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牛膝对肝脏损伤保护作用的研究
Protective Effects of Achyranthes Bidentata Bl. on Liver Injury
【作者】 陈坤;
【导师】 李金贵;
【作者基本信息】 扬州大学 , 临床兽医学, 2010, 硕士
【摘要】 牛膝是我国常用的大宗中药材之一,具有补肝肾、强筋骨、通淋利尿、引血下行的功效。现代药理学研究表明牛膝具有抗炎镇痛、活血化瘀、抗氧化、抗衰老、抗肿瘤及增进记忆力、促进蛋白同化等多种作用。本研究通过小鼠部分肝切除肝再生模型,卡介苗加脂多糖尾静脉注射诱导小鼠免疫性肝损伤模型和体外H2O2诱导大鼠肝细胞损伤模型,分别探讨牛膝在对抗肝损伤,改善肝功能方面的作用,为牛膝的临床的应用提供科学依据。为了探讨牛膝对小鼠肝再生的影响,本文用手术方法建立了ICR小鼠68%肝叶切除动物模型。给小鼠灌服牛膝水煎液(2、4、6 g/kg.bw),观测小鼠肝再生过程中不同时间点(0、24、48、72、120、192 h)的生理和病理学的变化。结果显示:与模型组相比,牛膝对小鼠围手术期体重无明显的增重效应,部分肝切除后24h牛膝水煎液组小鼠血浆ALT、IL-6、CB的含量显著降低,在48h小鼠血浆CB的含量仍低于模型组,在其他时间点的作用规律不明显;在术后192h时牛膝水煎液中、低剂量组(4、2 g/kg.bw)小鼠的肝指数和脾指数显著高于模型组,肝组织切片H.E.染色镜检显示术后192h,牛膝水煎液组肝细胞有丝分裂相明显高于模型组。这提示牛膝能够降低因部分肝切除引起的小鼠血浆ALT、IL-6和CB水平的升高,减轻手术引起的肝损伤,而有利于肝再生。为了探讨牛膝对免疫性肝损伤小鼠的影响,本文以BCG加LPS尾静脉注射法诱导小鼠免疫性肝损伤模型,小鼠尾静脉注射BCG(1mg/0.2ml),然后灌服牛膝水煎液每天一次连续10天,10天后以相同方式给予LPS(7.5μg/0.2ml)。检测结果显示:牛膝水煎液高、中、低剂量(4、2、1 g/kg.bw)可明显降低免疫性肝损伤引起的小鼠血清ALT的升高,减轻肝组织病理性损伤,使肝组织匀浆中的GSH的含量和SOD活性升高,但不能降低因肝损伤引起的肝指数和血清AST、NO的升高。这提示牛膝对BCG联合LPS诱导的小鼠免疫性肝损伤有保护作用。为了进一步探讨牛膝的抗氧化作用机制,本文通过体外原位灌流法分离大鼠肝细胞,采用MTT法检测牛膝水煎液(6.4、3.2、1.6、0.8 mg/ml)和牛膝多糖(200、100、50、25μg/ml)对肝细胞的毒性,结果表明受试浓度药物没有毒性或毒性极低。在此基础上建立体外H2O2损伤大鼠肝细胞模型,探讨牛膝的抗氧化作用。结果显示受试药物浓度均能降低因H2O2损伤大鼠肝细胞悬液的AST和ALT活性。这提示牛膝和牛膝多糖对H2O2引起的肝细胞损伤均具有保护作用。
【Abstract】 Achyranthes bidentata Bl. (Ab) is one of commonly used Chinese herbal medicine, which has the activities of nourishing liver and kidney, strengthening bones and tendons, promoting diuresis and relieving stranguria, promoting blood circulation. Modern pharmacological studies shows that Achyranthes bidentata Bl. has a variety of pharmacological effects such as alleviate inflammatory pain, activating blood circulation to dissipate blood stasis, anti-oxidant, anti-caducity, anti-tumor and to increase the ability of memory and promote assimilation of protein. Three experimental model were established respectlively to explore the effects of Ab water decoction on reducing liver damage and improving liver function in this study, the three models are liver regeneration model after partial hepatectomy in mice, mouse immunological liver injury model induced by injecting BCG plus LPS via tail vein and model of rat hepatocytes injury induced by H2O2.To investigate the effects of Achyranthes bidentata Bl. on mice liver regeneration after partial hepatectomy, 68% hepatectomy model was established by surgical method in ICR mice in this study. Ab water decoction(2、4、6 g/kg.bw) were given daily via gavage, the changes of physiology and pathology were observed. The results showed that: comparing with model group, Ab water decoction had no significant weight gain effects during perioperative period, the level of mice plasma ALT, IL-6, CB in Ab water decoction group were significantly lower than those of model group during 24h after PH, the plasma CB was still significantly lower, however, no obvious difference could be observed in other times; liver index and spleen index in middle and low dose group(4、2 g/kg.bw)was significantly higher than those of model group at 192h after PH, H.E. staining of liver tissue slice showed that the numbers of hepatocytes with mitosis were significantly higher than the model group at 192h after PH. This data suggested Ab could reduce the increased plasma ALT, IL-6 and CB levels caused by PH, alleviate the liver injury and beneficial to liver regeneration.In order to study the effects of Ab on immunological liver injury in mice, mouse liver immunological injury was established by injection BCG (1mg/0.2ml) via tail vein and 10d after injecting LPS (7.5μg/0.2ml), during the 10d period Ab water decoction were given daily via gavage. Physiological indexes and pathological changes showed that: Ab water decoction (1,2,4 g/kg.bw) could significantly reduce higher serum ALT activity caused by the liver injury and alleviate the pathological liver injury, increase liver homogenates contents of GSH and SOD, but these dose of Ab water decoction could not reduce the higher liver index, serum AST and NO level induced by liver damage. The results showed that Ab had some protective effects on mouse immunological liver injury.In order to further study the mechanism of the antioxidant function of Ab, rat hepatocytes were isolated by in situ perfusion in this study,the toxicity of Ab water decoction (6.4,3.2,1.6,0.8 mg/ml) and the Achyranthes bidentata polysaccharides (ABPS) (200,100,50,25μg/ml) to rat hepatocytes were measured by MTT assay. The results showed that the tested drugs had no or very low toxicity. On base of this data, the model of rat hepatocytic injury was established by explring to H2O2 in vitro, all concentrations of tested drugs could reduce the activity of AST and ALT in hepatocytes suspensions. Both Ab and ABPS have protective effect on the hepatocytic injury caused by H2O2.
【Key words】 Achyranthes bidentata Bl.; liver regeneration; partial hepatectomy; immunological liver injury; hepatocytes; antioxidant;
- 【网络出版投稿人】 扬州大学 【网络出版年期】2011年 02期
- 【分类号】R285.5
- 【被引频次】3
- 【下载频次】220