目的 探讨姜黄素（curcumin）联合有氧运动对大鼠肝脏脂肪变性的改善作用及其潜在分子机制。方法 50只5 w龄雄性SD大鼠随机等分为对照组（普通饲料+无特殊运动干预,C组）、模型组（高脂饲料+无特殊运动干预,M组）、姜黄素组（高脂饲料+姜黄素+无特殊运动干预,Cu组）、有氧运动组（高脂饲料+有氧运动训练,A组）和联合组（高脂饲料+姜黄素+有氧运动训练,CuA组）。高脂饲料喂养8 w建立肝脏脂肪变性动物模型,高脂饲料喂养2 w后进行6 w姜黄素[ 400 mg/（kg·d）,5 ml/kg ]和/或70%最大摄氧量有氧运动干预。末次运动24 h后处死大鼠,取血液、肝。油红O、苏木素-伊红/染色后光学显微镜下观察肝内脂质沉积情况、肝组织形态及脂肪变性程度,透射电子显微镜下观察肝细胞中自噬体形态特征及数量,全自动生化分析仪检测血清中谷丙转氨酶（alanine aminotransferase,ALT）及谷草转氨酶（aspartate aminotransferase,AST）含量,免疫印迹法检测肝内沉默信息调节因子1（sirtuin1,SIRT1）、叉头盒蛋白O1（forkhead box O1,FoxO1）、乙酰化叉头盒蛋白O1（acetylated forkhead box O1,Ac-FoxO1）、微管相关蛋白1轻链3（microtubule-associated protein 1 light chain 3,LC3）、选择性自噬接头蛋白1（sequestosome 1 protein,P62）、自噬相关蛋白7（autophagy related protein 7,ATG7）蛋白质表达水平。结果 与M组相比,Cu、A、CuA组大鼠体重及血清ALT、AST含量显著降低（P<0.01）;肝细胞内自噬体数量增加,肝内脂质沉积明显改善,肝细胞脂肪变性分级评分、小叶炎症评分和NAFLD活动度评分（NAFLD activity score,NAS）及Ac-FoxO1和P62蛋白质表达水平均显著降低（P<0.05或P<0.01）,SIRT1、ATG7蛋白质表达水平、LC3-II/LC3-I蛋白质表达比值均显著升高（P<0.05或P<0.01）,FoxO1蛋白质表达水平无显著变化（P>0.05）;CuA组大鼠Ac-FoxO1/FoxO1蛋白质表达水平比值显著降低（P<0.01）,Cu、A组大鼠无显著差异（P>0.05）。2×2析因方差分析显示,姜黄素与有氧运动对血清ALT、AST含量、肝细胞脂肪变性分级评分和NAS、肝内P62、ATG7、SIRT1、Ac-FoxO1蛋白质表达水平均具有显著交互效应。结论 姜黄素联合有氧运动可通过激活SIRT1信号通路,促进FoxO1脱乙酰化,诱导自噬,改善大鼠肝脏脂肪变性和肝损伤,且二者具有协同效应,联合干预效果更佳。
Abstract
Objective To explore the effects of curcumin combined with aerobic exercise on hepatic steatosis in rats and the possible mechanisms. Methods Fifty 5 w old male SD rats were randomly divided into control group (regular diet+no special exercise intervention, group C), model group (high-fat diet+no special exercise intervention, group M), curcumin group (high-fat diet+curcumin+no special exercise intervention, group Cu), aerobic exercise group (high-fat diet+aerobic exercise, group A), and combination group (high-fat diet+curcumin+aerobic exercise, group CuA). Animal model of hepatic steatosis was established by feeding a high-fat diet for 8 weeks. After two weeks of high-fat diet feeding, oral administration of curcumin (400 mg/(kg · d), 5 ml/kg) and/or 70% VO2max aerobic exercise were carried out for six weeks. Rats were euthanized 24 hours after the last exercise, and blood and liver samples were collected. After staining with Oil Red O and Hematoxylin Eosin, observation of lipid deposition, liver tissue morphology, and degree of lipid degeneration in the liver under an optical microscope, observation of autophagosome morphology and quantity in liver cells under a transmission electron microscope. The serum alanine transaminase (ALT) and aspartate transaminase (AST) contents were detected by a biochemical analyzer. Western blotting was used to determine the protein levels of sirtuin1 (SIRT1), forkhead box O1 (FoxO1), acetylated forkhead box O1 (Ac-FoxO1), microtubule-associated protein 1 light chain 3 (LC3), sequestosome 1 protein (P62) and autophagy related protein 7 (ATG7). Results Compared with the M group, the body weight and serum ALT and AST levels of the rats in the Cu, A, and CuA groups were significantly reduced (P<0.01). The number of autophagosomes in the liver was increased, with a significant improvement in lipid deposition in the liver. Hepatic steatosis grading score, lobular inflammation score and the NAFLD activity score (NAS), Ac-FoxO1 and P62 protein expression levels were significantly reduced (P<0.05 or P<0.01). The protein expression levels of SIRT1 and ATG7, as well as the LC3-II/LC3-I protein expression ratio, were significantly increased (P<0.05 or P<0.01), while the protein expression level of FoxO1 showed no significant change (P>0.05). The ratio of Ac-FoxO1/FoxO1 protein expression was significantly reduced in the CuA group (P<0.01), while there was no significant difference in the Cu and A groups (P>0.05).The 2×2 factorial analysis showed that curcumin and aerobic exercise had significant interactive actions on serum ALT and AST levels, hepatic steatosis grading score and NAS scores, as well as liver P62, ATG7, SIRT1, Ac-FoxO1 protein expression. Conclusion Curcumin combined with aerobic exercise can improve liver steatosis and liver injury by promoting FoxO1 deacetylation via activating SIRT1 signaling pathway, inducing autophagy in rats, and the two have a synergistic effect, with joint intervention having a better effect.
关键词
姜黄素 /
有氧运动 /
肝脏脂肪变性 /
沉默信息调节因子1(SIRT1) /
叉头盒蛋白O1(FoxO1) /
自噬
Key words
curcumin /
aerobic exercise /
hepatic steatosis /
sirtuin1 (SRIT1) /
forkhead box O1 (FoxO1) /
autophagy
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基金
北京联合大学科研项目（No.ZK20202503）
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