N-glycome and genome in comprehension of type 1 diabetes

N-glikozilacija proteina plazme povezana je s komplikacijama šećerne bolesti tipa 1, dok je uloga ove enzimske modifikacije proteina u nastanku same bolesti neistražena. S razvojem šećerne bolesti tipa 1 do sada je povezano više od 50 gena, među kojima su i glikoziltransferazni geni. Glavni ciljevi ovog istraživanja su identifikacija N-glikana ukupnih proteina plazme i IgG-a karakterističnih za ranu fazu šećerne bolesti tipa 1 te gena kojima su N-glikani ukupnih plazmatskih proteina i IgG-a djece i adolescenata sa šećernom bolesti tipa 1 regulirani. Istraživanje je uključilo 1917 djece i adolescenata (0,6 – 19,1 godina) čiji su uzorci krvne plazme prikupljeni unutar tri mjeseca od dijagnoze šećerne bolesti tipa 1 kroz Danski registar dječjeg i adolescentnog dijabetesa, kao i 265 zdravih srodnika. N-glikani ukupnih proteina plazme i IgG-a analizirani su upotrebom tekućinske kromatografije, dok je genotipizacija 1105 djece i adolescenata sa šećernom bolesti tipa 1 provedena pomoću komercijalnog čipa (Immunochip, Illumina Infinium) koji sadrži 183 546 polimorfizama pojedinačnih nukleotida važnih za razvoj bolesti posredovanih imunološkim sustavom. Nakon navedenih analiza provedena je genomska asocijacijska studija. Ovo istraživanje pokazalo je da je šećerna bolest tipa 1 povezana s porastom oligomanoznih N-glikana i N-glikana s račvajućim N-acetilglukozaminom te padom monogalaktoziliranih N-glikana ukupnih plazmatskih proteina i IgG-a, kao i s porastom disijaliniziranih N-glikana IgG-a. Modeli temeljeni na N-glikanima ukupnih plazmatskih proteina i IgG-a su pokazali jako dobru diskriminacijsku moć između djece i adolescenata sa šećernom bolesti tipa 1 i njihovih zdravih srodnika (AUC > 0,9). Tri prethodno identificirana N-glikozilacijska lokusa (MGAT3, MGAT5 i ST6GAL1) povezana su s razinama N-glikana u ovom istraživanju, kao i C3 lokus na kromosomu 19 koji do sada nije bio povezan s N-glikozilacijom. C3 kodira glavni protein sustava komplementa te su dva C3 SNP-a (sinonimni i nesinonimni) povezana s razinama Man9 glikana ukupnih plazmatskih proteina. S obzirom da se takav Man9 glikan nalazi na C3 domeni uključenoj u interakciju patogena i C3 proteina, promjene u razinama Man9 mogle bi potencijalno interferirati s aktivacijom sustava komplementa u šećernoj bolesti tipa 1. Navedena otkrića pružaju temelje za daljnja istraživanja mehanizama koji reguliraju N-glikozilaciju u šećernoj bolesti tipa 1.Alterations of plasma N-glycosylation have mostly been studied in association to diabetes complications, whereas the role of these changes in type 1 diabetes onset is largely unknown. The present study was undertaken to determine plasma N-glycans representative of the type 1 diabetes onset and to gain the knowledge of genes regulating these changes. Plasma and IgG N-glycans were chromatographically analysed in a recent-onset 1917 type 1 diabetes cases (0.6-19.1 years) and their 244 unaffected siblings whose plasma samples were collected through the Danish Registry of Childhood and Adolescent Diabetes. This study identified an increase in the proportion of plasma and IgG high-mannose and bisecting GlcNAc structures, a decrease in monogalactosylation, and an increase in IgG disialylation characteristic of the early phase of type 1 diabetes. A notable discriminative power between children with type 1 diabetes and their healthy siblings was yielded with models including age, gender, and N-glycans with AUCs of 0.915 and 0.869 for addition of plasma and IgG N-glycans, respectively. Increasing the efficacy of models to determine individuals at risk of disease development would be a considerable asset for type 1 diabetes prevention trials. 183,546 single nucleotide polymorphisms (SNPs) on the commercially available Immunochip that covers all major autoimmune diseases were determined for 1105 study participants and genetic association study on plasma and IgG N-glycome data in type 1 diabetes was conducted. Three of the previously established N-glycosylation loci (MGAT3, MGAT5, and ST6GAL1) were significantly associated with N-glycosylation in the discovery cohort, as well as a novel locus on chromosome 19 encoding the C3, the pivotal protein of the complement activation pathway. This study of type 1 diabetes cases revealed a new genetic association of the C3 gene with N-glycosylation, namely of two C3 SNPs (synonymous and non- synonymous) with the Man9 glycan of total plasma proteins. Since such Man9 glycan is located on the domain implicated in pathogen binding on the C3 protein, the identified N-glycan alterations might potentially be implicated in the complement activation in type 1 diabetes. These findings offer starting points for further functional follow-up studies of mechanisms regulating N-glycosylation in type 1 diabetes