Research On Mechanism Of Female Impaired Glucose Regulation

BackgroundDiabetes mellitus (DM) is an epidemic disease all over the world with the rapid growth of incidence and prevalence. Diabetes and its related diseases have become an important public health problem. Epidemiological survey of 14 cities in China in 2008 has found that the prevalence of diabetes in people over the age of 20 has reached 9.7% and the pre-diabetes prevalence rate of 15.5%, which are 148 million people by estimation. Pre-diabetes, also known as impaired glucose regulation refers to impaired glucose tolerance (IGT) and impaired fasting glucose (IFG). Daqing Diabetes Prevention Study in China showed that in IGT people the average annual incidence of diabetes was 11%, and 93% of the cumulative incidence of diabetes in 20 years. Hereby, intervention initiated in pre-diabetes is critical for prevention of diabetes. Gestational diabetes mellitus (GDM) is a special type of diabetes, defined as varying degrees of abnormal glucose metabolism onset or first found during pregnancy. According to reports, GDM incidence rate in China is from 1% to 5%. In recent years, with in-depth study of diabetes, the GDM patient and the offspring exposed to high glucose environment suffered from significantly increased risk to develop into type 2 diabetes. The risk of GDM patients with post-natal incidence of type 2 diabetes is 7.43 times of the general population, and their offsprings are in higher risk of impaired glucose regulation than non-GDM offsprings (33% versus 14% at age 15-19 years). This cycle increased the population of impaired glucose regulation. Other studies found that in case of parental diabetes history, the incidence of diabetes in the offspring whose mother has diabetes higher than the offspring whose father has diabetes (35% vs.7%). Similar situation is also observed in type 2 diabetes. Presumption is then made that mothers play a more important role in the genetic background of diabetes pathogenesis. According to the Western medicine, the glucose regulation results from insulin resistance and secretion defects on the basis of genetic susceptibility and environmental influence. However, the exact pathogenesis is not clear. Metabolomics is the use of modern analytical techniques for high-throughput, high efficiency, high sensitivity, small molecule metabolites in biological systems (generally refers to the overall composition of the relative molecular mass is less than the 1000 low molecular weight compounds) to provide dynamic information for clinical follow-up analysis and make access to multidimensional analysis of physiological and pathological conditions, the environment and genetic makeup, etc. It is very appropriate for research of glucose metabolic disorder.Therefore, this paper is focused on metabolic disorder of glucose in women with different constitutions (yin deficiency and yang deficiency constitution) and different physiology (non-gestational and gestational). The research is based on 1H-NMR metabolomics methods, aiming to explore the disordered glucose metabolic profile and variation and to provide new ideas for prevention and treatment. Further research is carried out on microRNA difference in expression pattern of abnormal glucose metabolism during pregnancy.PurposeThrough Metabonomics model discriminates analysis method, we try to analyse and compare plasma metabolic changes of Yin-deficiency females and Yang-deficiency females in gestation period and non pregnant period. We also try to elucidate the metabolic mechanism through work on microRNA differential expression profiling of pregnant woman who are IGT.Method(1)We separate the yin-deficiency female into various groups of Gestation Period and Non Pregnant Period to Normal Glucose Tolerance, Impaired Glucose Tolerance and Diabetes Mellitus according to OGTT results. We obtain the original group of plasma metabolic fingerprint information through application of 1H-NMR technique. By data processing access to the sample information, we analyse these data with PCA, PLS-DA, and OPLS-DA pattern recognition. Then we review literatures on metabolite identification by combining HMDB database and further analyse the change of metabolic pathways.(2) We take the RNA extracted from yin-deficiency female in Gestation Period to chip experiments, according to the Cluster analysis and Ratio analysis of resort, we found the microRNA which are showing in the different way in IGT in pregnant period.Resultplasma metabolomics of impaired glucose tolerance Women of yin-deficiency and yang-deficiency: ①Comparison of YIN-IGT group and NGT group:Lipids, beta-glucose, beta-alanine, choline, proline, glycine, glutamine, glycerol content in plasma increase. And lactic acid, N-acetyl glutamic acid, alanine material content decrease. These material changes prompt that Triglycerides Catabolism、protein catabolism、Gluconeogenesis and Urea synthesis in Ornithine cycle metabolism is exuberant in YIN-IGT group.②omparison of YANG-IGT group and NGT group:most of Metabolite contents in plasma decrease, like Valine, arginine, lysine, glutamic acid, aspartic acid, creatine, phenylalanine, choline, arginine, inositol, threonine,6-Phosphorylation of glucose, Tyrosine networkacid, indole, dog uric acid, adenosine triphosphate, cytosine,5-hydroxy tryptophan,1-methyl-histidine, coenzyme 2.only lipid content increase. That prompts protein、 amino acid metabolism are slow, and Energy metabolism are disordered.③omparison of YANG-IGT group and YIN-IGT group:Two groups are different in metabolism, which are mainly related to disorder of amino acid metabolism and nucleotide metabolism.Impaired glucose regulation of Female plasma metabolomics: ①The NGT group, the IGR group and the DM group are different in metabolic map. After using OPLS-DA method for pattern recognizing, differences among each group are clear in sample distribution of the integral matrix. We can completely distinguish the NGT group, the IGR group and the DM group.②Comparison of YIN-IGT group and NGT group:Compared with NGT group, levels of VLDL, LDL, Cholesterol, unsaturated fatty acids,2-hydroxy-isobutyric acid and glucose rise in IGR group, which prompt that the differences are mainly to metabolic disorders.③Comparison of DM group and IGR group:Levels of Lysine, histidine, glycerol and sugar rise in DM group. Lipids, Levels of choline, leucine, valine, alanine fall; There are differences in glucose metabolism and amino acid metabolism between DM group and IGR group, mainly related to the metabolic pathway of the citric acid cycle, alanine-glucose cycle, amino acid oxidation. ④omparison of DM group and NGT group:Levels of Lipids, sugar,2-hydroxy-isobutyric acid and lysine content increase, leucine, proline, arginine, beta-alanine, choline, alpha-ketoglutarate, and glycerol fall. There are differences in glucose metabolism fat metabolism and amino acid metabolism between DM group and IGR group.Glucose regulating abnormal of Female plasma metbolomics in gestation period: ①There are differences in plasma 1H CPMG spectrum among the GN group, the GIGR group and the GDM group. After using PCA, PLS-DA PLS-DA and OPLS-DAD method for pattern recognizing, we can not distinguish the differences among three groups.② Comparison of GIGT group and GN group:After using PCA method for pattern recognizing, although there are overlap in individual samples, we can still distinguish the samples between GIGT group and GN group with clusters formed, the mainly different contents are LDL/VLDL, Isoleucine, alginic acid, threonine, lactate, choline, choline phosphate, which are involved in fat metabolism and glucose metaboliam.③Comparison of GIGT group and GDN group:Compared with GIGT group, levels of Isoleucine, proline, threonine fall in GDM group. Levels of Lipid and fatty acids rise. Levels of sugar are rise or fall according to different types. Beyond the differences of fat and sugar metabolism, there are differences in the amino acid metabolism too. ④omparison of GN group and GDM group:Compared with GN group, levels of Putrescine, glutamate, glutamine, methionine, lysine, tyrosine, ornithine, methyl histidine, creatine, oxidation-pentanedione, citric acid, succinic acid, unsaturated fatty acids, folic acid, fumaric acid, Hypoxanthine, ACP, GCP, formic acid, and pyrimidine rise, while levels of inositol and asparagine ammonia acid fall. Comparison of GDM and GN shows that not only the Sugar, fat, amino acid metabolism are different, the metabolism of nucleotides is different too.The research on microRNA Differentially expression map of abnormal Glucose metabolism during pregnancy: ①Comparison of GIGT group and GN group:We satiate 64 micro-RNAs in micro-RNA chip by SAM method, based on fold-change greater than 1.2(rise in GIGT), or fold-change less than 0.8(down-regulated in GIGT); Also based on T-test of the signal values between the two groups, P> 0.05 for the standard, then we satiate 7 differential expression of microRNAs:Levels of hsa-miR-19b, hsa-miR-1469, hsa-miR-106b, hsa-miR-17, hsa-miR-24, and hsa-miR-26a expression rise, only levels of U83B1 expression fall. Cluster analysis showed that these 7 differential expressions of microRNAs can distinguish GIGT group and GN group. All the differential expression of microRNAs in GIGT can affect Transforming growth factor-beta (TGF-β) signal pathway. TGF-β signal pathway is closely related to the early pancreatic development and beta cell function, we think that abnormal TGF-P signal may activate beta cell dysfunction.②Comparison of GDM group and GN group:We Satiate 59 micro-RNAs in micro-RNA chip by SAM method. Based on fold-change greater than 1.2(rise in GIGT), or fold-change less than 0.8(down-regulated in GIGT); Also based on T-test of the signal values between the two groups, P> 0.05 for the standard, then satiated 10 differentially expression microRNAs:Levels of hsa-miR-140-3p、 hsa-miR-107、hsa-miR-191, hsa-miR-103, hsa-let-7b, and hsa-miR-1469 expression rise; Levels of U95_x, SNORD119, vll_hsa-miR-923, U83B expression fall. These different microRNAs are closely related to insulin resistance, metabolic regulation and impaired glucose tolerance.ConclusionMetabolic disorder differs in women identified as YIN-IGT and Yang IGT.Metabolic disorder progresses in non-pregnant women from stage NGT through IGR to DM; similar condition is found in pregnant women from stage from GN through GIGT to GDM. The difference of glucose tolerance is significant between pregnant and non-pregnant women and metabolites vary between the two groups of women. CHE is a possible link between pregnant and non-pregnant women.In stage of GIGT, the miRNA in difference in involved with disorder of β cells; while the miRNA in difference is referred to insulin resistance and metabolic disorder of glucose and lipid. Therefore, we presume that injury toβ cells may be prior to insulin resistance and metabolic disorder.With metabonomic method, miR-103 and miR-107 are proved to be essential to the onset and progression of GDM. They may regulate insulin sensitivity and interfere with metabolic processes of glucose and lipid. This can provide new thoughts for further study.