Ondřej Cinek

HLA-DQA1*05-DQB1*0201 positivity predisposes to coeliac disease in Czech diabetic children.

The aims of this study were to estimate the prevalence of coeliac disease (CD) in Czech children with insulin dependent diabetes mellitus (IDDM), and to determine the contribution of HLA‐DQA1 and DQB1 to CD susceptibility among diabetic children. We screened 345 children with IDDM (186 boys and 159 girls, aged 0 to 18 y) for coeliac disease using the IgA endomysial antibodies (EMA) test. In all EMA‐positive children, small bowel biopsy was performed to confirm CD. To determine the role of the HLA‐DQA1*05‐DQB1*0201 (DQ2) and the DQA1*03‐DQB 1*0302 (DQ8) molecules in CD susceptibility among diabetic children, the HLA‐DQA1‐DQB1 was genotyped in all EMA‐positive, and in 186 of EMA‐negative diabetic patients. EMA positivity was found in 15/345 (4.3%) diabetic children. The diagnosis of CD was established in 14/345 (4.1%) children based on a bioptic finding of villous atrophy, while the remaining EMA‐positive patient had a normal bioptic finding, being diagnosed as a potential CD. The HLA DQA1*05‐DQB1*0201 (DQ2) molecule conferred a significant risk of CD among diabetic children (odds ratio = 4.1, CI 95% 1.1–15), being found more frequently in diabetic children with CD (80%) than in diabetic children without CD (49%).

Direct PCR Detection of Burkholderia cepacia Complex and Identification of Its Genomovars by Using Sputum as Source of DNA

ABSTRACT We developed a nested PCR assay that detects the recA gene of the Burkholderia cepacia complex in sputum. The product of the first PCR round is also used to identify the genomovar of the pathogen. The protocol achieves high sensitivity and specificity with simple interpretation of genomovar status.

Type 1 diabetes mellitus in Czech children diagnosed in 1990–1997: a significant increase in incidence and male predominance in the age group 0–4 years

Aims To overview total, age‐and sex‐specific incidence rates of type 1 diabetes mellitus and their trends in Czech children 0–14 years of age in the period of 1990–1997.

Effectiveness of sequencing selected exons of DNAH5 and DNAI1 in diagnosis of primary ciliary dyskinesia.

Primary ciliary dyskinesia (PCD) is a rare genetically heterogenous condition. Mutations in DNAH5 or DNAI1 genes can be found in about a third of the patients with PCD. Increased occurrence of mutations was described in several exons of these long genes. The objective of the study was to test the sensitivity of sequencing of selected 13 exons (as compared to costly sequencing of all 100 exons of the two genes), and to determine the prevalence of the DNAH5 or DNAI1 mutations in the Czech PCD database.

An effective combination of sanger and next generation sequencing in diagnostics of primary ciliary dyskinesia

Primary ciliary dyskinesia (PCD) is a multigenic autosomal recessive condition affecting respiratory tract and other organs where ciliary motility is required. The extent of its genetic heterogeneity is remarkable. The aim of the study was to develop a cost‐effective pipeline for genetic diagnostics using a combination of Sanger and next generation sequencing (NGS).

Sulphonylurea treatment does not improve psychomotor development in children with KCNJ11 mutations causing permanent neonatal diabetes mellitus accompanied by developmental delay and epilepsy (DEND syndrome)

© 2007 The Authors. 1176 Journal compilation

Tolerogenic Dendritic Cells from Poorly Compensated Type 1 Diabetes Patients Have Decreased Ability To Induce Stable Antigen-Specific T Cell Hyporesponsiveness and Generation of Suppressive Regulatory T Cells

Tolerogenic dendritic cells (tolDCs) may offer an interesting intervention strategy to re-establish Ag-specific tolerance in autoimmune diseases, including type 1 diabetes (T1D). T1D results from selective destruction of insulin-producing β cells leading to hyperglycemia that, in turn, specifically affects a patient’s immune system. In this study, we prepared monocyte-derived tolDCs modulated by dexamethasone and vitamin D2 from 31 T1D patients with optimal glycemic control and 60 T1D patients with suboptimal glycemic control and assessed their tolerogenic properties in correlation with metabolic state of patients. tolDCs differentiated from both groups of patients acquired a regulatory phenotype and an anti-inflammatory profile. Interestingly, tolDCs from well-controlled patients expressed higher levels of inhibitory molecules IL-T3 and PD-L1. Additionally, glutamic acid decarboxylase (GAD)65–loaded tolDCs from well-controlled patients decreased significantly primary Th1/Th17 responses, induced stable GAD65-specific T cell hyporesponsiveness, and suppressed markedly control DC-induced GAD65-specific T cell activation compared with poorly controlled patients. The ability of tolDCs from poorly controlled patients to induce durable GAD65-specific T cell hyporesponsiveness was reversed once the control of glycemia improved. In both groups of patients, tolDCs were able to induce regulatory T cells from autologous naive CD4+ T cells. However, regulatory T cells from well-controlled patients had better suppressive abilities. The functionality of tolDCs was confirmed in the adoptive transfer model of NOD-SCID mice where tolDCs delayed diabetes onset. These results suggest that metabolic control of T1D affects the functional characteristics of tolDCs and subsequent effector T cell responses. Metabolic control may be relevant for refining inclusion criteria of clinical trials in the settings of T1D.

HbA1c‐based diabetes diagnosis among patients with glucokinase mutation (GCK‐MODY) is affected by a genetic variant of glucose‐6‐phosphatase (G6PC2)

Aims  Genetic variation at the rs560887 locus of the glucose‐6‐phosphatase, catalytic 2 gene (G6PC2) is known to affect regulation of fasting glycaemia. We determined the rs560887 genotype of patients with monogenic diabetes and glucokinase gene mutations (GCK‐MODY) and correlated the genotypes with HbA1c levels.

Substantial proportion of MODY among multiplex families participating in a Type 1 diabetes prediction programme

Patients with maturity‐onset diabetes of the young (MODY) might be over‐represented in families with histories of Type 1 diabetes. Our aim was to re‐evaluate families participating in the Czech T1D Prediction Programme (PREDIA.CZ) with at least two members affected with diabetes to assess the proportion of MODY among these families and determine its most significant clinical predictors.

Lack of PAX4 mutations in 53 Czech MODYX families

While the most common causes for maturity-onset diabetes of the young (MODY) are heterozygous mutations in the genes HNF1A, GCK and HNF4A, other genes have also been implicated in MODY aetiology in isolated cases. One such gene is the PAX4 gene, an encoding transcription factor essential for development of insulin-producing cells [1]. The body of evidence supporting the role of PAX4 in the MODY aetiology is, however, limited. Only two MODY probands, both of Thai origin, have been found to carry heterozygous mutations in the PAX4 gene [1]. Of note, although the first mutation (p.Arg164Trp) impaired the molecule function in vitro, the pedigree of its carrier included mutation-negative phenocopies; thus, clear evidence is lacking for co-segregation with the phenotype. For the mutation IVS71G>A found in the other proband, no functional study could be performed and no relatives were available for genetic testing. The only patients with MODY tested for PAX4 outside the Thai population were 11 French and eight British subjects [2,3]—all negative for mutations. Interestingly, a Japanese study tested PAX4 in 393 individuals with Type 2 diabetes and found that approximately 3% of patients carried the p.Arg121Trp mutation [4]. However, the published data do not allow conclusions on the possibility that MODY was misclassified as Type 2 diabetes. As, to the best of our knowledge, no other published study of patients with MODY has focused on the PAX4 gene, we investigated this gene in a carefully selected and clinically welldocumented cohort of 53 Czech probands with MODY who were negative for mutations in the GCK, HNF1A, HNF4A, INS and NEUROD genes, and for structural variants in four common MODY genes (MODYX subjects). The procedure for identification of these MODYX patients among the nationwide Czech MODY registry is shown in Figure 1. The protocol was approved by the Ethics Committee of the University Hospital Motol, Prague. We first re-evaluated the clinical characteristics of all 281 Czech probands with MODY against the consensus for diagnosis of monogenic diabetes [5]. Briefly, they had to have a positive family history of diabetes in at least two generations, positive C-peptide and no islet autoantibodies. Then, all common MODY genes were tested for mutations as well as for structural rearrangements using techniques reported previously [6], with the exception of amplification of the exon 8 of the PAX4 gene, where long-range PCR (Qiagen, Hilden, Germany) was utilized to amplify the longer stretch of DNA flanking the exon. This test procedure identified 53 MODYX probands with median age at investigation of 33 years [interquartile range (IQR) 21.0–48.0] and median age at diagnosis of diabetes 19 years (IQR 13.5–30.5). The patients were on the edge of the normal and overweight categories, with a median BMI of 24 kg ⁄ m. The median fasting serum glucose was 6.7 mmol ⁄ l (IQR 6.3–8.3), the median C-peptide was 797 pmol ⁄ l (IQR 595.0–1190.0) and the median of HbA1c was 6.1% (IQR 4.5–8.2) and 43 mmol ⁄ mol (Diabetes Control and Complications Trial and International Federation of Clinical Chemistry, respectively). Most of the MODYX probands were treated with insulin (n = 30). Four probands were on oral hypoglycaemic agents and those with mild and stable hyperglycaemia were treated with diet (n = 8) or did not require any treatment (n = 4). PAX4 testing of these MODYX patients revealed no mutations. Our negative result, reached in a considerably larger group, confirms the previous findings from two other Caucasian populations [2,3]. This indicates that PAX4 is not an important contributor to MODY aetiology in Caucasians, yet it still may be important in the Asian populations: the genetic composition of MODY in Asia is clearly different to the Caucasian populations, as indicated by the lack of mutations in the three most common MODY genes (GCK, HNF1A or HNF4A): this was observed, for example, in studies from Korea and China [7,8]. In conclusion, we did not detect causal mutations in the PAX4 gene in a large and clinically well-characterized group of Czech MODYX probands, which may suggest—together with data from other European populations—that MODY in Caucasians FIGURE 1 Identification of the MODYX probands within the Czech MODY registry. DIABETICMedicine