胞浆游离钙和组织ATP,ADP,AMP在脑复苏中变化及药物的保护作用

目的：为了探讨胞浆游离钙、ＡＴＰ、ＡＤＰ、ＡＭＰ在脑缺血和再灌流损伤中的作用及相互关系以及药物的影响。方法：应用沙鼠建立脑缺血模型。缺血前１５分钟腹腔注射药物，分别阻断双侧颈动脉５０分钟，阻断颈动脉５０分钟再灌流１０分钟，再灌流６０分钟及１２０分钟。检测脑组织中ＡＴＰ、ＡＤＰ和ＡＭＰ的变化以及应用尼莫地平及东莨菪碱后的影响。结果：（１）胞浆游离钙在脑缺血５０分钟后大幅度升高，再灌流时再度升高，然后缓慢下降。（２）在缺血期ＡＴＰ迅速下降，再灌流期先上升，此后再度下降，而ＡＤＰ及ＡＭＰ在缺血及再灌流期积聚。（３）尼莫地平及东莨菪碱对减缓胞浆游离钙升高，减缓ＡＴＰ的耗竭均有作用。结论：对脑缺血和再灌流损伤有治疗意义。     

沙鼠脑缺血和再灌流后胞浆游离钙和ATP的相关性及东莨菪碱的影响

探讨胞浆游离钙及ＡＴＰ在脑缺血和再灌流损伤中的作用及相互关系以及药物的影响。方法应用沙鼠建立脑缺血模型，缺血前１５ｍｉｎ腹腔注射药物，分别阻断双侧颈总动脉３０ｍｉｎ，５０ｍｉｎ，阻断颈动脉５０ｍｉｎ再灌流１０ｍｉｎ、再灌注６０ｍｉｎ及１２０ｍｉｎ。     

尼莫地平及参麦注射液对脑复苏效应的实验研究

猪经股动脉放血降压至术前的50％，同时阻断其双侧颈总动脉和推动脉。10min后出现平坦脑电图（EEG）且伴随全身抽搐。制成上述急性完全性脑缺血模型5min后，分三组分别给予尼莫地平（0．2mg／kg）、参麦注射液（1ml／kg）及等量生理盐水静滴，比较尼莫地平、参麦注射液对脑缺血再灌流损伤的复苏效应。与对照组相比，结果表现为尼莫地平促进脑缺血再灌流猪EEG幅度的有效恢复且降低了EEG的异常率（P＜0．01）；参麦注射液有类似效果，但作用软弱（P＜0．05）；尼莫地平和参麦注射液均可抑制再灌流损伤后脑环磷酸腺苷（cAM）的升高（P＜0．05）；减轻脑水肿和大脑皮层损伤的程度。提示尼莫地平注射液和参麦注射液对缺血再灌流脑组织损伤有明显拮抗作用。     

盐酸戊乙奎醚对大鼠脑缺血-再灌流损伤的保护作用

目的 探讨盐酸戊乙奎醚对大鼠三动脉阻断法急性全脑缺血．再灌流损伤的保护作用。方法 144只雄性SD大鼠随机分为假手术组、缺血-再灌流组（生理盐水1ml）、东莨菪碱组（0．01mg／kg）和盐酸戊乙奎醚组（0．01mg／kg），缺血前40min分别腹腔注射相应药物，缺血时间为10min，于再灌流24h、3d和7d测定其行为学（开阔法、平衡木法、攀绳和肌力试验），再灌流2h、12h、24h、3d和7d取材测定海马CA1区存活神经元数量（HE染色）、凋亡神经元数量（TUNEL染色）、bcl-2和bax蛋白表达（免疫组化染色），部分大鼠于再灌流4d测定脑梗死体积（TTC法）。结果 与缺血-再灌流组比较，盐酸戊乙奎醚组和东莨菪碱组海马CA1区凋亡神经元数量减少（P〈0．01），bcl-2提前并延长表达（P〈0．01），盐酸戊乙奎醚组bax表达下降（P〈0．05或P〈0．01）。同时，盐酸戊乙奎醚组和东莨菪碱组存活神经元数量增加（P〈0．05），脑梗死体积缩小（P〈0．05或P〈0．01），行为学检测指标改善（P〈0．05）。盐酸戊乙奎醚组较东莨菪碱组作用更加明显（P〈0．05）。结论 盐酸戊乙奎醚对脑缺血．再灌流损伤大鼠具有脑保护作用，并且优于同样剂量东莨菪碱。改变bcl-2／bax的比例可能是其机制之一。     

山莨菪碱对全脑缺血-再灌流后脑线粒体损伤的保护作用

目的：探讨山莨菪碱对急性全脑缺血－再灌流后脑线粒体损伤的保护作用。方法：采用家兔全脑缺血－再灌流损伤模型。缺血20min，再灌流2h,观察脑线粒体呼吸功能、呼吸缺氧化酶活性、线粒体内Ca^2 和丙二醛含量的变化。结果：脑缺血－再灌流后，脑线粒体呼吸控制率、磷氧化、氧化磷酸化效率及烟酰胺腺嘌呤二核苷酸氧化酶、琥珀酸氧化酶、细胞色素C氧化酶活性明显降低（P＜0．01），而线粒体Ca^2 、丙二醛含量明显升高（P＜0．01）；再灌流早期给予山莨菪碱治疗后，上述线粒体损伤性改变明显减轻。结论：山莨菪碱对脑缺血－再灌流后线粒体损伤具有一定的保护作用，其机制可能与Ca^2 拮抗、抑制脂质过氧化及保护呼吸链酶活性有关。     

山莨菪碱对急性完全性脑缺血再灌流后脑组织Ca2+、MDA含量、SOD活性及超微结构的影响

40只家兔随机分为假手术组（A组）、缺血组（B组）、缺血再灌流组（C组）、治疗组（D组）。采用闭塞双侧颈总动脉和椎动脉及体循环低血压法建立急性完全性脑缺血再灌流损伤模型，缺血时限20min，再灌流2h，观察了脑组织Ca2＋、脂质过氧化产物一丙二醛（MDA）含量、超氧歧化酶（SOD）活性及脑组织超微结构改变。结果发现：治疗组于再灌流前1min注射山莨菪碱10mg／kg体重，并以5mg／h维持2h，脑组织Ca2＋、MDA含量较缺血组及缺血再灌流组明显降低，（P＜0．05，P＜0．01），SOD活性较缺血再灌流组明显增加（P＜0．05）同时脑组织超微结构损伤明显减轻。结果表明；再灌流早期给予山莨菪碱治疗对完全性脑缺血再灌注损伤具有明显保护作用。膜稳定作用、Ca2＋拮抗作用、抗脂质过氧化是其重要的作用环节。     

脑缺血小鼠脑和血中血小板活化因子的变化

本文对小鼠脑缺血期、再准流期脑和血中血小板活化因子（PAF）进行了测定，结果提示：缺血期及再灌流期脑和血中PAF均高于正常对照组（P＜0．01），以再灌流期升高为明显，本文就PAF对脑缺血的影响进行了讨论。     

脑缺血再灌流损伤实验研究

目的：探讨低温、ATP、醒脑净对脑缺血再灌流损伤的保护作用。方法：选纯种白兔通过“四管闭塞法”制成兔全脑缺血再灌流损伤模型，分四组测定丙二醛（MDA）和谷胱甘肽过氧化物酶（GSH－PX），同时电镜观察脑组织切片。结果：再灌流30min后实验各组与对照组相比，低温组和ATP组MDA显著低于对照组（P＜0．01），醒脑静组MDA显著低于对照组（P＜0．05）；低温组和ATP组GSH－Px显著高于对照组（P＜0．01），醒脑静组GSH－Px显著高于对照组（P＜0．05）。其中低温组的32～34℃组与26～28℃组之间MDA和GSH－Px无显著差别（P＞0．05）。电镜观察显示实验各组的脑损伤比对照组轻。结论：低温、ATP、醒脑净对脑缺血再灌流损伤有保护作用。     

脑缺血再灌注对大鼠不同脑区丙二醛、谷胱甘肽含量及谷胱甘肽过氧化物酶活性的影响

目的：探讨脑缺血再灌注时自由基代谢变化在迟发性神经元损伤中作用。方法：采用闭塞大鼠４条动脉全脑缺血模型，在全脑缺血３０分钟和缺血后再灌注不同时间，分别观察某些脑区丙二醛（ＭＤＡ）、谷胱甘肽（ＧＳＨ）含量以及谷胱甘肽过氧化物酶（ＧＳＨＰｘ）活性变化。结果：在大脑皮层和海马中，缺血３０分钟后，ＧＳＨ、ＧＳＨＰｘ显著下降，细胞膜ＭＤＡ有所增加，但无显著性差异。随再灌注时间延长，胞浆中ＧＳＨ逐渐回升，而ＧＳＨＰｘ进一步下降，并且细胞膜ＭＤＡ显著升高。在丘脑和下丘脑，各组ＧＳＨ、ＭＤＡ变化均无显著性差异，ＧＳＨＰｘ变化则与大脑皮层和海马中相似，但下降幅度较小。结论：脑缺血引发的自由基损伤主要发生在缺血再灌注期，且海马是脑缺血再灌注损伤中最易损伤区。     

氨基胍对大鼠脑缺血-再灌流损伤后脑微血管通透性的影响

目的：研究选择性诱导型一氧化氮合酶（iNOS)抑制剂氨基胍对脑缺血－再灌流损伤时脑微血管通透性的作用。方法：通过缺血60min,再灌流60min,造 成缺血－再灌流（I／R）损伤,并制成脑窗模型,观察氨基胍对脑微循环通透性的影响。结果：脑缺血－再灌流损伤后脑微循环通透性明显升高 ,并在110s后血管外荧光物质浓度就显著高于血管内,而运用氨基胍后,在80s后血管外的荧光物质浓度就显著高于血管内,结论：I／R后脑微循环通透性升高,iNOS激活产生的少量NO在I／R早期可能具有维持微血管通透性的作用。     

山莨菪碱对家兔急性完全性脑缺血及再灌流后脑组织[Ca2+]i和超微结构影响

目的　研究山莨菪碱对急性完全性脑缺血及再灌注后脑组织游离Ca2 ([Ca2 ]i)及超微结构的影响。方法　采用闭塞双侧颈总动脉和椎动脉及体循环低血压法建立家兔急性完全性脑缺血及再灌流损伤模型 ,缺血 2 0min ,再灌流 2h。40只家兔随机分为假手术组、缺血组、缺血再灌流组、治疗组 ,观察了脑组织 [Ca2 ]i和超微结构及山莨菪碱对其变化的影响。结果　缺血组及缺血再灌流组脑组织 [Ca2 ]i 浓度明显高于假手术组 (P <0 0 1) ,而且缺血再灌流组 [Ca2 ]i 明显高于缺血组(P <0 0 1) ,随着 [Ca2 ]i 增加 ,脑组织超微结构损伤明显加重。治疗组 [Ca2 ]i 浓度较缺血组及缺血再灌流组明显降低 (P <0 0 1) ,同时脑组织超微结构损伤明显减轻。结论　山莨菪碱通过阻断Ca2 内流对完全性脑缺血及再灌损伤具有保护作用。     

全脑缺血再灌注后脑线粒体功能变化及山莨菪碱的保护作用

目的：探讨全脑缺血再灌注后脑线粒体功能变化及山莨菪碱的保护作用。方法：采用家兔全脑缺血再灌注损伤模型，缺血20min、再灌注2h观察脑线粒体呼吸功能、呼吸链氧化酶活性、线粒体内Ca^2 和MDA含量的变化。结果:脑缺血再灌注后，脑线粒体呼吸控制率、磷氧比、氧化磷酸化效率及烟酰胺腺喋呤二核苷酸氧化酶、琥珀酸氧化酶、细胞色素C氧化酶活性明显降低(P＜0、01)，而线粒体Ca^2 、MDA含量明显升高(P＜0．01)；再灌注早期给予山莨菪碱治疗后，上述线粒体损伤性改变明显减轻。结论：山莨菪碱对脑缺血再灌注后线粒体损伤具有一定的保护作用，其机制可能与Ca^2 拮抗、抑制脂质过氧化及保护呼吸链酶活性有关。     

Levosimendan: effects of a calcium sensitizer on function and arrhythmias and cyclic nucleotide levels during ischemia/reperfusion in the Langendorff-perfused guinea pig heart.

The majority of clinically used inotropes act by increasing cytosolic calcium levels, which may hypothetically worsen reperfusion stunning and provoke arrhythmias. We tested the hypothesis that the calcium sensitizer levosimendan (levo) given during ischemia alone or ischemia and reperfusion would improve reperfusion function without promoting arrhythmias. The Langendorff-perfused guinea pig heart, subjected to 40-min low-flow ischemia (0.4 ml/min) with or without levo (10-300 nM) given during ischemia or ischemia/reperfusion was used. Left ventricular developed pressure (LVDP) was used as an index of mechanical function. The effect of levo (300 nM) or dobutamine (0.1 microM) on the incidence of ischemia/reperfusion arrhythmias was also investigated. Control hearts (vehicle-perfused) had LVDPs of 69.4 +/- 2.1 mm Hg whereas hearts treated with levo during ischemia and reperfusion (300 nM) had LVDPs of 104.5 +/- 2.7 mm Hg (p <.05). Hearts treated with levo during ischemia alone (10 nM) had reperfusion LVDPs of 95.8 +/- 4.2 mm Hg (p <.05) after 30-min reperfusion. Hearts treated with both levo and 10 microM glibenclamide (K(ATP) channel blocker) during ischemia had reperfusion LVDPs of 73.4 +/- 4.3 mm Hg after 30-min reperfusion. Of control hearts, 25% developed reperfusion ventricular tachycardia but not ventricular fibrillation. Levo-treated hearts had no ischemia/reperfusion arrhythmias whereas 83% (p <.05 versus control) of dobutamine-treated hearts developed ventricular tachycardia and 33% (p <.05 versus levo) developed reperfusion ventricular fibrillation. Levo improved reperfusion function without promoting arrhythmias in this model. This was possibly achieved by opening the K(ATP) channels during ischemia and sensitizing myocardial contractile apparatus instead of elevating cytosolic calcium levels in reperfused hearts.     

The beneficial effect of 2′-deoxycoformycin in renal ischemia-reperfusion is mediated both by preservation of tissue ATP and inhibition of lipid peroxidation

Renal ischemia injures the renal tubular cell by disrupting the vital cellular metabolic machinery. Further cell damage is caused when the blood flow is restored by oxygen free radicals that are generated from xanthine oxidase. Oxygen radicals cause lipid peroxidation of cell and organelle membranes, disrupting the structural integrity and capacity for cell transport and energy metabolism. In the present study, the possible therapeutic usefulness of the adenosine deaminase inhibitor, 2'-deoxycoformycin (DCF), during renal ischemia and reperfusion injury was investigated. The effects of DCF on renal malondialdehyde (MDA) and ATP levels were studied after 45 min ischemia and 15 min subsequent reperfusion in rat kidneys. MDA levels remained unchanged during ischemia, but increased after the subsequent reperfusion. DCF pretreatment (2.0 mg/kg i.m.) decreased MDA and increased ATP levels during the ischemia-reperfusion period. DCF exerts a dual protective action by facilitating purine salvage for ATP synthesis and inhibiting oxygen radical-induced lipid peroxidation. These results suggest that DCF therapy could be beneficial in the treatment of ischemia-reperfusion renal injuries.     

Neutrophil elastase inhibitor (ONO-5046) attenuates reperfusion-induced hepatic microcirculatory derangement, energy depletion and lipid peroxidation in rats

Microcirculatory derangement, energy depletion, and lipid peroxidation are associated with the development of ischemia-reperfusion injury in the liver. This study investigated the effects of a neutrophil elastase inhibitor (ONO-5046) on hepatic ischemia-reperfusion injury. Adult, male Sprague-Dawley rats were divided into four treatment groups: 1) sham-operated control (laparotomy only, no ischemia) and saline injection (1 mL/kg), n = 6; 2) ischemia control (1-h ischemia, 2-h reperfusion) and saline injection (1 mL/kg), n = 6; 3) intravenous injection with ONO-5046 at a dose of 1 mg/kg 5 min before ischemia and immediately after reperfusion plus 1-h ischemia and 2-h reperfusion, n = 6; and 4) intravenous injection with ONO-5046 at a dose of 10 mg/kg 5 min before ischemia and immediately after reperfusion plus 1-h ischemia and 2-h reperfusion, n = 6. A laser-Doppler flowmeter and in vivo microscopy were used to investigate hepatic microcirculation. Tissue malondialdehyde (MDA) and adenosine triphosphate (ATP) levels were determined at the end of the experiment. RESULTS: Compared with ischemia alone, ONO-5046 significantly reduced the extent of microcirculatory and hemodynamic derangement after ischemia-reperfusion. ONO-5046 at both doses significantly attenuated decreases in mean arterial pressure. ONO-5046 lessened adherent leukocyte count and improved flow velocity in the sinusoids and postsinusoidal venules. ONO-5046 at the dose of 10m/kg reduced MDA (1.97 +/- 0.54 micromol/g protein vs. 3.58 +/- 1.21 micromol/g protein in the ischemia and reperfusion group) and increased ATP levels (2.62 +/- 0.19 micromol/g wet wt vs. 0.57 +/- 0.37 pmol/g wet wt in the ischemia and reperfusion group), whereas ONO-5046 at a smaller dose (1 mg/kg) had lesser but significant effects on MDA and ATP alterations. This study demonstrates that treatment with ONO-5046, a neutrophil elastase inhibitor, can ameliorate ischemia-reperfusion injury of the rat liver.     

The possible role of endogenous amphiphiles in the membrane abnormalities of ischemic and reperfused myocardium

Calcium entry into cardiac cells is believed to be controlled by transmembrane-voltage dependent, protein regulated "channels." The sarcoplasmic reticulum participates in the regulation of cytosolic calcium by ATP dependent Ca2+ sequestration during diastole, and by action potential stimulated calcium release. Massive calcium overloading occurs during reperfusion following myocardial ischemia. Calcium overloading activates phospholipases, which may activate another mechanism involved in lethal cellular injury, that is, the accumulation of long chain fatty acids and their derivatives. These compounds are soluble amphiphiles, and once liberated, they may insert into biological membranes and change membrane composition, physiology, and response to ions and drugs. Sarcoplasmic reticulum vesicles were used as an in vitro model to study the effects of palmitic acid, oleic acid, and palmitylcarnitine on the ability of this membrane system to sequester calcium within the vesicles. In the absence of phosphate, palmitic acid enhanced the ability of the vesicles to sequester calcium. Oleic acid and palmitylcarnitine inhibited calcium sequestration. In the presence of phosphate palmitic acid also inhibited the sequestration of calcium by sarcoplasmic reticulum, although not as severely as oleic acid and palmitylcarnitine. These results suggest that the disturbances in cellular calcium homeostasis following ischemia may be due, in part, to the incorporation of accumulated long chain fatty acids into membranes.     

茶氨酸对脑缺血损伤大鼠自由基代谢的影响

目的:从对自由基代谢影响方面探讨茶氨酸对脑缺血损伤的保护作用机制。方法:选用30只SD雄性大鼠,线栓阻塞大鼠大脑中动脉制备局灶性脑缺血模型,观察脑缺血损伤大鼠神经症状改变,测定脑组织含水量、脑组织病理及钙离子含量;观察其血清和脑组织超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量变化以及茶氨酸对上述变化的影响。结果:脑缺血组大鼠的神经症状评分、脑组织含水量及钙离子含量均显著增高,病理损伤明显,正常神经元细胞数目减少;茶氨酸可显著降低神经症状评分、脑组织含水量及钙离子含量,增加正常神经元细胞数目。脑缺血组大鼠的血清、脑组织SOD活性显著降低,茶氨酸组大鼠血清、脑组织SOD活性较脑缺血组显著增高;脑缺血组大鼠的血清、脑组织MDA含量显著增高,茶氨酸组脑组织MDA含量较脑缺血组下降明显。结论:茶氨酸对大鼠脑缺血损伤具有显著的保护作用,可能与调节脑缺血损伤引起的自由基代谢失调有关。     

灯盏花素注射液对沙土鼠脑缺血-再灌注后能量代谢及脑水肿的影响

背景脑缺血-再灌注后能量代谢障碍和脑水肿是脑缺血-再灌注损伤重要因素之一.中药灯盏花素注射液(其有效成分为黄芩素甙)可以防止脑缺血-再灌注诱发的蛋白激酶C的激活、减轻钙超载,且可减小缺血梗死灶的体积,从而减轻脑缺血-再灌注损伤.可灯盏花素注射液对脑缺血-再灌注后能量代谢和脑水肿有何影响?目的观察灯盏花素注射液对沙土鼠脑缺血-再灌注后能量代谢和脑水肿的影响.设计随机对照实验.单位济宁医学院附属医院及徐州医学院附属医院江苏省麻醉学重点实验室.材料实验于1999-02/08在江苏省麻醉学重点实验室完成.选用雄性沙土鼠72只.方法将实验动物随机分成3组,即假手术组、常温对照组和灯盏花素组,假手术组8只,常温对照组和灯盏花素组每组各32只.根据再灌注时间将常温对照组和灯盏花素组分4个亚组,即缺血末组、再灌注10 min组、再灌注30 min组和再灌注60 min组.每亚组8只动物.常温对照组和灯盏花素组制备前脑缺血-再灌注损伤模型,脑缺血时间为10 min.假手术组仅游离双侧颈总动脉但不予阻断.灯盏花素组于缺血前15 min给予灯盏花素注射液90 mg/kg腹腔注射,假手术组和常温对照组则给予等量的生理盐水.脑水的测定采用干湿重法,应用高效液相及紫外检测仪测定海马三磷酸腺苷,二磷酸腺苷,磷酸腺苷的含量.主要观察指标①实验动物海马三磷酸腺苷、二磷酸腺苷、磷酸腺苷的含量.②实验动物脑皮质水含量.结果纳入实验的动物为72只,均进入结果分析,无实验动物脱失.①实验动物海马三磷酸腺苷、二磷酸腺苷、磷酸腺苷的含量常温对照组在缺血末、再灌注10 min、再灌注30 min、再灌注60 min时,海马组织三磷酸腺苷和腺苷酸池含量明显降低,三磷酸腺苷含量分别为假手术组的68%,56%,49%和50%(P均＜0.01),腺苷酸池含量为假手术组的62%,50%,51%和52%(P均＜0.01).而灯盏花素组各时间点三磷酸腺苷含量分别为假手术组的84%,69%,64%和63%,腺苷酸池含量为假手术组的86%,72%,68%和69%,均明显高于常温对照组(P均＜0.05).②实验动物脑皮质水含量脑缺血-再灌注后常温对照组脑皮质水含量明显高于假手术组(P＜0.05),并且随再灌注时间的延长逐渐加重.灯盏花素组脑皮质水含量虽明显高于假手术组,但明显低于常温对照组(P＜0.05).结论灯盏花素可能通过抑制能量代谢障碍、减轻脑水肿而发挥脑保护作用.     

老龄大鼠脑缺血再灌注肺损伤机制研究

目的从ATP酶活性变化和自由基损伤方面研究老龄大鼠脑缺血再灌注肺脏损伤机制。方法青年(5月龄)和老龄(20月龄以上)大鼠均分为模型组和对照组,观察大鼠全脑缺血30min再灌注60min后肺脏组织形态和丙二醛(MDA)含量及超氧化物岐化酶(SOD)、ATP酶活性的变化。结果青年和老龄模型组大鼠肺脏组织均出现明显的病理改变,老龄模型组较青年模型组严重。老龄对照组肺组织MDA/SOD比值高于青年对照组组。青年模型组肺Ca2+-ATP酶活性低于青年对照组和老龄模型组。结论脑缺血再灌注肺损伤老龄大鼠较青年大鼠严重,Ca2＋-ATP酶活性的降低和自由基损伤可能是肺损伤发生的机制之一。