Yousr Gorgi

Distribution of HLA-B*27 subtypes in Tunisians and their association with ankylosing spondylitis.

OBJECTIVE The human leukocyte antigen HLA-B27 is a class I antigen of the major histocompatibility complex strongly associated with ankylosing spondylitis (AS) and other related spondyloarthropathies (SpAs). The mechanism of this association remains unknown. HLA-B27 is a serologic specificity that represents a family of at least 25 different HLA-B27 alleles (2701-2725). These alleles are closely related by nucleotide sequence homology, but differ in ethnic distribution. The purpose of the present study is to investigate the distribution of HLA-B27 alleles in healthy controls and in patients with ankylosing spondylitis (AS). METHODS We selected 160 HLA-B27-positive individuals (39 controls and 121 patients with ankylosing spondylitis). Typing of HLA-B27, and Cw alleles were performed by polymerase chain reaction amplification with sequence specific primers (PCR-SSP), and by serological typing (microlymphocytotoxicity). RESULTS Seven B27 subtypes were identified: B*2702, 03, 04, 05, 07, 09 and B*2714. The distribution of these alleles in the population of patients was B*2702 (47.1%) and B*2705 (47.1%). These subtypes were also detected in 16 (41%) and 16 (41%), respectively of the 39 control subjects. HLA-B*2707 was detected in 4 (3.31%) patients and in 3 (7.6%) controls. B*2704, 09 and B*2714 were relatively rare and were detected in one subject each. No significant differences were noticed in the frequencies and distribution of HLA-B27 alleles between patients and controls. CONCLUSIONS Our results show a restricted number of HLA B27 subtypes associated with AS. B*2702 and B*2705 were equally common in patients and controls. The most prominent B27/Cw haplotypes in the patient groups and controls were B*2702/Cw02022 and B*2705/Cw02022.

Polymorphism in ICAM-1, PECAM-1, E-selectin, and L-selectin genes in Tunisian patients with inflammatory bowel disease.

Background Ulcerative colitis (UC) and Crohns disease (CD) are chronic intestinal disorders characterized by immune dysregulation and leukocytes recruitment into gastrointestinal tract. Cell adhesion molecules (CAM) mediate the extravasation of leukocytes and their accumulation in inflamed intestinal mucosa. Recently, CAM genes have been implicated in determining susceptibility to UC and CD. We investigate seven mutations in CAM: G241R and K469E in ICAM-1, V125L in PECAM-1, G98T, S128R, and L554F in E-selectin and F206L in L-selectin in 197 Tunisian patients (73 with UC and 124 with CD) and 194 controls. These polymorphisms were detected by polymerase chain reaction sequence-specific primers and restriction enzyme analysis. Results A significant increase in allele frequencies of 206L of L-selectin and the associated genotype F/L was observed in both patients with UC and CD compared with controls. Subgroup analysis showed that the L206 allele and F/L206 genotype frequencies were significantly increased in UC patients with left-sided type; whereas, the F/L206 genotype was significant in CD patients with ileocolonic location and stricturing behavior compared with controls. No significant differences in allele or genotype frequencies were observed for ICAM-1 K469E, E-selectin, and PECAM-1 polymorphisms between UC patients, CD patients, and controls. Conclusion We found an association of inflammatory bowel disease with allele L206 of L-selectin gene, whereas genotype L/F was associated with a subgroup of UC (left-sided type) and CD patients with more extensive location of disease and stricturing behavior. However, further studies are needed to confirm our findings.

Cytokine Gene Polymorphisms in Kidney Transplantation

BACKGROUND Acute and chronic rejections remain an important cause of graft loss after renal transplantation. It has been suggested that cytokine genotyping may have a predictive role to identify patients at greater risk of rejection regardless of human leukocyte antigen (HLA) compatibility and/or the presence of anti-HLA antibodies before the renal allograft. OBJECTIVES We sought to investigate polymorphisms of tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, interleukin (IL)-10, IL-6, and interferon (IFN)-γ as indices of differential cytokine production in kidney transplantation and to examine their predictive roles for acute or chronic rejection. PATIENTS AND METHODS TNF-α (G/A -308), TGF -β1 (haplotype codon 10/codon 25), IL-10 (haplotype-1082, -819, -592), IL-6 (C/G -174), and IFN-γ (T/A +874) single nucleotide polymorphisms (SNPs) were detected using polymerase chain reaction (PCR)-specific sequence primers (SSP) in 231 kidney transplant recipients (KTR) including 106 treated with mycophenolate mofetil (MMF+). RESULTS We observed no significant associations of any of investigated polymorphism taken alone with acute rejection episodes (ARE) or chronic allograft dysfunction (CAD). Nevertheless, TGF-β1 Low (L) production was correlated with greater graft survival at 20 years (81.8%) compared with intermediate (L) or high (H) levels (56.1%), affect that the difference was not significant (P = .2). Cytokine haplotype analysis in KTR (MMF-) without HLA-mismatches or presynthesized anti-HLA antibodies (n = 32) showed ARE to be significantly more prevalent among the TNF-α*H/TGF- β1*H/IL-10*H production haplotype (75%) compared with the other haplotypes (16%; P = .03). CONCLUSION The presence of TGF-β1-H secretion profile may protect the kidney graft. TNF-α*H/TGF-β1*H/IL-10*H haplotype was associated with a higher risk of ARE and with poorer graft survival.

Polymorphisms of Toll-like receptor-4 and CD14 in systemic lupus erythematosus and rheumatoid arthritis.

BackgroundToll-like receptor 4 (TLR4) and its co-receptor CD14 play a major role in innate immunity by recognizing PAMPs and signal the activation of adaptive responses. These receptors can recognize endogenous ligands mainly auto-antigens. In addition, TLR4 (Asp299Gly) and CD14 (C/T -159) polymorphisms (SNPs) may modify qualitatively and/or quantitatively their expression. Therefore, they could be implied in autoimmune diseases and can influence both susceptibility and severity of systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA).Patients and methodsTLR4 (Asp299Gly) and CD14 (C/T -159) SNPs were genotyped using polymerase chain reaction (PCR)-RFLP in 127 SLE patients, 100 RA patients, and 114 healthy controls matched in age and gender.ResultsCD14*T allele was significantly more frequent in SLE patients (0.456) comparatively to controls (0.355), p = 0.02 OR (95% CI) = 1.53 [1.04-2.24]. In RA patients, the higher frequency of CD14*T allele (0.405) failed to reach significance, p = 0.28. Investigation of the TLR4 (Asp299Gly) SNP showed no significant association neither with SLE nor with RA.Analysis of these SNPs according to clinical and biological features showed a significant higher frequency of arthritis in SLE patients carrying CD14*T/T genotype (92%) comparatively to those with C/C and C/T genotypes (72.5%), p = 0.04. Moreover, SLE patients carrying CD14*T/T/TLR4*A/A haplotype had significantly more arthritis (91.3%) than the rest of SLE group (73%), p = 0,044 and confirmed by multivariable analysis after adjustment according to age and gender, p = 0.01.ConclusionThe CD14 (-159)*T allele seems to be associated with susceptibility to SLE and arthritis occurrence.

THE INVOLVEMENT OF HLA -DRB1, DQA1, DQB1 AND COMPLEMENT C4A LOCI IN DIAGNOSING SYSTEMIC LUPUS ERYTHEMATOSUS AMONG TUNISIANS

Background Genetic susceptibility to systemic lupus erythematosus (SLE) varies among populations. Few data exist on associations of HLA class II and class III alleles of the major histocompatibility complex (MHC) and susceptibility to SLE in Tunisians. Patients and Methods We compared HLA-DRB1*, DQA1, DQB1* and C4 allotypes in 62 Tunisian SLE patients and 100 matched controls. We also assessed the association of specific alleles with distinct autoantibody profiles in SLE patients. Results HLA-DRB1*0301, -DRB1*1501 and C4AQO alleles were increased in the SLE patients, while the frequencies of HLA-DRB1*04 and DQB1*03 were decreased. HLA-DQA1*0102 and DQA1*0501 were significantly increased in the SLE patients. HLA-DQB1*0201 and DQB1*0602 were more frequent in the SLE patients. C4A*QO and C4B*QO were increased in frequency in the SLE patients compared to the controls, but only C4A null was significantly increased. Eleven of 17 SLE patients with the C4 null allele were HLA-DRB1 *0301 positive. Three of 16 SLE patients with HLA-DRB1*1501 were associated with HLA-DQB1*0501 rather than DQB1 *0602, as has been reported in European SLE patients. Conclusions The MHC class II alleles (DRB1, DQA1, DQB1) and C4 null associations noted in other ethnic groups are also found in Tunisians, suggesting shared susceptibility factors across ethnic lines in predisposition to SLE. In contrast to other ethnic groups, MHC class II alleles are not associated with the presence of specific autoantibodies in Tunisian SLE patients.

Interleukin-18 gene polymorphisms in tunisian patients with inflammatory bowel disease.

Aim: Interleukin (IL)-18 can regulate the Th2-mediated immune response and it may be involved in the pathogenesis of Th1 and Th2 chronic inflammatory diseases. This study sought to detect a possible association between two single nucleotide polymorphisms (SNPs) (–137G/C and –607C/A) in the IL-18 gene promoter region and susceptibility to inflammatory bowel disease (IBD) including Crohn’s disease (CD) and ulcerative colitis (UC) in the Tunisian population. Methods: The (–137G/C and –607C/A) IL-18 polymorphism was analyzed in 105 patients with CD, 59 patients with UC, and 100 controls using the sequence-specific polymerase chain reaction method. Results: The distribution of allele and genotype frequencies illustrate that the –137G/G genotype frequency was significantly higher in UC than in controls (p value corrected (pc) = 0.038). On the other hand, we found a statistically significant association (pc = 0.033) between genotype AA of the IL-18 gene promoter (–607C/A) polymorphism in UC patients and the distal localization of the lesions. In CD, no significant differences were observed at positions –607 and –137. The analysis of IBD patients according to clinical behavior revealed no difference. Conclusion: The two SNPs at position –607 (C/A) and –137 (G/C) in the promoter region of the IL-18 gene was associated with the development of UC but not CD, providing a strong support for an IBD susceptibility gene in the region surrounding IL-18. It remains to be determined precisely how the IL-18 alleles influence the pathogenesis of IBD.

New C1q mutation in a Tunisian family

Hereditary C1q deficiency (C1qD) is the most penetrant genetic factor predisposing to the development of lupus pathology with more than 93% of C1q deficient patients developing this autoimmune pathology throughout their life. It is a rare autosomal recessive deficiency, with only 67 cases reported so far including one Tunisian girl who died at the age of three from complications resulting from severe systemic lupus erythematosus. Although C1qD was confirmed in the serum of this patient using C1q ELISA and classical pathway specific functional assays, no DNA sample had been obtained from this patient. Here we report the analysis of sera and DNA of members of this patients closer family. Our analysis identified a homozygous mutation within the gene encoding the C-chain of C1q leading to a deficiency of C1q in an older sister of our original patient. This mutation, termed g.5580G4C, represents a single basepair substitution in exon 1 of the C1q C chain gene which changes the codon of Gly61 to Arg 61. Amongst the other 14 mutations leading to C1qD, g.5580G4C represents the first reported transversion leading to human C1qD.

(AT) Repeat in the 3′ Untranslated Region of the CTLA-4 Gene and Susceptibility to Acute Allograft Rejection in Tunisian Renal Transplantation

Allograft rejection is an immune response relying on the proliferation and the differentiation of T cells. CTLA-4 is a co-stimulatory molecule, expressed on activated T lymphocytes, which has been shown to play a crucial role in the down-regulation of T-cell activation. Herein, we have examined the impart of a genetic marker in the CTLA-4 gene on renal transplant outcomes. A cohort of 144 renal recipients and 100 healthy subjects were genotyped by the fragments analysis method using an automated sequencer. Patients were classified into two groups: Group I included 31 HLA-identical haplotype allograft recipients and Group II, 113 showing one or more HLA haplotype mismatches. Forty patients (27.78%) developed at least one acute rejection episode (ARE): 9 in Group I and 31 in Group II. Before transplantation, 20 patients were lymphocytotoxic antibodies (LCT) positive: 4 Group I, 2 of whom developed an ARE, and sixty in Group II, including 8 with an ARE. The occurrence of an ARE was associated with the presence of LCT before transplantation among the entire cohort of patients (P = .032) and among Group II (P = .037). The allelic frequencies of (AT)n polymorphism did not reveal significant differences between patients and controls. The most prevalent alleles were the 88 bp (51% in controls and 44.44% in patients) and the 106 bp (8% and 10.76%, respectively). We noticed an increase of the 120 bp allele frequency among patients who had undergone an ARE compared with those who did not display this complication (8.75% vs 3.85%). Likewise, among LCT-negative Group I, recipients the incidence of the 120 bp allele was higher in ARE than non-ARE patients. Although the differences were not statistically significant, we propose that the 120 bp allele of the CTLA-4 gene (AT)n microsatellite a predisposes to acute rejection episodes in renal transplantation.

Phylogenetic Analysis of Isolated HCV Strains from Tunisian Hemodialysis Patients

The present study describes the strains of hepatitis C virus (HCV) isolated from Tunisian hemodialysis patients. Thirty-three HCV strains isolated from different dialysis centers in Tunis City were amplified by RT-PCR in a region of the NS5b gene, genotyped by sequencing, and compared to international sequences by phylogenetic analysis. The phylogenetic tree showed that 16 HCV isolates have been identified as subtype 4k (48.5%), 7 as unspecified HCV-4 subtype (21.2%), 5 as subtype 4a et 1b (each 15.2%). The analysis of this tree revealed that the HCV-1b strains were closely related to Anglo-Saxon and European isolates, while the HCV-4 isolates are genetically similar to Egyptian and African strains. Phylogenic analysis of 33 Tunisian isolates with international HCV strains on a region of the NS5b gene demonstrated that the subtype 4k submerged the Tunis city and a new subtype of HCV4 seems to be suspect in this area.

Prediction of T Cell Epitopes from Leishmania major Potentially Excreted/Secreted Proteins Inducing Granzyme B Production.

Leishmania-specific cytotoxic T cell response is part of the acquired immune response developed against the parasite and contributes to resistance to reinfection. Herein, we have used an immune-informatic approach for the identification, among Leishmania major potentially excreted/secreted proteins previously described, those generating peptides that could be targeted by the cytotoxic immune response. Seventy-eight nonameric peptides that are predicted to be loaded by HLA-A*0201 molecule were generated and their binding capacity to HLA-A2 was evaluated. These peptides were grouped into 20 pools and their immunogenicity was evaluated by in vitro stimulation of peripheral blood mononuclear cells from HLA-A2+-immune individuals with a history of zoonotic cutaneous leishmaniasis. Six peptides were identified according to their ability to elicit production of granzyme B. Furthermore, among these peptides 3 showed highest affinity to HLA-A*0201, one derived from an elongation factor 1-alpha and two from an unknown protein. These proteins could constitute potential vaccine candidates against leishmaniasis.