Abdellatif Maalej

Analysis of skewed X-chromosome inactivation in females with rheumatoid arthritis and autoimmune thyroid diseases

IntroductionThe majority of autoimmune diseases such as rheumatoid arthritis (RA) and autoimmune thyroid diseases (AITDs) are characterized by a striking female predominance superimposed on a predisposing genetic background. The role of extremely skewed X-chromosome inactivation (XCI) has been questioned in the pathogenesis of several autoimmune diseases.MethodsWe examined XCI profiles of females affected with RA (n = 106), AITDs (n = 145) and age-matched healthy women (n = 257). XCI analysis was performed by enzymatic digestion of DNA with a methylation sensitive enzyme (HpaII) followed by PCR of a polymorphic CAG repeat in the androgen receptor (AR) gene. The XCI pattern was classified as skewed when 80% or more of the cells preferentially inactivated the same X-chromosome.ResultsSkewed XCI was observed in 26 of the 76 informative RA patients (34.2%), 26 of the 100 informative AITDs patients (26%), and 19 of the 170 informative controls (11.2%) (P < 0.0001; P = 0.0015, respectively). More importantly, extremely skewed XCI, defined as > 90% inactivation of one allele, was present in 17 RA patients (22.4%), 14 AITDs patients (14.0%), and in only seven controls (4.1%, P < 0.0001; P = 0.0034, respectively). Stratifying RA patients according to laboratory profiles (rheumatoid factor and anti-citrullinated protein antibodies), clinical manifestations (erosive disease and nodules) and the presence of others autoimmune diseases did not reveal any statistical significance (P > 0.05).ConclusionsThese results suggest a possible role for XCI mosaicism in the pathogenesis of RA and AITDs and may in part explain the female preponderance of these diseases.

Lack of Association of VDR Gene Polymorphisms with Thyroid Autoimmune Disorders: Familial and Case/Control Studies

We study the association between three Vitamin D receptor gene polymorphisms (rs10735810, rs1544410, rs731236) and susceptibility to thyroid autoimmune diseases. Seventy-six affected subjects, belonging to a large family, as well as one hundred unrelated Tunisian patients and one hundred healthy Tunisian controls were genotyped. A family-based association test and a standard chi-square test were used to assess association in family and case-control data, respectively. Our results showed no significant association of the Vitamin D receptor gene polymorphisms with the susceptibility to thyroid autoimmune diseases in the family. Moreover, allele frequencies for the three polymorphisms in the Tunisian population were similar to those reported in the Tunisian control population and none was associated with the disease. These results suggest a lack of association between the Vitamin D receptor gene polymorphisms and susceptibility to thyroid autoimmune diseases in Tunisian population, in agreement with data from the UK, but in conflict with studies from the Far East.

Association study of CARD8 (p.C10X) and NLRP3 (p.Q705K) variants with rheumatoid arthritis in French and Tunisian populations

The objective of the study was to investigate the association of caspase activating and recruitment domain 8 (CARD8) and nucleotide‐binding oligomerization domain, leucine‐rich repeat and pyrin domain containing 3 (NLRP3) polymorphisms with rheumatoid arthritis (RA) in Tunisian and French populations. CARD8 (c.30T>A, rs2043211) and NLRP3 (c.2113C>A, rs35829419) single nucleotide polymorphisms (SNPs) were genotyped in 100 French RA trio families and 141 Tunisian patients with RA and 191 unrelated healthy controls, using TaqMan® allelic discrimination assay. The genetic analyses for the association and linkage in French families were performed using the comparison of allelic frequencies (AFBAC), the genotype relative risk (GRR) and the transmission disequilibrium test (TDT). Data for case and control samples were analysed by chi‐square‐test, GRR and odds ratio (OR). No significant differences between alleles and genotypes frequencies were detected in French trio and Tunisian patients with RA and controls, either with CARD8 or with NLRP3 SNPs both in French and in Tunisian populations. Moreover, stratifying patients according to the presence of rheumatoid factor (RF), anti‐cyclic peptides antibodies (ACPA), erosion, nodules, other autoimmune disease or HLA‐DRB1*04‐positive subgroups did not show any significant association with CARD8 or NLRP3 (P ≥ 0.05). This study suggests that variations in the innate immunity genes CARD8 (p.C10X) and NLRP3 (p.Q705K) have no effect on RA susceptibility either in the Tunisian or in the French population.

Association of IRF5 gene polymorphisms with rheumatoid arthritis in a Tunisian population

Objective: A strong genetic association of rheumatoid arthritis (RA) with the interferon regulatory factor 5 (IRF5) gene has been described previously in a Swedish population, although this result was not confirmed in a French population. We undertook an association study between IRF5 and the RA phenotype, as well as a study with serological markers of RA, in a Tunisian population. Methods: A single‐nucleotide polymorphism (SNP; rs2004640) was genotyped using a Taqman 5′ allelic discrimination assay on an ABI 7500 real‐time polymerase chain reaction (PCR) instrument in 140 RA patients and 185 controls. Rheumatoid factor (RF) and anti‐citrullinated protein/peptide antibodies (ACPA) were determined by enzyme‐linked immunosorbent assay (ELISA). Association was assessed based on the χ2 test and odds ratios (ORs) with 95% confidence intervals (CIs). Results: The frequency of the TT genotype of the IRF5 SNP rs2004640 differed significantly between patients and controls (p = 0.01). This difference was greater when a subgroup of patients with another ‘autoimmune’ disorder was considered (p = 0.007). A weak but significant association was also found in a subgroup of patients who were positive for ACPA (p = 0.04) or erosion (p = 0.01). Conclusions: Our results indicate that the TT genotype of the IRF5 (rs2004640) dimorphism is associated with RA in a Tunisian population.

Association study of human leukocyte antigen-DRB1 alleles with rheumatoid arthritis in south Tunisian patients

The aim of this study is to explore relationship between HLA-DRB1 alleles and the susceptibility and clinical features of rheumatoid arthritis (RA) in the south Tunisian population. We studied 142 RA patients and 123 controls matched for age, sex, and ethnicity. HLA-DRB1 genotyping and HLA-DRB1*04 subtypes were performed using polymerase chain reaction/sequence-specific primers. Association was assessed based on the χ2 test and odds ratio with 95% confidence interval. For multiple comparisons, p value was corrected (pc) with Bonferroni test. Two alleles, HLA-DRB1*04 (p = 0.045, pc = NS) and HLA-DRB1*10 (p = 0.021, pc = NS), were found to have increased frequencies in RA patients compared to controls. In contrast HLA-DRB1*08 allele was found to have a decreased frequency in patients compared to controls (p = 0.044, pc = NS). Molecular subtyping of the most prevalent allele (DRB1*04) revealed increased frequencies of HLA-DRB1*04:05 in patients compared to controls (p = 0.013, pc = NS) whereas HLA-DRB1*04:02 showed a protective effect (p = 0.005, pc = 0.04). Moreover, stratified analyses indicated statistically significant associations between HLA-DRB1*04 allele and anti-cyclic peptides antibodies positivity (ACPA+) and rheumatoid factor positivity (RF+; pc = 0.03, for both subgroups), HLA-DRBI*10 and ACPA+ and the presence of another autoimmune disease (pc = 0.05 and pc = 0.007, respectively), and HLA-DRB1*04:05 and RF+ and erosion (pc = 0.005 and pc = 0.049; respectively). A significant decrease in the frequency of the DRB1*04:02 allele was observed in patients with ACPA+ and RF+ subgroups (pc = 0.04 and pc = 0.02, respectively). Our results showed that there was a trend of positive association of HLA-DRB1*04 and HLA-DRB1*10 with RA as such and significant associations with the disease severity in the south Tunisian population.

Allelic structure and distribution of 103 STR loci in a Southern Tunisian population.

Genotypes of 103 short tandem repeat (STR) markers distributed at an average of 40 cM intervals throughout the genome were determined for 40 individuals from the village of BirEl Hfai (BEH). This village of approximately 31.000 individuals is localized in the south-west of Tunisia. The allele frequency distributions in BEH were compared with those obtained for individuals in the CEPH (Centre d’Etude du Polymorphisme Humain) data using a Kolmo-gorov-Smirnov two-sample test. Fourteen out of the 103 markers (13.2%) showed significant differences (P<0.05) in distribution between the two populations. Population heterogeneity in BEH was indicated by an excess of observed homozygosity deviations from Hardy-Weinberg equilibrium at 3 loci (P<0.0005). No evidence for genotypic disequilibrium was found for any of the marker pairs. This demonstrated that in spite of a high inbreeding level in the population, few markers showed evidence for a different pattern of allelic distribution compared to CEPH.

Anticyclic citrullinated peptide antibody and rheumatoid factor in south Tunisian patients with rheumatoid arthritis: association with disease activity and severity.

To explore relationships between immunological status, clinical features, radiographic damage, disease activity, and functional disability in Tunisian patients with rheumatoid arthritis (RA).

Association study of MICA gene polymorphisms with rheumatoid arthritis susceptibility in south Tunisian population

The aim of this study was to investigate the role of major histocompatibility complex (MHC) class I chain‐related gene A (MICA) polymorphisms, important in natural killer (NK) cell function, in patients with rheumatoid arthritis (RA). A transmembrane (TM) alanine‐encoding GCT repeats, termed A4, A5, A5.1, A6 and A9 in the MICA gene, and single‐nucleotide polymorphisms (SNPs): the Met129Val polymorphism (rs1051792) and the nonsynonymously coding SNP (rs1051794) were genotyped in 142 patients with RA and 123 unrelated healthy individuals using, respectively, PCR fluorescent method, nested PCR‐RFLP and allele specific PCR (ASP). Association was assessed based on the χ2 test, genotype relative risk (GRR) and odds ratio (OR) with 95% confidence intervals (CIs). Our results show a trend of association of the different MICA genotypes G/G, G/A and A/A (P = 0.029) which did not attain the significance after Bonferronis correction (pc = 0.08). Although, we revealed a significant association of the genotype A/A of MICA‐250 in patients with RA compared to healthy controls (pc = 0.033). In contrast, no significant differences between alleles and genotypes frequencies were found either with MICA‐TM or MICA met129 val (P > 0.05) in our sample. Moreover, stratification of patients with RA according to clinical and immunological data for the different polymorphisms studied shows a significant association of both MICA‐250 G allele (pc = 0.0075) and MICA‐250 GG genotype (pc = 0.008) and both allelic (val) (pc = 0.021) and genotypic (val/val) distribution (pc = 0.0095) for MICA met129 val in the RF‐positive subgroup compared to RF‐negative patients with RA. In contrast, we found a strong association of the MICA‐TM A9 allele in RF‐negative patients with RA (pc = 0.0003). This study indicates the involvement of the MICA‐250 polymorphism in the genetic susceptibility and severity to RA and suggests that variations in MICA‐TM and MICA met129 val may have an effect on RA severity in our south Tunisian sample.

A Tunisian case-control association study of a 6q polymorphism in rheumatoid arthritis

Rheumatoid arthritis (RA), the most common autoimmune inflammatory disease of the joints, is a multifactorial disease, involving both genetic and environmental risk factors. TNFAIP3 (tumor necrosis factor, alpha-induced protein 3) gene located in a region of genetic susceptibility in RA is an attractive candidate to be involved in autoimmunity disorders. Our aim was to test the single nucleotide polymorphism rs6920220 located near TNFAIP3 in a case–control study in Tunisian population. The rare allele rs6920220-A was reported to have a risk effect on RA in several genome-wide association studies. Our results revealed a trend of an association of rs6920220-A allele with RA and genotypes containing this allele were in a higher proportion in RA patients than in matched controls. These findings have to be confirmed by a replication in largest RA and control groups of the same ethnic origin. TNFAIP3 gene may have a key role in autoimmunity through its action as a negative regulator of the NF-κB pathway. Further functional investigations are required to understand the mechanism by which this gene is involved in the RA pathogenesis.

No major genes in autoimmune thyroid diseases: complex segregation and epidemiological studies in a large Tunisian pedigree

1Unité Cibles pour le Diagnostic et la Thérapie, Centre de Biotechnologie Sfax, BP ‘1177’ 3018, Universiteé de Sfax, Sfax, Tunisia 2Unité d’ Epidémiologie Génétique et Moléculaire, Faculté de Médecine de Tunis, La Rabta-1007 Bab Saâdoun Tunis, Tunisia 3Unité de Bioinformatique, Centre de Biotechnologie Sfax, BP ‘1177’ 3018, Sfax, Tunisia 4Service Endocrinologie, CHU Hédi Chaker, Route el Ain, 3000, Sfax, Tunisia 5Laboratoire de Génétique Moléculaire Humaine, Faculté de Médecine, Sfax, Avenue Majida Bou Leila 3029, Sfax, Tunisia 6Cabinet d’Endocrinologie, Sfax, Avenue Majida Bou Leila 3029, Sfax, Tunisia 7Dispensaire Bir Hfay, 24005, Sidi Bouzid, Tunisia