脂多糖诱导小鼠肝损伤介质水平变化及五灵胶囊的作用

                         作者：刘明义，王胜春，赵慧萍

【摘要】  　　目的探讨脂多糖（LPS）诱导小鼠肝损伤介质水平变化及五灵胶囊的作用。方法小鼠尾静脉注射（iv ）LPS 3.5mg/kg ，酶法检测3，6，12和24 h小鼠血清ALT、 AST、 ALP、TRIG，化学法测定肝组织内MDA、NOS水平，免疫组化检测肝组织内TNF-α，CD14，iNOS，eNOS的阳性表达率。结果与正常组比较,小鼠iv 注射LPS 后3 h血清ALT，AST水平明显升高并持续至12 h，24 h下降仍显著高于正常组，ALP水平明显持续下降。在12 h MDA含量显著升高，24 h下降但明显高于正常组，而甘油三酯（TG）水平各时间点明显升高，TNF-α，CD14，iNOS，eNOS阳性表达率上调。五灵胶囊能明显降低ALT，AST水平和MDA含量，对ALP和 TRIG水平无作用，抑制肝组织内TNF-α，CD14，iNOS，eNOS阳性表达率。结论MDA，TNF-α，CD14，iNOS，eNOS参与了LPS诱导肝损伤，五灵胶囊抑制LPS诱导肝损伤的作用机制与抑制活性递质生成有关。

【关键词】  肝损伤 一氧化氮合酶 活性介质 脂多糖

　　Abstract：ObjectiveTo explore the change of mediator levels in hepatic injured mice induced by Lipoplysaccharide (LPS) and effects of Wuling capsule.MethodsLPS was given to mice by injection with a dose of LPS 3.5mg/kg via the tail vein, 3,6,12,24 hours later, the levels of ALT, AST, ALP in serum were detected by enzyme method and the contents of MDA, NOS in hepatic tissue of mice were determined by chemistry, immunohistochemistry method was employed  to determine the expression of TNF-α, CD14, iNOS and eNOS in hepatic tissue. Results3 hours after LPS was given to mice, compared with normal group, the levels of ALT and AST were obviously elevated and persisted for 12 hours, and which were reduced in 24 hours but still high obviously as compared with normal group. The level of ALP were evidently decreased, and the content of MDA was significantly increased at 6 hour and lowered at 24 hours but still higher than normal. The level of triglycerides was increased and the expression of TNF-α , CD14, iNOS ,eNOS in liver injury tissue was markedly up-regulated.ConclusionThe mediators as MDA, TNF-α, CD14, iNOS ,eNOS participated in liver injury induced by LPS, the protective mechanism  Wuling capsule liver injury might be related with inhibiting the release of mediators.    

　　Key words：Liver injury;  Nitric oxide synthase;  Activity mediators;  Lipoplysaccharid
   
　　内毒素血症与肝损害可互为因果，从而对肝病的发生发展及预后产生重要的影响［1］。脂多糖（lipopolysaccharide  LPS）致肝损伤机制诸多学者认为：主要是通过激活枯否细胞（Kupffer cells KC）释放细胞因子而介导肝细胞损害，LPS促进枯否细胞合成释放TNF-α, 进而促进NO生成加重肝损伤［2］。早期研究表明五灵胶囊对LPS诱导枯否细胞释放细胞因子有明显抑制作用［3］，五灵胶囊具有疏肝健脾，扶正活血的作用，对内毒素血症肝损伤及介质TNF-α，CD14，iNOS，eNOS表达有无影响还未经证实，本文对此进行了观察，以便为临床用药提供依据。

　　1  材料
   
　　动物：昆明种小鼠，雄性，体质量（25±2）g，中国人民解放军第四军医大学动物中心提供，动物合格证号：SCK(军)2002-05；丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）、碱性磷酸酯酶（ALP），甘油三酯（TG）,胆固醇(Chol)试剂盒（北京中生）；丙二醛（MDA）,一氧化氮合酶 （NOS）试剂盒（南京建成）； Rabbit anti-CD14，TNF-α，NOSⅡ（iNOS），NOSⅢ（eNOS）一抗（武汉博士德）；S-P超敏试剂盒（福州迈新）；脂多糖（LPS）（Sigma公司）；五灵胶囊（中国人民解放军第四军医大学西京医院药剂科提供，批号：20040122），称取五灵胶囊2 g，蒸馏水加热溶解至10 ml为经口给药供试液。

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