程序性坏死在自体原位肝移植术大鼠肝脏损伤中的作用

Role of necroptosis in liver injury in rats undergoing autologous orthotopic liver transplantation

目的评价程序性坏死在自体原位肝移植术大鼠肝损伤中的作用。方法SPF级成年雄性SD大鼠24只,8~10周龄,体重250~280g,采用随机数字表法分为3组(n=8):假手术组(S组)、自体肝移植组(T组)和程序性坏死抑制剂necrostatin-1组(N组)。T组和N组采用自体肝移植术制备大鼠肝移植模型。N组和T组术前30min分别腹腔注射necrostatin-1 1.0mg/kg和等容量二甲基亚砜。于门静脉开放6h(S组术毕后6h)取下腔静脉血样,测定血清AST和ALT浓度,取大鼠肝组织,光镜下观察肝脏形态学,采用Suzuki病理学评分评估肝脏损伤程度,分光光度计法检测肝组织MDA含量和SOD活性,Western blot法检测受体相互作用蛋白激酶1(RIPK1)、RIPK3和混合系列蛋白酶样结构域(MLKL)的表达。结果与S组比较,T组血清AST和ALT浓度、肝组织病理学评分和MDA含量升高,SOD活性降低,肝组织RIPK1、RIPK3和MLKL表达上调(P<0.05);与T组比较,N组血清AST和ALT浓度、肝组织病理学评分和MDA含量降低,SOD活性升高,肝组织RIPK1、RIPK3和MLKL表达下调(P<0.05)。结论程序性坏死参与了自体肝移植术大鼠肝脏损伤的过程。

Objective To evaluate the role of neeroptosis in liver injury in rats undergoing autologous orthotopic liver transplantation (AOLT).Methods Twenty-four SPF adult male Sprague-Dawley rats,weighing 250-280g,were divided into 3 groups (n=8 each)using a random number table method: sham operation group (group S),AOLT group (group T) and neeroptosis inhibitor neerostatin-1 group (group N).Rats were anesthetized using intraperitoneal 5% chloral hydrate 0.5ml/100g.A standard model of OALT was established as previously deseribed by Yao et al.in T and N groups.Necrostatin-1 1.0 mg/kg and the equal volume of dimethyl sulfoxide were intraperitoneally injected at 30 min before surgery in N and T groups,respectively.Blood samples were collected from the inferior vena eava at 6h after opening the portal vein (at 6h after operation in group S)for determination of serum aspartate amino-transferase (AST)and alanine amino-transferase (ALT)concentrations.The livers were removed for examination of the pathological changes (with a light mioroscope)and for determination of the malondialdehyde (MDA) contents and superoxide dismutase (SOD)activity (using a spectrophotometer)and expression of receptorinteracting protein kinase-1(RIPK1),RIPK3 and mixed lineage kinase domain-like (MLKL)in liver tissues (by Western blot).The damage to livers was evaluated using Suzuki's scoring system.Results Compared with group S,the serum AST and ALT concentrations,Suzuki's score of liver tissues and MDA content were significantly increased,the SOD activity was decreased,and the expression of RIPK1,RIPK3 and MLKL was up-regulated in group T (P<0.05).Compared with group T,the serum AST and ALT concentrations,Suzuki's score of liver tissues and MDA content were significantly decreased,the SOD activity was increased,and the expression of RIPK1,RIPK3 and MLKL was down-reguIated in group N (P < 0.05).Conclutsion Necroptosis is involved in liver injury in rats undergoing AOLT.