Agnès Hamzaoui

Expression of Th-17 and RORgammat mRNA in Behçet’s Disease

Summary Background To investigate plasma IL-17 level and the expression of Th17 cell transcription factor RORγt in the pathogenesis of Behçet’s Disease (BD). Material/Methods Blood samples were collected from 73 patients with BD (45 patients were in active stage), 20 systemic lupus erythematosus (SLE) and 12 multiple sclerosis patients (MS). Twelve patients with BD were investigated both in their active and remission stages. Samples were processed to detect IL-17A level in plasma by enzyme-linked immunosorbent assay (ELISA). Related gene expression was assessed by real-time reverse transcription polymerase chain reaction. Function of Th17 cells in active BD patients with erythema nodosum (EN)-like eruption was studied in relation to human umbilical vein endothelial cells (HUVECs). Results We demonstrated the presence of Th17 cells and RORγt among the peripheral blood mononuclear cells (PBMC). The percentage of circulating Th17 cells and the ability to produce interleukin-17A (IL-17A) were increased in samples derived from patients with active BD, MS and SLE patients. We observed that IL-17A from patients with active BD could induce adhesion molecule messenger RNA expression in HUVECs. Conclusions RORγt determined Th17 cell might be involved with increased IL-17A in BD. Our results indicate that IL-17 contributes to the active proinflammatory pattern that is characteristic of inflammatory diseases and patients with active BD.

Induced sputum levels of IL-33 and soluble ST2 in young asthmatic children

Abstract Objective: Interleukin-33 is an IL-1 family cytokine which signals via its T1/ST2 receptor, and acts as a key regulator of inflammation, notably the type-2 response implicated in asthma. This study aims to measure the expression of soluble ST2 (sST2) and IL-33 in asthmatic children, depending on disease activity. Methods: Thirty-seven children with well-defined asthma (20 moderate and 17 mild asthmatics) were studied. IL-33 and sST2 were measured by ELISA in serum and induced sputum (IS) samples, and compared with 22 age- and sex-matched healthy controls. Real-time quantitative PCR was used to determine IL-33 and TNF-α mRNA expression in IS. Results: sST2 and IL-33 levels in IS and serum were significantly higher in patients compared with healthy controls (p = 0.0001). The increase in sST2 and IL33 was significantly more important in moderate cases than in mild asthma. A significant correlation was observed between serum and IS IL-33 levels (r = 0.497; p = 0.0018). Higher levels of IL-33 mRNA were detected in IS from asthmatics than those observed in controls. A significant correlation was found between TNF-α and IL-33 mRNA expression in the asthmatic subjects (r = 0.772, p = 0.0001). Conclusions: Values of sST2 and IL-33 observed in IS were found to correlate with disease activity. Elevated IL-33 mRNA expression in IS and its correlation with TNF-α reflected the inflammatory process observed in the lung of young asthmatics.

Contribution of VDR polymorphisms to type 1 diabetes susceptibility: Systematic review of case–control studies and meta-analysis

Vitamin D receptor (VDR) polymorphisms have been inconsistently investigated in type 1 diabetes (T1D). However, the results are inconsistent and inconclusive. The current study aimed to investigate the role of TaqI, BsmI, ApaI and FokI VDR polymorphisms in T1D disease. Following the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines, a systematic search and meta-analysis of the literature, since 1998 until december 2013, was conducted. Subgroup analyses were performed to detect potential sources of heterogeneity from selected study characteristics. Meta-analyses yielded a non-significant association of TaqI polymorphism with T1D [OR=1.014 (0.783-1.312); P=0.918] in the recessive model. The BsmI polymorphism was not associated with T1D [OR=1.44 (0.944-1.386); P=0.171] in the dominant model. Also, ApaI polymorphism was not associated with T1D risk [OR=0.996 (0.859-1.155); P=0.960] in the homozygous model. The FokI polymorphism was not associated with T1D risk [OR=0.968 (0.743-1.263); P=0.813] in dominant model. Stratification according to study characteristics showed that publication year, age, gender, estimated vitamin D levels and latitude moderated significantly association between VDR polymorphisms and T1D disease. Meta-analysis on haplotypes revealed that BAT might be a significant risk factor for T1D [OR=1.331 (0.957-1.850; P=0.089]. However, the bAT was found to be a significant protective factor [OR=0.639 (0.460-0.887); P=0.007]. As conclusion, individual VDR polymorphisms seemed not to be associated with T1D risk. However, haplotypes contributed significantly to disease susceptibility. Study characteristics moderated the association between VDR polymorphisms and T1D. These results suggested that, in T1D pathogenesis, VDR polymorphisms interact with each other and with environmental factors.

Vitamin D reduces the differentiation and expansion of Th17 cells in young asthmatic children.

Vitamin D [25(OH)D3] deficiency has been associated with asthma as in many inflammatory and autoimmune pathologies; however, there is still a lack of data about the effects of administration of vitamin D in immune regulation in young asthmatic patients. In this study, we investigated its inhibitory effect on the immune response in young asthmatic patients and the possible mechanisms involved. Peripheral blood CD4(+) T cells from 10 asthmatic patients and 10 healthy controls were cultured under Th17 polarizing conditions in the presence or absence of [25(OH)D3], IL-17 cytokine production was determined by ELISA and flow cytometry. Messenger RNA (mRNA) expression of several factors related to Th17 cell function was determined by real-time PCR. The effect of [25(OH)D3]-treated dendritic cells (DCs) on CD4(+) T cell response was determined by ELISA and flow cytometry. Stimulation of naive CD4(+) T cells under Th17 polarizing conditions showed a higher Th17 cell differentiation in asthmatic patients than healthy controls. The addition of [25(OH)D3] significantly inhibited Th17 cell differentiation both in patients [P<0.001] and in normal controls [P=0.001] in a dose-dependent way. [25(OH)D3] was able to inhibit the gene expression of RORC, IL-17, IL-23R, and CCR6. [25(OH)D3]-treated DCs significantly inhibited IL-17 production [P=0.002] and decreased the percentage of CD4(+)IL-17(+) [P=0.007] in young asthmatics. The findings suggest that the inhibitory effect of [25(OH)D3] on the Th17 response was mediated via both T cells and DCs. DCs pathway is involved in the direct inhibition of 25(OH)D3 on Th17 cell differentiation in young asthmatics.

NOD2 is highly expressed in Behçet disease with pulmonary manifestations

BackgroundExcessive Th1 cells and TLRs functions are involved in the pathogenesis of Behcets disease (BD) in response to bacterial antigens. NOD2, an intracellular pathogen recognition sensor, modulates innate defence to muropeptides derived from various bacterial species. To further define a role for NOD2 in BD, we analysed NOD2 transcriptional responses in BAL-MNC from BD patients with pulmonary manifestations.MethodsWe analysed NOD1, NOD2, T-bet and TLRs mRNA expression with real-time polymerase chain-reaction in BAL cells obtained from 23 BD patients with pulmonary manifestations and their matched controls.ResultsWe found that NOD2 mRNA expression was highly up-regulated in BAL cells from BD and sarcoidosis patients compared to healthy control group (P = 0.001). In BD patients, significant correlation was found between NOD2 and T-bet mRNA expression (r = 0.602; P = 0.0009). In BAL from BD patients, NOD2 and T-bet mRNA expression were significantly correlated with BAL-lymphocytes (r = 0.485, P = 0.010; r = 0684, P = 0.0001 respectively). NOD2 in BD was also correlated with TLR 2(r = 0.444; P = 0.021) and TLR 4 (r = 0.574; P = 0.001) mRNA expression.ConclusionOur results indicate that BAL-MNC from BD patients expressed NOD2 as a result of lung inflammation. TLRs and NOD2 synergize for the induction of proinflammatory cytokines. BAL inflammatory cells showed an increased Th1 situation as indicated by increased T-bet mRNA expression.

Immunological responses in patients with Behçet’s disease: advances in understanding

Behçet’s disease (BD) is an inflammatory, multisystemic disease possibly induced by susceptible microbiological pathogens. BD is attributed to genetic and environmental factors that cumulatively elicit the dysfunction of T and B lymphocytes and dendritic cells. Immunological responses in BD were investigated in most of the inflammatory sites and have relied upon the classic paradigm of distinct innate and adaptive parts of the immune system. However, recent advances in inflammatory and autoimmune diseases suggest that this division may be overly simplistic, with emerging evidence of a breakdown in conventional hallmarks of each system. Molecular markers revealing polymorphisms at the DNA level, Treg and Th17 cells permitted an increasing understanding in the knowledge of the disease. This review will focus on the immune mechanisms and give an overview of the major new immunological hypothesis and the role of the main cellular populations that drive the immune responses in BD.

Vitamin D receptor TaqI and ApaI polymorphisms: A comparative study in patients with Behçet’s disease and Rheumatoid arthritis in Tunisian population

Recent genetic surveys have identified vitamin D receptor (VDR) as a susceptibility gene for several autoimmune diseases. This study was designed to investigate the association of VDR gene polymorphisms with Behçets disease (BD) and Rheumatoid arthritis (RA). A case-control study including 151 BD, 106 RA patients and an appropriate number of healthy control subjects were performed using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) techniques. Association between TaqI polymorphism and BD was marginal under codominant and recessive models (P=0.078 and P=0.058, respectively). After stratification, we found evidence for a significant association between TaqI polymorphism and BD in the elderly subjects (P=0.037). The minor ApaI a allele tended to confer an increased risk for BD susceptibility (P=0.087). BD patients with VDR homozygous AA or aa genotypes were at increased risk for development of erythema nodosum (EN) skin manifestation (P=0.038). No significant association was observed for VDR ApaI and TaqI polymorphisms with RA risk (P>0.05). TaqI and ApaI polymorphisms might be modestly implicated in BD pathogenesis. They could be considered as potential biomarkers in BD rather than susceptibility genes. However, TaqI and ApaI seemed not to be implicated in RA pathogenesis.

Pulmonary manifestations in Behçet disease: impaired natural killer cells activity

BackgroundBehçet’s disease (BD) is a systemic vasculitis with unknown aetiology, where, besides genetic predisposition, an immune dysregulation involving T and B lymphocytes and hyperactive neutrophils contribute to disease pathogenesis. The aim of this study was to determine the cytotoxicity of natural killer (NK) cells in bronchoalveolar lavage (BAL) from BD patients with pulmonary manifestations.MethodsBAL was performed in 27 patients with BD and pulmonary manifestations, 14 patients with Rheumatoid Arthritis (RA) and 23 healthy controls (HC). Related orphan receptor C (RORC) and forkheadbox P3 (FOXP3) mRNA transcript were determined in BAL by reverse transcription–polymerase chain reaction (RT-PCR). NK cells, NK cell cytotoxicity, and lymphokine-activated killer (LAK) activity against K562 cells were measured by flow cytometry. Proportions of NK precursors and expression of genes for IL-2 receptor β (IL-2Rβ; CD122), perforin, and granzyme in NK cells were measured by flow cytometry or RT-PCR.ResultsThe analysis of transcription factors revealed an increase in the RORC/FOXP3 ratio (Th17/Treg cells) in BAL from BD patients. Percentages of NK were significantly lower in BD than in RA patients and healthy controls. Purified NK cells derived from BD patients were found to have lower cytotoxicity and LAK activity than those from controls. This defect of NK cells in BD patients was related to down-regulation of perforin and granzyme expression in NK cells.ConclusionIn BD patients, the increased RORC/FOXP3 ratio indicated an inflammatory state of the lung. NK cells were decreased together with an impairment of their activity due to a defective expression of granzyme and perforin. These abnormalities possibly contribute to immune system dysregulation found in BAL of BD patients with pulmonary manifestations.

Human Leukocyte Antigen G Polymorphism and Expression Are Associated with an Increased Risk of Non-Small-Cell Lung Cancer and Advanced Disease Stage.

Human leukocyte antigen (HLA)-G acts as negative regulator of the immune responses and its expression may enable tumor cells to escape immunosurveillance. The purpose of this study was to investigate the influence of HLA-G allelic variants and serum soluble HLA-G (sHLA-G) levels on risk of non-small-cell lung cancer (NSCLC). We analyzed 191 Caucasian adults with NSCLC and 191 healthy subjects recruited between January 2009 and March 2014 in Ariana (Tunisia). Serum sHLA-G levels were measured by immunoassay and HLA-G alleles were determined using a direct DNA sequencing procedures. The heterozygous genotypes of HLA-G 010101 and -G 010401 were associated with increased risks of both NSCLC and advanced disease stages. In contrast, the heterozygous genotypes of HLA-G 0105N and -G 0106 were associated with decreased risks of NSCC and clinical disease stage IV, respectively. Serum sHLA-G levels were significantly higher in patients with NSCLC and particularly in those with advanced disease stages compared to healthy subjects. The area under the receiver-operating characteristic (ROC) curves was 0.82 for controls vs patients. Given 100% specificity, the highest sensitivity achieved to detect NSCLC was 52.8% at a cutoff value of 24.9 U/ml. Patients with the sHLA-G above median level (≥ 50 U/ml) had a significantly shorter survival time. This study demonstrates that HLA-G allelic variants are independent risk factors for NSCLC. Serum sHLA-G levels in NSCLC patients could be useful biomarkers for the diagnostic and prognosis of NSCLC.

Update on Asthma Genetics: Results From Meta-Analyses of Candidate Gene Association Studies

Several studies have investigated the risk factors associated with asthma. Both genetic and environmental factors are considered to contribute to asthma susceptibility. Individual genetic association studies usually suffer from small sample size leading to biased results. Meta-analysis is a powerful tool that has the potential to resolve this limitation by increasing the statistical power of analyses. The current review summarizes the recent knowledge concerning genetic factors involved in asthma predisposition based on meta-analyses. Using the keywords: asthma, meta-analysis, polymorphism, we searched Pubmed, Medline, Embase and Google Scholar databases for the associated articles. Genetic polymorphisms in twenty-three genes are associated with asthma risk in meta-analyses. However, polymorphisms in nine genes showed none significant association. These findings are used to assess the genetic risk factors and to understand the molecular pathways related to asthma.