Assuntos
Dedos/anormalidades , Glaucoma de Ângulo Fechado/etiologia , Deficiência Intelectual/complicações , Microcefalia/complicações , Hipotonia Muscular/complicações , Miopia/complicações , Obesidade/complicações , Degeneração Retiniana/complicações , Adulto , Deficiências do Desenvolvimento/complicações , Feminino , Glaucoma de Ângulo Fechado/patologia , Glaucoma de Ângulo Fechado/cirurgia , Humanos , Pressão Intraocular , PrognósticoRESUMO
BACKGROUND: Copy number variants (CNVs) have recently been reported to be associated with several autoimmune conditions. Moreover, loci involved in immunity are enriched in CNVs. Therefore, we hypothesized that CNVs in immune genes associated with Graves' disease (GD) may contribute to the etiology of disease. METHODS: One hundred ninety-one North American Caucasian GD patients and 192 Caucasian controls were analyzed for CNVs in three major immune regulatory genes: CD40, PTPN22, and CTLA-4. Copy number was determined using quantitative-PCR (Q-PCR) assays specifically designed for determining copy numbers in genomic DNA. Additionally, a well-characterized CNV in the amylase gene was typed in a separate dataset of DNA samples that were derived from cell lines or blood. RESULTS: No CNVs could be confirmed in the CD40 and CTLA-4 genes, even though a CD40 CNV is cataloged in the Database of Genomic Variants. Only the PTPN22 CNV was confirmed in our cohort, but it was rare and appeared in only two individuals. A key finding was that the source of DNA has a significant effect on CNV typing. There was a statistically significant increase in amylase locus deletions in cell line-derived DNA compared to blood-derived DNA samples. CONCLUSIONS: We conclude that CNV analysis should be performed only using blood-derived DNA Samples. Additionally, the CTLA-4, CD40, and PTPN22 loci do not harbor CNVs that play a role in the etiology of GD.
Assuntos
Antígenos CD/genética , Antígenos CD40/genética , Dosagem de Genes , Doença de Graves/genética , Doença de Graves/imunologia , Proteína Tirosina Fosfatase não Receptora Tipo 22/genética , Adulto , Amilases/genética , Antígeno CTLA-4 , Estudos de Casos e Controles , Linhagem Celular , Predisposição Genética para Doença , Variação Genética , Genoma Humano , Humanos , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNARESUMO
Previously, we have shown that statistical synergism between amino acid variants in thyroglobulin (Tg) and specific HLA-DR3 pocket sequence signatures conferred a high risk for autoimmune thyroid disease (AITD). Therefore, we hypothesized that this statistical synergism mirrors a biochemical interaction between Tg peptides and HLA-DR3, which is key to the pathoetiology of AITD. To test this hypothesis, we designed a recombinant HLA-DR3 expression system that was used to express HLA-DR molecules harboring either AITD susceptibility or resistance DR pocket sequences. Next, we biochemically generated the potential Tg peptidic repertoire available to HLA-DR3 by separately treating 20 purified human thyroglobulin samples with cathepsins B, D, or L, lysosomal proteases that are involved in antigen processing and thyroid biology. Sequences of the cathepsin-generated peptides were then determined by matrix-assisted laser desorption ionization time-of-flight-mass spectroscopy, and algorithmic means were employed to identify putative AITD-susceptible HLA-DR3 binders. From four predicted peptides, we identified two novel peptides that bound strongly and specifically to both recombinant AITD-susceptible HLA-DR3 protein and HLA-DR3 molecules expressed on stably transfected cells. Intriguingly, the HLA-DR3-binding peptides we identified had a marked preference for the AITD-susceptibility DR signatures and not to those signatures that were AITD-protective. Structural analyses demonstrated the profound influence that the pocket signatures have on the interaction of HLA-DR molecules with Tg peptides. Our study suggests that interactions between Tg and discrete HLA-DR pocket signatures contribute to the initiation of AITD.
