α-硫辛酸对大鼠肝缺血再灌注后缺血型胆道病变防治作用的实验研究

Experimental study on preventive and therapeutic effect of alpha-lipoic acid on ischemic type biliary lesion after ischemia-reperfusion in rat liver

目的:探讨大鼠肝缺血再灌注(ischemia-reperfusion,I/R)后缺血型胆道病变的发生机制及α-硫辛酸(alpha-lipoic acid,α-LA)对其的防治作用。方法:成年健康雄性SD大鼠90只,随机分为正常对照组(N组)、I/R组、α-LA处理组(LA组),建立全肝缺血再灌注损伤(ischemia-reperfusion injury,IRI)模型。检测血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)、γ-谷氨酰转肽酶(γ-glu-tamyl transferase,GGT)、总胆红素(total bilirubin,TBIL)水平;电子显微镜观察胆小管变化;FITC-Phal-loidin显示胆小管F-actin微丝的分布变化,激光共聚焦显微镜采集图像并定量分析。结果:I/R组血清ALT、AST、GGT和TBIL水平与N组相比明显升高(P<0.05),LA组血清ALT、AST、GGT和TBIL水平与I/R组相比明显降低(P<0.05)。F-actin荧光变化和电子显微镜超微结构改变相符,I/R各组F-actin荧光染色紊乱、减弱,强度明显低于N组(P<0.05);胆小管微绒毛大量脱落,管腔扩张。LA组病理改变较轻,F-actin荧光染色减弱,但程度较I/R组轻,荧光紊乱不甚明显,平均荧光强度高于I/R组(P<0.05);胆小管微绒毛部分脱落,管腔扩张亦不明显。结论:肝脏I/R可造成胆小管F-actin微丝破坏、微绒毛丧失,导致胆小管收缩减弱,胆汁排泄功能受损,这可能是大鼠肝脏I/R后缺血型胆道病变发生的主要机制;α-硫辛酸通过清除氧自由基,降低脂质过氧化反应,调节细胞内氧化剂-抗氧化剂的含量,保护胆小管F-actin微丝结构免遭缺血再灌注损伤破坏,进而预防和治疗缺血型胆道病变。

Objective： To investigate the mechanism of ischemic type biliary lesion after ischemia-reperfusion （I/R）in rat liver and the role of α-Iipoic acid in the prevention and treatment. Methods： Ninety Sprague Dawley rats were divided randomly into three groups： normal control （N）, ischemia reperfusion （I/R） and alpha-lipoic acid pretreatment （LA） groups. The model of rat hepatic ischemia-reperfusion was established. The activity of serum ALT、AST、γ-GGT and TBIL were measured. The changes of bile canaliculus were observed by transmission electron microscope. The modification of F-actin microfilaments was quantified using FITC-Phalloidin and analysised by confocal laser scanning microscopy （CLSM）. Results： The ALT,AST, γ-GGT and TBIL levels of the ischemia reperfusion group was significantly increased （P〈0.05）, while the alpha-lipoic acid pre- treatment group was significantly decreased （P〈0.05） compared with the normal control group. Bile canalicular F-actin was destroyed after reperfusion and this modification of F-actin staining was consistent with the observation by transmission electron microscopy. The staining of F-actin in the ischemia reperfusion group was decreased significantly （P〈0.05） compared with the normal control group. There was a marked loss of canalicular microvilli and distension of canalicular lumen in the ischemia reperfusion group. The staining of F-actin in the LA group was significantly increased than those in the I/R group （P〈0.05）, Conclusioons： Ischemia reperfusion induces an disruption of bile canalicular F-actin microfilaments and a loss of microvilli. Alpha-lipoic acid can protect bile canalicular F-actin microfilaments against IRI.