基于LC-MS的代谢组学技术分析孜亚比提片的降糖作用机制
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摘要
目的:观察孜亚比提片对2型糖尿病大鼠的降糖效果,并基于代谢组学技术分析其降糖作用机制。方法:根据临床调研结果对不同组分来源的孜亚比提片进行筛选,筛选出最佳组方配比;并以2型糖尿病大鼠为实验对象进行药效学实验研究,正常组和模型组分别给予相同剂量的生理盐水、孜亚比提片组给予300 mg·kg-1孜亚比提片、二甲双胍组给予300 mg·kg-1盐酸二甲双胍片,每组8只,记录给药后实验组的空腹血糖值及体质量变化并进行数据分析;同时利用超高效液相色谱-线性离子阱/静电场轨道阱组合式高分辨质谱联用(UHPLC-LTQ Orbitrap MS)技术对大鼠血清进行代谢组学分析,采用主成分分析(PCA),偏最小二乘判别分析(OPLS-DA)对不同组别大鼠血清代谢物进行分析,鉴定出潜在生物标记物,并分析维吾尔族药孜亚比提片的降糖作用机制。结果:与正常组比较,模型组大鼠的空腹血糖值升高(P <0. 01);与模型组比较,孜亚比提片组大鼠状态整体较好,毛色光滑,其空腹血糖值也有所下降(P <0. 05),表明孜亚比提片对2型糖尿病大鼠有一定的降糖作用;根据LC-MS代谢组学数据结果最终得到8个生物标记物。与模型组相比孜亚比提片组中L-valine,propionylcarnitine含量均降低(P <0. 05),sphingosine-1-phosphate,LPC(16∶1/0∶0),LPC(18∶0/0∶0),LPC(18∶2/0∶0),LPC(20∶1/0∶0),PC(19∶0/0∶0)的含量均升高(P <0. 05)。结论:孜亚比提片能够降低2型糖尿病大鼠血糖值,并能改善2型糖尿病大鼠的一般生理特征,其作用机制可能与改善氨基酸代谢,脂质代谢有关。
Objective: To observe the hypoglycemic effect of Uygur medicine Ziya Biti tablet on the type2 diabetic rats,and analyze the hypoglycemic mechanism based on metabolomic techniques. Method: According to the results of clinical research about different origins of Ziya Biti tablet,the optimal composition was screened out;type 2 diabetic rats were taken as an experimental object in the pharmacodynamic experiments; the control group and model group were given the same dose of normal saline,Ziya Biti bablet group was given 300 mg·kg-1,the metformin group was given 300 mg·kg-1 metformin hydrochloride. The fasting blood and weight changes of the experimental group after the treatment were recorded and compared with normal group; ultra-high performance liquid chromatography-linear ion trap/electrostatic field orbit trap combined-type high resolution mass spectrometry( UHPLC-LTQ/Orbitrap MS) technology was used to conduct the metabolomics analysis on the rat serum,and principal component analysis( PCA),partial least squares discriminant analysis( OPLS-DA) on different groups of rat serum metabolites were performed to identify potential biomarkers. Result: Compared with the model group,the rats in the Uygur medicine Ziya Biti tablet showed a healthy states,and the blood glucose were decreased( P <0. 05),which indicated that Uygur medicine Ziya Biti tablet had a certain hypoglycemic effect on type 2 diabetic rats. Eight biomarkers were finally obtained according to the results. Compared with model group,L-valine and propionylcarnitine in the Uygur medicine group were decreased( P < 0. 05),while sphingosine-1-phosphate,LPC( 16 ∶ 1/0 ∶ 0),LPC( 18 ∶ 0/0 ∶ 0),LPC( 18 ∶ 2/0 ∶ 0),LPC 20 ∶ 1/0 ∶ 0),PC( 19 ∶ 0/0 ∶ 0) were elevated( P <0. 05). Conclusion: The experimental results showed that Uygur medicine Ziya Biti tablet can reduce the blood glucose of type 2 diabetic rats and allivate general physiological characteristics. The mechanism of action may be related to the improvement of amino acid metabolism and lipid metabolism.
Objective: To observe the hypoglycemic effect of Uygur medicine Ziya Biti tablet on the type2 diabetic rats,and analyze the hypoglycemic mechanism based on metabolomic techniques. Method: According to the results of clinical research about different origins of Ziya Biti tablet,the optimal composition was screened out;type 2 diabetic rats were taken as an experimental object in the pharmacodynamic experiments; the control group and model group were given the same dose of normal saline,Ziya Biti bablet group was given 300 mg·kg-1,the metformin group was given 300 mg·kg-1 metformin hydrochloride. The fasting blood and weight changes of the experimental group after the treatment were recorded and compared with normal group; ultra-high performance liquid chromatography-linear ion trap/electrostatic field orbit trap combined-type high resolution mass spectrometry( UHPLC-LTQ/Orbitrap MS) technology was used to conduct the metabolomics analysis on the rat serum,and principal component analysis( PCA),partial least squares discriminant analysis( OPLS-DA) on different groups of rat serum metabolites were performed to identify potential biomarkers. Result: Compared with the model group,the rats in the Uygur medicine Ziya Biti tablet showed a healthy states,and the blood glucose were decreased( P <0. 05),which indicated that Uygur medicine Ziya Biti tablet had a certain hypoglycemic effect on type 2 diabetic rats. Eight biomarkers were finally obtained according to the results. Compared with model group,L-valine and propionylcarnitine in the Uygur medicine group were decreased( P < 0. 05),while sphingosine-1-phosphate,LPC( 16 ∶ 1/0 ∶ 0),LPC( 18 ∶ 0/0 ∶ 0),LPC( 18 ∶ 2/0 ∶ 0),LPC 20 ∶ 1/0 ∶ 0),PC( 19 ∶ 0/0 ∶ 0) were elevated( P <0. 05). Conclusion: The experimental results showed that Uygur medicine Ziya Biti tablet can reduce the blood glucose of type 2 diabetic rats and allivate general physiological characteristics. The mechanism of action may be related to the improvement of amino acid metabolism and lipid metabolism.
引文
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[19]皮子凤,门丽慧,张静,等.五味子治疗大鼠糖尿病肾病作用机制的血清代谢组学研究[J].分析化学,2015,43(2):169-175.
[20]陈瑞,马宁宁,范姗姗,等.基于代谢组学分析金芪降糖片改善2型糖尿病大鼠脂代谢紊乱的作用机制[J].中国实验方剂学杂志,2018,24(20):102-107.
[21] Wishart D S. Metabolomics:the principles and potential applications to transplantation[J]. Am J Transplant,2010,5(12):2814-2820.
[22] James E L,Parkinson E K. Serum metabolomics in animal models and human disease[J]. Curr Opin Clin Nutr Metab Care,2015,18(5):478-483.
[23]陈瑞,马宁宁,范姗姗,等.基于代谢组学分析金芪降糖片改善2型糖尿病大鼠脂代谢紊乱的作用机制[J].中国实验方剂学杂志,2018,24(20):102-107.
[24]常晋霞,刘文虎,王仕宝,等.基于GC-MS代谢组学分析刺五加总苷提取物的降糖作用[J].中国实验方剂学杂志,2017,23(16):101-107.
[25]冯小可,刘佳莅,王岚,等.香砂六君丸对糖尿病胃轻瘫大鼠的血清代谢组学研究[J].中国实验方剂学杂志,2015,21(23):87-91.
[26] XING F. Lysophosphatidylcholine up-regulates human endothelial nitric oxide synthase gene transactivity by cJun N-terminal kinase signalling pathway[J]. J Cell Mol Med,2010,13(6):1136-1148.
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[28] WANG L, LIU J, MO Y Y, et al.Lysophosphatidylcholine-induced surface redistribution regulates signaling of the murine G protein-coupled receptor G2A[J]. Mol Biol Cell,2005,16(5):2234-2247.
[29] Yea K,Kim J,Yoon J H,et al. Lysophosphatidylcholine activates adipocyte glucose uptake and lowers blood glucose levels in murine models of diabetes[J]. J Biol Chem,2009,284(49):33833-33840.
[30] Barber M N,Risis S,YANG C,et al. Plasma lysophosphatidylcholine levels are reduced in obesity and type 2 diabetes[J]. PLo S One,2012,7(7):41456.
[31]王中华.基于代谢组学的2型糖尿病和环境污染物暴露生物标志物研究[D].北京:北京协和医学院,2015.
[32]宋丹军,潘家琪,李鹏旭,等.溶血磷脂酰胆碱在肝脏疾病中的研究进展[J].中国药理学通报,2014,30(12):1642-1646.
[2] Dedoussis G V,Kaliora A C,Panagiotakos D B. Genes,diet and type 2 diabetes mellitus:a review[J]. Rev Diabet Stud,2007,4(1):13-24.
[3] Yach D,Stuckler D,and Brownell K D. Epidemiologic and economic consequences of the global epidemics of obesity and diabetes[J]. Nat Med,2006,12(1):62-66.
[4]朱禧星.现代糖尿病学[M].上海:上海医科大学出版社,2000:125.
[5]李桂荣,李桂红,王烨,等.孜亚比提片对血糖和血脂作用的实验研究[J].中国新药与临床杂志,2008,27(7):495-499.
[6]国家药典委员会.中华人民共和国卫生部药品标准·维吾尔药分册[M].乌鲁木齐:新疆科技卫生出版社,1999:162.
[7] Nicholson J K,Lindon J C,Holmes E. Metabonomics':understanding the metabolic responses of living systems to pathophysiological stimuli via multivariate statistical analysis of biological NMR spectroscopic data[J].Xenobiotica,1999,29(11):1181-1189.
[8] Kim H K,Choi Y H, Verpoorte R. NMR-based metabolomic analysis of plants[J]. Nat Protoc,2010,5(3):536-549.
[9] Lanza I R,ZHANG S,Ward L E,et al. Quantitative metabolomics by 1H-NMR and LC-MS/MS confirms altered metabolic pathways in diabetes[J]. PLo S One,2010,5(5):10538.
[10] Wishart D S. Quantitative metabolomics using NMR[J].Trends Anal Chem,2008,27(3):228-237.
[11] Patterson J,Focant J F. Comprehensive two-dimensional gas chromatography time-of-flight mass[J]. J High Resolut Chromatogr,2003,23(3):208-214.
[12] YI L Z,HE J,LIANG Y Z,et al. Plasma fatty acid metabolic profiling and biomarkers of type 2 diabetes mellitus based on GC/MS and PLS-LDA[J]. FEBS Lett,2006,580(30):6837-6845.
[13] Fiehn O. Erratum:metabolite profiling for plant functional genomics[J]. Nat Biotechnol,2000,18(11):1157-1161.
[14] Lindon J C,John C,Holmes E,et al. Contemporary issues in toxicology the role of metabonomics in toxicology and its evaluation by the COMET project[J].Toxicol Appl Pharmacol,2003,187(3):137-146.
[15]许国旺,杨军.代谢组学及其研究进展[J].色谱,2003,21(4):316-320.
[16] Stringer K A,Mckay R I, Karnovsky A, et al.Metabolomics and its application to acute lung diseases[J]. Front Immunol,2016,7(4):44.
[17] Matthiesen R. Bioinformatics methods in clinical research[J]. Methods Mol Biol,2010,646(3):67-76.
[18]王静,袁子民,李云兴,等.基于GC-MS代谢组学法研究黄连、生地黄治疗Ⅱ型糖尿病的配伍机制[J].中国中药杂志,2014,39(3):526-530.
[19]皮子凤,门丽慧,张静,等.五味子治疗大鼠糖尿病肾病作用机制的血清代谢组学研究[J].分析化学,2015,43(2):169-175.
[20]陈瑞,马宁宁,范姗姗,等.基于代谢组学分析金芪降糖片改善2型糖尿病大鼠脂代谢紊乱的作用机制[J].中国实验方剂学杂志,2018,24(20):102-107.
[21] Wishart D S. Metabolomics:the principles and potential applications to transplantation[J]. Am J Transplant,2010,5(12):2814-2820.
[22] James E L,Parkinson E K. Serum metabolomics in animal models and human disease[J]. Curr Opin Clin Nutr Metab Care,2015,18(5):478-483.
[23]陈瑞,马宁宁,范姗姗,等.基于代谢组学分析金芪降糖片改善2型糖尿病大鼠脂代谢紊乱的作用机制[J].中国实验方剂学杂志,2018,24(20):102-107.
[24]常晋霞,刘文虎,王仕宝,等.基于GC-MS代谢组学分析刺五加总苷提取物的降糖作用[J].中国实验方剂学杂志,2017,23(16):101-107.
[25]冯小可,刘佳莅,王岚,等.香砂六君丸对糖尿病胃轻瘫大鼠的血清代谢组学研究[J].中国实验方剂学杂志,2015,21(23):87-91.
[26] XING F. Lysophosphatidylcholine up-regulates human endothelial nitric oxide synthase gene transactivity by cJun N-terminal kinase signalling pathway[J]. J Cell Mol Med,2010,13(6):1136-1148.
[27] Murugesan G,Sandhya Rani M R,Gerber C E,et al.Lysophosphatidylcholine regulates human microvascular endothelial cell expression of chemokines[J]. J Mol Cell Cardiol,2003,35(11):1375-1384.
[28] WANG L, LIU J, MO Y Y, et al.Lysophosphatidylcholine-induced surface redistribution regulates signaling of the murine G protein-coupled receptor G2A[J]. Mol Biol Cell,2005,16(5):2234-2247.
[29] Yea K,Kim J,Yoon J H,et al. Lysophosphatidylcholine activates adipocyte glucose uptake and lowers blood glucose levels in murine models of diabetes[J]. J Biol Chem,2009,284(49):33833-33840.
[30] Barber M N,Risis S,YANG C,et al. Plasma lysophosphatidylcholine levels are reduced in obesity and type 2 diabetes[J]. PLo S One,2012,7(7):41456.
[31]王中华.基于代谢组学的2型糖尿病和环境污染物暴露生物标志物研究[D].北京:北京协和医学院,2015.
[32]宋丹军,潘家琪,李鹏旭,等.溶血磷脂酰胆碱在肝脏疾病中的研究进展[J].中国药理学通报,2014,30(12):1642-1646.