脑出血后凝血酶与脑水肿关系及水蛭粉的疗效

目的　探讨凝血酶在脑出血后脑水肿形成机制中的作用以及水蛭粉治疗脑水肿的疗效。方法　分别制作大鼠自体全血、凝血酶、肝素抗凝血、生理盐水及假手术脑血肿模型 ,观察不同时间血肿周围脑组织含水量 ,Na+ 、K+ 含量的变化以及病理改变 ;并观察水蛭粉 (2 5 0mg/kg口服 ,每天 2次 ,治疗 5d)治疗脑水肿的疗效。结果　大鼠脑内注射全血或凝血酶后 ,脑组织的含水量、电解质含量的变化相一致 ;在 2 4h时 ,脑水含量、Na+ 含量较其它 3组高 ,而K+ 含量则较低 (P <0 0 5 )。水蛭粉治疗后脑水含量、Na+ 、Ca2 + 含量较对照组降低 ,而K+ 含量升高 (P <0 0 5 )。结论　凝血酶可能在脑出血后脑水肿形成机制中起重要作用 ;水蛭粉对大鼠全血和凝血酶导致脑水肿有显著影响。     

脑出血后水蛭素干预对水通道蛋白4表达的影响

目的:在脑出血后不同时段应用凝血酶抑制剂水蛭素干预,观察其对脑水肿、水通道蛋白4(aquaporin-4,AQP4)及其mRNA表达的影响。方法:将40只健康雄性大鼠随机分为4组,每组10只,采用自体动脉血注入尾状核法制成大鼠脑出血模型,其中1组作为对照组,造模后不予水蛭素干预,另3组分别于造模后6h、12h、18h在血肿内注入水蛭素进行干预,32h后提取脑组织,应用反转录聚合酶链反应(RT-PCR)检测AQP4mRNA在脑水肿组织的表达,并采用Western blot和免疫组化检测AQP4蛋白在脑水肿组织中的表达,同时用干重、湿重法计算脑组织含水量来表示脑水肿程度。结果:与对照组相比,凝血酶抑制剂水蛭素能减轻脑出血后脑组织的水肿程度,并降低脑组织中AQP4mRNA及AQP4蛋白的表达,在6h和12h干预组中其抑制作用最为明显(P<0.05),且AQP4mRNA及AQP4蛋白的表达减少与脑水肿变化趋势一致。18h干预组的脑水肿程度、AQP4mRNA及AQP4蛋白的表达虽有低于对照组的趋势,但差异无统计学意义。结论:脑出血后,凝血酶抑制剂水蛭素能减少脑组织水肿的程度、降低AQP4mRNA和AQP4蛋白的表达,水蛭素干预组中以6h及12h干预组的作用最为明显。     

凝血酶与神经细胞骨架蛋白的研究

目的：观察凝血酶与神经细胞骨架蛋白的关系及使用凝血酶抑制剂（如水蛭素）及钙离子拮抗剂（尼莫地平）是否能减轻凝血酶对脑细胞的毒性损伤，是否存在治疗的有效时间。方法：分别以不同剂量的凝血酶注入SD大鼠尾状核，建立SD大鼠模，采用免疫组化法观察神经细胞骨架蛋白的改变。SD大鼠随机分为6组。1组：生理盐水组；2组：小剂量凝血酶；3组：大剂量凝血酶组；4组：小剂量凝血酶与小剂量水蛭素同时注入组；5组：大剂量凝血酶与大剂量水蛭素同时注入组；6组：尼莫地平组。每组每时相（4h,24h,48h,3d,7d）和5只SD大鼠。结果：大剂量凝血酶的早期（4h）即能引起细胞骨架蛋白的变化，24h-48h，可进一步导致可逆或不可逆损伤，3至7d则主要造成神经细胞不可逆损伤即坏死，而生理盐水，小剂量凝血酶无此作用。水蛭素可拮抗凝血酶的这种作用，尼莫地平组4h神经骨架蛋白无明显变化，24h-48h可导致神经细胞可逆损伤，3至7d则主要造成神经细胞不可逆损伤，结论：钙离子拮抗剂（尼莫地平）能减轻或延缓因酶对脑细胞的毒性作用，有保护神经元的作用。为进一步治疗嬴得时间，由以上结果可推测早期脑水肿是由于凝血酶对脑细胞的毒性作用所致，这种作用（可能通过凝血酶受体）可引起神经细胞突起的改变而造成细胞损伤。     

尼莫地平治疗急性高血压性脑出血的疗效初步探讨

目的:探讨尼莫地平治疗急性高血压性脑出血的临床疗效。方法:64例急性高血压性脑出血患者被随机分成尼莫地平组30例和对照组34例,2组均接受传统脑出血治疗,尼莫地平组同时静脉滴注尼莫地平,于治疗前及治疗后第7、14天测量脑血肿和脑水肿带体积,于入院时、治疗后第30、90天随访时采用欧州卒中评分量表(ESS)评定神经功能。结果:尼莫地平组较对照组脑血肿、脑水肿体积明显减小,ESS评分明显增高。结论:在高血压性脑出血早期可在传统脑出血治疗的同时使用尼莫地平,以进一步减轻脑水肿、促进脑血肿吸收、改善神经功能。     

高血压性脑出血微创术后加用尼莫地平口服疗效观察

目的　探讨尼莫地平对脑出血微创颅内血肿清除术后的治疗效果。方法　对 2 5例脑基底节出血的患者微创术后口服尼莫地平治疗 ,观察治疗前后神经功能、头颅CT血肿大小及周围水肿的变化 ,并与对照组进行比较。结果　尼莫地平治疗组神经功能恢复、血肿吸收及周围水肿减少较对照组显著 (P <0 .0 5、0 .0 5、0 .0 1)。结论　尼莫地平能促进神经功能的恢复 ,促进血肿吸收 ,抑制脑水肿形成     

凝血酶对脑微血管内皮细胞的损害及水蛭素的保护作用

目的：探讨凝血酶对大鼠脑微血管内皮细胞（BMVEC）的损伤以及凝血酶抑制剂水蛭素的保护作用。方法：分离新生大鼠脑微血管内皮细胞并进行培养,取第二或第三代内皮细胞进行实验。实验分成3组：对照组、凝血酶组和水蛭素组。通过透射电镜和流式细胞术检测3组细胞凋亡的情况。利用TRITC-鬼笔环肽对细胞进行染色观察3组细胞骨架的形态变化。结果：凝血酶可以增加血管内皮细胞凋亡并导致细胞骨架重排、紊乱,而水蛭素可以减轻凝血酶的这种作用。结论：凝血酶可以损害血管内皮细胞骨架,引起细胞收缩、变形,同时增加内皮细胞凋亡,破坏血脑屏障。水蛭素可以减轻这种改变。     

水蛭素对大鼠DIC模型脑组织水通道蛋白4表达的干预作用

目的 探讨水蛭素对DIC模型的大鼠脑组织水通道蛋白4(AQP4)表达的干预作用.方法 通过鼠尾静脉滴注脂多糖制作DIC模型,将大鼠分为三组即对照组、DIC组和水蛭素组,选取4个时间点(4,6,8,12 h)测定血小板计数,PT,APTT,纤维蛋白原和D-D水平的变化;同时用免疫印迹杂交法检测大鼠脑组织水通道蛋白4的表达.结果 与对照组相比,DIC组及水蛭素组血小板计数,PT,APTT,纤维蛋白原和D-D差异具有统计学意义(P＜0.05);水蛭素组上述指标变化明显低于DIC组(P＜0.05);AQP4表达在水蛭素组明显低于DIC组(P＜0.05).结论 DIC时产生的凝血酶可能通过上调AQP4蛋白的表达,参与脑水肿形成和发展;水蛭素通过抑制凝血酶的形成,减轻脑水肿.     

脑出血后脑水肿的治疗

按病理形态和发病机制脑水肿分细胞毒性、血管源性和间质性三种 ,脑出血后脑水肿一般以前二者为主。脑出血后血肿压迫引起周围脑组织缺血以及血液循环障碍、酸中毒和和代谢紊乱 ,血浆和细胞中的血管活性物质引起血管痉挛、血管扩张或通透性改变 ,红细胞毒性效应和血红蛋白裂解产物导致的水肿形成与血脑屏障破坏促进了出血区脑水含量的增加[1] ,血浆中的白蛋白、膜性成分及胞内大分子物质经组织间隙渗透入脑组织 ,加重脑水肿 ,促进颅内压增高 ,甚至产生脑疝。因此 ,积极治疗脑水肿是改善脑出血预后的关键。现就脑出血后脑水肿的治疗综述如下…     

脑出血后脑内血肿成分的毒性作用

脑出血是常见的脑血管意外之一 ,而脑内血肿作为脑出血后的直接产物 ,是影响患者生存率和生存质量的重要因素。既往认为 ,出血后凝血块的占位效应产生的机械性压迫是造成继发性脑损害的主要原因。自从 1980年Ｓｕｚｕｋｉ和Ｅｂｉｎａ提出血凝块的毒性作用以来 ,血肿成分的毒性作用逐渐被人们所认识并日益受到重视。认为其对神经细胞的毒性作用和对血脑屏障的破坏可能是脑出血后脑水肿形成的重要因素之一。本文就脑出血后脑内血肿成分的主要毒性物质及其作用机制作一综述。1　凝血酶1.1　凝血酶的基本特性　凝血酶是一种血清丝氨酸蛋白酶 ,…     

凝血酶致大鼠脑水肿与脑内MMP-9、MMP-2蛋白表达关系的实验研究

目的探讨凝血酶对大鼠脑内MMP-9、MMP-2蛋白表达的影响。方法 Wistar大鼠随机分为假手术组、对照组及凝血酶实验组。实验组脑内注入凝血酶,在不同时间点采用干湿重法测脑水含量,Western blog方法检测脑内MMP-9、MMP-2蛋白的表达。对照组注入等量生理盐水。结果脑组织水含量在凝血酶注入后12 h开始增加,3 d达高峰。MMP-9在注射凝血酶后12 h开始明显表达增加,与对照组相比差异显著(P0.01),3 d达高峰,随后持续下降。MMP-2在注射凝血酶后24h才开始有明显表达,后持续增多,5 d达高峰,随后有所下降,与对照组相比差异显著(P0.05)。结论凝血酶在脑出血后脑水肿及脑组织损伤中起了关键的作用,MMP-9、MMP-2参与了急性期脑水肿过程。     

Thrombin functions as an inflammatory mediator through activation of its receptor

A rat model of inflammation was used to investigate the biological effects of thrombin. The thrombin-specific inhibitor Hirulog markedly attentuated the carrageenin-induced edema of the paw of the rat. Injection of thrombin into the paw also produced edema. The effect of thrombin was due to activation of its receptor; a thrombin receptor activating peptide (TRAP) reproduced the effects of thrombin in causing edema. TRAP also increased vascular permeability as demonstrated by extravasation of Evans blue and 125I-labeled serum albumin. The release of bioactive amines played an important role in mediating the TRAP- induced edema; the serotonin/histamine antagonist cryproheptadine and the histamine H2 receptor antagonist cimetidine reduced significantly the edema caused by TRAP. Treatment of rats with the mast cell degranulator 48/80 to deplete these cells of their stores of histamine and serotonin abolished completely the ability of TRAP to produce edema. Histochemical examination confirmed that TRAP treatment led to mast cell degranulation. Thus, it has been possible to demonstrate that thrombin acts as an inflammatory mediator in vivo by activating its receptor, which in turn leads to release of vasoactive amines from mast cells.     

Protease-activated receptor 1 mediates thrombin-dependent, cell-mediated renal inflammation in crescentic glomerulonephritis

Protease-activated receptor (PAR)-1 is a cellular receptor for thrombin that is activated after proteolytic cleavage. The contribution of PAR-1 to inflammatory cell-mediated renal injury was assessed in murine crescentic glomerulonephritis (GN). A pivotal role for thrombin in this model was demonstrated by the capacity of hirudin, a selective thrombin antagonist, to attenuate renal injury. Compared with control treatment, hirudin significantly reduced glomerular crescent formation, T cell and macrophage infiltration, fibrin deposition, and elevated serum creatinine, which are prominent features of GN. PAR-1-deficient (PAR-1(-/-)) mice, which have normal coagulation, also showed significant protection from crescentic GN compared with wild-type mice. The reductions in crescent formation, inflammatory cell infiltration, and serum creatinine were similar in PAR-1(-/-) and hirudin-treated mice, but hirudin afforded significantly greater protection from fibrin deposition. Treatment of wild-type mice with a selective PAR-1-activating peptide (TRAP) augmented histological and functional indices of GN, but TRAP treatment did not alter the severity of GN in PAR(-/-) mice. These results indicate that activation of PAR-1 by thrombin or TRAP amplifies crescentic GN. Thus, in addition to its procoagulant role, thrombin has proinflammatory, PAR-1-dependent effects that augment inflammatory renal injury.     

Platelet thromboembolism

Abstract. Platelet-dependent thrombosis and subsequent embolization are major causes of cerebral ischaemia. Beside aspirin which irreversibly blocks platelet cyclo-oxygenase, several other substances interfere in different platelet metabolic pathways and block platelet adhesion and aggregation. We found in an experimental model using non-human primates that a specific peptide inhibitor blocking GP IIb/IIIa platelet receptor which binds fibrinogen completely, prevents the retention of embolized platelet aggregates in the cerebral circulation. As thrombin may play a key role for platelet activation in vivo leech-derived hirudin, a direct thrombin inhibitor as well as activated protein C which limits thrombin production and also prevents platelet dependent thrombus formation very effective. We demonstrated in the same non-human primate model of platelet embolization that the amount of retention of platelet emboli in the vascular bed depends on the nature of the vasculature. For example, platelet emboli were cleared very quickly from brain microcirculation, whereas platelet embolization into the lower limb via the femoral artery caused a significantly longer retention of the embolized material. Such specific mechanisms may be caused by different levels of local vasodilators as PGI₂ or EDRF.     

Effect of ifenprodil, a polyamine site NMDA receptor antagonist, on brain edema formation following asphyxial cardiac arrest in rats

OBJECTIVE: Brain edema occurs in experimental and clinical cardiac arrest (CA) and is predictive of a poor neurological outcome. N-Methyl--aspartate (NMDA) receptors contribute to brain edema elicited by focal cerebral ischemia/reperfusion (I/R). Ifenprodil, a NMDA receptor antagonist, attenuates brain edema and injury size in rats after focal cerebral I/R. We assessed the hypothesis that ifenprodil reduces CA-elicited brain edema. METHODS: Eighteen male Sprague-Dawley rats were assigned to group 1 (normal control, n=6), group 2 (placebo-treated CA, n=6), or group 3 (ifenprodil-treated CA, n=6). CA was induced by 8 min of asphyxiation and the animals were resuscitated with cardiopulmonary resuscitation (CPR), ventilation, epinephrine (adrenaline), and sodium bicarbonate (NaHCO3). Ifenprodil of 10 mg/kg or a placebo vehicle was given intraperitoneally 5 min before CA. Brain edema was determined by brain wet-to-dry weight ratio at 1 h after resuscitation. RESULTS: There were no differences between groups 2 and 3 in all physiological variables at baseline. Time from asphyxiation to CA was 201.5 +/- 7.5 s in group 2 and 160.7 +/- 10.4 s in group 3 (P<0.001). Resuscitation time was 68.2 +/- 13.3 s in group 2 and 92.8 +/- 18.2 s in group 3 (P<0.05). Ifenprodil decreased mean arterial pressure (MAP) before asphyxiation, from 128 +/- 7 in group 2 to 82 +/- 15 mmHg in group 3 (P<0.001), and negated immediate post-resuscitation hypertension. Brain wet-to-dry weight ratio was 5.64 +/- 0.44 in group 1, 7.34 +/- 0.95 in group 2 (P<0.01 versus group 1), and 5.93 +/- 0.40 in group 3 (P<0.05 versus group 2). CONCLUSIONS: Ifenprodil reduces CA-elicited brain edema. In addition, we observed significant hemodynamic changes caused by ifenprodil.     

Morphine-induced analgesia in rats withdrawn from concurrent nimodipine and morphine treatment

The effect of repeated administration of dihydropyridine calcium channel antagonist, nimodipine, given concurrently with morphine on the development of tolerance to the antinociceptive actions of morphine in rats was studied. In acute experiments nimodipine (1 mg/kg i.p.) enhanced the antinociceptive effect of morphine (2.5 mg/kg s.c.) in the hot-plate and tail immersion tests. Daily administration of morphine either for 10 days (increasing the daily dose from 20 to 35 mg/kg) or for 24 days (increasing the daily dose from 20 to 70 mg/kg) induced tolerance to the antinociceptive effect of a challenge dose of morphine (10 mg/kg) administered 24 h after the withdrawal from chronic morphine. Concurrent administration of nimodipine (1 mg/kg per day) with morphine for 10 or 24 days augmented the reduction of the antinociceptive effect of morphine. Neither acute nor repeated administration of nimodipine with morphine altered the concentrations of morphine or its metabolite morphine 6-glucuronide in the brain tissue or in the plasma. The observed further reduction in the nociceptive response in morphine tolerant animals pre-treated with nimodipine is, most probably, due to the adaptive changes in the central dihydropyridine calcium channels induced by the withdrawal from repeated nimodipine treatment.     

Effects of inhibitors of arachidonic acid metabolism and calcium entry on responses to acetylcholine, potassium and norepinephrine in the isolated canine saphenous vein

The present study was designed to determine whether interference with the metabolism of arachidonic acid or the entry of extracellular calcium affects the responses of the canine saphenous vein to acetylcholine, potassium or norepinephrine. Rings of canine saphenous vein, with or without endothelium, were suspended in organ chambers filled with physiological salt solution and set at their optimal length for isometric tension recording. Removal of the endothelium, confirmed by the absence of the characteristic relaxation induced by thrombin in intact rings, did not affect concentration-response curves to acetylcholine or norepinephrine. The cyclooxygenase inhibitor, indomethacin, augmented the response to acetylcholine. This effect was comparable in rings with and without endothelium and in rings pretreated with phentolamine. Indomethacin did not alter the response to norepinephrine but augmented that to potassium. Similar results were obtained with the cyclooxygenase inhibitors, acetylsalicylic acid and meclofenamate. The antioxidant and lipoxygenase inhibitor nordihydroguaiaretic acid and the cyclooxygenase/lipoxygenase inhibitor phenidone blocked the ability of indomethacin to augment acetylcholine- and potassium-induced contractions. Arachidonic acid-induced contractions were not blocked by indomethacin but were inhibited by nordihydroguaiaretic acid, phenidone and the calcium entry blockers diltiazem and nimodipine. Diltiazem and nimodipine inhibited responses to potassium and acetylcholine without affecting those to norepinephrine. The augmentation by indomethacin of potassium- and acetylcholine-evoked contractions was inhibited by diltiazem and nimodipine. In rings of femoral artery denuded of endothelium, indomethacin had no effect on the responses to acetylcholine, norepinephrine or potassium. These results suggest that, in the canine saphenous vein but not in the femoral artery, activation of the lipoxygenase pathway for the metabolism of arachidonic acid augments preferentially contractions which depend upon the entry of extracellular calcium.     

银杏叶制剂对大鼠蛛网膜下腔出血后脑水肿的防治研究

目的探讨银杏叶制剂对蛛网膜下腔出血后脑水肿的防治作用。方法应用非开颅性大鼠模型,检测蛛网膜下腔出血前及出血后１ｈ、６ｈ及２４ｈ时脑组织含水率和电解质含量的变化,观察银杏叶制剂和尼莫地平对其影响。结果蛛网膜下腔出血后６ｈ、２４ｈ脑组织含水率逐渐增加,Ｎａ含量增多,Ｋ含量减少,应用银杏叶制剂和尼莫地平均可使上述病理变化程度减轻。结论银杏叶制剂可有效防治大鼠蛛网膜下腔出血后脑水肿。     

水蛭素融合蛋白研究进展

水蛭素是一种高效特异的凝血酶抑制剂，临床上用于血栓病的预防和治疗，但它半衰期短、出血副作用强和功能单一。近年来，通过把水蛭素与某些功能蛋白融合表达来解决这些问题，得到的融合蛋白或在体内延长水蛭素的半衰期，或降低其出血副作用，或带来新的功能，如：溶栓、抑制血小板凝集、特异寻靶等。本文对近年来国内外水蛭素融合蛋白研究状况进行综述。