Interleukin-4 Gene Polymorphisms in Romanian Patients with Inflammatory Bowel Diseases: Association with Disease Risk and Clinical Features.

1. INTRODUCTION: Multiple cytokines have been studied for their role in the propagation of the inflammatory process related to inflammatory bowel diseases (IBD), but the role of interleukin-4 remains controversial. The aim of this study was to evaluate the role of two IL-4 gene single nucleotide polymorphisms (SNPs) in disease susceptibility and phenotypic expression. 2. MATERIALS AND METHODS: A group of 160 patients with IBD (86CD/74UC) and 160 healthy controls were genotyped for IL-4 rs2243250/-590C/T and rs2070874/-34C/T using real-time polymerase chain reaction with TaqMan assay. 3. RESULTS: The analysis of IBD patients and controls revealed a significantly reduced frequency of the minor allele T of both SNPs in CD patients (p = 0.03, OR 0.55 and p = 0.02, OR 0.52) and for the entire IBD group (p = 0.01, OR 0.57 and p = 0.01, OR 0.55). Haplotype analysis identified the most frequent haplotype (rs2243250/rs2070874 CC) associated with a high risk for developing IBD (either UC or CD) (p = 0.003). IBD patients with extraintestinal manifestations had significantly increased frequency of the minor alleles T. We also found an association between the presence of allele C of rs2070874 and response to antiTNF treatment. 4. CONCLUSIONS: This is the first study to investigate the IL-4 gene's relation to IBD susceptibility conducted in Romania. Both SNPs were found to be associated with disease susceptibility and phenotypic features, such as extraintestinal manifestations and response to antiTNF agents.

Genetic Variants of Interleukin-4 in Romanian Patients with Idiopathic Nephrotic Syndrome.

Background and objectives: One of the most frequent glomerular diseases in the pediatric population is represented by the idiopathic nephrotic syndrome (INS). The exact mechanisms mediating the disease are still unknown, but several genetic factors have been studied for possible implications. Cytokines are considered to play a pivotal role in mediating INS disease progression, interleukin-4 (IL-4) exhibiting particular interest. The objective of this research project was to investigate the association between two IL-4 gene single-nucleotide polymorphisms (SNPs) and INS susceptibility as well as response to steroid therapy, in a group of Romanian children. Materials and Methods: In total, 75 patients with INS and 160 healthy controls of Romanian origin were genotyped for IL-4 rs2243250/-590C/T and rs2070874/-34C/T using real-time polymerase chain reaction. Association tests were performed using the DeFinetti program and Plink 1.07 software and p-values < 0.05 were considered statistically significant. Results: The analysis of INS patients and controls revealed a similar genotype distribution of the studied SNPs. The minor T alleles were less frequent in the INS group, but not statistically significant (p = 0.1, OR = 0.68 and p = 0.2, OR = 0.74). Regarding the response to steroids, a low frequency of 590*T allele in steroid-resistant patients (7.7%), compared with steroid-sensitive patients (14%) and controls (17.5%), was obtained, but the difference did not reach the statistical significance threshold. The same result was obtained for -34C/T SNP. Conclusions: This is the first study examining the relationship between the IL-4 gene and INS susceptibility conducted in a European population, and particularly in Romania. The investigated SNPs were found to not be associated with disease susceptibility or response to the steroid treatment of pediatric INS.

Preliminary Study Regarding the Association between Tumor Necrosis Factor Alpha Gene Polymorphisms and Childhood Idiopathic Nephrotic Syndrome in Romanian Pediatric Patients.

BACKGROUND: Childhood idiopathic nephrotic syndrome (INS) is one of the most common glomerular diseases, characterized by heavy proteinuria, hypoalbuminemia, dyslipidemia and generalized edema. Although some progresses were made regarding the pathogenesis of this disease, there are a lot of questions still left unanswered. Some of them involve the implications of several cytokines, including tumor necrosis factor alpha (TNF-alpha), in the development and clinical course of INS. OBJECTIVE: Our objective was to analyze the role of two single nucleotide polymorphisms of TNF-alpha gene in the development of pediatric INS and their implication in the response to corticosteroid therapy. MATERIAL AND METHODS: Seventy patients with INS and 159 healthy controls were included in this study. They were analyzed for TNF-alpha gene polymorphisms by using polymerase chain reaction. The two SNPs (rs1799724/-857C/T and rs1800629/-308G/A) were genotyped by TaqMan Genotyping Assays, association tests were performed and p values <0.05 were considered significant. RESULTS: Minor alleles frequencies were 15.72% in INS patients versus 18.55% in controls for 857*T allele and 11.43% in INS versus 13.2% in controls for 308*A allele. Although the minor alleles were more frequent in controls than in patients, the difference was not statistically significant (p=0.46, OR=0.818 and p=0.59, OR=0.848).Analyzing the response to corticosteroid therapy, we found a low frequency of 857*T allele in steroid resistant patients (9.09%) compared to steroid sensitive patients (16.95%) and controls (18.55%). Regarding 308*A allele, the frequencies were 18.18% in the corticoresistant group and 10.17% in the corticosensitive one. None of them was statistically significant (p>0.05). CONCLUSIONS: We conclude that neither -857C/T, nor-308G/A polymorphisms of TNF-alpha gene are associated with the susceptibility and response to steroid treatment of INS in our population. Given the small sample size used, future studies are necessary to clarify the results observed in the present study.

ERAP1 and ERAP2 Gene Variations Influence the Risk of Psoriatic Arthritis in Romanian Population.

Psoriatic arthritis (PsA) is a chronic inflammatory disorder that belongs to the group of spondyloarthritis (SpA). It was found that single nucleotide polymorphisms (SNPs) of endoplasmic reticulum aminopeptidase (ERAP1 and ERAP2) genes influence the risk of ankylosing spondylitis, the most common form of SpA and the risk of psoriasis. Our purpose was to investigate the possible association of ERAP1 and ERAP2 gene SNPs with psoriatic arthritis susceptibility in Romanian population. Subsequent analyses included patients' subgroups according to HLA-B27 status. Psoriatic arthritis patients (N = 98) and random healthy controls (N = 139) were genotyped for ERAP1/2 genes SNPs rs30187, rs27044, rs2910686, and rs2248374 by TaqMan Allelic Discrimination Assays. An additional control group (N = 108; 100% HLA-B27 positive) was used for subsequent analyses. The results showed the association of rs2248374 SNP of ERAP2 gene with the risk of PsA, especially for HLA-B27 negative disease (p = 0.02; OR 1.59). ERAP2 haplotype GT (rs2248374/rs2910686) was significantly under-represented in PsA patients than in controls (43 vs. 55%; p = 0.02). The analysis of ERAP1 SNPs in HLA-B27 positive controls and PsA subgroup showed strong evidence of association for rs30187 (p = 0.005; OR 2.73) and for CC rs30187/rs27044 haplotype (47% in patients vs. 70.5% in controls; p = 0.006). In conclusion, we found a significant association of ERAP2 with PsA and HLA-B27 negative PsA, while ERAP1 association was restricted only to HLA-B27 positive disease. To our knowledge, this is the first study that investigated ERAP2 polymorphisms in relation to PsA susceptibility.

Association of Leukotriene C4 Synthase A-444C Polymorphism with Asthma and Asthma Phenotypes in Romanian Population.

INTRODUCTION: Leukotriene C4 synthase (LTC4S) gene -444A/C polymorphism has been implicated in susceptibility to asthma, but a large number of studies have reported inconclusive results. The aim of this study was to investigate the association between the -444A/C polymorphism of LTC4S gene and asthma, asthma phenotypes (aspirin intolerant/tolerant asthma) and different characteristics of the patients. MATERIAL AND METHODS: We included 106 patients with asthma (60 with aspirin tolerant asthma - ATA, 46 with aspirin intolerant asthma - AIA) and 103 controls. All the subjects were genotyped for LTC4S-444 A/C by Real-Time PCR. We assessed the association of LTC4S promoter polymorphism with asthma and its phenotypes and with clinical and biological characteristics of asthmatic patients. RESULTS: We did not find a significant association between the studied polymorphism and asthma, but the minor allele tended to be more frequent in AIA patients. We found a significant association between the minor allele C and lower levels of serum total immunoglobulin E and eosinophils, suggesting a possible role of -444A/C LTC4S polymorphism as modulating factor of allergic inflammation in asthma. CONCLUSION: The results show that LTC4S -444A/C SNP is not associated with susceptibility to asthma in Romanian patients, but could influence asthma phenotype, namely aspirin intolerant asthma.

The clinical value of human leukocyte antigen HLA-DRB1 subtypes associated to Graves' disease in Romanian population.

The aim of this study was to identify the primary susceptibility HLA-DRB1 alleles associated with GD in Romanian population and to seek whether specific HLA-DRB1 haplotypes are associated with differences in the clinical presentation of GD at diagnosis. Molecular typing of HLA-DRB1 alleles was performed in 77 Romanian Caucasian GD patients and 445 racially matched controls. In GD patients, age, presence of eye disease, goiter grade, autoantibody status and titer, TSH, FT4, FT3, TT3 levels were recorded at diagnosis. The allelic frequencies of HLA-DRB1*03 (41.55% vs. 17.75%, p < 0.0001, χ(2) = 20.81) and DRB1*11 (42.85% vs. 30.56%, p = 0.045, χ(2) = 3.98)were higher, whereas those of HLA-DRB1*01(3.89% vs. 16.40%, p = 0.007, χ(2) = 7.281) and DRB1*15 (10.38% vs. 21.34%, p = 0.038, χ(2) = 4.309)were lower in GD patients than in controls. FT4/TT3 ratio (p = 0.015) and anti-thyroglobulin antibodies (p = 0.024) were higher in *03/11 patients compared to *X/X, *11/Z, *03/Y patients (where X is any other allele than *03 and *11, Y is any other allele than *11, Z is any other allele than *03). In conclusion, HLA-DRB1*03 and DRB1*11 may be the primary susceptibility HLA-DRB1 alleles associated with GD in Romanian population, whereas HLA-DRB1*01 and DRB1*15 seem to be protective. At diagnosis, HLA-DRB1*03/11 GD patients had higher FT4/TT3 ratio and anti-thyroglobulin antibody levels.

Association study in Romanians confirms IL23A gene haplotype block rs2066808/rs11171806 as conferring risk to psoriatic arthritis.

BACKGROUND: The cytokines IL12 and IL23 have been recently implicated in the pathogenesis of psoriatic arthritis (PsA). In this study we investigated the genetic variations in the genes coding for IL12, IL23 and IL23 receptor as a plausible source of susceptibility and modification of clinical symptoms of PsA in Romanian population. METHODS: Twenty five SNPs mapping to IL12A, IL12B, IL23A, IL23R and IL12RB1 genes were genotyped in 94 PsA patients and 161 healthy controls of Romanian ethnicity using the Sequenom genotyping platform. RESULTS: The exonic SNP rs11171806 from IL23A gene was significantly underrepresented in patients versus controls (p=0.03, OR 0.391) and the carriers of rs11171806/rs2066808 AC haplotype had decreased risk for PsA (p=0.03). The two SNPs of the highly conserved gene IL23A are in complete LD in our population. Genetic variants of IL12B gene were associated with polyarticular subtype of PsA. No associations were found between SNPs from IL12A, IL23R and IL12RB1 genes and susceptibility to PsA and its phenotypes. CONCLUSION: We confirm the previously described association of rs2066808 variant with psoriasis and PsA and we show evidence of an extended genomic region inside IL23A gene as carrier of true disease susceptibility factors. These data suggest a role for IL23 in the PsA pathogenesis in Romanians.

Prospective antibody case finding of coeliac disease in type-1 diabetes children: need of biopsy revisited.

AIM: To evaluate whether coeliac disease (CD) can be diagnosed by measuring autoantibodies without small-intestinal mucosal biopsies in children with type 1 diabetes. METHODS: Case finding of CD was undertaken in 181 consecutive IgA-competent children with type 1 diabetes using transglutaminase 2 (TG2) and endomysial IgA antibody (EMA) tests in serum and the rapid point of care test in fingertip whole-blood sample. Endoscopy with intestinal biopsies was recommended for patients with high TG2-IgA titres (>96 U) and in children with lower positive tests if either the EMA test or the rapid point of care test was additionally positive. The duodenal mucosal biopsies were graded according to the Marsh classification. RESULTS: The TG2-IgA test had a 15.5% and the EMA test a 6.0% seropositivity. All seven biopsied high-titre TG2-IgA-positive children were symptom free and found to have CD (Marsh 3 type lesion). These patients were also positive for EMA and in the rapid point of care test. Lower titre TG2-IgA-positive children had histological Marsh 1 to 3a lesions. CONCLUSIONS: None of the type 1 diabetes children with high TG2-IgA titres would have needed endoscopy with duodenal biopsies to reach a CD diagnosis. Lower TG2-IgA-positive patients need to be biopsied.

A pilot study of the association of tumor necrosis factor alpha polymorphisms with psoriatic arthritis in the Romanian population.

Tumor necrosis factor alpha (TNF-alpha) is an important pro-inflammatory cytokine implicated in the pathogenesis of psoriatic arthritis. We have performed a case-control association study of three TNF-alpha gene polymorphisms in a group of Romanian psoriatic arthritis patients versus ethnically matched controls. A second group of patients with undifferentiated spondyloarthritis was used in order to look for similarities in the genetic background of the two rheumatic disorders. The -857C/T polymorphism was associated with susceptibility to psoriatic arthritis in our population at the individual level (p = 0.03, OR 1.65, 95% CI 1.05-2.57) and in combined haplotypes with the -238G/A and -308G/A SNPs. Regarding the investigated polymorphisms and derived haplotypes, no potential association was found with the susceptibility to undifferentiated spondyloarthritis in Romanian patients.