【摘要】 目的:研究血红素加氧酶-1一/氧化碳(HO-1/CO)系统对四氯化碳(CC l4)诱导大鼠急性肝损伤的保护作用及其机制。方法:随机将30只SD大鼠分成6组,即正常对照组、四氯化碳染毒组(CC l4)、hem in组、hem in+CC l4组、CO组和CO+CC l4组,每组5只大鼠。采用W estern b lot法测定HO-1蛋白的表达情况;测定各组大鼠血清丙氨酸氨基转移酶(ALT)和天门冬氨酸氨基转移酶(AST)水平,肝组织丙二醛(MDA)浓度和超氧化物歧化酶(SOD)活性以及caspase-3活性和肿瘤坏死因子(TNF)α-水平变化;以HE染色观察肝组织病理形态学改变以及采用TUNEL法观察肝细胞凋亡情况。结果:CC l4成功诱导了大鼠急性肝损伤,表现为染毒24 h后血清ALT,AST水平(2 136.3±163.4 U,1 422.7±221.7 U)以及肝组织MDA浓度(5.28±0.93μmol/g)、肝组织caspase-3活性(光密度值4.69±1.02)和TNFα-水平(256.3±27.3 ng/L)均显著升高,肝组织SOD活性(45.9±14.8 U/mg)下降,和正常对照组相比差异有统计学意义(P<0.01);病理学和TUNEL法检测结果均显示肝有严重损伤,大量肝细胞发生凋亡。给予hem in预处理能显著诱导大鼠肝HO-1的表达,并对肝产生明显的保护作用,表现为大鼠血清ALT,AST水平(287.1±24.3 U,246.2±21.7 U)和肝组织MDA浓度(3.27±1.34μmol/g)明显降低,肝组织SOD活性(71.4±22.6 U/mg)升高,肝组织TNFα-水平(132.6±19.5 ng/L)和caspase-3活性(光密度值2.49±1.47)明显低于CC l4组;此外,hem in预处理亦使染毒大鼠的病理学指标得到明显改善,细胞凋亡的数目显著减少。腹腔注射低浓度CO亦能降低染毒大鼠ALT,AST水平和MDA浓度,抑制caspase-3活性和TNFα-水平,并使组织SOD活性升高,同时改善肝损伤程度,对肝产生明显保护作用。结论:HO-1诱导表达和给予外源性CO均对急性肝损伤大鼠产生明显的保护作用,提示HO-1/CO系统在防护急性肝损伤的病理生理过程中占有重要地位;HO-1/CO系统的保护机制可能与其减轻脂质过氧化反应,抑制caspase-3活性和降低TNFα-水平有关。更多还原

【Abstract】 Objective:To investigate the protective role of heme oxygenase-1 and its reaction product,carbon monoxide against acute liver injury induced by carbon tetrachloride in rats.Methods: Thirty male Sprague-Dawley rats were randomly divided into six groups with five in each.The control group received a single dose of corn oil injection.Carbon tetrachloride was injected intraperitoneally（i.p） to establish acute liver injury models in rats.Hemin（50 μmol/kg） was administered i.p.12 hours before CCl₄ treatment,with an aim to induce HO-1 protein expression in the liver of rats.Carbon monoxide was injected i.p.12 hours prior to CCl₄ injection,resulting in about 8%-12% carboxyhemoglobin concentration in vivo.The expression of HO-1 in the liver of hemin-treated rats was determined by western blot method at different time points.At 24 h after carbon tetrachloride administration,all rats were sacrificed to collect blood samples for the examination of ALT,AST levels and to remove liver tissues for analysis of MDA concentration,SOD activity and caspase-3 activity as well as TNF-a contents.In addition,histopathological changes were investigated and hepatocyte apoptosis was detected by TUNEL method.Results: The administration of carbon tetrachloride to rats caused a marked hepatic damage,characterized by significant elevation of serum ALT,AST levels（2 136.3±163.4 U,1 422.7±221.7 U） and liver MDA con-tent（5.28±0.93 μmol/g）,caspase-3 activitiy（optical density value（4.69）±1.02） and TNF-α level（256.3±27.3 ng/L） combined with a remarkable reduction in liver SOD activity（45.9±14.8 U/mg） as compared with the control rats.Histopathological observations revealed severe damage in the liver and prominent hepatocyte apoptosis took place in CCl₄treated rats.However,pretreatment with hemin could induce high expression of HO-1 protein and exert potent protective effects against liver injury,as demonstrated by a significant decrease in ALT,AST levels（287.1±24.3 U,246.2±21.7 U） and MDA concentration（3.27±1.34 μmol/g）,reduction in caspase-3 activity（optical density value 2.49±1.47） and TNF-α level（132.6±19.5 ng/L）,as compared with the CCl₄-treated rats.Moreover,hepatocyte apoptosis and liver injury were both attenuated remarkably in the liver of rats pretreated with hemin.In contrast to hemin administration,single injection of exogenous CO produced the same protective effects,as indicated by the remarkable reduction of ALT,AST levels and caspase-3 activity and TNF-a levels.Conclusion: The above results suggest that HO-1/CO system has a potent protective effect on acute liver injury induced by carbon tetrachloride in rats.Induction of HO-1 expression and low concentration of CO can inhibit the progress of hepatic damage,which might be due to the alleviation of lipid peroxidation and reduction of caspase-3 activity or inhibition of TNF-α level.更多还原