基于HMGB1/TLR4/NF-κB信号通路探讨忍冬苷对脓毒症肝损伤大鼠的影响

Study on Effect of Lonicerins on Sepsis Liver Injury in Rats Based on the HMGB1/TLR4/NF-κB Signaling Pathway

目的:基于高迁移率族蛋白B1(HMGB1)/Toll样受体4(TLR4)/核因子κB(NF-κB)信号通路探讨忍冬苷对脓毒症肝损伤大鼠的影响。方法:60只雄性SD大鼠随机分为模型组、对照组、阳性对照组(地塞米松10 mg/kg)、忍冬苷低剂量(7.5 mg/kg)、忍冬苷中剂量(15 mg/kg)、忍冬苷高剂量(30 mg/kg)组,每组10只。采用盲肠结扎穿刺法建立大鼠脓毒症模型。实验结束后麻醉大鼠取血制备血清,检测血清中超氧化物歧化酶(SOD)活性和丙二醛(MDA)、白介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、白介素-10(IL-10)含量;分离肝组织称重,计算肝脏指数,一部分用于HE染色观察肝组织病理变化,一部分用于制备组织匀浆检测组织中肝功能指标谷丙转氨酶(ALT)、天冬氨酸转氨酶(AST)活性及HMGB1、TLR4、NF-κB蛋白表达。结果:与对照组比较,模型组大鼠血清中MDA、TNF-α、IL-6含量、肝脏指数以及肝组织中AST、ALT活性、HMGB1、TLR4、NF-κB蛋白表达显著增加(P<0.05),SOD活性与IL-10含量显著下降(P<0.05);且肝组织出现明显病灶,肝细胞水肿变性,大量炎性细胞浸润。与模型组比较,阳性对照组与忍冬苷各剂量组大鼠血清中MDA、TNF-α、IL-6含量、肝脏指数以及肝组织中AST、ALT活性、HMGB1、TLR4、NF-κB蛋白表达显著降低(P<0.05),SOD活性与IL-10含量显著升高(P<0.05);忍冬苷低、中剂量组仍可见病灶和水肿,但病变减轻;地塞米松组与忍冬苷高剂量组肝细胞结构趋于正常,未发现病灶。与阳性对照组比较,忍冬苷低、中剂量组大鼠血清中MDA、TNF-α、IL-6含量、肝脏指数以及肝组织中AST、ALT活性、HMGB1、TLR4、NF-κB蛋白表达显著升高(P<0.05),SOD活性与IL-10含量显著降低(P<0.05);忍冬苷高剂量组上述指标无显著差异(P>0.05)。结论:忍冬苷通过下调HMGB1/TLR4/NF-κB信号通路的表达,抑制氧化应激,减轻炎症反应,改善肝功能,发挥对脓毒症肝损伤的保护作用。

Objective:To investigate the effect of lonicerin on septic liver injury in rats based on the high mobility group protein B1(HMGB1)/Toll-l-like receptor 4(TLR4)/nuclear factor-κB(NF-κB)signaling pathway.Methods:60 male SD rats were divided into model group,control group,positive control group(dexamethasone 10 mg/kg),lonicerin low dose(7.5 mg/kg),lonicerin middle dose(15 mg/kg),lonicerin high dose(30 mg/kg)groups,each group had 10 rats.The cecal ligation and puncture method was used to establish a rat sepsis model.After the experiment,the rats were anesthetized and blood was collected to prepare serum,and the superoxide Dismutase(SOD)activity and the contents of malondialdehyde(MDA),interleukin-6(IL-6),tumor necrosis factorα(TNF-α),interleukin-10(IL-10)in the serum were detected.The liver tissues were separated and weighed,the liver index was calculated,one part was used for HE staining to observe the pathological changes of liver tissue,and the other part was used to prepare tissue homogenate to detect the activities of liver function indexes alanine transaminase(ALT),aspartate Transaminase(AST)and the protein expression of HMGB1,TLR4 and NF-κB in the tissue.Results:Compared with the control group,the serum MDA,TNF-α,IL-6 contents,liver index,and liver tissue AST,ALT activity,HMGB1,TLR4 and NF-κB protein expression were increased significantly in the model group(P<0.05),the SOD activity and IL-10 content decreased significantly(P<0.05).Obvious lesions appeared in liver tissue,hepatocytes were edema and degeneration,and a large number of inflammatory cells infiltrated.Compared with the model group,the serum MDA,TNF-α,IL-6 contents,liver index,and liver tissue AST,ALT activity,HMGB1,TLR4,and NF-κB protein expression in the positive control group and the lonicerin each dose groups decreased significantly(P<0.05),the SOD activity and IL-10 content increased significantly(P<0.05).The lesions and edema were still seen in the lonicerin low and middle dose groups,but the lesions were alleviated.The liver cell structure of the positive control group and the lonicerin high dose group tended to be normal,and no lesions were found.Compared with the positive control group,the serum MDA,TNF-α,IL-6 contents,liver index,and liver tissue AST,ALT activity,HMGB1,TLR4,and NF-κB protein expression in the lonicerin low and middle dose groups increased significantly(P<0.05),the SOD activity and IL-10 content reduced significantly(P<0.05).There were no significant differences in the above indicators in the lonicerin high dose group(P>0.05).Conclusion:Lonicerin can down-regulate the HMGB1/TLR4/NF-κB signaling pathway,reduce inflammation reaction and oxidative stress,improve liver function,and play a protective effect on septic liver injury.