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Development of measurement for hepatic isc hemia-repetfusion in jiry accoidirg to are gulation of activin-follistin system

肝脏缺血再灌注测量的发展

基本信息

批准号：
18591516
负责人：
MORINE Yuji
金额：
$ 2.41万
依托单位：
The University of Tokushima
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

项目摘要

Background)Ischemia-reperfusion injury to the liver are of clinical importance in humans after hemorrhagic and cardiogenic shock, liver surgery, or liver transplantation. It was reported that blockade of the action of activin with administration of follistatin accelerates recovery from ischemia renal injury by accelerating regeneration after ischemia. In this study we evaluated the role of activin-follistatin on hepatic ischemia reperfusion injury and develop the new measurement for hepatic ischemia reperfusion injury.Method and Results)1) Effects of preactivation by portal vein ligation on liver regeneration (assessment according to activin)This study evaluated the effect of regenerating livers preactivated by PVL following massive hepatectomy. The mRNA expression levels of activin A and ActR II A were significantly higher in regenerating liver group than in normal liver group at late pahse. The regenerating liver preactivated by PVL is restricted late-phase liver regeneration after m … More assive hepatectomy.2) Beneficial Effects of follistatin in hepatic ischemia reperfusion injury Total hepatic ischemia for 30 min was performed followed by reperfusion. Follistatin (1mg/ body) was administered at the time of reperfusion. Four of 5 animals died in control group within 3days after reperfusion, whereas 80% of animals survived in follistatin group (P<0.05). AST was significantly lower at 3 hr after reperfusion in follistatin group (P<0.05). LDH was also lower at 6 hr after reperfusion in follistatin group (P<0.05). Follistatin inhibited the mRNA expression of bA subunit of activin. Moreover the expression of IL-6, which is inflammatory cytokine, was suppressed at 6 hr after reperfusion (P<0.05).3)Beneficial Effects of fluvastatin on Hepatic Ischemia-Reperfusion InjuryRats were divided into 3 groups: fluvastatin group; oral administration of fluvastatin (20mg/kg/day)for 2 days before the operation, control group, sham group. Fluvastatin and control groups were given total hepatic warm ischemia for 30 min. After reperfusion, an expression of mRNA level of endotherial NO synthase (eNOS) and NOX4 was determined by reverse transcriptase (RT)-PCR, tumor necrosis factor- a (TNF-α)and interleukine-1 β(IL-1β) mRNA by real-time RT-PCR. In the fluvastatin group, the expression of eNOS mRNA was significantly higher, and NOX4 mRNA was significantly lower (p<0.05). Furthermore, the expression of inflammatory cytokines mRNA were suppressed in the fluvastatin group. In particular, TNF- α was significantly lower in the fluvastatin group (P<0.05).4) Beneficial Effects of fluvastatin on Liver Microcirculation and Regeneration The liver regeneration rate in fluvastatin group was significantly higher at 72 hours after hepatectomy (p<0.05). Sinusoidal area in fluvastatin group was maintained histologicaly. Furthermore, the expression of-SMA protein in the liver was inhibited in fluvastatin group at 48 hours after hepatectomy.Summary) We were not able to elucidate the regulation of liver regneration using activin, because we had difficulty with the purification of activin. In this study, we clarified the regulatory mechanism of hepatic ischemia-reperfusion injury and effect of several modulators including follistatin for hepatic ischemia-reperfusion injury using molecular biological method. Less

背景）肝脏缺血再灌注损伤在人类出血性和心源性休克、肝脏手术或肝脏移植后具有重要的临床意义。据报道，通过施用卵泡抑素阻断激活素的作用，通过加速缺血后的再生来加速缺血性肾损伤的恢复。本研究旨在探讨激活素-卵泡抑素在肝脏缺血再灌注损伤中的作用，并建立一种新的肝脏缺血再灌注损伤的检测方法。方法与结果1）门静脉结扎预激活对肝再生的影响（以激活素为指标）本研究评价了PVL预激活对大面积肝切除术后再生肝的影响。再生肝组激活素A和ActR Ⅱ A的mRNA表达水平在晚期明显高于正常肝组。PVL预激活的再生肝对移植后晚期肝再生有抑制作用， ...更多信息 2）卵泡抑素在肝缺血再灌注损伤中的有益作用进行30分钟的全肝缺血，然后再灌注。再灌注时给予卵泡抑素1 mg/只。再灌注后3d内对照组5只动物中有4只死亡，而卵泡抑素组动物存活率为80%（P<0.05）。再灌注后3 h，卵泡抑素组AST明显低于对照组（P<0.05）。再灌注6 h时，卵泡抑素组的LDH也明显低于对照组（P<0.05）。卵泡抑素抑制激活素bA亚基mRNA的表达。3）氟伐他汀对肝脏缺血再灌注损伤的保护作用将大鼠随机分为3组：氟伐他汀组、对照组、假手术组，氟伐他汀组于术前2d灌胃氟伐他汀20 mg/kg/d，对照组灌胃氟伐他汀20 mg/kg/d，假手术组灌胃氟伐他汀20 mg/kg/d，假手术组灌胃氟伐他氟伐他汀组和对照组给予全肝热缺血30 min，再灌注后用逆转录酶（RT）-PCR法检测内皮型一氧化氮合酶（eNOS）和一氧化氮合酶4（NOX 4）mRNA的表达，实时荧光定量PCR法检测肿瘤坏死因子-α（TNF-α）和白细胞介素-1 β（IL-1β）mRNA的表达。氟伐他汀组eNOS mRNA表达明显高于对照组（P <0.05），而NOX 4 mRNA表达明显低于对照组（P<0.05）。此外，氟伐他汀组的炎症细胞因子mRNA的表达受到抑制。4）氟伐他汀对肝微循环和再生的影响术后72小时，氟伐他汀组肝再生率明显高于对照组（p<0.05）。氟伐他汀组窦状隙区在组织学上得以维持。此外，在肝切除术后48小时，在氟伐他汀组中，-SMA蛋白在肝脏中的表达被抑制。本研究采用分子生物学方法阐明了肝脏缺血再灌注损伤的调控机制以及卵泡抑素等多种调节剂对肝脏缺血再灌注损伤的影响。少