CAJ | 학술논문

目的观察急性肝衰竭(ALF)中决定肝细胞能量代谢特点的线粒体内关键酶:柠檬酸合成酶(CS)、肉毒碱棕榈酰转移酶1(CPT-1)、细胞色素C氧化酶(COX)的活性变化.方法 40只雄性SD大鼠分为对照组、造模4h组、造模8h组、造模12 h组及造模24 h组.模型组给予D-氨基半乳糖800 mg/kg和脂多糖100μg/kg联合腹腔注射构建大鼠ALF模型,对照组给予0.9％的等渗盐水1ml腹腔注射.检测各组大鼠ALT、AST、总胆红素、血清白蛋白、凝血酶原时间和凝血酶原活动度;观察肝脏病理和线粒体超微结构变化;检测各组大鼠肝脏线粒体中的CS、CPT-1和COX的活性及mRNA水平,并分析其与肝细胞凋亡及线粒体超微结构之间的关系.方差分析进行各组间总体差异的比较,独立样本t检验进行两组之间差异的比较.结果通过生物化学检查和肝脏病理检查证实本实验的动物模型构建成功.大鼠ALF造模4h时可见肝细胞凋L的出现,8h时凋亡非常明显,12h时可见肝细胞的坏死,24 h时肝脏组织内肝细胞坏死更为显著,并有炎症细胞的浸润.电镜观察线粒体的超微结构发现,模型4h组就已经发生了变化,到24 h时线粒体的外膜结构几乎完全破坏,线粒体崩解.CS、CPT-1、COX的活性在4h均开始升高,8h达到高峰,12h开始下降,24 h下降更为明显.造模8h组与正常组相比,CS[(0.030 8±0.008 2)μmol/min对比(0.004 0±0.001 6)μmol/min]、CPT 1[(0.130 3±0.075 8)μmol/min对比(0.033 4±0.008 3)μmol/min]和COX[(0.022 9±0.002 9)μmol/min对比(0.004 8±0.0007)μmol/min]差异均有统计学意义(t值分别为1.481、2.619和1.014,P＜0.05或P＜ 0.01).结论大鼠ALF过程中,CS、CPT-1和COX活性在肝细胞凋亡时升高,其参与了ALF的发生和发展,尤其是促进了肝细胞凋亡的发生.
목적 관찰급성간쇠갈(ALF)중결정간세포능량대사특점적선립체내관건매:저몽산합성매(CS)、육독감종려선전이매1(CPT-1)、세포색소C양화매(COX)적활성변화.방법 40지웅성SD대서분위대조조、조모4h조、조모8h조、조모12 h조급조모24 h조.모형조급여D-안기반유당800 mg/kg화지다당100μg/kg연합복강주사구건대서ALF모형,대조조급여0.9％적등삼염수1ml복강주사.검측각조대서ALT、AST、총담홍소、혈청백단백、응혈매원시간화응혈매원활동도;관찰간장병리화선립체초미결구변화;검측각조대서간장선립체중적CS、CPT-1화COX적활성급mRNA수평,병분석기여간세포조망급선립체초미결구지간적관계.방차분석진행각조간총체차이적비교,독립양본t검험진행량조지간차이적비교.결과 통과생물화학검사화간장병리검사증실본실험적동물모형구건성공.대서ALF조모4h시가견간세포조L적출현,8h시조망비상명현,12h시가견간세포적배사,24 h시간장조직내간세포배사경위현저,병유염증세포적침윤.전경관찰선립체적초미결구발현,모형4h조취이경발생료변화,도24 h시선립체적외막결구궤호완전파배,선립체붕해.CS、CPT-1、COX적활성재4h균개시승고,8h체도고봉,12h개시하강,24 h하강경위명현.조모8h조여정상조상비,CS[(0.030 8±0.008 2)μmol/min대비(0.004 0±0.001 6)μmol/min]、CPT 1[(0.130 3±0.075 8)μmol/min대비(0.033 4±0.008 3)μmol/min]화COX[(0.022 9±0.002 9)μmol/min대비(0.004 8±0.0007)μmol/min]차이균유통계학의의(t치분별위1.481、2.619화1.014,P＜0.05혹P＜ 0.01).결론 대서ALF과정중,CS、CPT-1화COX활성재간세포조망시승고,기삼여료ALF적발생화발전,우기시촉진료간세포조망적발생.
Objective To determine the roles of mitochondrial apoptosis and energy metabolism in hepatocytes during the pathogenic process of acute renal failure (ALF) by assessing disease-related differential activities of several key mitochondrial enzymes,including citrate synthase (CS),carnitine palmitoyltransferase-1 (CPT-1) and cytochrome c oxidase (COX).Methods Thirty-two male SpragueDawley rats were given D-galactosamine followed by and lipopolysaccharide (LPS) to induce acute liver failure and sacrificed after 4 (4 h group),8 (8 h group),12 (12 h group) and 24 hours (24 h group) of treatment.Eight unmodeled rats served as controls.Effects related to apoptosis were evaluated by pathological analysis of hepatic tissues and TUNEL staining.Ultrastructural changes in mitochondria were assessed by electron microscopy.The activity and expression of CS,CPT-1 and COX were measured.Results Hepatocyte apoptosis was present in the 4 h treatment group and was increased obviously in the 8 h treatment group.Hepatocyte necrosis was first observed in the 12 h treatment group and was significantly higher in the 24 h treatment group,with inflammatory cell invasion.Ultrastructural changes in mitochondria were present in the 4 h treatment group,and the 24 h treatment group showed mitochondria with completely destroyed outer membranes,which resulted in mitochondrial collapse.Activity and protein expression ofCS,CPT-1 and COX were increased in the 4 h group (vs.controls),were at their peak in the 8 h group (CS:t =1.481,P＜ 0.01; CPT-1:t =2.619,P＜ 0.05; COX:t =1.014,P＜ 0.01) and showed a decreasing trend in the 12 h group.In addition,the activities of CS,CPT-1 and COX were enhanced at the stage of hepatocyte apoptosis,suggesting that these enzymes were involved in the initiation and development of ALF.Conclusion Energy metabolism plays an important role in hepatocyte apoptosis during ALF.