reperfusion. Isolated, spontaneously beating rat hearts were used. The ischaemic episodes ranged from 15 to 60 min, and prazosin (0.01 to 10 micro mol/1) was added both before inducing ischaemia and upon reperfusion. When 0.01 to 1 micro mol/1 prazosin was present before and after the ischaemic episode the reperfusion-induced gain in Ca2+ was attenuated, but not abolished. Pretreatment with 0.01 to 1 micro mol/1 prazosin slowed the ischaemic-induced rise in resting tension, enhanced mechanical recovery after 30 but not 60 min ischaemia, and exerted a dose-dependent slowing effect on the ischaemia-induced depletion of ATP and CP, with 1 micro mol/1 being the optimal dose. Adding 0.01 to 1 micro mol/1 prazosin at the time of reperfusion neither prevented excess Ca2+ accumulation upon reperfusion nor did it exert an energy-sparing effect. 5 to 10 micro mol/1 prazosin did not attenuate the reperfusion-induced gain in Ca2+, irrespective of whether it was added before or only at the time of reperfusion. These results show that the dose-response curve for the inhibitory effect of prazosin on Ca2+ overload is complex, and that adding prazosin coincident with the reperfusion of isolated ischaemic hearts does not attenuate Ca2+ gain.     

硒谷胱甘肽过氧化物酶保护心肌作用机制

通过人工半合成及天然低成饲料喂养豚鼠，在出现组织硒水平及谷胱甘肽过氧化物酶（ＧＰ）活性明显降低同时，动物红细胞、心肌细胞等的膜结构出现异常，膜脂组成中的磷脂减少．代表细胞膜老化指数的胆固醇／磷脂的分子比增加，作为心肌线粒体内膜必需界面脂的心磷脂明显减少，线粒体能量转换的关键呼吸酶．细胞色素氧化酶（ＣＣＯ）活性明显降低，同时代表其二级结构的园二色谱异常，２０８ｎｍ峰及２２４ｎ峰明显降低．各型脂质过氧化产物，如代表膜磷脂自由基氧化损伤初级产物的脂氢过氧化物、续发产物的丙二醛、呼气烷烃和脂过氧化物聚集产物的荧光色脂均明显增加。这可能正是３０多年前Ｎｅｕｂｅｒｔ等（１９６２）提出ＧＰ对细胞膜的保护作用，尤其所谓线粒体“收缩因子”（“Ｃｏｎｔｒａｃｔｉｏｎ　ｆａｃｔｏｒ”）作用的机制所在。     

The protective role of heat stress in the ischaemic and reperfused rabbit myocardium.

Cells subjected to increases in temperature induce the expression of several proteins known as heat shock or stress proteins. This process enhances the cell's ability to overcome the effects of further stress. In this respect, the effects of heat stress have been reported to protect the hearts of rats following ischaemia and reperfusion. We have confirmed and extended this observation, not only using different indices of myocardial injury but also in another species, namely the rabbit. Animals were anaesthetized and the body temperature raised to 42 degrees C for a 15-min period. Controls were treated in the same way but without heating. Twenty-four hours later the rabbits were re-anaesthetized and the hearts removed for either heat stress protein analysis or perfusion with Krebs buffer using an isolated perfused heart apparatus. Hearts were subjected to 60 min of low flow (1 ml/min) ischaemia followed by 30 min of reperfusion. All hearts subjected to heat stress showed an enhanced recovery of function upon reperfusion as measured by improvements in developed pressure (27.3 +/- 3.6 vs 16.3 +/- 3.0 mmHg) and diastolic pressure (37.3 +/- 7.4 vs 54.7 +/- 3.1 mmHg). In addition, creatine kinase release, associated with reperfusion, was significantly reduced in the heat-stressed hearts (532 +/- 102 vs 1138 +/- 73 mU/min/g wet wt). Myocardial accumulation and release of oxidized glutathione, an index of oxidative stress, was significantly reduced in the heat-stressed group (0.003 +/- 0.003 vs 0.376 +/- 0.113 nmol/min/g wet wt). The improved metabolic status of the reperfused heat-stressed hearts was further demonstrated by a significant conservation in the levels of ATP (6.1 +/- 0.9 vs 2.8 +/- 0.8 mumol/g dry wt) and CP (36.9 +/- 6.4 vs 16.4 +/- 5.1 mumol/g dry wt). Finally, isolated mitochondrial function in terms of respiratory control index (RCI) was maintained in the heat-stressed hearts (9.2 +/- 0.9 vs 5.7 +/- 0.2) and overloading with calcium was reduced. These data extend the hypothesis that heat stress protects the heart following ischaemia and reperfusion in this in vitro model, in a way as yet undetermined.     

吡格列酮对大鼠缺血再灌注诱导的内质网应激相关的心肌保护作用

目的观察吡格列酮对大鼠心肌缺血再灌注损伤（MIRI）时JNK、p-JNK及caspasc-12蛋白表达的影响,探讨吡格列酮通过JNK通路对内质网应激途径的心肌保护作用。方法Wistar大鼠40只随机分为假手术组（sham组）、缺血再灌注组（I／R组）、I／R＋Pio（吡格列酮）组及I／R＋Pi0＋sP600125组各10只。制作大鼠MIRI模型;TUNEL检测心肌细胞凋亡,免疫组织化学检测各组caspase-12表达变化,westernBlot法检测各组JNK、P-JNK的表达。结果吡格列酮预处理组大鼠心肌细胞凋亡、JNK磷酸化率及caspase-12蛋白表达水平明显比I／R组降低（P〈0．05）,加用JNK抑制剂（SP600125）后上述指标进一步下降,与吡格列酮组比较差异有统计学意义（P〈0．05）。结论缺血再灌注损伤可激活JNK通路,诱导过度的ERS,增加ER凋亡信号介导的细胞凋亡。吡格列酮预处理可减少ER凋亡信号介导的细胞凋亡,JNK信号途径在吡格列酮预处理抑制ER凋亡信号分子活化的机制中发挥重要作用。     

三磷酸肌醇受体参与心肌钙调神经磷酸酶信号通路的激活

目的:研究激活心肌细胞三磷酸肌醇受体（IP3R）是否触发钙调神经磷酸酶（CaN）介导的心肌肥厚信号通路。方法:培养的乳鼠心肌细胞检测心肌细胞蛋白合成,细胞内钙变化,CaN、活化T细胞核因子3（NFAT3）及锌指转录因子（GATA4）,胚胎基因（αskeletalactin,βMHC）及即刻早期基因（cfos,cmyc）蛋白表达。结果:三磷酸肌醇（IP3）能显著致原代培养的心肌细胞时间和剂量依赖性地增加心肌细胞蛋白合成,能显著致心肌细胞的早期即刻基因和胚胎基因表达,能显著增加心肌细胞内游离钙。IP3剌激IP3受体,能显著激活心肌细胞CaN/NFAT3/GATA4信号通路,使心肌细胞早期即刻基因（cfos、cmyc）及胚胎基因（αskeletalactin,βMHC）表达增加。结论:激活IP3R介导的CaN/NFAT3/GATA4信号通路能显著地促使心肌细胞肥大,这条信号通路不同于已知的G蛋白偶联受体介导的心肌肥厚信号转导途径。     

依达拉奉对大鼠心肌再灌注损伤的保护作用及机制

目的探讨自由基清除剂依达拉奉对大鼠心肌再灌注损伤的保护作用及机制。方法将造模成功的24只Wistar大鼠随机分为对照组、缺血再灌注组(再灌注组)、保护组,每组8只。实验结束后检测各组大鼠血清肌酸激酶同工酶(CK-MB)、谷胱甘肽过氧化物酶(GSH-PX)活性及丙二醛水平;光学显微镜观察心肌形态;采用TUNEL和免疫组织化学法分别检测心肌细胞凋亡及心肌凋亡蛋白Bcl-2、Bax的表达。结果与对照组比较,再灌注组大鼠CK MB、丙二醛水平明显升高,GSH-PX活性明显降低,Bcl-2、Bax、Bax/Bcl-2明显升高(P0.01);与再灌注组比较,保护组大鼠CK-MB、丙二醛水平明显降低,GSH-PX活性明显升高,Bcl-2明显升高,Bax、Bax/Bcl-2明显降低(P0.01)。与保护组比较,再灌注组大鼠可见大面积心肌梗死,凋亡细胞数明显增加(P0.01)。结论依达拉奉能够减轻大鼠心肌损伤程度,其机制与减少自由基损伤、抑制心肌细胞凋亡有关。     

Activation of mitogen-activated protein kinases in the non-ischemic myocardium of an acute myocardial infarction in rats

As one of the signal transduction pathways related to myocardial remodeling, mitogen-activated protein kinases (MAPKs) possibly play an important role in ischemic heart disease, but it is still unknown whether myocardial MAPKs are activated in the non-ischemic region of an acute myocardial infarction (AMI). Therefore, the present study investigated the myocardial activity of extracellular signal-regulated kinases (ERKs), c-Jun NH2 terminal kinases (JNKs) and p38MAPK during the acute phase of an infarction of the rat heart, and measured the geometrical ventricular changes by echocardiography. All MAPKs were significantly activated in the ischemic myocardium (IM), non-ischemic septal wall (SW), and right ventricular wall (RV). Furthermore, the activation patterns of MAPKs differed in each region. The activation of p44ERK, JNKs and p38MAPK in the IM occurred rapidly after myocardial ischemia, followed by those in the SW and RV. The activator protein-1 DNA binding activities of the IM, SW and RV increased significantly at I day after coronary ligation. Echocardiography showed increased SW motion and RV dilatation. In conclusion, this is the first in vivo evidence that myocardial MAPKs are activated in the non-ischemic region of an AMI. Echocardiographic results suggest that acceleration of workload and/or stretch may partially induce the activation of MAPKs.     

缺血预适应及侧支循环对缺血再灌注心肌的保护作用

目的 探讨急性心肌梗死前心绞痛产生的缺血预适应独立于或协同冠状动脉侧支循环对缺血-再灌注心肌的保护作用。方法 44例发病6 h内接受直接冠状动脉介入治疗(PCI)的急性心肌梗死患者,按梗死前48 h内有无心绞痛分成2组,有梗死前48 h内心绞痛(PA)组24例、无梗死前48 h内心绞痛(NPA)组20例,每组再按有无冠状动脉侧支循环血管分成两亚组,PCI完成后对梗死相关血管(IRA)血流进行TIMI分级评价,同时左心室造影,记录室壁运动记分(WMS)及左心室舒张末期压力(LVEDP),监测心肌酶48 h,PCI完成后1周、4周末分别行99mTc-MIBI心肌灌注显像(SPECT),比较前后显像缺损程度记分(SS)的变化,计算并比较两组及亚组间心肌挽救指数(MSI),第2周时行平衡法核素心室造影(ERNA),比较两组及亚组间左心室收缩功能参数、左心室舒张功能参数、心室收缩同步性(LVSS)的差异。结果 PA组心肌损伤标记物肌酸激酶(CK)及其同工酶(CK-MB)峰值水平显著低于NPA组[(1172±985)U/L比(2291±1267)U/L,P<0.05;(197±102)U/L比(316±144)U/L,P<0.05],冠状动脉造影(CAG)结果显示两组侧支循环Rentrop分级无差异,PCI完成时PA组IRA无再流(no-reflow)发生率显著低于NPA组(8.3％比20％,P<0.05),WMS和INEDP也显著低于NPA组[分别为(5.39±0.91比7.11±1.27),P<0.0     

Regional desensitization of beta-adrenergic receptor signaling in swine with chronic hibernating myocardium

Contractile reserve during submaximal beta-adrenergic stimulation is attenuated in patients and swine with hibernating myocardium. We tested the hypothesis that this arises as a regional adaptive response in beta-adrenergic adenylyl cyclase coupling. Pigs (n=8) were studied 3 months after instrumentation with a left anterior descending artery (LAD) stenosis when flow (LAD, 0.7+/-0.2 versus 1.2+/-0.1 mL/min per gram in normal remote; P<0.05) and wall thickening (LAD, 15.5 [corrected]+/-3.2% versus 40.0+/-5.5% in remote; P<0.05) were reduced in the absence of infarction. Whereas basal cAMP production was normal (LAD, 87+/-18 versus 91+/-19 pmol/mg per minute; P=NS), responses to isoproterenol were blunted (LAD, 83+/-6 versus 146+/-25 pmol/mg per minute in remote; P<0.05). beta-receptor density and subtype were unchanged, but there was a reduction in the number of high-affinity binding sites (LAD, 40+/-4% versus 53+/-7% in normal remote; P<0.05). The Gialpha2/Gsalpha ratio increased (LAD, 1.8+/-0.3 versus 0.99+/-0.3 in remote myocardium; P<0.05), although GppNHp-stimulated cAMP production was equivocally reduced. Forskolin responses were unchanged and similar to shams. These data indicate regional attenuation of beta-receptor adenylyl cyclase signaling in hibernating myocardium. This blunts the local contractile response to beta-adrenergic stimulation and may serve to protect against a myocardial supply/demand imbalance when external determinants of myocardial workload increase during sympathetic activation.     

Redox regulation of angiotensin II preconditioning of the myocardium requires MAP kinase signaling

A recent study documented reactive oxygen species (ROS), generated through NADPH oxidase by angiotensin II (Ang II) with the activation of NADPH oxidase subunits, p22phox and gp91phox, to be responsible for the preconditioning effect of Ang II. The present study was designed to determine if similar to ischemic preconditioning (PC), mitogen-activated protein (MAP) kinases are also involved in Ang II PC of the heart. Isolated working rat hearts were perfused for 15 min with KHB (Krebs-Henseleit bicarbonate) buffer containing Ang II in the absence or presence of an Erk (1/2) inhibitor, PD 098059, a p38MAPK inhibitor, SB 202190, a JNK inhibitor, SP 600125 or a ROS scavenger, N-acetyl cysteine (NAC). All hearts were subsequently subjected to 30 min global ischemia followed by 2 h reperfusion with KHB buffer only. Cardioprotection was examined by determining infarct size, cardiomyocyte apoptosis and ventricular recovery. Redox and MAP kinase regulation were studied by determining the survival signaling mediated by Akt and Bcl-2. In consistent with previous results, Ang II preconditioned the heart as evidenced by improved postischemic ventricular recovery and reduced infarct size and decreases cardiomyocyte apoptosis. Ang II phosphorylated both Akt, Bcl-2 and Bad, which was blocked by NAC, PD 098059 or SP 600125, but not by SB 202190. NAC, PD 098059 and SP600125, but not SB202190, also abolished the cardioprotective effect of Ang II preconditioning. The results indicate that Ang II preconditioning is potentiated through MAP kinases that are regulated by redox signaling.     

Apoptosis signaling pathways in osteoarthritis and possible protective role of melatonin

Osteoarthritis (OA) is a degenerative joint disease characterized by progressive erosion of articular cartilage. As chondrocytes are the only cell type forming the articular cartilage, their gradual loss is the main cause of OA. There is a substantial body of published research that suggests reactive oxygen species (ROS) are major causative factors for chondrocyte damage and OA development. Oxidative stress elicited by ROS is capable of oxidizing and subsequently disrupting cartilage homeostasis, promoting catabolism via induction of cell death and damaging numerous components of the joint. IL‐1β and TNF‐α are crucial inflammatory factors that play pivotal roles in the pathogenesis of OA. In this process, the mitochondria are the major source of ROS production in cells, suggesting a role of mitochondrial dysfunction in this type of arthritis. This may also be promoted by inflammatory cytokines such as IL‐1β and TNF‐α which contribute to chondrocyte death. In patients with OA, the expression of endoplasmic reticulum (ER) stress‐associated molecules is positively correlated with cartilage degeneration. Melatonin and its metabolites are broad‐spectrum antioxidants and free radical scavengers which regulate a variety of molecular pathways such as inflammation, proliferation, apoptosis, and metastasis in different pathophysiological situations. Herein, we review the effects of melatonin on OA, focusing on its ability to regulate apoptotic processes and ER and mitochondrial activity. We also evaluate likely protective effects of melatonin on OA pathogenesis.     

细胞外调节蛋白激酶通路对胃癌细胞化疗效果的影响

目的:观察细胞外调节蛋白激酶(extraeelluar regulated protein kinase,ERK)信号通路对胃癌细胞化疗效果的影响并探讨其机制.方法:足叶乙甙作用于胃癌SGC7901和BGC823细胞,采用MTT比色法检测细胞的生存率,采用流式细胞仪和Hoechst33258荧光染色检测细胞周期分布和凋亡,Western杂交法检测ERK1/2的磷酸化以及c-Myc和P53蛋白表达水平.同时采用PD98059抑制ERK信号通路后观察足叶乙甙对细胞增殖、凋亡、c-Myc和P53表达的影响.结果:足叶乙甙呈时间-剂量依赖性抑制SGC7901和BGC823细胞的生长并明显诱导细胞的凋亡,同时上调ERK1/2的活性(磷酸化水平),并增强c-Myc和P53的表达,与对照组比较,足叶乙甙组凋亡率明显增高(19.48%±1.57% vs 5.67%±0.81%.17.38%±1.49% vs4.97%±0.73%,均P<0.01),PD98059可明显增强足叶乙甙的细胞生长抑制作用并提高细胞的凋亡水平,与足叶乙甙组比较,足叶乙甙组 PD98059组凋亡率(34.35%±2.84%,32.11%±3.25%)明显增加(P<0.01);同时上调足叶乙甙诱导的P53表达,并抑制c-Myc表达的上升趋势.结论:足叶乙甙可活化胃癌细胞ERK信号通路而影响胃癌细胞的化疗效果,其机制可能是通过抑制P53并上调c-Myc的表达,从而抑制细胞的凋亡实现.