Iranian Journal of Allergy, Asthma and Immunology
Volume:17 Issue: 6, Dec 2018

Allergy poses a heavy health burden in modern society. Other than symptom-relieving medications, the only available treatment approach is allergen-specific immunotherapy, which in spite of offering a potential cure, requires a long treatment duration with multiple doses of allergen administration and carries a risk of anaphylaxis. Gene therapy has shown advantages in experimental studies for treatment of tumors, genetic diseases, chronic infections, and allergy. To date, adenovirus has been the most extensively used gene transfer vector, and offers high efficiency and safety. Here, we review studies of adenovirus-mediated gene therapy targeting different steps in the development of allergic diseases. Adenovirus-mediated gene therapy might be a promising add-on therapy for allergy treatment.

 Asthma is a chronic disorder of the airways characterized by reversible airflow obstruction, inflammation and bronchial hyperresponsiveness. Different immune cells and molecules have been attributed to involve in pathogenesis of asthma. In the current case-control study, the expression of T cell Ig and mucin domain-containing molecule-3 (Tim-3) and programmed death-1 (PD-1) was studied on CD4+ T cells of patients with asthma and normal controls. The frequency of Tim-3+/PD-1+/CD4+ T cells was determined by a three color flow cytometry method in 37 patients with asthma and 32 healthy controls. To evaluate the Th1/Th2 ratio, peripheral blood mononuclear cells were isolated from all samples and stimulated with phorbol 12- myristate 13- acetate ( PMA)/ionomycin for 18 h. IFN-γ) and Interleukin-4 (IL-4) were measured in culture supernatants by-(ELISA). Serum total immunoglobulin E (IgE) was also measured in all samples. Significant increase in percentage and absolute count of Tim-3+/PD-1+/CD4+, Tim-3+/CD4+ and PD-1+/CD4+ T cells was found in asthmatic patients compared to healthy controls (p=0.02 and p=0.003, respectively). The IFN-γ/IL-4 ratio (Th1/Th2 ratio) was significantly higher in healthy controls than that of asthmatic patients (p=0.029). Our data regarding the increased expression of PD-1 and Tim-3 on CD4+ T cells of patients with asthma suggest the potential roles of these immune checkpoint receptors in immune dys-regulation of asthma.

 Orosomucoid 1-like 3 (ORMDL3) gene, located on chromosome 17q21, is an asthma candidate gene that encodes ORMDL3. This molecule has been reported to play a role in airway remodeling and bronchial hyper-responsiveness. In this study, we aimed to investigate the possible association of ORMDL3 single nucleotide polymorphism (SNP) (rs12603332) with susceptibility to allergic asthma in Iranian Northwestern Azeri population. 193 asthmatic patients and 185 normal individuals were included. Genomic DNA was extracted and genotyping was performed by standard restriction fragment length polymorphism-polymerase chain reaction RFLP-PCR method using BstUI restriction enzyme. Our results showed dominant presence of TC genotype and C allele in both patients (49.2% and 59.8%, respectively) and controls (48.6% and 60%, respectively). Frequency of genotypes and alleles showed no significant difference between two groups (p=0.994 and p=1.00, respectively). None of alleles could be defined as risk allele for allergic asthma (OR=0.99, 0.88-1.12, 95% CI). We failed to show significant association between ORMDL3 rs12603332 with predisposition to allergic asthma in Iranian Northwestern Azeri population. More studies with larger number of participants should be done to find more reliable results for such association.

Eosinophilic esophagitis (EoE) is a chronic immune-mediated disease isolated to the esophagus Food allergy is thought to play an important role in the pathophysiology of EOE. The aim of this study is to evaluate demographic features and sensitivity of patients with reference to common food allergens. Children with documented EoE were enlisted for this study. Skin prick test and patch test were done for all participants. Each test contained common food allergens. Other tests, such as evaluation of total IgE and eosinophil count, were also done. A total of 30 patients (the mean age of participants was 3±6 years) with 6 months to 15 years documented EoE participated in this study. The mean duration of symptoms was 2 years. Association with other allergic disorders was seen in 16 (32%) patients [Asthma in 8 (26.7%), allergic rhinitis in 5 (16.7%) and eczema in 3 (10%)]. The mean level of total IgE was 413.5±505.5 (IU/ML): total IgE level was above normal range for age in 17 children. The mean level of eosinophil was 372.2±305.2, and eosinophilia was seen in 11 patients (36.7%). The skin prick test and patch test findings showed that 28 patients (93%) and 17 of 30 patients (56.6%) tested positive to foods respectively. The most common positive skin prick test was for sesame and walnut. Patch test showed that the most common positive test was for fish. Evaluating the symptoms indicated that vomiting (70%) is the most prevalent symptom in patients; the less prevalent symptom was dysphagia (3.3%). Other prevalent symptoms were subsequent abdominal pain (33.3%), gaining weight failure (33.3%), heartburn (16.6%), cough (10%), fecal impaction (10%), and constipation (6.66%). Our series confirms the high degree of atopy in Iranian children with EoE. These patients seem to be polysensitized to several food allergens Because of different eating cultures in different countries, considering special food in selecting allergens for allergic tests is needed.

 Regulatory T cells are of utmost importance for tolerating the fetus. In some pregnancy complications such as pre-eclampsia, the frequency of CD4+CD25+Foxp3+ regulatory T cells is altered, but there is no consistency regarding the results. Besides, little is known about the frequency of CD8+CD25+Foxp3+ Treg cells in pregnancy complications. Therefore, we aimed to investigate the frequency of both CD4+ and CD8+ regulatory T cells in the peripheral blood of women afflicted by preeclampsia. Ten non-pregnant, ten healthy pregnant, and ten preeclamptic women participated in this study. Four colors flow cytometry method was used to identify the frequency of the CD4+ and CD8+ regulatory T cells in the peripheral blood. Results indicated that the frequencies of CD4+CD25+Foxp3+ and CD8+CD25+Foxp3+ cells were significantly lower in preeclamptic women compared to healthy pregnant and non-pregnant ones (p<0.05). A positive correlation was also observed between CD4+ and CD8+ regulatory T cells (R= 0.532, p=0.002). Moreover, CD4+ regulatory T cells negatively correlated with systolic and diastolic blood pressures (R=-0.760 and -0.753, respectively; p<0.001). CD8+ regulatory T cells also had a negative correlation with systolic (R=-0.503, p=0.001) and diastolic (R=-0.590, p=0.005) blood pressures. In conclusion, a reduction in the frequencies of both CD4+ CD25+ Foxp3+ and CD8+CD25+Foxp3+ regulatory T cells might be important in the pathogenesis of pre-eclampsia.

Multiple sclerosis (MS) is a chronic inflammatory disorder of the central nervous system (CNS), in which axonal damage is a deteriorative factor. Brain-Derived Neurotrophic Factor (BDNF) is described as a neuronal-survival gene, also capable of exerting pleiotropic effects on the immune cells. Here, we aimed to investigate expression levels of BDNF and its antisense RNA, BDNF-AS, in Iranian MS patients. Our case-control study was based on collecting 50 whole blood samples of relapsing-remitting MS patients and 50 healthy controls. Then, expression analysis of BDNF and BDNF-AS was performed by Real-time quantitative PCR. We found a strong and positive correlation between BDNF and BDNF-AS in MS patients. This is while no significant difference in BDNF and BDNF-AS expression levels was seen between MS patients and controls (p>0.05). A significant and strong positive correlation was found between the expression levels of BDNF-AS and BDNF (r=0.785, p<0.0001). Further, significant positive moderate correlations of BDNF and BDNF-AS with other lncRNAs (GSTT1-AS1 and IFNG-AS1) and genes (TNF and IFNG) were revealed (p<0.0001). Additionally, there was no correlation between the BDNF and BDNF-AS expressions and disease duration, age at onset, and Expanded Disability Status Scale of Kurtzke (EDSS) (p>0.05). BDNF and BDNF-AS expression levels revealed insignificant discrepancies in patients and controls. We found a strong and positive correlation between BDNF and BDNF-AS in MS patients, which is, based on previous studies, a quit novel finding and can be further discussed by future works to unravel its possible application in MS. We suggest evaluation of different leukocytes subsets separately along with large cohort studies comprising a higher number of individuals from different ages to unravel the effects of other possible aspects.

Type 1 diabetes (T1D) is an autoimmune disease resulting from the damage of pancreatic B-cells mediated by autoreactive CD4+ and CD8+ T cells. In recent years, follicular T helper (Tfh) cells have been recognized as a subpopulation of CD4+ T cells providing help for B cells differentiation and antibody production. In this study, we examined the frequency of circulating CD4+CXCR5+ and CD4+CXCR5+ICOS+ (representing Tfh) cells as well as serum levels of anti-glutamic acid decarboxylase 65 (GAD65) and islet cell autoantibodies (ICA) in children with type I diabetes. We analyzed the percentage of Tfh cells within peripheral blood mononuclear cells in 20 children with T1D (≤300 days from disease onset; Mean age 6.8±4.6 years) and 18 healthy individuals (Mean age 8.8±2.2 years) using flow cytometry. Anti-glutamic acid decarboxylase (GAD) and islet-cell cytoplasmic autoantibodies (ICA) levels were determined by ELISA and indirect immunofluorescence respectively. We found that the frequency of CD4+CXCR5+ and CD4+CXCR5+ICOS+ (Tfh) cells were significantly increased in the peripheral blood of patients compared with healthy controls (p<0.001). Furthermore, elevated levels of anti-GAD and ICA antibodies were detected in children with T1D (p=0.001 and p=0.02 respectively). There was no correlation between Tfh cells frequency and the autoantibody levels. The results of our study indicate an increased frequency of Tfh cells in children With T1D that could suggest a possible role of these cells in the disease pathogenesis.

Type 1 diabetes (T1D), a spontaneous autoimmune disease, is associated with destruction of insulin-producing β-cells in the pancreas. Since some heat shock proteins (HSP), such as HSP70 exert a protective effect in both tissues and cells, the present study was conducted to elucidate the effects of carbenoxolone (CBX) as an HSP70 inducer on T1D. The disease was induced in male C57BL/6 mice using streptozotocin (STZ) and subjects were allocated to therapeutic 1 and therapeutic 2 groups, as well as negative and positive control groups. The treated mice (therapeutic 1 and therapeutic 2 groups) received 50 mg/kg CBX intraperitoneally every 24 hours, in the therapeutic 1 group the drug was injected before and after disease induction whereas in the therapeutic 2 group the drug was injected only after disease induction. Serum fasting blood sugar (FBS) level, cytokines production (Interferon-gamma (IFN-γ), Interleukin 10 (IL-10), and IL-17), serum HSP70 level and CD4+CD25+Foxp3+ regulatory T cell (Treg) frequency measurements were outperformed 14 days after the last STZ injection. Our results showed that in the treated groups, serum HSP70, IFN-γ, and IL-17 levels were increased in contrast to the untreated groups. The IL-10 level was markedly decreased in comparison to untreated diabetic mice (p<0.05). Moreover, it was found that the frequency of Tregs in treated mice was lower in comparison to the untreated mice but the difference was not significant (p>0.05). Our results confirm that CBX might through HSP70 induction, followed by increasing IFN-γ level leads to suppression of IL-10 production in diabetic mice resulted in toxic effects on pancreatic islet beta cells and deteriorating of disease.

Among the many pneumococcal antigens, choline-binding proteins (CPBs) display high immunogenicity in animal models. This study aims to determine the immunogenicity of CbpM, CbpG and CbpL proteins in a mice model. The genes were cloned into pET21a expression vector and the recombinant proteins were produced. Mice and rabbits were immunized with the purified recombinant proteins. Subsequently, the mice were challenged with S. pneumoniae and their survival as well as bacterial clearances were followed. The antibody responses of the mice immunized with recombinant proteins were determined by ELISA assay. The opsonophagocytosis assay was performed using rabbit’s sera. Passive immunization was also carried out in another group of mice using two doses of anti-CbPs antibodies. Finally, these mice were challenged and their survival as well as bacterial clearance were determined. The mice actively immunized with CbpM, CbpG and CbpL recombinant proteins showed survival rate of 100%, 85% and 75%, respectively. The survival rates among passively immunized mice groups which received100µg/ml dose of anti-CbpM, anti-CbpG and anti- CbpL were 50%, 50% and 25%, respectively. For all groups, however, only 25% of mice which received 10µg/ml dose of antibody survived after bacterial challenge. The rates of opsonization with anti-CbpM, anti-CbpG, and anti-CbpL antibodies at 100 and 10µg/ml doses, were found to be 45.6 and 14.7%, 82.3 and 12.9% and 12.2 and 9.35%, respectively. Our findings suggest that the recombinant proteins particularly CbpM and CbpG can protect the mice against pneumococcus19F serotype and effectively induce a protective antibody response. Thus, CbpG and CbpM proteins might be used as suitable vaccine candidate in pneumococcal vaccine formulations.

Preparation of the indigenous varicella zoster vaccine could significantly reduce the disease burden of varicella zoster virus especially in immunosuppressed children. To achieve this goal, the varicella zoster virus was isolated from an 8 years boy infected with chicken pox. The virus was cultivated in sensitive cell line and determined varicella zoster. The adaptation and attenuation of virus was carried out after several passages in MRC-5 cell culture, Primary Guinea pig embryo fibroblast cell culture and again switching in MRC-5 cell culture. The challenged of vaccine dose was found 3LogCCID50. Following two doses of immunization in guinea pigs via inoculated cell culture-fluid attenuated- local isolated VZV at zero and 14 day, the humoral immune response, varicella-zoster virus (VZV) IgG and IgM were determined using enzyme-linked Immunosorbent and seroneutralization assays at 7, 14, 21, 30, 60, 90.120 days after receiving of the first and second dose of vaccine. The results of immunization showed good 93% seroconversion in guinea pig which compared with vOKa vaccine was not significant (p<0.05). The prepared attenuate varicella zoster virus promising a candidate Virus for our future plan to vaccine production.

Major histocompatibility complex (MHC) class II deficiency is a rare primary immunodeficiency disorder (PID) with less than 200 cases worldwide. Here, we report an 8 month–old girl with MHC class II deficiency with a novel homozygous mutation in RFXANK gene (NM_001278728: exon 5: c.495G>A: p.Trp165*) and normal CD4+ T cell counts, diagnosed by whole exome sequencing (WES) and negative HLA–DR proteins on peripheral blood mononuclear cell (PBMC) in flow cytometry. She was referred with pneumonia, prolonged fever, resistance to antibiotics (ceftriaxone, clindamycin, and vancomycin), and low serum immunoglobulin (IG) levels, while natural killer (NK), B, and T cells were normal. She received intra-venous immune-globulin (IVIG) replacement, broad spectrum antibiotics, and anti-fungal treatments. The presented case report is interesting not only because of the rarity of the PID but also due to normal CD4+ T cell counts. According to our experience, we suggest that physicians consider MHC class II deficiency in families with consanguineous marriages, even with normal CD4+ T cell counts. At the first, the diagnosis of the disease could be successfully perform using WES, and finally, treatment with hematopoietic stem cell transplantation can save the patients’ lives.

Asthma is one of the most common chronic diseases all over the world, which impacts socioeconomics of families and health care systems. We need to estimate the costs of asthma to set health policies and structural adjustments to control burden of asthma in societies. The aim of present study was to evaluate direct costs of asthma in children in Tehran-Iran. In a descriptive-analytical study, 100 children who were referred to Children’s Medical Center of Tehran University of Medical sciences, from March 2014 to March 2015, were selected randomly and studied. In an interview with parents we evaluated direct costs of asthma including physician fees, admission fees, medication costs, costs of transportation, and others. In present study, 72 patients (72.0%) were male and 28 patients (28.0%) were female with mean age of 6.9±0.3 years old. Annual mean of direct costs of outpatient care were calculated 5,179,000±620,000 Rials (159.3±19$), and with considering the admission costs, direct medical expenditure were calculated 6,069,600±140,000 Rials (186.7±4$) throughout the year. Non-medical annual costs of asthma in children were calculated 4,439,700±437,540 Rials (136.6±13$) during 2014-2015 in Tehran-Iran. Based on findings of present study, total costs of asthma with the help of close physician follow ups and effective usage of medications has decreased in recent decades; furthermore, in our study, outpatient management costs which accounted for 85.5% of total direct costs of asthma in children, had the largest share of direct costs of asthma management.