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

肝缺血再灌注损伤在人类出血性、心源性休克、肝手术或肝移植后具有重要的临床意义。据报道，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肾损伤的恢复。本研究旨在评价激活素-卵泡抑素在肝缺血再灌注损伤中的作用，并建立新的肝缺血再灌注损伤测量方法。方法与结果)1)门静脉结扎预激活对肝脏再生的影响（以激活素评价）本研究评价了PVL预激活肝切除术后肝脏再生的效果。晚期再生肝组激活素A和ActR II A mRNA表达量显著高于正常肝组。PVL预激活的再生肝在大范围肝切除术后晚期肝再生受到限制。2)卵泡抑素对肝缺血再灌注损伤的有益作用，肝全缺血30min后再灌注。再灌注时给予卵泡listatin （1mg/体）。对照组5只动物中4只在再灌注后3d内死亡，而卵泡抑素组80%的动物存活（P<0.05）。再灌注后3 h，卵泡抑素组AST明显降低（P<0.05）。再灌注后6 h，卵泡抑素组LDH明显降低（P<0.05）。卵泡抑素抑制激活素bA亚基mRNA表达。炎症因子IL-6的表达在再灌注后6小时受到抑制（P<0.05）。3)氟伐他汀对肝缺血再灌注损伤的有益作用大鼠分为3组：氟伐他汀组；术前2天口服氟伐他汀（20mg/kg/天），对照组、假手术组。氟伐他汀组和对照组给予全肝热缺血30 min。再灌注后，采用逆转录酶(RT)- pcr检测内皮NO合成酶（eNOS）和NOX4 mRNA的表达水平，实时RT- pcr检测肿瘤坏死因子- a （TNF-α）和白细胞介素-1β (IL-1β) mRNA的表达水平。氟伐他汀组eNOS mRNA表达量显著升高，NOX4 mRNA表达量显著降低（p<0.05）。此外，氟伐他汀组炎症细胞因子mRNA的表达受到抑制。特别是氟伐他汀组TNF- α显著降低（P<0.05）。4)氟伐他汀对肝脏微循环和再生的有益作用氟伐他汀组在肝切除术后72 h肝再生率显著高于对照组（p<0.05）。氟伐他汀组窦区维持组织学。此外，氟伐他汀组在肝切除术后48小时肝脏中sma蛋白的表达受到抑制。我们无法阐明激活素对肝脏再生的调控作用，因为我们在纯化激活素方面有困难。本研究采用分子生物学方法，阐明肝缺血再灌注损伤的调控机制，以及卵泡抑素等几种调节剂对肝缺血再灌注损伤的作用。少

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