乌司他丁联合粉防己碱预处理对大鼠肝缺血再灌注损伤的保护作用及机制研究

摘要

目的 观察乌司他丁(Ulinastatin,UTI)联合粉防己碱(Tetrandrine,TET)预处理对大鼠肝缺血再灌注损伤(hepatic ischemia-reperfusion injury,HIRI)的保护作用,并分析其作用机理.方法 将40只雌性SD大鼠随均分为5组:假手术组(Sham组)、缺血再灌注损伤组(HIRI组)、乌司他丁预处理组(UTI组)、粉防己碱预处理组(TET组)及乌司他丁联合粉防己碱预处理组(UTI+TET组),每组8只.五组分别于缺血前30 min腹腔注射生理盐水(5 mL/kg)、生理盐水(5 mL/kg)、UTI(5 mL/kg)、TET(5 mL/kg)以及UTI(5 mL/kg)+TET(5 mL/kg).于肝缺血再灌注6 h后收集大鼠血清及肝组织标本.检测大鼠血清中乳酸脱氢酶(lactate dehydrogenase,LDH)、碱性磷酸酶(alkaline hosphatase,ALP)活性.检测大鼠肝组织中髓过氧化物酶(myeloperoxidase,MPO)、谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)活性及大鼠肝组织中核转录因子kappa B(nuclear transcription factor kappa B,NF-κB)p65的表达水平.结果 (1)HIRI组、UTI组、TET组及UTI+TET组血清LDH活性均较Sham组显著升高(P<0.01),HIRI组及TET组血清ALP活性均较Sham组升高(P<0.05,P<0.01),但UTI组、TET组及UTI+TET组血清LDH及ALP活性均明显低于HIRI组(P<0.01);UTI+TET组血清LDH及ALP活性明显低于TET组及UTI组(P<0.01)(.2)HIRI组、UTI组、TET组及UTI+TET组肝组织中MPO活性均较Sham组显著升高(P<0.01)、GSH-Px活性均较Sham组显著降低(P<0.01);但UTI组、TET组及UTI+TET组肝组织中MPO活性均显著低于HIRI组(P<0.01)、GSH-Px活性均显著高于HIRI组(P<0.01);UTI+TET组肝组织中MPO活性均显著低于TET组及UTI组(P<0.01)、GSH-Px活性均显著高于TET组及UTI组(P<0.01)(.3)HIRI组、UTI组、TET组及UTI+TET组肝组织中NF-κB p65相对表达量明显高于Sham组(P<0.01);UTI组、TET组及UTI+TET组肝组织中NF-κB p65相对表达量明显低于HIRI组(P<0.01);UTI+TET组肝组织中NF-κB p65相对表达量明显低于UTI组及TET组(P<0.01).结论 UTI联合TET对HIRI具有保护作用,其机制可能与其抑制肝细胞脂质过氧化作用、增强体内抗氧化能力及下调肝组织中NF-κB p65表达有关.%Objective To observe the protective effect and to analyze the mechanism of ulinastatin (UTI) combined with tetrandrine (TET) pretreatment of hepatic ischemia-reperfusion injury (HIRI) in rats.MethodsA total of 40 female SD rats were randomly divided into 5 groups, each group 8 rats: Sham operation group (Sham group), hepatic ischemia reperfusion injury group (HIRI group), ulinastatin pretreatment group (UTI group), tet-randrine pretreatment group (TET group), and ulinastatin combined with tetrandrine pretreatment group (UTI+ TET group). Five groups were respectively injected with normal saline (5 mL/kg), normal saline (5 mL/kg), UTI (5 mL/kg), TET (5 mL/kg), and UTI (5 mL/kg) combined with TET (5 mL/kg) by intraperitoneal injection 30 min before ischemia. The serum and liver tissue samples were collected 6 h after hepatic ischemia reperfusion. The activities of lactate dehydrogenase (LDH) and alkaline hosphatase (ALP) in serum were detected. The activities of myeloperoxidase (MPO), glutathione peroxidase (GSH-Px) and the expression level of nuclear transcription fac-tor kappa B (NF-κB) p65 in liver tissue were tested.Results (1)The activities of LDH in group HIRI, UTI, TET and UTI+TET were significantly higher than that in Sham group (P<0.01). The activities of ALP in group HIRI and TET were higher than those in Sham group (P<0.05,P<0.01). The activities of LDH and ALP in group UTI, TET and UTI+TET were significantly lower than those in HIRI group (P<0.01). The activities of LDH and ALP in UTI+TET group were significantly lower than those in group UTI and TET (P<0.01). (2)Compared with Sham group, the activities of MPO were significantly increased, the activities of GSH-Px were significantly decreased in group HIRI, UTI, TET and UTI+TET (P<0.01). In group UTI, TET and UTI+TET, the activities of MPO were significantly lower, and the activities of GSH-Px were significantly higher than that in HIRI group (P<0.01). The activities of MPO were significantly lower, the activities of GSH-Px were significantly higher in UTI+TET group than those in group UTI and TET (P<0.01). (3)The relative expressions of NF-κB p65 in group HIRI, UTI, TET and UTI+TET were higher than those in Sham group (P<0.01). The relative expressions of NF-κB p65 in group UTI, TET and UTI+TET were lower than that in HIRI group (P<0.01). The relative expressions of NF-κB p65 in UTI+TET group were lower than that in group UTI and TET (P<0.01).ConclusionUTI combined with TET has a protective effect on HIRI, the mechanism may be related to inhibiting hepatic lipid peroxidation in hepatic cells, enhancing antioxidant capacity and down regulating the expressions of NF-κB p65 in liver tissue.