Expression of tolerogenic dendritic cells in the small intestinal tissue of patients with celiac disease.

Tolerogenic dendritic cells (tolCDs) play an important role in the regulation of inflammation in autoimmune diseases such as celiac disease (CeD). Dendritic cells express CD207, CD11c, and CD103 on their surface. In addition to the receptors mentioned above, tolCDs can express the immune-regulating enzyme indoleamine 2,3-dioxygenase (IDO). This study aimed to determine the mRNA and protein expression of CD11c, CD103 and CD207 markers, and also IDO gene expression in intestinal tissues of CeD patients in comparison to the healthy individuals. Duodenal biopsies were collected from 60 CeD patients and 60 controls. Total RNA was extracted and gene expression analysis was performed using Real-time PCR SYBR® Green method. Additionally, biopsy specimens were paraffinized and protein expression was evaluated using immunohistochemistry (IHC) for expression of CD11c+, CD207+and CD103+. Gene expression levels of CD11c (P = 0.045), CD103 (P < 0.001), CD207 (P < 0.001) and IDO (P = 0.01) were significantly increased in CeD patients compared to the control group. However, only CD103 protein expression was found to be significantly higher in CeD patients in comparison to the control group (P < 0.001). The result of this study showed that the expresion levels of CD11c, CD103, CD207 and IDO markers were higher in CeD patients compared to the controls, indicating the effort of dendritic cells to counterbalance the gliadin-triggered abnormal immune responses in CeD patients.

Gold-based nanoplatform for a rapid lateral flow immunochromatographic test assay for gluten detection.

BACKGROUND: Gluten, a food allergen, is available in foods derived from wheat, rye and barley. It damages the small intestine and causes celiac disease. Herein, we designed a rapid immunochromatographic lateral flow test assay for detecting the gluten contents of raw materials. In this rapid test, the presence of gluten was screened through the capturing of gliadin (a toxic component of gluten) by two identical gliadin monoclonal antibodies. One of the antibodies was immobilized on the membrane in the test zone as a capture reagent. The other antibody was labeled with gold nanoparticles (AuNPs) as a detector reagent. RESULTS: Gold nanoparticles with a size of about 20 nm were synthesized and conjugated to the gliadin monoclonal antibodies. The detection limit of the experimental assay was 20 ppm and positive results were visualized after 15 min using only 40 µL of the extracted sample for each test. Analysis of different flour samples identified the best sensitivity and specificity of the lateral flow test strip (LFTS). CONCLUSION: The experimental LFTS is an easy-to-use and rapid method for the screening of gluten level in raw materials. The LFTS may be employed to ensure the safety of foods.

Development of a nano biosensor for anti-gliadin detection for Celiac disease based on suspension microarrays.

Celiac disease is an autoimmune disorder represented by the ingestion of the gluten protein usually found in wheat, barley and rye. To date, ELISA has been the most accurate method for determining the presence of anti-gliadin, which is cumbersome, expensive (compared to a suspension microarray technique), and requires extensive sample preparation. In this study, in order to establish a more accurate assay to identify gliadin at lower concentrations, optical nano biosensors using an indirect immunoassay method for gliadin detection was designed and fabricated. For this, polycaprolactone (PCL) nano- to micro-beads were fabricated as a platform for the gliadin antigen which were optimized and nano functionalized with amine groups for such purposes. The gliadin antibody, which is selective to gliadin, was then added to the beads. Static light scattering tests were conducted to determine PCL particle size distribution and sizes were found from 0.1 to 30 µm, which is suitable for flowcytometry detection devices. Anti-gliadin detection was performed using an anti IgG mouse antibody conjugated with FITC in a flow cytometry device to detect the smallest particle. Fluorescence intensity was investigated at different concentrations of anti-gliadin and a standard curve used to determine gluten concentration based on fluorescence intensity. Results showed that the fluorescence intensity increased with greater concentrations of anti-gliadin providing a very effective method of detection due to selectivity at a 5 ppm detection limit. This represents a new highly sensitive and fast method for anti-gliadin detection. Further, the disuse of a cross linker and the use of a dedicated antibody at a very low level (1 µl) made this new method very economical to identify anti-gliadin concentrations at the nano level. In summary, this study provides a new, more accurate and sensitive, as well as less expensive system to detect anti-gliadin for the improved diagnosis of celiac disease.

The role of CXCR3 and its ligands CXCL10 and CXCL11 in the pathogenesis of celiac disease.

The chemokine receptor CXCR3 and its ligands CXCL10 and CXCL11 have been suggested to give rise to the most relevant chemokine axis able to facilitate the entrance of immune cells into inflamed tissues and be activated in different inflammatory disorders, such as celiac disease (CD).The aim of this study was to investigate the expression level of CXCR3, CXCL10, and CXCL11 genes in celiac patients compared to healthy controls. Both cohorts have been recruited from the Iranian population.In this case-control study, biopsy specimens were collected from 71 celiac patients (60.5% female) and 90 control subjects (57% female) during 2016. Total RNA was extracted and mRNA expression levels of CXCR3, CXCL10, and CXCL11 genes were investigated by SYBR green qPCR.Based on qPCR and relative quantification method, the mRNA expression levels of CXCR3, CXCL10, and CXCL11 were significantly higher in duodenal biopsies of celiac patients compared to healthy controls in the study population (P = .038, P = .021, and P = .012 respectively).The result of this study showed that CXCR3/CXCL10/CXCL11 signaling axis is overexpressed in the small intestinal mucosa of CD patients compared to controls. This finding might explain the specific enrollment of the main cell populations that infiltrate the epithelium.

Contribution of HLA-DQ2/DQ8 haplotypes in type one diabetes patients with/without celiac disease.

BACKGROUND: Based on lack of data on the distribution of the related alleles in the T1D population in Iranian population, we assessed the frequency of HLA DQ2 and DQ8 haplotypes in patients with T1D with/without CD compared to healthy population. MATERIALS AND METHODS: 70 patients with T1D without celiac disease, 60 T1D cases with CD were compared to 150 healthy individuals during 2016. Ten mililiter Gheparinized blood samples were collected, genomic DNA was extracted and alleles were genotyped by Real-time PCR using SYBR Green as a low-resolution method. RESULTS: HLA-DQ2 and/or HLA-DQ8 genotypes was presented in 51% and 23% of T1D patients without CD respectively. Twenty one percent of those patients carried both alleles and 5% were negative for both alleles. T1D patients with CD had much higher DQ2 frequency (72%) and lower DQ8 (11.6%), than T1D patients without CD and controls, 14% carried both alleles and 3% were negative for both. The frequencies of DQ2 and DQ8 alleles in Iranian healthy population were 19 and 5% respectively. CONCLUSION: According to the same genetic background for CD and T1D we suggest that HLA-typing can be a very useful screening tool for CD in patients with type one diabetes.

The relationship between 174 G/C and -572 G/C of IL-6 gene polymorphisms and susceptibility of celiac disease in the Iranian population.

INTRODUCTION: Celiac disease (CD) is a chronic inflammatory intestinal disorder. Different immunological factors, including inflammatory cytokines, may play an important role in disease susceptibility. AIM: To investigate the relationship between -174G/C and -572G/C gene polymorphisms and the serum level of interleukin 6 (IL-6) and susceptibility to CD in the Iranian population. MATERIAL AND METHODS: In this case-control study blood samples were collected of 105 patients with CD and 106 healthy subjects randomly in 2016 and evaluated by polymerase chain reaction-restriction fragments length polymorphism (PCR-RFLP) method. A sequence was also used to confirm the results of both polymorphisms. The IL-6 concentration was measured using ELISA. RESULTS: The results showed a significant relationship between polymorphism -572G in CD patients when compared with control subjects by genotype (p = 0.001) and alleles (p = 0.022), respectively. There was no significant relationship between polymorphism 174G and frequency of genotype, but an association of this polymorphism with the frequency of alleles (p = 0.034), age (p = 0.001), and body mass index (p = 0.003) was seen. The serum level of interleukin-6 was significantly associated only with rs1800796 (p < 0.001). CONCLUSIONS: The results confirm previous studies in different parts of the world and indicate that IL-6 (572G/C) polymorphism may play a role in susceptibility to CD in the Iranian population.

Serum cytokines profile in treated celiac disease compared with non-celiac gluten sensitivity and control: a marker for differentiation.

BACKGROUND AND AIMS: There is increasing evidence regarding elevated serum levels of inflammatory cytokines in patients with celiac disease (CD), but little is known about their levels in patients with non-celiac gluten sensitivity (NCGS). The aim of this study was to evaluate the serum levels of inflammatory cytokines in Iranian patients with CD and NCGS and to compare them with those of healthy individuals. METHODS: A total of 110 treated CD, 15 with NCGS, and 46 healthy subjects were enrolled during 2016. Serum levels of IL-1, IL-6, IL-8, IL-15 and IFN-γ were measured using ELISA, and compared between groups. The correlation of the severity of mucosal damage and clinical symptoms with serum levels of cytokines was also assessed. RESULTS: The mean serum levels of IFN-γ (p = 0.04) and IL-6 (p = 0.007) were significantly different between the patients in the CD and control groups, and IL-8 was significantly higher in the CD group compared with patients in the NCGS group (p = 0.04). Statistically significant correlations were observed between the serum levels of IFN-γ and abortion (p = 0.01), IL-1 and weight loss (p = 0.043) and infertility (p = 0.0001) in CD patients, and between IFN-γ and abortion (p = 0.01) and infertility (p = 0.01) in the NCGS patients. Moreover, no significant relationship was observed between the severity of mucosal damage and the serum level of the studied cytokines. CONCLUSIONS: Inflammatory cytokines are implicated in the pathogenesis of CD, and their serum levels might help to identify a diagnostic marker to differentiate CD from NCGS. However, further studies with a larger sample size are recommended.

Study of prevalence, distribution and clinical significance of Blastocystis isolated from two medical centers in Iran.

AIM: This study aimed to survey prevalence and clinical significance of Blastocystis among symptomatic and asymptomatic groups. BACKGROUND: Blastocystis is a prevalent microorganism that is found in intestine of human and majority of animals. However, most studies have failed to establish correlation between the presence of the parasite and clinical manifestations. METHODS: from Dec 2016 to Jun 2017, 554 stool samples were collected from symptomatic and asymptomatic subjects referred to Imam Hossein Hospital and Gastroenterology and Liver Diseases Research Institute, Tehran, Iran. All samples were concentrated using conventional formalin-ethyl acetate concentration and then were microscopically examined using Lugol's iodine staining and light microscope. The fresh stool samples were also cultivated in DMEM medium and were examined for growth of Blastocystis every 48 hours with direct smear slides for 10 days. RESULTS: Blastocystis was observed among 93 (16.8%) of stool samples cultivated in DMEM. The findings represented that 64/398 (16.08%) and 29/156 (18.58%) of asymptomatic and symptomatic patients were infected with Blastocystis, respectively. In addition, there was no significant correlation between presence of symptoms and carrying Blastocystis (P=0.528), although statistically significant association was observed between presence of urticaria and carrying Blastocystis (P<0.05). Furthermore, a statistically significant correlation between observing the parasite and different age groups was seen (P<0.05). CONCLUSION: Blastocystis is a prevalent parasitic eukaryote among symptomatic and asymptomatic populations despite the higher prevalence among symptomatic group that suggests the chance of infection with Blastocystis raises with age.