桑叶对糖尿病小鼠肝脏Toll样受体基因表达的影响
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摘要
目的:探讨桑叶水提物对糖尿病小鼠肝组织Toll样受体(Toll-like receptors,TLRs)基因表达量的影响,并探讨其相关的作用机制。方法:ICR小鼠随机分为正常组(6只),糖尿病组。糖尿病组小鼠利用高脂高糖饲料喂养10周联合多次注射链脲佐菌素(STZ)建立高脂饲养和STZ共同诱导的小鼠糖尿病模型。造模成功小鼠随机分为模型组,二甲双胍组(600 mg·kg~(-1)),桑叶高、中、低剂量组(8,4,2 g·kg~(-1)),每组6只,灌胃(ig)给药,每周测空腹血糖,于实验中后期行口服糖耐量实验,ig给药10周后,测定小鼠血清胰岛素、总胆固醇、甘油三酯和肿瘤坏死因子-α(TNF-α),并应用实时荧光定量PCR(Real-time PCR)检测小鼠肝组织中TLR1~9表达量。结果:糖尿病模型小鼠经桑叶水提物治疗后,进水量减少,显著降低空腹血糖,改善糖耐量,显著降低血脂,增加胰岛素含量,改善胰岛素抵抗,同时降低血清中TNF-α(P<0.05,P<0.01)。Realtime PCR结果显示,糖尿病小鼠肝脏组织TLRs受体中TLR1,TLR2,TLR3,TLR7,TLR8和TLR9表达明显高于正常组(P<0.05,P<0.01),TLR5表达明显低于正常组;桑叶给药组后,桑叶中剂量组肝组织中的TLR7,TLR8和TLR9表达量,显著降低于模型组(P<0.05,P<0.01)。结论:桑叶水提物的降糖作用可能与抑制糖尿病模型小鼠肝组织中TLRs受体有关。
Objective: To investigate the regulatory effect of the water extract from Mori Folium on the gene expressions of Toll-like receptors( TLRs) in the liver from diabetic mice,and explore its action mechanism.Method: ICR mice were randomly divided into normal group( n = 6) and diabetic group. The mice in diabetic group received high-fat and high-sugar fodder for 10 weeks and injection with 100 mg·kg~(-1)streptozotocin( STZ)for 3 days continuously to establish the diabetic mice models. The successfully modeled mice were then randomly divided into model group,metformin gorup( 600 mg·kg~(-1)),Mori Folium groups( high dose 8 g·kg~(-1),middle dose 4 g·kg~(-1),and low dose 2 g·kg~(-1)) for 10 weeks,n = 6 in each group. The drugs were given by ig administration,and the fasting blood glucose was detected every week. Oral glucose tolerance test was conducted in the mid and later phases of experiment. After ig administration for 10 weeks,serum insulin( INS),total cholesterol( T-CHO),triglyceride( TG) and tumor necrosis factor-α( TNF-α) were detected,and the expression level of TLR 1-9 in the liver was measured with quantitative real-time PCR. Result: After the diabetic model mice were treated with Mori Folium water extract,their water intake was reduced; fasting blood glucose was significantly reduced; glucose tolerance was improved; blood fat was significantly reduced; insulin concentration was dramatically increased; insulin resistance was improved; and serum TNF-α level was reduced( P < 0. 05,P <0. 01). Quantitative Real-time PCR data showed that TLR1,TLR2,TLR3,TLR7,TLR8 and TLR9 mRNA expression levels were significantly increased in livers of the diabetic rats( P < 0. 05,P < 0. 01,as compared with normal group),and TLR5 expression level in model group was significantly lower than that in normal group. In addition,the expression levels of TLR7,TLR8 and TLR9 in liver of Mori Folium middle dose group were significantly lower than those in model group( P < 0. 05,P < 0. 01). Conclusion: The water extract of Mori Folium could ameliorate diabetes,and its mechanism may be partly related to inhibiting the TLRs expression in the liver.
Objective: To investigate the regulatory effect of the water extract from Mori Folium on the gene expressions of Toll-like receptors( TLRs) in the liver from diabetic mice,and explore its action mechanism.Method: ICR mice were randomly divided into normal group( n = 6) and diabetic group. The mice in diabetic group received high-fat and high-sugar fodder for 10 weeks and injection with 100 mg·kg~(-1)streptozotocin( STZ)for 3 days continuously to establish the diabetic mice models. The successfully modeled mice were then randomly divided into model group,metformin gorup( 600 mg·kg~(-1)),Mori Folium groups( high dose 8 g·kg~(-1),middle dose 4 g·kg~(-1),and low dose 2 g·kg~(-1)) for 10 weeks,n = 6 in each group. The drugs were given by ig administration,and the fasting blood glucose was detected every week. Oral glucose tolerance test was conducted in the mid and later phases of experiment. After ig administration for 10 weeks,serum insulin( INS),total cholesterol( T-CHO),triglyceride( TG) and tumor necrosis factor-α( TNF-α) were detected,and the expression level of TLR 1-9 in the liver was measured with quantitative real-time PCR. Result: After the diabetic model mice were treated with Mori Folium water extract,their water intake was reduced; fasting blood glucose was significantly reduced; glucose tolerance was improved; blood fat was significantly reduced; insulin concentration was dramatically increased; insulin resistance was improved; and serum TNF-α level was reduced( P < 0. 05,P <0. 01). Quantitative Real-time PCR data showed that TLR1,TLR2,TLR3,TLR7,TLR8 and TLR9 mRNA expression levels were significantly increased in livers of the diabetic rats( P < 0. 05,P < 0. 01,as compared with normal group),and TLR5 expression level in model group was significantly lower than that in normal group. In addition,the expression levels of TLR7,TLR8 and TLR9 in liver of Mori Folium middle dose group were significantly lower than those in model group( P < 0. 05,P < 0. 01). Conclusion: The water extract of Mori Folium could ameliorate diabetes,and its mechanism may be partly related to inhibiting the TLRs expression in the liver.
引文
[1]Donath M Y,Shoelson S E.Type 2 diabetes as an inflammatory disease[J].Nat Rev Immunol,2011,11(2):98-107.
[2]袁婷,关晓菲,卢文冉,等.Toll样受体的内源性激活在糖尿病及糖尿病肾病中的作用——TLRs表达、激活促进DM和DN的发生、发展[J].中国免疫学杂志,2015,31(8):1124-1128.
[3]吴珺,鲁一兵,刘超.Toll样受体和糖尿病[J].国际内分泌代谢杂志,2008,28(2):116-118.
[4]章超群,武晓旭,吴永贵,等.白芍总苷对糖尿病大鼠肾组织Toll样受体信号通路调节的研究[J].中国药理学通报,2014,30(3):58-63.
[5]WU L H,HUANG C C,Adhikarakunnathu S,et al.Loss of toll-like receptor 3 function improves glucose tolerance and reduces liver steatosis in obese mice[J].Metabolism,2012,61(11):1633-1645.
[6]白华.《神农本草经》桑叶考证[J].内蒙古中医药,2016,35(1):102-103.
[7]曾聪彦,李乐愚,梅全喜,等.桑叶降血糖作用研究进展[J].中华中医药学刊,2016,34(1):192-194.
[8]王娅,李嘉盈.桑叶抗糖尿病活性成分研究[J].安徽医药,2016,20(3):425-428.
[9]Vichasilp C,Nakagawa K,Sookwong P,et al.Development of high 1-deoxynojirimycin(DNJ)content mulberry tea and use of response surface methodology to optimize tea-making conditions for highest DNJ extraction[J].Food Sci Technol,2012,45(2):226-232.
[10]唐明敏,刘洋,田思敏,等.桑叶不同提取部位对α-葡萄糖苷酶抑制活性体外研究[J].新疆医科大学学报,2015,38(6):711-714.
[11]MA X,Iwanaka N,Masuda S,et al.Morus alba leaf extract stimulates 5'-AMP-activated protein kinase in isolated rat skeletal muscle[J].J Ethnopharmacol,2009,122(1):54-59.
[12]ZHANG Y,REN C,LU G,et al.Anti-diabetic effect of mulberry leaf polysaccharide by inhibiting pancreatic islet cell apoptosis and ameliorating insulin secretory capacity in diabetic rats[J].Int Immunopharmacol,2014,22(1):248-257.
[13]Ann J Y,Eo H,Lim Y.Mulberry leaves(Morus alba L.)ameliorate obesity-induced hepatic lipogenesis,fibrosis,and oxidative stress in high-fat diet-fed mice[J].Genes Nutr,2015,10(6):1-13.
[14]Naowaboot J,Chung C H,Pannangpetch P,et al.Mulberry leaf extract increases adiponectin in murine3T3-L1 adipocytes[J].Nutr Res,2012,32(1):39-44.
[15]赵保胜,高晓燕,刘洋,等.桑叶治疗2型糖尿病药效及其机制研究[J].中国实验方剂学杂志,2012,18(20):263-266.
[16]Park M Y,Lee K S,Sung M K.Effects of dietary mulberry,Korean red ginseng,and banaba on glucose homeostasis in relation to PPAR-alpha,PPAR-gamma,and LPL mRNA expressions[J].Life Sciences,2005,77(26):3344-3354.
[17]Batra A,Pietsch J,Fedke I,et al.Leptin-dependent tolllike receptor expression and responsiveness in preadipocytes and adipocytes[J].Am J Pathol,2007,170(6):1931-1941.
[18]Pfaffl M W.A new mathematical model for relative quantification in real-time RT-PCR[J].Nucleic Acids Res,2001,29(9):e45.
[19]Esser N,Legrand-Poels S,Piette J,et al.Inflammation as a link between obesity,metabolic syndrome and type 2diabetes[J].Diabetes Res Clin Pract,2014,105(2):141-150.
[20]Lappas M.NOD1 expression is increased in the adipose tissue of women with gestational diabetes[J].J Endocrinol,2014,222(1):99-112.
[21]李丽燕.Toll样受体的研究进展[J].临床医学工程,2011,18(3):93-96.
[22]Dasu M R,Devaraj S,Park S,et al.Increased toll-like receptor(TLR)activation and TLR ligands in recently diagnosed type 2 diabetic subjects[J].Diabetes Care,2010,33(4):861-868.
[23]Shimizu S,Kurashige Y,Nishimura M,et al.Involvement of toll-like receptors in autoimmune sialoadenitis of the non-obese diabetic mouse[J].J Oral Pathol Med,2012,41(7):517-523.
[24]Ghanim H,Mohanty P,Deopurkar R,et al.Acute modulation of toll-like receptors by insulin[J].Diabetes Care,2008,31(9):1827-1831.
[25]Schwabe R F,Seki E,Brenner D A.Toll-like receptor signaling in the liver[J].Gastroenterology,2006,130(6):1886-1900.
[26]Lee A S,Ghoreishi M,CHENG W K,et al.Toll-like receptor 7 stimulation promotes autoimmune diabetes in the NOD mouse[J].Diabetologia,2011,54(6):1407-1416.
[27]WONG F S,HU C,ZHANG L,et al.The role of Toll-like receptors 3 and 9 in the development of autoimmune diabetes in NOD mice[J].Ann N Y Acad Sci,2008,1150(2):146-148.
[28]Patra M C,Choi S.Recent progress in the development of Toll-like receptor(TLR)antagonists[J].Expert Opin Ther Pat,2016,26(6):719-730.
[2]袁婷,关晓菲,卢文冉,等.Toll样受体的内源性激活在糖尿病及糖尿病肾病中的作用——TLRs表达、激活促进DM和DN的发生、发展[J].中国免疫学杂志,2015,31(8):1124-1128.
[3]吴珺,鲁一兵,刘超.Toll样受体和糖尿病[J].国际内分泌代谢杂志,2008,28(2):116-118.
[4]章超群,武晓旭,吴永贵,等.白芍总苷对糖尿病大鼠肾组织Toll样受体信号通路调节的研究[J].中国药理学通报,2014,30(3):58-63.
[5]WU L H,HUANG C C,Adhikarakunnathu S,et al.Loss of toll-like receptor 3 function improves glucose tolerance and reduces liver steatosis in obese mice[J].Metabolism,2012,61(11):1633-1645.
[6]白华.《神农本草经》桑叶考证[J].内蒙古中医药,2016,35(1):102-103.
[7]曾聪彦,李乐愚,梅全喜,等.桑叶降血糖作用研究进展[J].中华中医药学刊,2016,34(1):192-194.
[8]王娅,李嘉盈.桑叶抗糖尿病活性成分研究[J].安徽医药,2016,20(3):425-428.
[9]Vichasilp C,Nakagawa K,Sookwong P,et al.Development of high 1-deoxynojirimycin(DNJ)content mulberry tea and use of response surface methodology to optimize tea-making conditions for highest DNJ extraction[J].Food Sci Technol,2012,45(2):226-232.
[10]唐明敏,刘洋,田思敏,等.桑叶不同提取部位对α-葡萄糖苷酶抑制活性体外研究[J].新疆医科大学学报,2015,38(6):711-714.
[11]MA X,Iwanaka N,Masuda S,et al.Morus alba leaf extract stimulates 5'-AMP-activated protein kinase in isolated rat skeletal muscle[J].J Ethnopharmacol,2009,122(1):54-59.
[12]ZHANG Y,REN C,LU G,et al.Anti-diabetic effect of mulberry leaf polysaccharide by inhibiting pancreatic islet cell apoptosis and ameliorating insulin secretory capacity in diabetic rats[J].Int Immunopharmacol,2014,22(1):248-257.
[13]Ann J Y,Eo H,Lim Y.Mulberry leaves(Morus alba L.)ameliorate obesity-induced hepatic lipogenesis,fibrosis,and oxidative stress in high-fat diet-fed mice[J].Genes Nutr,2015,10(6):1-13.
[14]Naowaboot J,Chung C H,Pannangpetch P,et al.Mulberry leaf extract increases adiponectin in murine3T3-L1 adipocytes[J].Nutr Res,2012,32(1):39-44.
[15]赵保胜,高晓燕,刘洋,等.桑叶治疗2型糖尿病药效及其机制研究[J].中国实验方剂学杂志,2012,18(20):263-266.
[16]Park M Y,Lee K S,Sung M K.Effects of dietary mulberry,Korean red ginseng,and banaba on glucose homeostasis in relation to PPAR-alpha,PPAR-gamma,and LPL mRNA expressions[J].Life Sciences,2005,77(26):3344-3354.
[17]Batra A,Pietsch J,Fedke I,et al.Leptin-dependent tolllike receptor expression and responsiveness in preadipocytes and adipocytes[J].Am J Pathol,2007,170(6):1931-1941.
[18]Pfaffl M W.A new mathematical model for relative quantification in real-time RT-PCR[J].Nucleic Acids Res,2001,29(9):e45.
[19]Esser N,Legrand-Poels S,Piette J,et al.Inflammation as a link between obesity,metabolic syndrome and type 2diabetes[J].Diabetes Res Clin Pract,2014,105(2):141-150.
[20]Lappas M.NOD1 expression is increased in the adipose tissue of women with gestational diabetes[J].J Endocrinol,2014,222(1):99-112.
[21]李丽燕.Toll样受体的研究进展[J].临床医学工程,2011,18(3):93-96.
[22]Dasu M R,Devaraj S,Park S,et al.Increased toll-like receptor(TLR)activation and TLR ligands in recently diagnosed type 2 diabetic subjects[J].Diabetes Care,2010,33(4):861-868.
[23]Shimizu S,Kurashige Y,Nishimura M,et al.Involvement of toll-like receptors in autoimmune sialoadenitis of the non-obese diabetic mouse[J].J Oral Pathol Med,2012,41(7):517-523.
[24]Ghanim H,Mohanty P,Deopurkar R,et al.Acute modulation of toll-like receptors by insulin[J].Diabetes Care,2008,31(9):1827-1831.
[25]Schwabe R F,Seki E,Brenner D A.Toll-like receptor signaling in the liver[J].Gastroenterology,2006,130(6):1886-1900.
[26]Lee A S,Ghoreishi M,CHENG W K,et al.Toll-like receptor 7 stimulation promotes autoimmune diabetes in the NOD mouse[J].Diabetologia,2011,54(6):1407-1416.
[27]WONG F S,HU C,ZHANG L,et al.The role of Toll-like receptors 3 and 9 in the development of autoimmune diabetes in NOD mice[J].Ann N Y Acad Sci,2008,1150(2):146-148.
[28]Patra M C,Choi S.Recent progress in the development of Toll-like receptor(TLR)antagonists[J].Expert Opin Ther Pat,2016,26(6):719-730.