姜黄素对慢性酒精肝损伤保护作用的研究

  本文关键词：姜黄素对慢性酒精肝损伤保护作用的研究，由笔耕文化传播整理发布。

        第一部分姜黄素对乙醇诱导的大鼠原代肝细胞氧化损伤的保护作用目的：以Sparague-Dawley大鼠（简称SD大鼠）原代肝细胞为研究对象，建立酒精肝损伤模型，探讨酒精对肝细胞的氧化损伤以及姜黄素对其的保护作用，并探讨姜黄素对肝细胞HO-1活性表达的诱导。方法：1、分离并培养SD大鼠原代肝细胞，用不同浓度乙醇（0~200mmol/L）对大鼠原代肝细胞染毒8h，或者100mmol/L无水乙醇染毒不同时间（0~24h），检测细胞的LDH释放水平、AST水平以及氧化/抗氧化系统水平（MDA水平、GSH含量和SOD活性）。2、100mmol/L无水乙醇染毒细胞前1h用不同剂量姜黄素（0~50μmol/L）预处理，或者乙醇染毒前不同时间（0~5h）用15μmol/L姜黄素预处理，检测细胞LDH、AST、MDA水平，GSH含量和SOD活性。3、梯度离心法分离提取各组肝细胞微粒体，检测HO-1酶的活性。结果：1、随着乙醇浓度的增加和作用时间的延长，大鼠原代肝细胞LDH释放水平和AST水平显著增高，MDA水平升高，GSH含量和SOD活性下降，，在浓度为100mmol/L和时间为8h时影响程度明显，P值均<0.05。2、与乙醇对照组相比，提前用姜黄素干预可有效抑制LDH的释放、抑制AST和MDA水平的升高，提高GSH含量以及SOD活性，预作用时间为1h、剂量为15μmol/L时差异有统计学意义（P <0.05）。3、姜黄素能上调HO-1蛋白表达，提高肝细胞内HO-1酶的活性，以15μmol/L的剂量预作用1h时效果最明显（P <0.05）。结论：乙醇诱导的大鼠原代肝细胞氧化损伤程度与给予乙醇的浓度及其作用时间呈正相关，姜黄素能够有效降低乙醇诱导的肝细胞氧化损伤，并能诱导HO-1发挥抗氧化作用。第二部分姜黄素对BALB/c小鼠慢性酒精性肝损伤的干预作用目的：以BALB/c小鼠为研究对象，建立慢性酒精性肝损伤模型，并探讨姜黄素对酒精诱导的肝脏氧化损伤的保护作用。方法：1、将实验小鼠随机分为4组（A：正常对照组，B：姜黄素组，C：酒精组，D：酒精+姜黄素组），C、D两组梯度酒精暴露6周（前四周灌胃量为2.4g/kg/day，后两周增加到4g/kg/day）建立慢性酒精性肝损伤模型，B、D两组给予75mg/kg/d的姜黄素行灌胃干预。2、实验结束后收集小鼠血液及肝组织标本，制作肝组织病理切片，检测血清转氨酶（AST、ALT）水平和脂质代谢（TC、TG、HDL-C）水平。3、检测肝组织ROS含量和氧化/抗氧化系统水平（T-AOC、GSH、GPx、GST、MDA）。结果：1、正常对照组和姜黄素组小鼠体重增长明显大于酒精灌胃的两组（P值均<0.05），但姜黄素的补充并未明显改善由酒精灌胃所造成的体重增长缓慢这一情况。2、与正常对照组相比，酒精灌胃组肝组织切片可见明显的脂肪变性、血清转氨酶AST、ALT水平提高（P <0.05），以及血清TC、TG含量升高（P <0.05）、HDL-C含量降低（P>0.05）。给予姜黄素干预后，可见肝脏脂肪变性有明显减轻，AST、ALT水平下降，血脂紊乱恢复正常。3、与正常对照组相比，酒精灌胃组小鼠肝组织ROS含量显著上升，T-AOC、GSH、GPx以及GST水平都显著降低，MDA水平明显升高，P值均<0.05。而酒精+姜黄素组小鼠肝组织ROS含量以及各项抗氧化、脂质过氧化指标均恢复正常。结论：姜黄素能减轻酒精诱导的肝脏脂肪变性、抑制血清转氨酶的释放、改善脂质代谢紊乱，并且抑制ROS在肝脏中的蓄积、减缓肝组织的氧化损伤，从而对酒精诱导的肝损伤起保护作用。

    Part1Protective Effects of Curcumin on Oxidative DamageInduced by Ethanol in Rat Primary HepatocytesObjective: To establish alcoholic liver injury model in Sparague-Dawley rat primaryhepatocytes, and study the protective effects of curcumin on oxidative damage induced byethanol.Methods:1. Rat primary hepatocytes were treated with ethanol of different dosage (0-200mmol/L) for8h, or100mmol/L in different time (0-24h) to detect cell viability (LDH,AST levels) and oxidative/antioxidative system (MDA, GSH and SOD levels).2. On the basis of this, the hepatocytes were pre-treated with curcumin of differentdoses (0-50μmol/L) for1h, or15μmol/L in different time (0-5h) before ethanol exposure(100mmol/L,8h).Then detect LDH, AST, MDA, GSH and SOD levels.3. Extract the hepatocellular microsomes by gradient centrifugation and detect theHO-1enzyme activity.Results:1. There is remarkable increase of LDH and AST levels along with the increase ofethanol dosage and the extend of the time, as well as MDA formation in hepatocytes, and adecrease of GSH and SOD in ethanol group, it is significant difference at the dosage of100mmol/L or the time of8h (P <0.05).2. Compared with the alcohol group, pre-treatment of curcumin could significantlyprohibit the release of AST and LDH, the rise of MDA level and the decline of SODactivities and GSH content of hepatocytes exposed to ethanol (P <0.05).3. Compared with the control group, pre-treatment of curcumin could improve theprotein level and enzyme activity of HO-1at the dose of15μmol/L and time for1h.Conclusions: Ethanol-induced oxidative damage may occurred in rat primary hepatocytesat dose-and time-dependent manner. Curcumin could prevent its damage, as well asup-regulate HO-1protein expression and improve its enzyme activity. Part2Effects of Curcumin on Chronic Alcoholic Liver Injuryin BALB/c MiceObjective: To establish chronic alcoholic liver injury model in BALB/c mice, andinvestigate the protective effects of curcumin on oxidative damage.Methods:1. BALB/c mice were randomly divided into4groups (A: normal control group,B: curcumin group, C: ethanol group and D: ethanol+curcumin group), the chronicalcoholic liver injury model was established with gradient dosages of ethanol exposure by6weeks (from2.4g/kg/day to4g/kg/day). Group B and D were administered with curcuminof75mg/kg/day.2. Blood samples and liver tissues were collected at the end of the experiment, thehistopathological changes of the liver were observed, the enzyme activity of ALT, AST inserum and plasma levels of TC, TG, HDL-C were determined.3. The content of ROS in liver tissues was determined, as well as the level of T-AOC,GSH, GPx, GST and MDA.Results:1. The body weight gain in group A and B were significant higher than the othertwo groups (P <0.05), but no significant differences were found among group C and D (P>0.05).2. Compared with the control group, ethanol exposure significantly elevated the levelof serum AST, ALT, TC and TG (P>0.05), decline the level of HDL-C (P>0.05), andlipid droplet accumulation were observed. These were attenuated by curcuminsupplementation.3. Compared with the control group, ethanol exposure resulted in liver ROS generation,T-AOC, GSH, GPx and GST depletions and MDA elevention (P <0.05), which weresignificantly reversed by curcumin supplementation.Conclusion: Curcumin supplementation could alleviate pathological changes in the liverand hepatic enzymes release, attenuate the lipid disorder, improve antioxidant enzymeactivity and protect liver from ethanol-induced oxidative stress damage.

姜黄素对慢性酒精肝损伤保护作用的研究

缩略词5-7摘要7-9Abstract9-10前言11-14第一部分 姜黄素对乙醇诱导的大鼠原代肝细胞氧化损伤的保护作用14-29    1 实验材料14-16    2 实验方法16-20    3 实验结果20-26    4 讨论26-29第二部分 姜黄素对 BALB/c 小鼠慢性酒精性肝损伤的保护作用29-43    1 实验材料29    2 试验方法29-33    3 实验结果33-41    4 讨论41-43小结43-46参考文献46-51综述51-65    参考文献60-65附录65-67致谢67

  本文关键词：姜黄素对慢性酒精肝损伤保护作用的研究，由笔耕文化传播整理发布。