摘要
目的 探讨丹皮酚基于Kelch样环氧氯丙烷相关蛋白 1(Kelch-like ECH-associated protein 1,Keap1)/核因子 E2 相关因子 2(nuclear factor erythroid 2-related factor 2,Nrf2)/抗氧化反应元件(antioxidant response element,ARE)信号通路改善乙醇刺激所致小鼠肝、脑炎症和氧化应激损伤的作用机制.方法 将 C57BL/6 小鼠随机分为空白组,模型组,水飞蓟宾组(36.8 mg/kg),丹皮酚高、中、低(480、240、120 mg/kg)剂量组,每组 15 只;适应期各组小鼠自由饮用 Liber-DeCarli对照液体饲料,模型复制期空白组小鼠自由饮用 Lieber-DeCarli对照液体饲料,其他组小鼠自由饮用 Lieber-DeCarli乙醇液体饲料并连续灌胃相对应的药液 10d;测定小鼠血脂[三酰甘油(triglyceride,TG)、总胆固醇(total cholesterol,TC)]、肝功能[丙氨酸氨基转移酶(alanine aminotransferase,ALT)、天冬氨酸氨基转移酶(aspartate aminotransferase,AST)]、炎症因子[白细胞介素(interleukin,IL)-6、IL-1α、IL-1β、肿瘤坏死因子(tumor necrosis factor,TNF)-α)]以及氧化应激指标[过氧化氢酶(catalase,CAT)、还原型谷胱甘肽(glutathione,GSH)、超氧化物歧化酶(super oxide dismutase,SOD)、丙二醛(malondialdehyde,MDA)]水平;采用苏木精—伊红染色法、油红 O染色观察各组小鼠肝、脑组织形态变化;采用 Western blot法、免疫荧光法以及 qRT-PCR法检测小鼠肝、脑组织Keap1/Nrf2/ARE信号通路相关蛋白及 mRNA表达水平.结果 与模型组比较,丹皮酚高剂量组小鼠体质量显著增加(P＜0.05),血脂(TG、TC)、肝功能(ALT、AST)、炎症因子(IL-6、IL-1α、IL-1β、TNF-α)、氧化应激指标(CAT、GSH、SOD)水平显著升高(P＜0.05),MDA含量显著降低(P＜0.05);小鼠肝、脑组织 Nrf2、血红素氧合酶-1、醌 NADH脱氢酶 1、谷氨酸—半胱氨酸连接酶催化亚基蛋白及 mRNA表达水平显著升高(P＜0.05),Keap1 蛋白及 mRNA表达水平显著降低(P＜0.05);丹皮酚高剂量组小鼠肝、脑组织病理状态明显改善.结论 丹皮酚能显著减轻乙醇诱导的小鼠肝、脑炎症和氧化应激损伤,其机制可能是通过调控 Keap1/Nrf2/ARE信号通路实现的.
Abstract
Objective To investigate the mechanism of action of paeonol improving liver and brain inflammation and oxidative stress injury induced by acute alcohol stimulation in mice based on the on Kelch-like ECH-associated protein 1(Keap1)/nuclear factor erythroid 2-related factor 2(Nrf2)/antioxidant response element(ARE)signaling pathway.Methods C57BL/6 mice were randomly divided into blank group,model group,silybin group(36.8 mg/kg),and high-,middle-,and low-dose paeonol groups(480,240,and 120 mg/kg),with 15 mice in each group.During the adaptive phase,the mice in all groups were given Lieber-DeCarli control liquid feed freely;during modeling,the mice in the blank group were given Lieber-DeCarli control liquid feed freely,and those in the other groups were given Lieber-DeCarli alcohol liquid feed freely and the corresponding drug by ga-vage for 10 days.The mice were measured in terms of blood lipids[triglyceride(TG)and total cholesterol(TC)],liver func-tion[alanine aminotransferase(ALT)and aspartate aminotransferase(AST)],inflammatory factors[interleukin-6(IL-6),in-terleukin-1α(IL-1α),interleukin-1β(IL-1β),and tumor necrosis factor-α(TNF-α)],and oxidative stress indicators[catalase(CAT),glutathione(GSH),superoxide dismutase(SOD),and malondialdehyde(MDA)];HE staining and oil red O staining were used to observe the pathological changes of the liver and brain;Western blot,immunofluorescence assay,and qRT-PCR were used to measure the expression levels of Keap1/Nrf2/ARE signaling pathway-related proteins and their mRNA expression levels in mouse liver and brain.Results Compared with the model group,the high-dose paeonol group had a significant increase in body weight(P＜0.05),significant increases in blood lipids(TG and TC),liver function parameters(ALT and AST),in-flammatory factors(IL-6,IL-1α,IL-1β,and TNF-α),and oxidative stress indicators(CAT,GSH,and SOD)(P＜0.05),and a significant reduction in the content of MDA(P＜0.05);there were significant increases in the protein and mRNA expression levels of heme oxygenase-1,NAD(P)H quinone oxidoreductase 1,and glutamate-cysteine ligase catalytic subunit and significant reductions in the protein and mRNA expression levels of Keap1 in mouse liver and brain(P＜0.05);the high-dose paeonol group showed significant improvements in the pathological state of the mouse liver and brain.Conclusion Paeonol can signifi-cantly alleviate alcohol-induced liver and brain inflammation and oxidative stress injury in mice,possibly by regulating the Keap1/Nrf2/ARE signaling pathway.
基金项目
安徽省高等学校自然科学研究项目(2022AH050521)
安徽省自然科学基金(2008085MH300)
维普
万方数据