青苹果对高脂模型雄性大鼠AMPK-ACC信号通路的作用
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摘要
探究青苹果汁对高脂模型雄性大鼠AMPK-ACC信号通路的影响。将雄性SD大鼠70只随机抽取20只做空白对照组用基础饲料喂养。其余用高脂高糖饲料建立高脂模型,分别在第4周和第8周用ELISA法检测血清指标,与空白组比较,在确定高脂模型建立成功后随机分为5组:阴性对照组(高脂模型组),阳性对照组,青苹果高、中、低剂量组,每组10只。各组动物按各自药物和剂量灌胃给药,其中阳性对照组给予白藜芦醇0.1mg/10g,高、中、低剂量青苹果组分别给予0.3mL/10g、0.2mL/10g、0.1mL/10g剂量青苹果汁,空白对照组和阴性对照组给予0.1mL/10g蒸馏水灌胃。1次/d,连续给药8周后处死动物,采用RT-PCR检测各大鼠肝组织AMPK、ACCmRNA的表达,Westernblotting检测AMPK、ACC活性表达水平。与高脂模型(阴性对照)组比较,高、中剂量青苹果组AMPKmRNA与AMPK蛋白表达增强而ACCmRNA与ACC蛋白表达减弱且有统计学意义(p<0.05)。高、中剂量青苹果可通过激活AMPK-ACC信号通路,进而刺激脂肪酸的氧化,最终达到降脂效应。
To investigate the effect of green apple juice on AMPK-ACC signaling pathway in male rats with hyperlipidemia. 70 healthy SD male rats were enrolled in this study, of which 20 rats were randomly selected as blank control group fed with basal diet and the other 50 rats were fed with high fat and high sugar diet to establish hyperlipidemia model. Blood serum indexes including total cholesterol(TC) and triglyceride(TG) were detected by ELISA on the 4 th and 8 th week after feeding in order to determine whether or not hyperlipidemia model was successful. After hyperlipidemia model establishment, 50 rats with hyperlipidemia were randomly divided into 5 group: negative control group(hyperlipidemia model group), positive control group, high dose green apple group,middle dose green apple and low dose green apple, 10 rats per group. Positive control group received intragastric administration of resveratrol(0.1 mg/10 g) while green apple group received intragastric administration of green apple juice: high dose 0.3 m L/10 g, middle dose 0.2 m L/10 g and low dose 0.1 m L/10 g, once a day. Blank control and negative control group received daily intragastric administration of 0.1 m L/10 g distilled water. Rats were sacrificed after 8 weeks of consecutive drug deliveries. RT-PCR was adopted to detect the expression of m RNA of AMPK and ACC in rat liver tissue and Western blotting adopted to detect the protein expression of AMPK and ACC. The levels of AMPK m RNA and AMPK protein expression in high and middle dose group were significantly higher than their counterparts of hyperlipidemia model group. The levels of ACC m RNA and ACC protein expression in high and middle dose group were significantly lower than their counterparts of hyperlipidemia model group(p<0.05). High dose and middle dose of green apple juice may activate AMPK-ACC signaling pathway, in turn inhibit fatty acid synthesis and accelerate fatty acid oxidation, and finally lower the serum lipid level of hyperlipidemia rats.
To investigate the effect of green apple juice on AMPK-ACC signaling pathway in male rats with hyperlipidemia. 70 healthy SD male rats were enrolled in this study, of which 20 rats were randomly selected as blank control group fed with basal diet and the other 50 rats were fed with high fat and high sugar diet to establish hyperlipidemia model. Blood serum indexes including total cholesterol(TC) and triglyceride(TG) were detected by ELISA on the 4 th and 8 th week after feeding in order to determine whether or not hyperlipidemia model was successful. After hyperlipidemia model establishment, 50 rats with hyperlipidemia were randomly divided into 5 group: negative control group(hyperlipidemia model group), positive control group, high dose green apple group,middle dose green apple and low dose green apple, 10 rats per group. Positive control group received intragastric administration of resveratrol(0.1 mg/10 g) while green apple group received intragastric administration of green apple juice: high dose 0.3 m L/10 g, middle dose 0.2 m L/10 g and low dose 0.1 m L/10 g, once a day. Blank control and negative control group received daily intragastric administration of 0.1 m L/10 g distilled water. Rats were sacrificed after 8 weeks of consecutive drug deliveries. RT-PCR was adopted to detect the expression of m RNA of AMPK and ACC in rat liver tissue and Western blotting adopted to detect the protein expression of AMPK and ACC. The levels of AMPK m RNA and AMPK protein expression in high and middle dose group were significantly higher than their counterparts of hyperlipidemia model group. The levels of ACC m RNA and ACC protein expression in high and middle dose group were significantly lower than their counterparts of hyperlipidemia model group(p<0.05). High dose and middle dose of green apple juice may activate AMPK-ACC signaling pathway, in turn inhibit fatty acid synthesis and accelerate fatty acid oxidation, and finally lower the serum lipid level of hyperlipidemia rats.
引文
Chen G.Q.,Xie X.Y.,Yao X.B.,and Tang C.,2015,Analysis and identification of FOXM1 related mi RNA expression profiling on human ovarian cancer OVCAR3 cells,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(9):1812-1217(陈国庆,谢晓英,姚细保,唐琛,2015,人卵巢癌OVCAR3细胞FOXM1相关差异mi RNA表达谱分析鉴定,基因组学与应用生物学,34(9):1812-1817)
Ge L.,2013,Effects of apple pomace polyphenols on body weight and blood lipid in rats,Shipin Gongye Keji(Science and Technology of Food Industry),34(10):342-344(葛蕾,2013,苹果渣多酚对高脂大鼠体重及血脂的影响,食品工业科技,34(10):342-344)
Guo H.,2008,Effect of AMPK downstream packet and insulin secretion of INS-1 cells,Thesis for M.S.,Shandong University,Supervisor:Gao L.,pp.36-40(郭华,2008,AMPK对INS-1细胞下游分子及胰岛素分泌的影响,硕士学位论文,山东大学,导师:高聆,pp.36-40)
Guo J.,Piao S.H.,Shi Z.F.,and Huang P.,2013,Distribution of traditional chinese medical syndrome patterns in hyperlipidemia patients:an investigation of medical literature,Guangzhou Zhongyiyao Daxue Xuebao(Journal of Guangzhou University of Traditional Chinese Medicine),30(5):609-614(郭姣,朴胜华,石忠峰,黄珮,2013,高脂血症中医证候分布规律文献研究,广州中医药大学学报,30(5):609-614)
Hawley S.A.,Pan D.A.,Mustard K.J.,Ross L.,Bain J.,Edelman A.M.,Frenguelli B.G.,and Ha die D.G.,2005,Calmodulin-dependent protein kinase kinase-beta is an alternative upstream kinase for AMP-activated protein kinase,Cell Metab.,2(1):9-19
Hu D.Y.,and Wa ng J.H.,2009,The status quo of Chinese prevention and treatment of dyslipidemia,Zhongguo Shiyong Neike Zazhi(Chinese Journal of Practical Internal Medicine),29(1):2-4(胡大一,王家宏,2009,我国血脂异常防治现状,中国实用内科杂志,29(1):2-4)
Jiang X.D.,Hu L.Q.,Yi R.,and Li Q.B.,2015,Advances of adenylate-activated pr otein kinase and its activator in anti-tumor,Zhongguo Xinyao Zazhi(Chinese Journal of New Drugs),24(24):2803-2813(蒋晓丁,胡立庆,易锐,李乾斌,2015,腺苷酸活化蛋白激酶及其激活剂在抗肿瘤领域的研究进展,中国新药杂志,24(24):2803-2813)
Li F.,Li Y.F.,and Zhang J.H.,2015,Micro RNAs affect tumor metastasis through regulating tumor angiogenesis,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(11):2332-2337(李芳,李玉凤,张景华,2015,微小RNA通过调控肿瘤血管形成影响肿瘤转移,基因组学与应用生物学,34(11):2332-2337)
Li J.X.,Wang N.,Wang H.J.,Zhou X.H.,Li M.J.,Xie X.H.,and Ai Z.L.,2008,Effect of apple polyphenols on weight loss,Shipin Kexue(Food Science),29(8):597-599(李建新,王娜,王海军,周向辉,李美娟,谢新华,艾志录,2008,苹果多酚的减肥降脂作用研究,食品科学,29(8):597-599)
Lin Z.J.,Zhang B.,and Liu X.Q.,2013,Advances in AMPK-ACC signaling pathway and related metabolic diseases,Zhonghua Tangniaobing Zazhi(Chinese Journal of Diabetes),21(5):474-476(林志健,张冰,刘小青,2013,AMPK-ACC信号通路与相关代谢疾病的研究进展,中华糖尿病杂志,21(5):474-476)
Liu J.,Wang W.,Wang M.,Sun J.,Liu J.,Li Y.,Qi Y.,Wu Z.,and Zhao D.,2012,Impact of diabetes,high triglycerides and low HDL cholesterol on risk for ischemic cardiovascular disease varies by LDL cholesterol level:a 15-year follow-up of the Chinese multi-provincial cohort study,Diabetes Res.Clin.Pract.,96(2):217-224
Liu L.J.,He P.X.,Luo X.H.,and Ren Z.Q.,2010,The study of the healthy adult males the changes of blood lipids with age and the prevalence and distribution characteristics of dyslipidemia in normal men,Zhongguo Dangdai Yiyao(China Modern Medicine),17(2):9-11(刘丽君,贺佩祥,罗湘杭,任志强,2010,健康成年男性血脂随年龄的变化规律及血脂异常的分布特征,中国当代医药,17(2):9-11)
Qin Y.H.,2003,Study eggs in vivo AMPK activity distribution and stress on AMPK activity,Thesis for M.S.,Sichuan Agricultural University,Supervisor:Chen D.W.,pp.8-10(秦玉辉,2003,蛋鸡体内AMPK酶活分布及应激对AMPK酶活影响研究,硕士学位论文,四川农业大学,导师:陈代文,pp.8-10)
Van Vliet-Ostaptchouk J.V.,Nuotio M.L.,Slagter S.N.,Doiron D.,Fischer K.,Foco L.,Gaye A.,Gogele M.,Heier M.,Hiekkalinna T.,Joensuu A.,Newby C.,Pang C.,Partinen E.,Reischl E.,Schwienbacher C.,Tammesoo M.L.,Swertz M.A.,Burton P.,Ferretti V.,Fortier I.,Giepmans L.,Harris J.R.,Hillege H.L.,Holmen J.,Jula A.,Kootstra-Ros J.E.,Kvaly K.,Holmen T.L.,Mannisto S.,Metspalu A.,Midthjell K.,Murtagh M.J.,Peters A.,Pramstaller P.P.,Saaristo T.,Salomaa V.,Stolk R.P.,Uusitupa M.,van der Harst P.,van der Klauw M.M.,Waldenberger M.,Perola M.,and Wolffenbuttel B.H.,2014,The prevalence of metabolic syndrome and metabolically healthy obesity in Europe:a collaborative analysis of ten large cohort studies,BMC Endocrine Disorders,14(9):13-15
Zhang K.J.,2013,Effect of resistance training in the elderly and hepatic lipid metabolism related gene expression in mice,Thesis for M.S.,East China Normal University,Supervisor:Chen C.Z.,pp.43-46(张凯杰,2013,抗阻训练对老年小鼠肝脏脂代谢及其相关基因表达的影响,硕士学位论文,华东师范大学,导师:陈彩珍,pp.43-46)
Zhang Y.Y.,Liu X.,He J.,Jiang X.L.,and Yang Z.W.,2011,The role of AMPK gene in glucose and lipid metabolism in C57BL/6J mice,Zhongguo Tangniaobing Zazhi(Chinese Journal of Diabetes),19(12):935-938(张远远,刘星,何君,姜晓亮,杨志伟,2011,AMPK基因在C57BL/6J小鼠糖脂代谢中的作用,中国糖尿病杂志,19(12):935-938)
Ge L.,2013,Effects of apple pomace polyphenols on body weight and blood lipid in rats,Shipin Gongye Keji(Science and Technology of Food Industry),34(10):342-344(葛蕾,2013,苹果渣多酚对高脂大鼠体重及血脂的影响,食品工业科技,34(10):342-344)
Guo H.,2008,Effect of AMPK downstream packet and insulin secretion of INS-1 cells,Thesis for M.S.,Shandong University,Supervisor:Gao L.,pp.36-40(郭华,2008,AMPK对INS-1细胞下游分子及胰岛素分泌的影响,硕士学位论文,山东大学,导师:高聆,pp.36-40)
Guo J.,Piao S.H.,Shi Z.F.,and Huang P.,2013,Distribution of traditional chinese medical syndrome patterns in hyperlipidemia patients:an investigation of medical literature,Guangzhou Zhongyiyao Daxue Xuebao(Journal of Guangzhou University of Traditional Chinese Medicine),30(5):609-614(郭姣,朴胜华,石忠峰,黄珮,2013,高脂血症中医证候分布规律文献研究,广州中医药大学学报,30(5):609-614)
Hawley S.A.,Pan D.A.,Mustard K.J.,Ross L.,Bain J.,Edelman A.M.,Frenguelli B.G.,and Ha die D.G.,2005,Calmodulin-dependent protein kinase kinase-beta is an alternative upstream kinase for AMP-activated protein kinase,Cell Metab.,2(1):9-19
Hu D.Y.,and Wa ng J.H.,2009,The status quo of Chinese prevention and treatment of dyslipidemia,Zhongguo Shiyong Neike Zazhi(Chinese Journal of Practical Internal Medicine),29(1):2-4(胡大一,王家宏,2009,我国血脂异常防治现状,中国实用内科杂志,29(1):2-4)
Jiang X.D.,Hu L.Q.,Yi R.,and Li Q.B.,2015,Advances of adenylate-activated pr otein kinase and its activator in anti-tumor,Zhongguo Xinyao Zazhi(Chinese Journal of New Drugs),24(24):2803-2813(蒋晓丁,胡立庆,易锐,李乾斌,2015,腺苷酸活化蛋白激酶及其激活剂在抗肿瘤领域的研究进展,中国新药杂志,24(24):2803-2813)
Li F.,Li Y.F.,and Zhang J.H.,2015,Micro RNAs affect tumor metastasis through regulating tumor angiogenesis,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(11):2332-2337(李芳,李玉凤,张景华,2015,微小RNA通过调控肿瘤血管形成影响肿瘤转移,基因组学与应用生物学,34(11):2332-2337)
Li J.X.,Wang N.,Wang H.J.,Zhou X.H.,Li M.J.,Xie X.H.,and Ai Z.L.,2008,Effect of apple polyphenols on weight loss,Shipin Kexue(Food Science),29(8):597-599(李建新,王娜,王海军,周向辉,李美娟,谢新华,艾志录,2008,苹果多酚的减肥降脂作用研究,食品科学,29(8):597-599)
Lin Z.J.,Zhang B.,and Liu X.Q.,2013,Advances in AMPK-ACC signaling pathway and related metabolic diseases,Zhonghua Tangniaobing Zazhi(Chinese Journal of Diabetes),21(5):474-476(林志健,张冰,刘小青,2013,AMPK-ACC信号通路与相关代谢疾病的研究进展,中华糖尿病杂志,21(5):474-476)
Liu J.,Wang W.,Wang M.,Sun J.,Liu J.,Li Y.,Qi Y.,Wu Z.,and Zhao D.,2012,Impact of diabetes,high triglycerides and low HDL cholesterol on risk for ischemic cardiovascular disease varies by LDL cholesterol level:a 15-year follow-up of the Chinese multi-provincial cohort study,Diabetes Res.Clin.Pract.,96(2):217-224
Liu L.J.,He P.X.,Luo X.H.,and Ren Z.Q.,2010,The study of the healthy adult males the changes of blood lipids with age and the prevalence and distribution characteristics of dyslipidemia in normal men,Zhongguo Dangdai Yiyao(China Modern Medicine),17(2):9-11(刘丽君,贺佩祥,罗湘杭,任志强,2010,健康成年男性血脂随年龄的变化规律及血脂异常的分布特征,中国当代医药,17(2):9-11)
Qin Y.H.,2003,Study eggs in vivo AMPK activity distribution and stress on AMPK activity,Thesis for M.S.,Sichuan Agricultural University,Supervisor:Chen D.W.,pp.8-10(秦玉辉,2003,蛋鸡体内AMPK酶活分布及应激对AMPK酶活影响研究,硕士学位论文,四川农业大学,导师:陈代文,pp.8-10)
Van Vliet-Ostaptchouk J.V.,Nuotio M.L.,Slagter S.N.,Doiron D.,Fischer K.,Foco L.,Gaye A.,Gogele M.,Heier M.,Hiekkalinna T.,Joensuu A.,Newby C.,Pang C.,Partinen E.,Reischl E.,Schwienbacher C.,Tammesoo M.L.,Swertz M.A.,Burton P.,Ferretti V.,Fortier I.,Giepmans L.,Harris J.R.,Hillege H.L.,Holmen J.,Jula A.,Kootstra-Ros J.E.,Kvaly K.,Holmen T.L.,Mannisto S.,Metspalu A.,Midthjell K.,Murtagh M.J.,Peters A.,Pramstaller P.P.,Saaristo T.,Salomaa V.,Stolk R.P.,Uusitupa M.,van der Harst P.,van der Klauw M.M.,Waldenberger M.,Perola M.,and Wolffenbuttel B.H.,2014,The prevalence of metabolic syndrome and metabolically healthy obesity in Europe:a collaborative analysis of ten large cohort studies,BMC Endocrine Disorders,14(9):13-15
Zhang K.J.,2013,Effect of resistance training in the elderly and hepatic lipid metabolism related gene expression in mice,Thesis for M.S.,East China Normal University,Supervisor:Chen C.Z.,pp.43-46(张凯杰,2013,抗阻训练对老年小鼠肝脏脂代谢及其相关基因表达的影响,硕士学位论文,华东师范大学,导师:陈彩珍,pp.43-46)
Zhang Y.Y.,Liu X.,He J.,Jiang X.L.,and Yang Z.W.,2011,The role of AMPK gene in glucose and lipid metabolism in C57BL/6J mice,Zhongguo Tangniaobing Zazhi(Chinese Journal of Diabetes),19(12):935-938(张远远,刘星,何君,姜晓亮,杨志伟,2011,AMPK基因在C57BL/6J小鼠糖脂代谢中的作用,中国糖尿病杂志,19(12):935-938)