大鼠肝脏保存再灌注时羟自由基与细胞凋亡、Bcl-2蛋白表达的关系及丹参的保护作用

The relationship between hydroxyl radical contents,apoptosis and expression of Bcl-2 protein and the protection of salvia miltiorrhiz in a rat liver preservation-reperfusion model

目的 阐明羟自由基 (·OH)与细胞凋亡、Bcl 2蛋白表达在大鼠肝脏不同保存时相再灌注过程中的变化规律及相互关系 ,探讨丹参对肝脏保存再灌注损伤的影响。方法 以大鼠肝脏非循环灌注为模型 ,用高效液相色谱 (HPLC)紫外检测法测定·OH生成物二羟苯甲酸 (DHBA)含量 ,用原位末端标记法及电子显微镜观察细胞凋亡 ,用免疫组织化学法检测Bcl 2蛋白表达 ,观察了正常组 (未保存 )、对照组 (乳酸林格液即LR液 )及丹参组 (LR液 +丹参 )保存鼠肝不同时相的效果。结果 随着保存时间延长 (18h内 ) ,·OH水平及细胞凋亡指数 (AI)呈进行性升高 ,且两者呈显著正相关 (r =0 81) ,相同保存时间丹参组明显低于对照组 (P =0 0 4) ;而Bcl 2蛋白表达指数 (BI)呈进行性降低 ,BI与·OH水平及AI呈显著负相关 (r =- 0 84) ,相同保存时间丹参组BI显著高于对照组 (P =0 0 3)。结论 ·OH及细胞凋亡同时参与肝脏保存再灌注损伤 ,丹参对该损伤具有明显的保护作用 ,其作用机理可能主要与抗氧自由基、抑制细胞凋亡及提高Bcl 2蛋白表达作用有关。

Objective[WT5”BZ] To elucidate the dynamic changes of hydroxyl radical (·OH) and apoptosis in a rat liver preservation-reperfusion model, analyse their interrelations and the protective effects of salvia miltiorrhiza.[WT5”HZ]Methods[WT5”BZ] Using isolated nonrecirculated perfusion rat liver model, we observed the preservation effect of control group (with lactate Ringer's solution),DS group (with lactate Ringer's solution added with salvia miltiorrhiza) and normal group(without preservation) by measuring the content of dihydroxybenzoic acid (DHBA) which was formed after reaction of ·OH with salicylic acid,apoptosis index(AI) and the expression index of Bcl-2 protein(BI).[WT5”HZ]Results[WT5”BZ] ·OH levels expressed as DHBAs, and AI enhanced markedly as the time of preservation prolonged. There was a significantly positive correlation between DHBA contents and AI.Both parameters in DS group were significantly lower than those in control group with the same preservation time ( P <0 05). The dynamic changes of BI were just on the contrary when comparing with both parameters.[WT5”HZ]Conclusions[WT5”BZ] ·OH and apoptosis both participate in liver preservation-reperfusion injury, and salvia miltiorrhiza may play an important role on alleviating this injury by a mechanism which correlates to the effect of resisting free radical, inhibiting the occurrence of apoptosis and enhancing the expression of Bcl-2 protein. [WT5”HZ]