Genetic Etiology Of Idiopathic Type 1 Diabetes

Objectives: Idiopathic type 1 diabetes(T1D)is a subtype of T1D with an unknown etiology,which is a provisional clinical diagnosis.Our study was aimed to clarify the proportion of idiopathic T1D and the heterogeneity of clinical phenotypes,to investigate the etiology of idiopathic T1D from the perspectives of monogenic mutations,immune-related and β-cellrelated gene variants,ultimately to look forward to establishing the basis for individualized and precise treatment of idiopathic T1D.Methods: A total of 1724 patients diagnosed with T1D who visited the department of metabolism and endocrinology in the Second Xiangya Hospital of Central South University from January 2000 to December 2020 were included in this study.The diagnostic criteria should meet at least two of the following criteria:(i)age at onset ≤30 years old and >6 months old;(ii)insulin dependent at time of enrollment;(iii)at least one episode of diabetic ketosis or ketoacidosis.(1)All T1D patients were tested for glutamic acid decarboxylase autoantibody(GADA),protein tyrosine phosphatase autoantibody(IA-2A)and zinc transporter 8 autoantibody(Zn T8A)by using radioligand-binding assays,and those who were negative for all three autoantibodies were diagnosed with idiopathic T1D.Patients with disease duration within 1year were defined as newly diagnosed T1D,and the proportion and clinical characteristics of newly diagnosed idiopathic T1D were clarified.Logistic regression was used to analyze the possible factors influencing the occurrence of idiopathic T1D.(2)We determined a gene panel associated with monogenic diabetes by doing a literature review and investigation.Multiplex amplification polymerase chain reaction combined with next-generation sequencing was used to sequence targeted genes in all patients diagnosed with idiopathic T1D.The pathogenicity of variants was determined by referring to the American College of Medical Genetics and Genomics guidelines.Patients were diagnosed with monogenic diabetes when they carried a(likely)pathogenic variant consistent with the mode of inheritance or the heteroplasmy of mitochondrial 3243A>G in peripheral blood was greater than 3%.We aimed to explore the etiology of idiopathic T1D from the perspective of monogenic mutations.(3)The human leukocyte antigen(HLA)regions of non-monogenic diabetic patients with idiopathic T1D were sequenced using Illumina sequencing platform.The HLA-DRB1,DQA1 and DQB1 loci were genotyped,and haplotypes were constructed using HLA-HD and PHASE software.The susceptibility and protection of HLA haplotypes were judged by synthesizing relevant reports from domestic research teams.We aimed to explore the etiology of idiopathic T1D from the perspective of immunerelated genes by comparing the clinical phenotypes of patients with different HLA DR-DQ genotypes.(4)Among the remaining idiopathic T1D with unknown etiology,β-cell-associated gene variants were selected from the target gene panel sequencing data for analysis.The association between β-cell-related gene variants and the clinical phenotypes of idiopathic T1D was analyzed by using generalized linear model and mixed-effects score test.We aimed to explore the etiology of idiopathic T1D from the perspective of β-cellrelated gene variants.Results:(1)24.5%(295/1205)of newly diagnosed T1D were idiopathic.Among these idiopathic T1D,median onset age was 25.0(16.0-35.0)years,16.9% were overweight,15% had a family history of type 2diabetes in first-degree relatives,median fasting C-peptide and 2-hour postprandial C-peptide were 142.1(64.4-276.0)pmol/L and 288.0(86.7-596.6)pmol/L.The results of multinomial logistic regression analysis showed that factors influencing the occurrence of idiopathic T1D included overweight,better β-cell function and family history of T2D(P<0.05).(2)4.15%(20/482)of “idiopathic T1D” carried a monogenic mutation interpreted as(likely)pathogenic or mitochondrial 3243A>G variant,who should be classified as monogenic diabetes,with the proportions of maturity onset diabetes of the young(MODY)and mitochondrial diabetes being 2.49%(12/482)and 1.66%(8/482),respectively.The most frequent subtypes were mitochondrial diabetes(40%;8/20),HNF1A-MODY(25%;5/20)and INS-MODY(20%;4/20).The heterogeneity of mitochondrial heteroplasmy in peripheral blood was also great in patients diagnosed with mitochondrial diabetes,with heteroplasmy levels ranging from 10.72-62.02%.When compared with other idiopathic T1D patients with unknown etiology,MODY patients had a younger age of onset(P<0.05),patients with mitochondrial diabetes had an older age of onset and greater β-cell function(P<0.05)(3)HLA DR-DQ typing was performed in 462 patients with idiopathic T1D of unknown etiology(20 cases of monogenic diabetes were excluded).HLA results were available in 450 idiopathic T1D,included in the analysis.The results showed that the frequencies of HLA susceptible haplotypes(DR3/DR4/DR9)and protective haplotypes(DR8/DR11/DR12/DR15/DR16)were 46.3% and 25.4%,respectively.28.2%(127/450)of patients with idiopathic T1D carrying two susceptible haplotypes,whose clinical features were more similar to autoimmune T1D,exhibiting worse β-cell function(P<0.05),and could be classified as autoimmune predisposed diabetes.(4)A total of 17 genes associated with β-cell function in monogenic diabetes,including ABCC8,APPL1,BLK,GCK,HNF1 A,PAX4 and PAX6,were included in the analysis of β-cell-related genes in the remaining 323 cases of idiopathic T1D of unknown etiology(monogenic diabetes and autoimmune predisposed diabetes were excluded).Results revealed that common variants in ABCC8,APPL1,GCK and PAX6,low frequency variants in ABCC8,BLK and PAX4,and rare variants in ABCC8 and HNF1 A were associated with β-cell function in idiopathic T1D(P<0.05),with common variants explaining 2.7% and 5.2%variability of fasting C-peptide and 2-hour postprandial C-peptide.Patients carrying low frequency BLK and GATA4 variants required lower insulin doses(P<0.05),carriers of ABCC8 rare variants were more likely to have an onset in childhood(OR=4.52 [1.29-15.86],P<0.05),while carriers of HNF1 A rare variants were more likely to have a family history of T2D(OR=10.39 [1.84-58.6],P=0.008).Conclusions: Approximately 1/4 of newly diagnosed T1D with negative islet autoantibodies are classfied as idiopathic T1D.The clinical phenotypes of idiopathic T1D are greatly heterogeneous.Using nextgeneration sequencing technology combined with clinical phenotypes could classify 30% of idiopathic T1D patients as monogenic diabetes or autoimmune predisposed diabetes.The etiology of idiopathic T1D remains unclear,genetic variants related to β-cell function are associated with the clinical phenotype of patients,suggesting that they may be a potential cause of idiopathic T1D.