TLR2 and TLR4 expression on CD14(++) and CD14(+) monocyte subtypes in adult-onset autoimmune diabetes.

BACKGROUND: Peripheral blood monocytes are key effectors of innate immunity. Dysfunction, changes in their counts or altered expression of cytokines and pattern-recognition receptors on monocytes may contribute to the development of the autoimmune type of diabetes mellitus (AD). AIMS: We aimed to analyze the counts and proportions of the two main subtypes of monocyte cells, CD14(++) and CD14(+), and to look for potential changes in the expression of toll-like receptors 2 (TLR2) and 4 (TLR4) as well as cytokine prolactin (PRL) in adult-onset AD, including diabetes mellitus type 1 (T1DM) and latent autoimmune diabetes in adults (LADA). METHODS: We examined 21 T1DM patients, 9 patients with LADA, 16 control patients with type 2 diabetes mellitus (T2DM) and 24 healthy individuals. All diabetic patients were diagnosed after the age of 18 years. Expression at the mRNA level was determined by quantitative PCR. Flow cytometry was used to ascertain membrane expression and cell counts. RESULTS: T1DM patients had fewer CD14(++) (P < 0.01) and CD14(+) (P < 0.0001) monocytes whereas T2DM subjects showed decreased counts of CD14(+) monocytes compared to healthy controls (P < 0.001). TLR2 protein expression was significantly increased in T1DM CD14(+) monocytes compared to healthy controls (P < 0.05), while TLR4 expression in T1DM CD14(++) cells was significantly lower (P < 0.0001). There was no significant difference between the groups in terms of PRL mRNA expression in monocytes. CONCLUSIONS: The observed changes in the proportions of both immune cell types and in the expression of functional pattern-recognition receptors on monocytes in the subjects examined may arise as a consequence of chronic inflammation that accompanies long-term diabetes.

The HLA-Cw*06 allele and -1149 G/T polymorphism of extrapituitary promoter of PRL gene as a possible common genetic predisposing factors to psoriasis vulgaris and psoriatic arthritis in Czech population.

Psoriatic arthritis (PsA) affects approximately 30 % of patients suffering from psoriasis vulgaris (PsV), but the risk factors for its development have not been well elucidated yet. The HLA-Cw*06 allele was described as a predisposing factor to PsV. Prolactin is known as an immune response modulator, and its elevated levels present risk for PsV development. It is possible that these factors interact and together emphasize the predisposition to both diseases. We tested on an association of HLA-Cw alleles and functional polymorphism -1149 G/T in PRL gene extrapituitary promoter with PsV and PsA in Czech population. We found a statistically significant association between HLA-Cw*06 allele and PsV (P corrected = 0.0013) that was most prominent in early onset disease subtype (P corrected = 0.0013). The association between HLA-Cw*06 and PsA was low (P corrected = 0.0585) and restricted to PsA patients with early PsV onset (P corrected = 0.0195). We found no association of -1149 G/T PRL gene polymorphism with either PsV or PsA.

HLA class II, MICA and PRL gene polymorphisms: the common contribution to the systemic lupus erythematosus development in Czech population.

The genetic components contribute to the systemic lupus erythematosus development. This study for the first time determined the distribution of the polymorphisms and linkage disequilibrium in HLA class II, MICA and PRL gene among patients suffering from SLE and healthy Czech individuals. DNA was obtained from the peripheral blood cells of 123 SLE patients and 96 healthy people. Allele variants of the HLA class II, MICA transmembrane polymorphism and PRL extrapituitary promoter -1149G/T SNP were detected using the sequence-specific primers analysis, PCR-fragment analysis and PCR-RFLP, respectively. In Czech population, only DRB1*03-DQB1*0201 haplotype is significantly associated with increased risk for SLE development: the frequency in SLE group was 44.7% in comparison with 15.2% in controls, P (c) < 0.0001; OR 4.54 CI 95% (2.36-9.09). The MICA-A5.1 allele is present significantly more often in SLE (55.7%) than controls (39.9%), P (c) = 0.005; OR 1.88 CI 95% (1.29-2.77), and the combination of HLA DRB1 *03 together with MICA-A5.1 is strongly associated with SLE [P (c) < 0.000001; OR 9.71 CI 95% (3.4-27.7)]. On the other hand, the MICA-A6 allele is less frequent in SLE patients compared to controls, 10.6% and 19.7%, respectively [P (c) = 0.035; OR 0.48 CI 95% (0.28-0.82)], and the combination of absence both alleles MICA-A6 and HLA DRB*11 seems to be risk for SLE development compared to controls, 84.6 and 70.2%, respectively, [P (c) = 0.0003 OR 2.32 CI 95% (1.47-3.70)]. We found that only G allele of the -1149 G/T SNP is associated with specific clinical manifestation of SLE, arthritis [P (c) = 0.022; OR 2.63, CI 95% (1.45-4.81)]. HLA class II-MICA combinations may increase/decrease a risk for SLE development. Multiple studies focusing on the ethnical differences as well as genetic-epigenetic relationships are necessary for better understanding SLE pathogenesis.

HLA-Cw*06 class I region rather than MICA is associated with psoriatic arthritis in Czech population.

Psoriatic arthritis (PsA) is a chronic inflammatory joint disease which affects patients suffering from psoriasis. The genetic background especially the susceptibility loci on the short arm of the chromosome six contribute to PsA development. In our study, we looked for the role of the MICA and HLA-Cw genes polymorphisms in PsA pathogenesis. We investigated 100 PsA patients and 94 healthy Czech individuals. We found an association between HLA-Cw*06 and PsA namely PsA with psoriasis type I (age of psoriasis onset before 40 years) compared to healthy individuals (P (corrected) < 0.05, OR 2.56, CI 95% 1.33-4.76 and P (corrected) = 0.01, OR 3.03, CI 95% 1.53-5.88, respectively). The MICA-A9 allele of the transmembrane microsatelite MICA polymorphism occurred more frequently in PsA with psoriasis type II group (age of psoriasis onset after 40 years) than in controls, 58.6 versus 38.0%, respectively however, this finding did not reach a statistical significance after correction (P (corrected) = 0.085).

Autoimmune diabetes mellitus with adult onset and type 1 diabetes mellitus in children have different genetic predispositions.

Type 1 diabetes with manifestation after 35 years of age is defined by CP <200 pmol/L and institution of insulin therapy within 6 months after diagnosis. Latent autoimmune diabetes mellitus in adults (LADA) manifesting after 35 years of age is defined by minimum 6 months after diagnosis without insulin therapy and C peptide (CP) >200 pmol/L and antiGAD > 50 ng/mL. We aimed to find a possible genetic discrimination among different types of autoimmune diabetes. To accomplish this goal, we analyzed DNA samples from 31 LADA patients, 75 patients with adult-onset type 1 diabetes mellitus, 188 type 1 diabetic children, and 153 healthy adult individuals. We studied five genetic loci on chromosomes 6, 11, 4, and 14: HLA DRB1 and DQB1 alleles, major histocompatibility complex (MHC) class I-related gene-A (MIC-A) microsatellite polymorphism, interleukin (IL)-18 single nucleotide polymorphism, the microsatellite polymorphism of nuclear factor kappa B gene (NF-kappaB1), and the single nucleotide polymorphism of a gene for its inhibitor (NF-kappaBIA). HLA-DR3 was detected as the predisposition allele for LADA (OR = 4.94, P < 0.0001). Further we found a statistically significant increase of NF-kappaBIA AA genotype (OR = 2.68, P < 0.01). On the other hand, DRB1*04, which is linked with DQB1*0302, was observed as a risk factor in patients with type 1 diabetes mellitus (T1DM) onset after 35 years of age (OR = 10.47, P < 0.0001 and OR = 9.49, P < 0.0001, respectively). There was also an association with MIC-A5.1 (OR = 2.14, P < 0.01). Statistically significant difference was found in the distribution of IL-18 promoter -607 (C/A) polymorphism between LADA and T1DM in adults (P < 0.01). We conclude that all subgroups of autoimmune diabetes have partly different immunogenetic predisposition.

Amplification of NOTCH1 and ABL1 gene loci is a frequent aberration in enteropathy-type T-cell lymphoma.

We have shown previously that amplification of chromosomal region 9q34 is the most frequent aberration in enteropathy-type T-cell lymphoma (ETL). To determine the minimum amplified 9q34 region and identify possible candidate gene(s), we performed a detailed microsatellite screening and quantitative real-time PCR (QPCR) on 26 ETL cases. Microsatellite analysis revealed allelic imbalance in both ABL1 and NOTCH1 gene loci (microsatellites D9S290-D9S1847 and D9S158 flanking the former and latter genes, respectively) localized in the band 9q34. The results were confirmed by TaqMan-based QPCR showing amplification of ABL1 and NOTCH1 exons in 50% and 65% of cases, respectively. Amplifications of the NOTCH1 gene were more frequent than of the ABL1 gene; moreover, the analyzed NOTCH1 exon consistently displayed higher levels of amplification than ABL1 coding sequences. From 9q34 known genes, NOTCH1 could thus be the primary target of genomic DNA amplification in ETL.

Diabetes mellitus in adults: association of HLA DRB1 and DQB1 diabetes risk alleles with GADab presence and C-peptide secretion.

In our study, we investigated the relationship of HLA class II alleles to antibody production against glutamic acid decarboxylase (GADab) and to C-peptide secretion (CP) in diabetic patients. A group of 334 patients (190 women) diagnosed after 35 years of age and 99 control subjects were studied. Patients were divided into four groups according to concentrations of CP and GADab, respectively (CP high/low, GADab positive/negative). HLA DQB1 and DRB1 alleles were genotyped by SSP-PCR. The significance of DQB1 and DRB1 risk alleles was evaluated by examination of their odds ratios computed by testing 2x2 tables considering Bonferonis' corrected P<0.05 as significant. We found strong association between the HLA DRB1*03 risk allele and presence of GADab, and close relationship of the HLA DRB1*04 and HLA DQB1*0302 risk alleles with decreased CP level. Taken together we conclude that the DRB1*04 and DQB1*0302 alleles are associated with progressive decrease of CP level, while DRB1*03 is a significant genetic marker of autoantibody (GADab) development.