扶正养肝合剂抑制NLRP3炎症小体活化缓解肝损伤作用

Fuzheng Yanggan Fomula alleviates liver injury through inhibiting NLRP3 inflammasome

本研究采用痤疮丙酸杆菌(Propionibacterium acnes,P.acnes)和脂多糖(lipopolysaccharide,LPS)诱导小鼠急性肝损伤模型,研究扶正养肝合剂(Fuzheng Yanggan Fomula,FYF)缓解急性肝损伤的作用及机制。通过建立P.acnes/LPS小鼠急性肝损伤模型,考察FYF对小鼠血清中谷丙转氨酶(alanine transaminase,ALT)、谷草转氨酶(aspartate aminotransferase,AST)和白细胞介素-1β(interleukin-1β,IL-1β)含量,以及肝组织中丙二醛(malonaldehyde,MDA)、抗氧化能力指数(oxygen radical absorbance capacity,ORAC)和谷胱甘肽(glutathione,GSH)水平;应用免疫印迹法检测肝脏Nod样受体蛋白3(Nod-like receptor protein 3,NLRP3)炎症小体相关通路中NLRP3蛋白、半胱天冬酶募集结构域的凋亡相关斑点样蛋白(apoptosis-associated speck-like protein containing a CARD,ASC)、天冬氨酸特异性半胱氨酸蛋白酶-1(cysteinyl aspartate specific proteinase-1,caspase-1)和IL-1β蛋白水平;应用苏木素-伊红(hematoxylinendash-eosin,HE)染色和免疫组化实验方法检测肝脏切片病理学变化和炎症情况;应用末端脱氧核苷酸转移酶标记(terminal deoxynucleotidyl transferase dUTP nick end labeling,TUNEL)方法检测肝脏切片细胞凋亡情况。动物福利和实验过程均遵循暨南大学动物伦理委员会的规定。实验结果显示,FYF能够缓解P.acnes/LPS诱导的急性肝损伤小鼠肝组织病理损伤和炎症细胞浸润,降低其血清中ALT、AST和IL-1β含量,同时抑制MDA产生,升高ORAC及GSH水平,下调caspase-1和IL-1β蛋白水平。上述研究结果表明,FYF通过抑制NLRP3炎症小体通路的激活,缓解P.acnes/LPS诱导的小鼠急性肝损伤,可为肝损伤疾病的预防和治疗提供理论依据和新的药物作用靶点。

In this study,the model of Propionibacterium acnes/lipopolysaccharide(P.acnes/LPS)-induced acute liver injury in mice was employed to investigate the protective effects of Fuzheng Yanggan Fomula(FYF)on acute liver injury.The effects of FYF on the contents of alanine aminotransferase(ALT),aspartate aminotransferase(AST),and interleukin-1β(IL-1β)in the serum,and the levels of malondialdehyde(MDA),oxygen radical absorbance capacity(ORAC),and glutathione(GSH)were examined in the livers of mice treated with P.acnes/LPS;The protein expression levels of Nod-like receptor protein 3(NLRP3),apoptosis-associated speck-like protein containing a CARD(ASC),cysteinyl aspartate specific proteinase-1(caspase-1),and IL-1βin liver tissues were detected by Western blot;Furthermore,hematoxylinendash-eosin(HE)staining,terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)staining,and immunohistochemical assay were used to observe pathological changes,apoptosis index,and inflammation infiltration of the liver tissue sections.All animal welfare and experimental procedures were followed by the Animal Ethics Committee of Jinan University.We conclude that FYF could alleviate P.acnes/LPS induced pathological damage and inflammatory infiltration in the liver of mice.Meanwhile,FYF decreases the contents of ALT,AST,IL-1β,and MDA,increases the contents of ORAC and GSH,and downregulates the expression of caspase-1 and IL-1βproteins.Collectively,these findings suggested that FYF could alleviate P.acnes/LPS induced acute liver injury in mice by inhibiting the activation of NLRP3 inflammasome,which provides a theoretical basis and a new drug target for the prevention and treatment of liver injury.