Human Leucocyte Antigen (HLA) alleles, cytokine polymorphisms and the type of factor VIII (FVIII) gene mutation are among predisposing factors for inhibitors (inh) development in children with severe haemophilia A (HA). The aim was to investigate the correlations among (i) FVIII gene intron-22 inversion, (ii) HLA alleles and haplotypes and (iii) certain cytokine polymorphisms, with the risk for FVIII inhibitors development in 52 Greek severe HA children, exclusively treated with recombinant concentrates. We performed Long-Range PCR for detection of intron-22 inversion and PCR-SSP, PCR-SSO for genotyping of HLA-A, B, C, DRB1, DQB1 alleles and also for cytokine polymorphisms of TNF-α, TGF-β1, IL-10, IL-6 and IFN-γ. Chi-squared test and Fischer's exact test were used for statistical analysis. A total of 28 children had developed inhibitors (Group I), 71.4% high responding, while 24 had not (Group II). No statistically increased intron-22 inversion prevalence was found in Group I compared with Group II (P = 0.5). Comparison of HLA allele frequencies between the two groups showed statistically significant differences in the following genotypes (i) promoting inhibitors development: DRB1*01(P = 0.014), DRB1*01:01(P = 0.011) and DQB1*05:01 (P = 0.005) and (ii) possibly protecting from inhibitors development: DRB1*11 (P = 0.011), DRB1*11:01 (P = 0.031), DQB1*03 (P = 0.004) and DQB1*03:01 (P = 0.014). Analysis of cytokines revealed a higher incidence of inhibitor detection only in homozygotes of the haplotypes ACC and ATA for IL-10 polymorphisms (P = 0.05). There is evidence that HLA alleles and cytokine polymorphisms play an important role in FVIII inh development. On the contrary, no statistically significant results were obtained for intron-22 inversion and its impact on FVIII inhibitors formation.

Various polymorphisms in cytokine genes have recently been investigated as candidate risk factors in allogeneic hematopoetic stem cell transplantation (allo-HSCT). We retrospectively analyzed specific polymorphisms in genes for interleukin (IL)-10, IL-6, tumor-necrosis factor alpha (TNF-a) and interferon gamma (IFN-c) in a pediatric cohort of 57 histocompatibility leucocyte antigen (HLA)-identical sibling myeloablative transplants. Both recipient and donor genotypes were tested for association with graft-versus-host disease (GVHD) by statistical methods including Cox regression analysis. We found a significant association between the IL-10 promoter haplotype polymorphisms at positions -1082, -819 and -592 with the occurrence of severe (grades III-IV) acute GVHD (aGVHD). Recipients with the haplotype GCC had a statistically significant decreased risk of severe aGVHD (hazard risk (HR)50.20, 95% confidence interval (CI): 0.06-0.67) in comparison with patients with other IL-10 haplotypes (P50.008). Transplant-related mortality at 1 year was significantly lower in recipients with this haplotype (HR50.17, 95% CI: 0.012-0.320) versus other IL-10 haplotypes (P50.03), whereas overall survival was not influenced by IL-10 haplotype polymorphisms. In multivariate analysis, the presence of the IL-10 GCC haplotype was found as the only variable associated with a statistically significant decreased hazard of severe aGVHD development (P50.02, HR50.21, 95% CI: 0.05-0.78). These results suggest that pediatric patients possessing the IL-10 GCC haplotype may be protected from the occurrence of severe aGVHD in the setting of matched sibling HSCT.

This study confirms the existing data and provides additional evidence supporting a strong genetic predisposition for CD associated with the class II alleles DQB1*02 and DQA1*05 encoding the serological specificity DQ2.

Chronic Granulomatous Disease (CGD) is an uncommon primary immunodeficiency caused by the absence or dysfunction of one of NADPH oxidase subunits, with heterogeneous genetic aetiologies. The aim of this study was the CGD patient registry in Greece, the identification of the responsible genotype and the potential correlation with the patient's clinical phenotype. Medical charts of 24 CGD patients, investigated by NBT test or DHR for NADPH oxidase activity, Western blot analysis for NADPH oxidase component expression and DNA sequencing (pyro- and cycle sequencing) for mutation analysis, were reviewed. All patients, but one, were classified into the different types of CGD. Sixteen patients from 14 unrelated families had X-linked CGD (66.7 %), four had mutations in the NCF1 gene (19 %), and three, from two unrelated families, had mutations in NCF2 (9.5 %) [Corrected]. Fifteen mutations were detected in the CYBB gene, including nonsense (53.8 %), splice site (30.8 %) and missense mutations (7.7 %), and deletions (7.7 %). Two novel mutations were identified; one in CYBB and one in NCF1. Carrier detection for X-CGD revealed that the de novo mutation rate was about 7 %. Prenatal diagnosis identified one affected male in three male fetuses tested. In both the X-linked and the autosomal recessive (AR-CGD) group, the gastrointestinal and respiratory manifestations were more common, followed by lympadenopathy in X-CGD and skin infections in the AR-CGD group. The patients with a mutation in CYBB had a wider variability of clinical manifestations and earlier diagnosis (4.6 years) compared to the AR-CGD group (12.9 years). The incidence of CGD in Greece is estimated at 0.90 (95 % CI 0.89-0.91) per 100,000 live births for the last decade.

BackgroundCeliac disease is a complex chronic immune-mediated disorder of the small intestine. Today, the pathobiology of the disease is unclear, perplexing differential diagnosis, patient stratification, and decision-making in the clinic.MethodsHerein, we adopted a next-generation sequencing approach in a celiac disease trio of Greek descent to identify all genomic variants with the potential of celiac disease predisposition.ResultsAnalysis revealed six genomic variants of prime interest: SLC9A4 c.1919G>A, KIAA1109 c.2933T>C and c.4268_4269delCCinsTA, HoxB6 c.668C>A, HoxD12 c.418G>A, and NCK2 c.745_746delAAinsG, from which NCK2 c.745_746delAAinsG is novel. Data validation in pediatric celiac disease patients of Greek (n = 109) and Serbian (n = 73) descent and their healthy counterparts (n = 111 and n = 32, respectively) indicated that HoxD12 c.418G>A is more prevalent in celiac disease patients in the Serbian population (P < 0.01), while NCK2 c.745_746delAAinsG is less prevalent in celiac disease patients rather than healthy individuals of Greek descent (P = 0.03). SLC9A4 c.1919G>A and KIAA1109 c.2933T>C and c.4268_4269delCCinsTA were more abundant in patients; nevertheless, they failed to show statistical significance.ConclusionsThe next-generation sequencing-based family genomics approach described herein may serve as a paradigm towards the identification of novel functional variants with the aim of understanding complex disease pathobiology.Electronic supplementary materialThe online version of this article (doi:10.1186/s40246-016-0091-1) contains supplementary material, which is available to authorized users.