解毒护肝方对药物性肝损伤大鼠TLR3/TNF-α/JNK2信号通路的影响

Effects of Jiedu Hugan Prescription on TLR3/TNF-α/JNK2 Signaling Pathway of Drug-induced Hepatotoxicity Rats

目的观察解毒护肝方对药物性肝损伤大鼠TLR3/TNF-α/JNK2信号通路的影响。方法采用对乙酰氨基酚灌胃制作大鼠肝损伤模型。将实验大鼠按体质量随机分为正常组、模型组、阳性对照组和解毒护肝方高、中、低剂量组,各给药组给予相应药液灌胃,正常组和模型组给予生理盐水灌胃。生化分析仪检测血清AST、ALT活性和总胆红素(TBIL)、直接胆红素(DBIL)含量;RT-PCR和Western blot分别检测肝组织JNK2基因和蛋白的表达;酶标仪检测肝组织匀浆肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)含量;免疫组化染色观察Toll样受体3(TLR3)蛋白表达。结果与正常组比较,模型组大鼠ALT、AST、DBIL、TNF-α水平显著升高(P<0.01),JNK2 mRNA和p-JNK2、TLR3蛋白表达显著升高(P<0.01);与模型组比较,解毒护肝方中、低剂量组大鼠AST、ALT活性显著降低(P<0.05,P<0.01),解毒护肝方低剂量组大鼠DBIL含量显著降低(P<0.05),解毒护肝方高剂量组大鼠TNF-α含量显著降低(P<0.01),解毒护肝方高、中剂量组大鼠JNK mRNA和TLR3蛋白表达均显著下调(P<0.05,P<0.01),p-JNK2蛋白表达有降低趋势,但差异无统计学意义(P>0.05)。结论解毒护肝方对药物性肝损伤大鼠有明显的保护作用,以中、低剂量组效果最为明显,其机制可能与调控TLR3/TNF-α/JNK2信号通路有关。

Objective To observe the effects of Jiedu Hugan Prescription on TLR3/TNF-α/JNK2 signaling pathway of acetaminophen-induced hepatotoxicity rats. Methods Hepatotoxicity rat model was prepared by intragastric administration of acetaminophen. The rats were randomly divided into normal group, model group,positive control group, Jiedu Hugan Prescription high-, medium-and low-dosage groups. Each administration group was given relevant medicine infusions for gavage, normal group and model group were given normal saline for gavage. At the end of the experiment, the levels of AST, ALT, TBIL and DBIL were detected by biomedical analyzer.JNK2 gene and protein expression in liver tissue were detected by RT-PCR and Western blot. The contents of TNF-αand IL-6 in liver tissue homogenate were detected by enzyme microplate reader. The expression of Toll-like receptor 3(TLR3) protein was observed by immunohistochemistry. Results Compared with the normal group, the levels of ALT,AST, DBIL and TNF-α in the model group significantly increased(P<0.01), and the expressions of JNK2 m RNA,p-JNK2, and TLR3 protein significantly increased(P<0.01). Compared with the model group, the activities of AST and ALT in Jiedu Hugan Prescription medium-and low-dosage groups significantly decreased(P<0.05, P<0.01);the content of DBIL level in Jiedu Hugan Prescription low-dosage group significantly decreased(P<0.05);the content of TNF-α in Jiedu Hugan Prescription high-dosage group significantly decreased(P<0.01). The expressions of JNK mRNA and TLR3 protein in Jiedu Hugan Prescription high-and medium-dosage groups were significantly down-regulated(P<0.05, P<0.01), and the expression of p-JNK2 protein decreased, without statistical significance(P>0.05). Conclusion Jiedu Hugan Prescription has obvious protective effects on rats with drug-induced hepatotoxicity, and the effects are most obvious in the medium-and low-dosage groups. The mechanism may be related to the regulation of TLR3/TNF-α/JNK2 signaling pathway.