Preliminary Study On The Contribution And Mechanism Of A Single Gene On Endocrine And Metabolic Diseases

Obesity,diabetes and Graves'disease are common endocrine and metabolic diseases and all of them are triggered by a combination of genetic predisposition and environmental factors.Obesity and diabetes were affected by some specific genes while Graves'disease is a polygenic disease.The current thesis is focused on contributions of diffirent genes to obesity,diabetes and Graves'disease respectively.One is about an adipokine regulating obesity and diabetes.The other is about the contribution of the combination of specific genes and environmental triggers on Graves'disease.Adipose tissue is an important endocrine organ,secreting mutiple adipokines regulating obesity and insulin resistance.Mimecan was found highly expressed in adipose tissue and secreted into the circulation.The expression of mimecan in serum was reduced after fasting and in epididymal adipose tissue from HFD(high fat diet)-induced mice was induced.Adiministration of mimecan-MBP by either intraperitoneally or intracerebroventricularly injection siginificantly suppressed food intake whether mice were with ad libitum access to food or under fasting condition.But the specific anorexic effect was inhibited when mimeca-MBP was inactivated by heating or pretreatment with mimecan polyclonal antibody.Besides,we found the anorexic effect of mimecan was independent from leptin receptor or melanocortin pathway,but via increasing IL-6,IL-1?expression in microglia from hypothalamus.On the other hand,mimecan aggrevated insulin resistance in db/db mice,and mimecan knockout improved insulin sensitivity in mice fed either SD or HFD.Furthermore,mimecan promoted the expression of inflammatory factors,mainly IL-6 and Socs3,in liver,adipose tissue and skeletal muscle,and the pro-inflammatory effect was most significant in skeletal muscle.Thus C2C12 myotubes were selected as the target cell line in vitro in the subsequent experiment.Both mimecan adiminstration and genetically overexpressed in myotubes impaired insulin signaling pathway,activated IKK/NF-?B pathway,and increased IL-6,Socs3,Ccl2 expression.Moreover,the induction of IL-6,Socs3,Ccl2 by mimecan depended on IKK/NF-?B pathway because only BMS,the inhibitor of IKK/NF-?B,could reverse the increment of these inflammtory factors when adding various inhibitors of inflammatory pathway.According to the previous evidences,mimecan aggrevated insulin resistance probably mediating by increasing IL-6,Socs3 and Ccl2 in skeletal muscle.Graves'diease(GD)is a common and intractable autoimmune disease,affected by both genetic factors and environmental triggers.The genetic predisposition accounts for up to 79%,but the inheritance pattern of GD did not follow the simple mendel's law.Consistent with many other autoimmune diseases,GD has a strong female preponderance(female-to-male ratio,>5:1).However,the reason GD is more prevalent among women is unknown.A reasonable explanation is that genes located on the X-chromosome may be involved.Our previous genome-wide association study(GWAS)data from1442 GD cases and 1468 controls,272 SNPs with P_GWAS?1×10^-4 were identified in Xq21.1 region,a linkage disequilibrium(LD)block.And rs5912838 on GPR174-ITM2A at Xq21.1 exhibited the most significant difference(P_GWAS=4.15×10^-8)in the initial GWAS stage.Then we designed the current study to identify the causal SNPs for GD in the Xq21.1 region by fine-mapping in 4,600 GD cases and 4,683 controls from the Chinese Han population.Consequently,rs3827440 was confirmed as having the most significant association(P_Combined=1.00×10^-16),and rs1736645 exhibited the second highest association(P_Combined=6.19×10^-13)after excluding rs3827440and its highly linked SNPs.Additionally,rs3827440 and rs1736645 were identified two independent GD-associated SNPs in Xq21.1 by forward logistic analysis and two-locus logistic analysis.Besides,the rs1736645 risk allele C was associated with a dramatic reduction in ITM2A transcription levels in PBMCs compared with the A allele,and ITM2A expression in PBMC from GD patients was significantly lower than that from healthy individuals.Notably,these two SNPs,rs3827440 and rs1751107(the only SNP tagged by rs1736645),were correlated with ITM2A expression in monocytes after lipopolysaccharide(LPS)and interferon-?(IFN-?)stimulation by searching the expression quantitative trait locus(eQTL)database,which includes 63specific SNPs located on the Xq21.1 locus regulating ITM2A expression under na?ve or LPS and IFN-?stimulation.Previous studies have shown that viral and bacterial infections,especially YE,function as triggers in the pathogenesis of GD.LPS are well-known bacterial products,and virus infections induce an increase of cytokines,such as IFN in vivo.Among these 63 SNPs,eight SNPs were significantly associated with GD(P_GWAS?1.00×10^-4)including rs3827440 and rs17511107.ITM2A was identified associated with immune response and ITM2A overexpression in transgenic mice causes downregulation of CD8 in CD4⁺CD8⁺double-positive thymocytes.Considering that GD is an autoimmune disease and that CD8⁺T cells are diminished in both peripheral and thyroid tissues of patients with GD,leading to an increase in the ratio of CD4⁺/CD8⁺.This study strongly suggests that environmental factors,such as viral and bacterial infections,probably contributed to GD pathogenesis by interacting with the susceptibility gene ITM2A.This is the first report on the mechanisms of interaction between genetic and environmental factors in the development of GD.In conclusion,endocrine and metabolic diseases were affecte by multiple genetic and environmental factors.Mimecan suppresses food intake and aggrevates insulin resistance in mice via inflammatory factors.While ITM2A mediates the contribution of the interaction of genetic factors(rs3827440 and rs1751107)and environmental factors,such as viral and bacterial infections to GD.