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Extension of limitation on hepatic warm ischemia, with special reference to analysis of the microcirculatory disturbance and the mechanism of ischemic tolerance from the view points of stress responce protein

肝脏热缺血限制的延伸，特别是从应激反应蛋白的角度分析微循环障碍和缺血耐受机制

基本信息

批准号：
09671300
负责人：
NOGUCHI Takashi
金额：
$ 1.15万
依托单位：
Mie University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

项目摘要

<Purpose> This study was performed to investigate the cytoprotective effect of ischemic tolerance established by initial transient ischemia as a sub lethal stress or administration of Geranyl-geranyl-acetone (GGA),which are known to be one of heat shock protein (Hsp) inducers, make an extension of hepatic warm ischemia limit, and to evaluate the mechanism ischemic tolerance, hepatic microcirculation and endothelial cell function with in vivo to in vitro. Moreover, small intestinal model was evaluated as well as hepatic model.<Materials & Methods> Male wistar rats were subjected to 45 min of hepatic ischemia followed reperfusion : (1) 20 min liver ischemia 7 days prior to 45 min ischemia, or (2) oral administration of GGA 200 mg/kgBW for 7 or 14 days prior to 45 min ischemia. Examined were survival rate, liver function, hepatic tissue blood flow, histologic findings, expression of Hsp 70 and NF-kB (Western blot), and NOィイD22ィエD2 + NOィイD23ィエD2 concentration in isolated hepatocyte/non-par … More enchymal cell cultures 8Griess method). In intestinal models which were subjected to 90 min superior mesenteric artery clumped, preconditioning groups were (1) 30 min initial ischemia 7 days prior to 90 min ischemia and (2) oral administration of GGA 14 days prior to 90 min ischemia.<Results> 1. The 7-day survival rate : all animals of 45 min ischemia were died within 48 hrs after reperfusion, in contrast, preconditioning groups were significantly higher (83% and 75%) respectively. 2. In preconditioning groups, ALT & HA levels in the serum, hepatic blood flow, and histologic findings including increased apoptosis in hepatocytes and endothelial cells showed a significant improvement . 3. Hsp 70 induction was remarkably increased immediately after reperfusion to 6 hrs. 4. The expression and activation of NF-kB were significantly suppressed. 5. The NO production was inhibited by iNOS in preconditioning groups. 6. Intestinal ischemia limit was extended by preconditioning and apoptosis was regulated with increased Hsp 70 expression.<Conclusion> Ishemic preconditioning and oral administration of GGA protected liver sinusoidal endothelial cell dysfunction and improved the hepatic microcirculatory disturbance in the postischemic liver by inducing Hsp 70. This Hsp 70 suppresses NF-kB to regulate the overproduction of NO attribute to iNOS. Preconditioning by ischemia or drugs establishes ischemic tolerance in small intestine as well as liver. Therefore, performing the ischemic preconditioning is expected to be of much benefit to extend hepatectomy, or liver transplantation and so on. Less

<Purpose>本研究旨在探讨以亚致死性缺血为起始条件建立的缺血耐受和热休克蛋白（Hsp）诱导剂香叶基香叶基丙酮（GGA）对肝细胞的保护作用，并从体内外两方面探讨缺血耐受的机制、肝微循环和内皮细胞功能。此外，还对小肠模型和肝脏模型进行了评价。<材料与方法>雄性Wistar大鼠肝脏缺血45 min后再灌注：（1）缺血45 min前7天肝脏缺血20 min，或（2）缺血45 min前7天或14天口服GGA 200 mg/kgBW。检测存活率、肝功能、肝组织血流量、组织学改变、Hsp 70和NF-kB的表达（Western blot）以及离体肝细胞/非离体肝细胞中NO标记物D22 + NO标记物D23的浓度。 ...更多信息间充质细胞培养8 Griess法）。在肠模型中，进行90 min的上级肠系膜动脉丛聚，预处理组为（1）90 min缺血前7天30 min初始缺血和（2）90 min缺血前14天口服GGA。<Results>1. 7天存活率：缺血45 min组动物均在再灌注48 h内死亡，而预处理组动物分别为83%和75%。2.在预处理组中，血清中的ALT和HA水平、肝血流量以及肝细胞和内皮细胞凋亡增加的组织学发现显示出显著改善。3. Hsp 70的诱导在再灌注后即刻至6小时显著增加。4. NF-kB的表达和活化被显著抑制。5.诱导型一氧化氮合酶抑制预处理组NO的产生。6.肠缺血预处理延长肠缺血极限，并通过增加Hsp 70表达调节细胞凋亡. <Conclusion>缺血预处理和口服GGA可通过诱导Hsp 70的表达，保护肝窦内皮细胞功能障碍，改善缺血后肝脏微循环障碍。Hsp 70通过抑制NF-κ B的表达来调节iNOS引起的NO的过度生成。缺血或药物预处理可在小肠和肝脏中建立缺血耐受。因此，进行缺血预处理有望对扩大肝切除术或肝移植等有很大益处。