内皮素双受体拮抗剂在实验性急性肺栓塞中的应用研究

目的观察静脉内皮素双受体拮抗剂RO2610612在急性肺栓塞中的应用效果,研究内皮素与急性肺栓塞的关系。方法12只健康杂种犬随机分为对照组和实验组,每组6只。两组所有动物给予戊巴比妥静脉麻醉后分别于颈静脉插入热稀释漂浮导管连接生理监护仪;气管内插入气管插管连接呼吸描记器。采集栓塞前数据后于颈静脉注射自体血栓建立急性肺栓塞动物模型。实验组建立模型后1h开始持续静滴RO26106120.2mg/(kg.h),对照组同时输入等量生理盐水。对照组及实验组所有动物于栓塞前、栓塞后1,2,4,6h记录体、肺循环血流动力学指标、呼吸生理指标;取股动脉血3ml,采用放射免疫法测定动脉血内皮素-1水平,并测动脉血气。结果两组动物栓塞后较栓塞前血浆内皮素-1水平、肺动脉平均压、肺血管阻力、呼吸频率、动脉血二氧化碳分压和肺气道阻力显著性升高(P<0.05),心输出量、动脉血氧分压和肺动态顺应性显著性下降(P<0.05)。实验组于栓塞2h后肺动脉平均压、肺血管阻力、呼吸频率、肺气道阻力升高水平显著性低于同时期对照组(P<0.05),血浆内皮素-1水平、心输出量、和肺顺应性显著性高于对照组(P<0.05)。结论在急性肺栓塞病理过程中,内皮素参与了肺循环阻力升高与肺气道阻力升高和肺顺应性下降的形成。内皮素受体拮抗剂拮抗内皮素与其受体的结合,在急性肺栓塞中,可以减缓肺循环阻力、肺气道阻力的上升,因而可能成为急性肺栓塞临床治疗的新手段。     

The endothelin antagonist bosentan does not improve survival in severe experimental pancreatitis in rats

Summary Background. Severity of pancreatitis seems to be aggravated by impairment of vascular perfusion of the gland. Early mortality occurs within the first few days from the acute consequences of pancreatic injury with subsequent inflammatory response. Because vasoactive substances, including endothelin, seem to contribute to early mortality in acute pancreatitis, we tested the hypothesis that the inhibition of endothelin action could alter the outcome after severe experimental pancreatitis. Methods. In two groups of rats, pancreatitis was induced by intraductal infusion into the pancreatic duct of 1 μL/g body weight (b.w.) of either a 4% or a 5% sodium taurocholate solution. The mixed endothelin A and endothelin B receptor antagonist bosentan (20 mg/kg b.w.) or vehicle was injected intravenously in 12-h intervals for 3 d starting 1 h after induction of bile acid pancreatitis. This dose of bosentan is known to completely inhibit the effect of exogenous endothelin. The survival rate was monitored for 7 d. Thereafter, the surviving rats were sacrificed and the pancreas was prepared for histological and biochemical evaluation. Results. Irrespective of the treatment protocol (bosentan versus saline), survival was not different in animals challenged with either 4% or 5% sodium taurocholate. The corresponding survival rates were 62% with bosentan and 77% without bosentan in the 4% sodium taurocholate group. In the 5% sodium taurocholate group, the survival rates were 20% with and 27% without bosentan. Morphological and biochemical alterations were identical in control as well as in endothelin-antagonist-treated rats. Conclusion. Therapy with the mixed endothelin A and endothelin B receptor antagonist bosentan does not influence the outcome after severe experimental pancreatitis. Therefore, blockade of endothelin A and B receptor subtypes may not be of major importance as a therapeutic principle in this model of experimental pancreatitis.     

ACE-inhibition is superior to endothelin A receptor blockade in preventing abnormal capillary supply and fibrosis of the heart in experimental diabetes

Aims/hypothesis There is little information whether cardiac capillary supply is deranged in diabetes. Hyperglycaemia is a potent stimulus for endothelin-1 (ET-1) production. We therefore hypothesised that increased ET-1 production in Streptozotocin-induced Type 1 diabetes causes abnormalities of cardiac capillaries and the aorta. To this end we compared the effects of an ET receptor A blocker (ET_A-RB) with that of an ACE-inhibitor (ACE-i) or their combination in rats with Streptozotocin (STZ) diabetes.Methods Sprague Dawley rats were injected with 65 mg STZ i.v. and subsequently developed diabetes. Rats were left untreated or received daily either the ACE-i Trandolapril, the ET_A-RB Darusentan or a combination of both. After 6 months the experiment was terminated and the heart and the aorta were investigated using quantitative morphological techniques.Results ACE-i but not ET_A-RB lowered blood pressure in STZ Type 1 diabetic rats. Capillary length density was lower in untreated STZ diabetic rats (2932±128 mm/mm³) compared to non-diabetic control rats (3410±252 mm/mm³). Treatment with ACE-i (3568±431 mm/mm³), but not with ET_A-RB (2893±192 mm/mm³), prevented the decrease in capillary supply. Volume density of the myocardial interstitium was higher in untreated STZ diabetic rats (0.86±0.04%) compared to non-diabetic control rats (0.36±0.06%). In all three intervention groups the values were lower (ACE-i: 0.53±0.05%, ET_A-RB: 0.7±0.08% and combination: 0.69±0.1).Conclusion/interpretation Our study identifies a capillary defect of the heart in STZ diabetes, i.e. decreased capillary supply. This abnormality was reversed by ACE-i, but not by ET_A-R blockade. A similar trend, although not complete normalisation, was seen in cardiac fibrosis.Abbreviations ET-1 Endothelin1 - STZ streptozotocin - ACE-i ACE-inhibitor - ET_A-RB ET receptor A blocker - AT1 Ang II receptor type 1 blockers - SD Sprague Dawley     

Is there a role for serum endothelin in predicting the severity of acute pancreatitis?

BACKGROUND: Acute pancreatitis remains a common presentation to acute surgical units and carries significant morbidity and mortality. The progression of the disease to necrotizing pancreatitis and multi-organ dysfunction syndrome (MODS) is associated with a very poor clinical outcome, and persistendy high mortality. Increases in serum endothelin (ET) have been seen in animal models of acute pancreatitis and this study aims to investigate whether there is a change in serum ET-1 in patients with acute pancreatitis and whether any such change is linked to disease severity. METHODS: All patients admitted with acute pancreatitis were prospectively recruited from die emergency admissions at the Norfolk and Norwich University Hospital. Serum ET levels were determined on admission, at 24 hours and 5 days post admission. Healthy adult controls were recruited from dermatology outpatients. RESULTS: A total of 21 patients joined the trial after giving informed consent. There were 3 men and 18 women with a median age of 65 years (range 26-87 years). Serum ET levels were significantly higher in acute pancreatitis patients than in normal controls (P <0. 05). An association was seen between persistendy raised serum ET levels and progression to MODS. CONCLUSIONS: The study does demonstrate a correlation between the circulating levels of ET and acute pancreatitis in humans, although it does not elicit its involvement in the pathogenesis of the disease. The observation that a persistendy high level of circulating ET-1 is associated with progression to MODS may indicate a role for ET in the monitoring of acute pancreatitis patients for recovery or progression to MODS.     

Optimal route of administration of mixed endothelin receptor antagonist (TAK-044) in liver transplantation

It is well known that endothelin-1(ET-1) is a factor involved in the pathogenesis of ischemia-reperfusion injury. This study was undertaken to investigate the optimal route (intravenous vs intraportal) for administering mixed endothelin receptor antagonist (TAK-044) in a liver transplantation. First, in a rat isolated liver cold-perfusion model, the pharmacodynamics of TAK-044 and endothelin-1 (ET) in the liver tissue and the systemic circulation after cold perfusion were compared in the different administration routes. Next, in a rat orthotopic transplantation model, we compared the hepatoprotective effect of TAK-044 among different administration routes. In each model, there were three groups: IV group, intravenous injection of TAK-044 (10 mg/kg) immediately before cold perfusion or an-hepatic phase; IP group, intraportal administration with cold perfusion solution or with reflush solution for the graft; control group, no treatment. In the cold perfusion model, liver tissue ET level increased to a similar extent after reperfusion in the three groups, and the plasma and liver tissue TAK-044 concentrations after reperfusion were highest in the IV group. However, the increase in plasma ET was also greatest, and therefore, the ratio of liver tissue to plasma TAK-044 was lower in the IV group compared with the IP group. In the transplantation model, elevation of plasma ET was significantly higher in the IV group. Leakage of serum alanine aminotransferase (ALT), sinusoidal narrowing, and cell swelling after grafting were significantly suppressed in the IP group. We conclude that intraportal administration before reperfusion offers more efficient accumulation of TAK-044 in the liver tissue, without harmful systemic elevation of ET, and achieves a hepatoprotective effect on the graft compared with intravenous administration. Received: April 12, 1999 / Accepted: July 23, 1999     

Accelerated reversal of carbon tetrachloride-induced cirrhosis in rats by the endothelin receptor antagonist TAK-044

BACKGROUND: A multifunctional mediator, endothelin (ET)-1 is implicated in the pathophysiology of liver cirrhosis. Carbon tetrachloride (CCl4)-induced cirrhosis in rats resolves upon termination of CCl4 treatment. We determined the hepatic ET-1 system during such reversal and assessed whether ET-1 receptor antagonism enhances this process. METHODS: Cirrhosis was induced in rats by CCl4 treatment for 8 weeks. Treatment with an ETA/ETB antagonist TAK-044 (10 mg/kg per day) was then started and determinations were made at 1, 2 and 4 weeks. RESULTS: After termination of CCl4 treatment, accelerated normalization of liver architecture and portal hypertension occurred in TAK-044-treated rats compared with saline-treated rats. The increased hepatic hydroxyproline concentration and collagen I mRNA expression also declined to greater extents in the TAK-044-treated group. Higher collagenase activity in cirrhosis decreased in saline-treated rats, but did not reach basal values. In TAK-044-treated rats, collagenase activity tended to increase at weeks 2 and 4. Increased ET-1 concentration and ETA receptor density declined to normal values in both groups. In contrast, increased ETB receptor density did not change in saline-treated rats, but decreased to control values in TAK-044-treated rats. CONCLUSIONS: Our results emphasize the role of ET-1 in chronic liver disease and strongly indicate the potential for ET-1 receptor antagonists in its treatment.     

选择性内皮素受体拮抗剂对大鼠心肌纤维化模型细胞因子表达的影响

目的 检测心肌纤维化调控因子的表达水平差异,探讨选择性内皮素受体拮抗剂对心肌纤维化细胞因子表达的影响.方法 选择健康成年雄性Wistar大鼠[（250±50）g]24只,随机分为3组,每组8只.A组（模型组）背部皮下注射异丙肾上腺素＋胃灌生理盐水;B组（安立生坦组）背部皮下注射异丙肾上腺素＋胃灌安立生坦;C组（对照组）注射生理盐水＋胃灌生理盐水.A组及B组大鼠背部皮下注射异丙肾上腺素5 mg/d,C组背部皮下注射等体积生理盐水,连续8d.第9天开始,B组给予安立生坦40 μg/d胃灌,1次/d,连续4周;C组及A组给予等体积生理盐水胃灌,1次/d,连续4周.4周后超声心动图评估心功能,观察心肌超微结构,明确各组模型细胞因子的表达.结果 M型超声心动图提示,模型组与对照组相比LVIDd、LVIDs显著增加,FS和EF显著降低（P＜0.05）.对照组较安立生坦组LVIDd、LVIDs有所增加（P＜0.05）,FS和EF略降低,但差异无统计学意义（P＞0.05）.B组和A组的的细胞因子表达有显著性差异,B组和C组的AngⅡ、TGF-β1、CTGF、ET-1细胞因子表达均明显小于A组,但差异无统计学意义;B组和C组HGF明显高于A组.病理结果提示,B组心肌细胞肿胀、断裂及胶原纤维增生程度明显低于A组.结论 选择性内皮素受体拮抗剂能够通过影响血管生成因子的表达,改变心肌纤维化进程.     

Effect of microcirculatory perfusion on distribution of trypsinogen activation peptides in acute experimental pancreatitis

Extraintestinal trypsinogen activation peptides (TAP) have been shown to correlate with severity of acute pancreatitis in humans as well as in various animal models. Ischemia superimposed on experimental pancreatitis, however, increases acinar cell injury without increasing TAP in plasma. We speculated that TAP generated in the pancreas might not reach the circulation in necrotizing pancreatitis due to decreased pancreatic perfusion. To test the hypothesis that generation of TAP in plasma is related to pancreatic perfusion and that plasma TAP may therefore underestimate acinar cell injury in necrotizing disease, we correlated TAP in pancreatic tissue and body fluids with capillary pancreatic blood flow in necrotizing and edematous pancreatitis. The ratio between necrosis and TAP in tissue was similar in both models; the ratio between TAP in plasma and tissue, however, was significantly lower in necrotizing pancreatitis, indicating that a certain amount of TAP generated in the pancreas did not reach the circulation. Decreased pancreatic perfusion found in necrotizing pancreatitis was consistent with this finding. Our data suggest that TAP in tissue is most reliable to indicate severity of acute pancreatitis, whereas plasma TAP may underestimate pancreatic injury in necrotizing disease due to decreased pancreatic perfusion.     

Bosentan and the endothelin system in congestive heart failure

The endothelin system appears to play an important role in the pathophysiology of congestive heart failure (CHF). Endothelin receptor antagonists represent a novel class of agents that are being evaluated for their potential benefits in treating various cardiovascular disorders. Bosentan is an orally active endothelin receptor antagonist that has been studied for the treatment of CHF. Early clinical experience with bosentan has confirmed some benefits on hemodynamic parameters in patients with CHF. Its role in slowing the progression of the disease and improving survival remains to be elucidated.     

Influence of the CCK-antagonist loxiglumide on bile-induced experimental pancreatitis

The present study investigates the effect of CCK-receptor blockade on taurocholate-induced pancreatitis in rats using the potent antagonist loxiglumide. Intraperitoneal administration (50 mg/kg) of loxiglumide began 3 h before, or 10 min or 3 h after induction of pancreatitis. Mean survival times of the experimental groups were 31.2, 23.6, and 20.5 h, respectively, compared to 18.2 h for controls. Survival for 24 h after induction of pancreatitis was significantly improved when the antagonist was given 3 h before, but not in the time periods after induction. After 72 h, survival time was not significantly altered in any of the groups. Furthermore, amylase and lipase levels quantified 10 h after induction of pancreatitis in ascites, blood, or tissue did not indicate a significant difference, nor was improvement in survival seen when the CCK-antagonist was tested in rats receiving a basal treatment with intravenous volume substitution, peritoneal lavage, and protease inhibition. We conclude that CCK-receptor blockade does not improve the final outcome of bile-induced pancreatitis in the rat, even if treatment is started before induction of pancreatitis.     

ETA受体拮抗剂BQ123对心肌梗死局部心肌组织中ET-1和ET受体分布的影响

目的探讨在急性心肌梗死(AMI)时,使用内皮素受体A拮抗剂(ETA受体拮抗剂)后对局部心肌组织ET-1和ET受体分布的影响.方法将22只兔随机分为4组,1周实验组(n=6)和1周对照组(n=5),4周实验组(n=6)和4周对照组(n=5),结扎左冠状动脉前降支建立AMI模型.实验组每天静脉滴注ETA受体拮抗剂BQ123,对照组每天使用等量生理盐水作相同处理,分别于1周和4周后取出心脏.采用免疫组化染色和图像分析法检测1周和4周各组动物左心室非梗死区、梗死边缘区和梗死中心区心肌组织的中ET-1和ET受体的分布.结果免疫组化染色和图像分析结果显示:各实验组ET-1和ET受体的分布均明显少于各自对照组相应部位(P＜0.05).结论AMI后,使用ETA受体拮抗剂能减轻局部心肌组织ET-1和ET受体的上调程度.     

内皮素受体拮抗剂治疗糖尿病肾病的研究进展

内皮素参与糖尿病肾病的发生和发展.其受体拮抗剂能增加肾脏血流、抑制肾小球系膜增生,减少系膜外基质堆积、缓解肾脏的炎性反应,可延缓肾脏纤维化,改善糖尿病肾病预后.     

内皮素受体拮抗剂GF-063和BQ-485对低氧培养大鼠肺动脉平滑肌细胞增殖的影响

目的:探讨新型内皮素受体拮抗剂GF-063和BQ-485对低氧培养的大鼠肺动脉平滑肌细胞(PASMCs)增殖的影响。方法:贴壁原代培养PASMCs。实验分为4组:常氧组(氧浓度210ml/L)、低氧组(氧浓度20ml/L)、低氧+内皮素A(ETA)受体拮抗剂BQ-485组(BQ-485的终浓度分别为1×10-6、1×10-7、1×10-8和1×10-9mo1/L)和低氧+GF-063组(GF-063的终浓度分别为1×10-6、1×10-7、1×10-8和1×10-9mo1/L)。上述4组细胞均分别培养24、48和72h。采用MTT比色法(波长492nm)检测BQ-485和GF-063对PASMCs增殖的影响(A值)。用流式细胞仪测定细胞周期;放射免疫检查法测定细胞培养上清液中内皮素-1(ET-1)的含量。结果:培养24h时,各实验组的A值差异不明显;培养48h时,低氧组的A值明显增加(P0.01),低氧+BQ-485组和低氧+GF-063组在二者的终浓度为1×10-8、1×10-7、1×10-6mo1/L时A值下降,与低氧组比较具有统计学意义(P0.01);在培养72h时,低氧组的A值仍高于常氧组,但较培养48h显著下降(P0.05)。培养48h时,低氧组G2、S期细胞的比率及DNA合成增加,与常氧组比较有统计学意义(分别为P0.01,P0.05);与低氧组比较,低氧+BQ-485组和低氧+GF-063组G2、S期细胞的比率下降(P0.05),G1期细胞增多(P0.05),DNA合成减少。在培养48h时,与常氧组比,低氧组ET-1的水平增高(P0.01),给予BQ-485、GF-063后ET-1的水平降低,BQ-485为1×10-7mol/L、GF-063为1×10-9mol/L时,可明显降低ET-1的水平(P0.01)。结论:GF-063和BQ-485作为两种新型的内皮素受体拮抗都剂能抑制低氧培养的PASMCs增殖,减少ET-1的生成,且GF-063的其作用较BQ-485强。     

Endogenous endothelin in a rat model of acute colonic mucosal injury

BACKGROUND: Endothelin (ET) is involved in various biologic activities in non-vascular and vascular tissues. While ET has some significant effects on gastrointestinal functions, the possible role of endogenous ET in the host response to mucosal injury has not been well clarified. METHODS: The present study describes an investigation of the effects of an endothelin A receptor antagonist, BQ-123, on lactate dehydrogenase (LDH), mucus and albumin flux into the perfusate in a rat model of acute colonic injury, induced by acetic acid perfusion. The present study also examined localization of ET in damaged rat colons by using immunohistochemistry. RESULTS: A 4% acetic acid treatment induced mild mucosal damage of perfused rat colon and increased LDH as well as albumin and protein-bound hexose release into the perfusate. Pretreatment with BQ-123 significantly reduced LDH activity and protein-bound hexose concentration in the perfusate and delayed the reduction of albumin leakage from damaged mucosa. Vascular endothelial, neural and surface epithelial cells of the colon showed strong ET-like immunoreactivity. Mucosal damage markedly influenced ET expression by epithelial cells. Mild mucosal damage decreased the ET expression by surface epithelial cells while moderate mucosal damage induced a mosaic location of ET-positive epithelial cells in the crypt. Severe mucosal damage abolished the ET expression by epithelial cells. CONCLUSIONS: Endothelin may play a role in the host response to acute mucosal damage. Mucosal ET production is significantly affected by mucosal injury.     

Therapy for acute pancreatitis with platelet-activating factor receptor antagonists

Acute pancreatitis （AP） causes release of platelet-activating factor （PAF）, which induces systemic effects that contribute to circulatory disturbances and multiple organ failure. PAF is a cell surface secretion of bioactive lipid, which could produce physiological and pathological effects by binding to its cell surface receptor called platelet-activating factor receptor （PAF-R）. Studies showed that PAF participates in the occurrence and development of AP and administration of platelet-activating factor receptor antagonists （PAF-RAs） could significantly reduce local and systemic events after AP. PAF has also been implicated as a key mediator in the progression of severe AP, which can lead to complications and unacceptably high mortality rates. Several classes of PAF-RA show PAF- RAs significant local and systemic effects on reducing inflammatory changes. As a preventive treatment, PAF-RA could block a series of PAF-mediated inflammatory injury and thus improve the prognosis of AP. This review introduces the important role of PAF-RA in the treatment of AP.     

TLR4信号传导通路与胰腺炎

胰腺炎的发病机制长期以来一直是基础和临床研究的一个重要课题,然而至今尚不完全明确.研究证实TLRs(Toll-like receptors)家族成员中TLR4可与G-菌内毒素脂多糖(lipopolysaccharide,LPS)结合,通过NF-κB信号通路激发多种炎症因子的合成进而参与多种器官疾病的发病过程.在鼠类模型和临床研究中已经显示TLR4信号通路在急性胰腺炎(acute pancreatitis,AP)的发病过程中起着重要的作用;上调TLR4信号通路可诱导致炎细胞因子大量释放参与重症急性胰腺炎(severe acute pancreatitis,SAP)病程中多器官功能障碍综合征的形成.因此,进一步明确TLR4信号通路在胰腺炎发病机制的作用,有可能通过阻断TLR4信号通路使胰腺炎获得疗效.