大黄素通过减轻氧化应激和肝细胞凋亡对脂多糖诱导急性肝损伤的保护机制研究＊

目的 观察大黄素对脂多糖（lipopolysaccharide,LPS）诱导的急性肝损伤大鼠的影响,并探讨其可能的作用机制。方法 40只雄性清洁级SD大鼠随机分为空白组（Control）,模型组（Model）、大黄素低浓度组（Emodin L）,大黄素高浓度组（Emodin H）,每组10只。除空白组,其他组均腹腔注射LPS建立急性肝损伤模型,造模后四组均给予正常食水,另大黄素低浓度组给予浓度为20 mg·kg^-1的大黄素（ig,bid）,大黄素高浓度给予浓度为40 mg·kg^-1的大黄素（ig,bid）,空白组及模型组给予等体积蒸馏水（ig,bid）,苏木精-伊红（HE）染色观察大鼠肝组织病理变化,大鼠血清中天冬氨酸转氨酶（AST）、丙氨酸转氨酶（ALT）的变化,大鼠肝脏中氧化应激指标谷胱甘肽过氧化物酶（GSH-Px）、超氧化物歧化酶（SOD）和丙二醛（MDA）,运用脱氧核糖核苷酸末端转移酶介导的缺口末端标记法（TUNEL）检测肝细胞凋亡情况,免疫荧光标记法（IF）观察促凋亡相关因子半胱氨酸天冬氨酸蛋白酶-3（Caspase-3）的表达。免疫印迹试验（Western Blot）检测抗凋亡因子中B淋巴细胞瘤-2基因Bcl-2、促凋亡因子中抑癌基因p53的蛋白表达。结果 与Control组相比,LPS腹腔注射可以导致大鼠肝脏病理性损伤和肝功能异常,致使肝脏氧化应激明显增强,肝细胞凋亡数量明显增多,促凋亡因子半胱氨酸天冬氨酸蛋白酶-3（Caspase-3）分泌明显增加,抗凋亡因子B细胞淋巴瘤-2（Bcl-2）蛋白表达受到抑制,促凋亡因子肿瘤抑制基因（p53）蛋白表达明显增强;而与模型组相比,大黄素组可以改善肝功能水平及肝脏病理变化,调节肝脏氧化应激,减轻肝细胞凋亡,减少半胱氨酸天冬氨酸蛋白酶-3（Caspase-3）的分泌,增强抗凋亡因子Bcl-2的蛋白表达,降低促凋亡因子p53的蛋白表达。结论 大黄素对LPS诱导的急性肝损伤起保护作用,其作用机制可能通过调节氧化应激和减轻肝细胞凋亡途径来实现。

Protective Mechanism of Emodin on Lipopolysaccharide-induced Acute Liver Injury by Reducing Oxidative Stress and Hepatocyte Apoptosis

Objective To observe the effect of rhubarb on lipopolysaccharide (LPS)-induced acute liver injury and explore its possible mechanism.Methods Forty male SD rats were randomly divided into blank groups (Control), Model group (Model), emodin low concentration group (Emodin L), Emodin high concentration group (Emodin H), with 10 in each group. Except for the control group, all the other groups were injected intraperitoneally LPS, and model of acute liver injury was established, All 4 groups were given normal fresh water. The low concentration groups were given 20 mg·kg^-1 of emodin (ig, bid). The high concentration group of emodin was given emodin with a concentration of 40 mg·kg^-1 (ig, bid). Equal volume distilled water (ig, bid) were given in Control and Model groups. The pathological changes of liver tissue in rats, rat liver function level (AST, ALT), oxidative stress indicators glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and malondialdehyde (MDA) were observed by HE staining. TUNEL was used to detect hepatocyte apoptosis. The expression of apoptosis-related factor caspase-3 was observed by immunofluorescence (IF). Immunoblotting test (Western Blot) was used to detect the expression of tumor suppressor gene p53 in B lymphoma-2 gene Bcl-2, apoptosis promoting factor in anti-apoptotic factor.Results Compared with control group, LPS intraperitoneal injection can lead to liver pathological injury and abnormal liver function in rats, resulting in increased oxidative stress, increased apoptosis of hepatocytes and increased secretion caspase-3 pro-apoptotic factors. The expression of anti-apoptotic factors bcl-2 protein was inhibited, and the expression of pro-apoptotic factors p53 protein was significantly enhanced; Compared with the model group, emodin group can improve liver function and liver pathological changes, regulate liver oxidative stress, reduce hepatocyte apoptosis, reduce caspase-3 secretion, enhance protein expression bcl-2 anti-apoptotic factor, and reduce protein expression p53 pro-apoptotic factor.Conclusion Emodin plays a protective role in LPS induced acute liver injury, and its mechanism may be achieved by regulating oxidative stress and reducing hepatocyte apoptosis.