RESUMEN
Insulin is a critical autoantigen for the development of autoimmune diabetes in non-obese diabetic (NOD) mice. About 80% of NOD females and 30-40% of NOD males develop diabetes. However, Insulin2 (Ins2) knockout NOD mice develop autoimmune diabetes with complete penetrance in both sexes, at an earlier age, and have stronger autoimmune responses to insulin. The severe diabetes phenotype observed in NOD-Ins2-/- mice suggests that lack of Ins2 expression in the thymus may compromise immunological tolerance to insulin. Insulin is a prototypical tissue specific antigen (TSA) for which tolerance is dependent on expression in thymus and peripheral lymphoid tissues. TSA are naturally expressed by medullary thymic epithelial cells (mTEC), stromal cells in peripheral lymphoid tissues and bone marrow (BM)-derived cells, mainly CD11c(+) dendritic cells. The natural expression of TSA by mTEC and stromal cells has been shown to contribute to self-tolerance. However, it is unclear whether this also applies to BM-derived cells naturally expressing TSA. To address this question, we created BM chimeras and investigated whether reintroducing Ins2 expression solely by NOD BM-derived cells delays diabetes development in NOD-Ins2-/- mice. On follow-up, NOD-Ins2-/- mice receiving Ins2-expressing NOD BM cells developed diabetes at similar rates of those receiving NOD-Ins2-/- BM cells. Diabetes developed in 64% of NOD recipients transplanted with NOD BM and in 47% of NOD mice transplanted with NOD-Ins2-/- BM (P = ns). Thus, NOD-Ins2-/- BM did not worsen diabetes in NOD recipients and Ins2 expression by NOD BM-derived cells did not delay diabetes development in NOD-Ins2-/- mice.
Asunto(s)
Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/inmunología , Tolerancia Inmunológica , Insulina/genética , Insulina/inmunología , Animales , Células de la Médula Ósea/inmunología , Femenino , Masculino , Ratones , Ratones Endogámicos NOD , Ratones Noqueados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transcripción GenéticaRESUMEN
AIM/HYPOTHESIS: We investigated whether beta cell neoformation occurs in the transplanted pancreas in patients with type 1 diabetes who had received a simultaneous pancreas-kidney transplant (SPK) and later developed recurrence of autoimmunity. METHODS: We examined pancreas transplant biopsies from nine SPK patients with or without recurrent autoimmunity or recurrent diabetes and from 16 non-diabetic organ donors. Tissues were analysed by immunohistochemistry and immunofluorescence. RESULTS: Numerous cytokeratin-19 (CK-19)(+) pancreatic ductal cells stained for insulin in six SPK recipients with recurrent autoimmunity, in five of whom diabetes requiring insulin therapy recurred. These cells also stained for the transcription factor pancreatic-duodenal homeobox-1 (Pdx-1), which is implicated in pancreatic development and beta cell differentiation. The number of insulin(+) ductal cells varied, being highest in the patient with the most severe beta cell loss and lowest in the normoglycaemic patient. In the patient with the most severe beta cell loss, we detected insulin(+)CK-19(+)Pdx-1(+) cells staining for the proliferation-related Ki-67 antigen (Ki-67), indicating proliferation. We were unable to detect Ki-67(+) beta cells within the islets in any SPK patient. Some insulin(+)CK-19(-) ductal cells contained chromogranin A, suggesting further endocrine differentiation. Insulin(+) cells were rarely noted in the pancreas transplant ducts in three SPK patients without islet autoimmunity and in six of 16 non-diabetic organ donors; these insulin(+) cells were never CK-19(+). CONCLUSIONS/INTERPRETATION: Insulin(+) pancreatic ductal cells, some apparently proliferating, were found in the transplanted pancreas with recurrent islet autoimmunity/diabetes. Replicating beta cells were not detected within islets. The observed changes may represent attempts at tissue remodelling and beta cell regeneration involving ductal cells in the human transplanted pancreas, possibly stimulated by hyperglycaemia and chronic inflammation.
Asunto(s)
Autoinmunidad/inmunología , Diabetes Mellitus Tipo 1/cirugía , Trasplante de Páncreas , Páncreas/metabolismo , Adulto , Proliferación Celular , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Insulina/análisis , Antígeno Ki-67/análisis , Masculino , Persona de Mediana Edad , Páncreas/inmunología , Páncreas/patología , Conductos Pancreáticos/inmunología , Conductos Pancreáticos/metabolismo , Conductos Pancreáticos/patologíaRESUMEN
The major susceptibility locus for type 1 diabetes mellitus (T1D) maps to the human lymphocyte antigen (HLA) class II region in the major histocompatibility complex on chromosome 6p21. In southern European populations, like the Basques, the greatest risk to T1D is associated with DR3 homo- and heterozygosity and is comparable to that of DR3/DR4, the highest risk genotype in northern European populations. Celiac disease (CD) is another DR3-associated autoimmune disorder showing certain overlap with T1D that has been explained by the involvement of common genetic determinants, a situation more frequent in DR3-rich populations, like the Basques. As both T1D- and CD-associated HLA alleles are part of conserved extended haplotypes (CEH), we compared DR3-homozygous T1D and CD patients to determine whether CEHs were equally distributed between both disorders or there was a differential contribution of different haplotypes. We observed a very pronounced distribution bias (P<10(-5)) of the two major DR3 CEHs, with DR3-B18 predominating in T1D and DR3-B8 in CD. Additionally, high-density single nucleotide polymorphism (SNP) analysis of the complete CEH [A*30-B*18-MICA*4-F1C30-DRB1*0301-DQB1*0201-DPB1*0202] revealed extraordinary conservation throughout the 4.9 Mbp analyzed supporting the existence of additional diabetogenic variants (other than HLA-DRB1*0301-DQB1*0201), conserved within the DR3-B18 CEH (but not in other DR3 haplotypes) that could explain its enhanced diabetogenicity.
Asunto(s)
Enfermedad Celíaca/genética , Enfermedad Celíaca/inmunología , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/inmunología , Antígeno HLA-DR3/genética , Femenino , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Haplotipos , Homocigoto , Humanos , Masculino , Polimorfismo de Nucleótido Simple , EspañaRESUMEN
The contribution of HLA genes to the genetic risk for celiac disease (CD) has been known for a long time. Recent publications have pointed to the possibility that a second, independent susceptibility locus could be located in the same genomic region, and a triplet repeat polymorphism in exon 5 of the gene MHC class I chain-related protein A (MICA; located between TNFA and HLA-B) has been associated with several autoimmune disorders, including type 1 diabetes mellitus (DM1) and Addison's disease. On the other hand, a single amino acid change in exon 3 of MICA (M129V) has been shown to strongly reduce MICA binding to NKG2D, an activating natural killer receptor expressed also on T cells, and this could have significant effects on autoimmune reactions. In this study, we have analyzed the contribution of these polymorphisms to CD in 37 Basque families, and have constructed MICA-HLA-DRB1 haplotypes to determine whether MICA has an effect independent from the HLA class II conferred risk. In our population, HLA-DRB1*0301 was associated with an increased risk for CD, while HLA-DRB1*1501 conferred protection from the disease (OR: 7.38 and 0.06, respectively). On the other hand, MICA allele A4 was positively associated with the disease (OR: 4.69) whereas allele A9 showed a trend towards protection (OR: 0.18), although significance did not hold after correction. No association of the exon 3 biallelic polymorphism was observed. A positive allelic association was found for haplotypes A5.1-DRB1*0301 (associated with risk for disease), A4-DRB1*0301 and A6-DRB1*07. In view of our results, both HLA-DRB1 and MICA are associated with CD, but stratification analysis did not show any independent contribution of the MICA polymorphisms analyzed to CD risk. Besides, MICA allele A4 (also A5.1 was associated with risk for CD and other diseases) is in strong linkage disequilibrium with HLA-DRB1*0301. Finally, the major histocompatibility complex region's conferred susceptibility to CD, at least in Basque, is very similar to that observed for DM1, with shared risk and protective haplotypes.