竹节参总皂苷缓解CCl_(4)诱导的大鼠急性肝损伤:基于调控PI3K/Akt/NF-κB信号通路

Total saponins of Panax japonicus alleviates CCl_(4)-induced acute liver injury in rats by regulating the PI3K/AktNF-κB signaling pathway

目的探讨土家族药用植物竹节参提取物总皂苷对CCl_(4)致急性肝损伤的保护作用及潜在的药理学机制。方法将6周龄SPF级雄性SD大鼠随机分为正常组、模型组、联苯双酯组(100 mg/kg)、竹节参总皂苷低、中、高(50、100、200 mg/kg)剂量组,各组8只,除空白组外,其余各组采用CCl_(4)诱导大鼠急性肝损伤模型,处理组于造模中给予药物灌胃干预。比较各组大鼠的血清谷草转氨酶(AST)、谷丙转氨酶(ALT)、总胆红素(TBil)和碱性磷酸酶(ALP)水平;HE染色观察肝组织病理学改变;免疫组化检测肝脏组织PI3K/Akt/NF-κB信号通路相关分子的表达;酶联免疫法测定肝脏组织总超氧化物歧化酶(T-SOD)、谷胱甘肽过氧化物酶(GSH-Px)、丙二醛(MDA)水平;蛋白免疫印迹法检测肝脏组织PI3K-Akt和SIRT6-NF-κB通路相关蛋白表达情况。结果网络药理学分析显示,竹节参总皂苷对急性肝损伤的有治疗作用,其关键的通路为PI3K/Akt等信号通路。血清学和酶联免疫学实验结果显示,与正常组相比,模型组大鼠血清和肝组织的AST、ALT、ALP、TBil和MDA明显增高(P<0.01),T-SOD和GSHPx水平显著降低(P<0.01);与模型组相比,各治疗组ALT、AST、ALP、TBIL和MDA水平显著降低(P<0.01),T-SOD和GSH-Px水平显著升高(P<0.01)。免疫组化结果显示,正常组大鼠肝组织细胞内未见p-NF-κB表达阳性,与正常组相比,模型组中p-NF-κB阳性表达则明显增加,阳性细胞数增多(P<0.01);与模型组比较,各治疗组阳性表达明显减少(P<0.01)。免疫印迹结果显示,相对于正常组,模型组PI3K、p-Akt蛋白的表达水平下降,p-NF-κB、TNF-α和IL-6蛋白的表达水平升高(P<0.05);与模型组相比,各治疗组PI3K、p-Akt和SIRT6蛋白的表达水平显著升高,p-NF-κB p65、TNF-α和IL-6的蛋白的表达水平显著降低(P<0.05)。结论竹节参总皂苷可以通过调节PI3K/Akt和NF-κB通路,有效减缓CCl_(4)诱导的大鼠急性肝损伤,发挥其抗炎、抗氧化应激和保护肝损伤的作用。

Objective To investigate the protective effect of total saponins of Panax japonicus(TSPJ)against CCl_(4)-induced acute liver injury(ALI)in rats and explore the underlying pharmacological mechanisms.Methods Male SD rat models of CCl_(4)-induced ALI were given intraperitoneal injections of distilled water,100 mg/kg biphenyl bisabololol,or 50,100,and 200 mg/kg TSPJ during modeling(n=8).Liver functions(AST,ALT,TBil and ALP)of the rats were assessed and liver pathologies were observed with HE staining.Immunohistochemistry was used to detect the expressions of PI3K/Akt/NF-κB signaling pathway molecules in liver tissue;ELISA was used to determine the levels of T-SOD,GSH-Px,and MDA.Western blotting was performed to detect the expression levels of PI3K-Akt and SIRT6-NF-κB pathways in the liver tissue.Results Network pharmacological analysis indicated that the key pathways including PI3K/Akt mediated the therapeutic effect of TSPJ on ALI.In the rat models of ALI,treatments with biphenyl bisabololol and TSPJ significantly ameliorated CCl_(4)-induced increase of serum levels AST,ALT,ALP,TBil and MDA and decrease of T-SOD and GSH-Px levels(all P<0.01).The rat models of ALI showed significantly increased expression of p-NF-κB(P<0.01),decreased expressions of PI3K,p-Akt and SIRT6 proteins,and elevated expression levels of p-NF-κB,TNF-αand IL-6 proteins in the liver,which were all significantly improved in the treatment groups(P<0.05 or 0.01).Conclusion TSPJ can effectively alleviate CCl_(4)-induced ALI in rats by suppressing inflammatory responses and oxidative stress in the liver via regulating the PI3K/Akt and SIRT6/NF-κB pathways.