圣草酚调控MAPK和Nrf2/HO-1信号通路缓解非酒精性脂肪肝的作用及机制

Effect and mechanism of eriodictyol on non-alcoholic fatty liver disease by regulating MAPK and Nrf2/HO-1 signaling pathway

目的研究圣草酚缓解非酒精性脂肪肝(NAFLD)的作用及可能机制。方法将16只C57BL/6J小鼠按体重随机分为对照组、NAFLD模型组和圣草酚低、高剂量组(50、100 mg/kg),每组4只。除对照组外,其余各组均使用高脂饲料诱导NAFLD模型。在预处理4周后,采取腹腔注射方式(0.01 mL/g)进行给药,每日1次,连续6周。测定小鼠体重、肝脏质量,观察小鼠肝组织病理损伤情况,测定小鼠血清中天冬氨酸转氨酶(AST)、丙氨酸转氨酶(ALT)和甘油三酯(TG)水平及肝组织中核转录因子红系2相关因子2(Nrf2)、血红素加氧酶1(HO-1)蛋白表达。使用0.5 mmol/L油酸诱导HepG2细胞建立体外NAFLD模型,实验设置正常对照组、NAFLD模型组和圣草酚低、中、高浓度组(50、100、150μmol/L);给药组在诱导模型的同时加入圣草酚,培养24 h。观察细胞中脂质沉积情况,检测细胞中TG、丙二醛(MDA)、活性氧(ROS)水平以及丝裂原活化蛋白激酶(MAPK)信号通路相关蛋白[细胞外信号调节激酶(ERK)、c-Jun氨基末端激酶(JNK)]磷酸化水平和Nrf2、HO-1蛋白表达水平。结果体内实验中,与NAFLD模型组比较,圣草酚低、高剂量组小鼠体重、肝脏质量和血清中AST、ALT和TG的水平(圣草酚低剂量组小鼠血清AST水平除外)均显著降低(P<0.01),脂滴沉积形成的空泡数量显著减少,Nrf2、HO-1蛋白表达水平进一步升高(P<0.01)。体外实验中,与NAFLD模型组比较,圣草酚低、中、高浓度组细胞中脂质沉积减少(P<0.01),ROS、MDA、TG水平显著降低(P<0.05或P<0.01),ERK、JNK磷酸化水平均显著下调(P<0.01),Nrf2、HO-1蛋白表达水平显著上调(P<0.01)。结论圣草酚可能通过抑制MAPK信号通路,进而激活Nrf2/HO-1信号通路来缓解NAFLD。

OBJECTIVE To study the effect and potential mechanism of eriodictyol on non-alcoholic fatty liver disease(NAFLD).METHODS Sixteen C57BL/6J mice were randomly divided into control group,NAFLD model group,and eriodictyol low-dose and high-dose groups(50,100 mg/kg),with 4 mice in each group.Except for control group,the other groups were fed with high fat diet to induce NAFLD model.After four weeks of preprocessing,they were given relevant medicine intraperitoneally(0.01 mL/g),once a day,for 6 consecutive weeks.The body weight and liver weight of mice were measured,and the pathological damage of liver tissue in mice was observed.The levels of aspartate aminotransferase(AST),alanine aminotransferase(ALT),and triglycerides(TG)in serum,as well as the protein expressions of nuclear factor-erythroid 2-related factor 2(Nrf2)and heme oxygenase-1(HO-1)in liver tissue were determined.In vitro NAFLD model was established by using 0.5 mmol/L oleic acid(OA)in HepG2 cells.Normal control group,NAFLD model group and eriodictyol low-,mediumand high-concentration groups(50,100,150μmol/L)were set up. HepG2 cells in drug groups were treated with eriodictyol for 24 h at the time of modeling. The lipid deposition was observed in cells, and the levels of TG, malondialdehyde (MDA) and reactive oxygen species (ROS) as well as the phosphorylation levels of the mitogen-activated protein kinase (MAPK) signal pathway related proteins [extracellular signal-regulated kinase (ERK), c- Jun N-terminal kinase (JNK)] and the protein expressions of Nrf2 and HO-1 were all determined. RESULTS In the in vivo experiment, compared with the NAFLD model group, the body weight, liver weight, the serum levels of AST, ALT and TG were all decreased significantly in eriodictyol low- and high-dose groups (except for serum level of AST in eriodictyol low-dose group) (P<0.01);liver lipid deposition was reduced significantly and the protein expressions of Nrf2 and HO-1 in liver tissues were further up-regulated (P<0.01). In the in vitro experiment, compared with the NAFLD model group, the lipid deposition in hepatocytes was reduced in eriodictyol low-, medium- and high-concentration groups (P<0.01), and the levels of ROS, MDA and TG were down-regulated (P<0.05 or P<0.01);the phosphorylation levels of ERK and JNK were significantly down-regulated (P<0.01), while the protein expressions of Nrf2 and HO-1 were up-regulated significantly (P<0.01). CONCLUSIONS Eriodictyol can inhibit MAPK signaling pathway and activate Nrf2/HO-1 signaling pathway to alleviate NAFLD.