4-羟基-苯丙噁唑-2-酮对非酒精性脂肪肝病模型大鼠炎症和凋亡信号通路的影响

Effects of 4-hydroxy-2(3H)-benzoxazolone on Inflammatory and Apoptosis Signaling Pathways in Non alcoholic Fatty Liver Disease Model Rats

目的:探讨4-羟基-苯丙噁唑-2-酮对非酒精性脂肪肝病(NAFLD)模型大鼠炎症和凋亡信号通路的影响。方法:将SD大鼠分为正常对照组(10只)和造模组(50只),正常对照组大鼠给予基础饲料,造模组大鼠给予高脂饲料以复制NAFLD模型。造模成功后,将大鼠分为正常对照组、模型组、水飞蓟宾组(26.25 mg/kg)和4-羟基-苯丙噁唑-2-酮高、中、低剂量组(100、50、25 mg/kg),每组8只,正常对照组和模型组大鼠灌胃0.6%羧甲基纤维素钠溶液,其余各组大鼠灌胃相应药物,灌胃体积为10 mL/kg,每天1次,连续4周。末次给药后,检测大鼠血清中白蛋白(ALB)、总蛋白(TP)、球蛋白(GLB)水平,ALB/GLB比值以及游离脂肪酸(FFA)水平;采用TUNEL染色法观察大鼠肝细胞凋亡情况;采用Western blot法检测大鼠肝组织中炎症信号通路相关蛋白[Toll样受体4(TLR4)、髓样分化因子88(MyD88)、核因子κB p65(NF-κB p65)、NF-κB抑制蛋白(IκBα)]的表达水平或磷酸化水平以及凋亡信号通路相关蛋白[B细胞淋巴瘤2(Bcl-2)、Bcl-2相关X蛋白(Bax)、胱天蛋白酶3(caspase-3)]的表达水平。结果:与模型组比较,4-羟基-苯丙噁唑-2-酮各剂量组大鼠血清中TP(低剂量组除外)、GLB、FFA水平和肝组织中TLR4(低剂量组除外)、MyD88(中剂量组除外)、caspase-3蛋白表达水平以及NF-κB p65、IκBα的磷酸化水平均显著降低(P<0.05或P<0.01),血清中ALB/GLB比值和肝组织中Bcl-2/Bax比值均显著升高(P<0.05或P<0.01),肝细胞凋亡现象有所改善。结论:4-羟基-苯丙噁唑-2-酮可改善大鼠NAFLD状态,其作用机制可能与抑制肝组织中TLR4/MyD88/NF-κB信号通路相关蛋白和凋亡相关蛋白表达有关。

OBJECTIVE:To investigate the effects of 4-hydroxy-2(3H)-benzoxazolone on inflammatory and apoptosis signaling pathways in non-alcoholic fatty liver disease(NAFLD)model rats.METHODS:SD rats were divided into normal control group(10 rats)and modeling group(50 rats).Normal control group was given basic diet,and modeling group were given high-fat diet to induce NAFLD model.After modeling,the rats were divided into normal control group,model group,silibinin group(26.25 mg/kg),and 4-hydroxy-2(3H)-benzoxazolone high-dose,medium-dose and low-dose groups(100,50,25 mg/kg),with 8 rats in each group.Normal control group and modeling group were given 0.6%CMC-Na intragastrically,and other groups were given relevant medicine 10 mL/kg intragastrically,once a day,for consecutive 4 weeks.After last medication,the serum levels of albumin(ALB),total protein(TP),globulin(GLB),ALB/GLB and free fatty acid(FFA)were detected;TUNEL staining was used to observe the apoptosis of rat hepatocytes.Western blot assay was used to detect the protein expression and phosphorylation level of inflammatory signaling pathway related proteins[Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),nuclear factor-κB p65(NF-κB p65),NF-κB inhibitor protein(IκBα)]in liver tissue as well as the expression of apoptosis signaling pathway related proteins[B cell lymphoma 2(Bcl-2),Bax,caspase-3].RESULTS:Compared with model group,serum levels of TP(except for low-dose group),GLB and FFA,the protein expression of TLR4(except for low-dose group),MyD88(except for medium-dose group)and caspase-3,the phosphorylation levels of NF-κB p65 and IκBαprotein were decreased significantly(P<0.05 or P<0.01).The ratio of ALB/GLB in serum and the ratio of Bcl-2/Bax in liver tissue were significantly increased(P<0.05 or P<0.01),and the phenomenon of hepatocyte apoptosis was improved.CONCLUSIONS:4-hydroxy-2(3H)-benzoxazolone can ameliorate NAFLD in rats,and the mechanism may be associated with inhibiting the expression TLR4/MyD88/NF-κB signaling pathway-related proteins and apoptosis-related proteins in liver tissues.