DOCK8 functions as an adaptor that links TLR-MyD88 signaling to B cell activation.

The adaptors DOCK8 and MyD88 have been linked to serological memory. Here we report that DOCK8-deficient patients had impaired antibody responses and considerably fewer CD27(+) memory B cells. B cell proliferation and immunoglobulin production driven by Toll-like receptor 9 (TLR9) were considerably lower in DOCK8-deficient B cells, but those driven by the costimulatory molecule CD40 were not. In contrast, TLR9-driven expression of AICDA (which encodes the cytidine deaminase AID), the immunoglobulin receptor CD23 and the costimulatory molecule CD86 and activation of the transcription factor NF-κB, the kinase p38 and the GTPase Rac1 were intact. DOCK8 associated constitutively with MyD88 and the tyrosine kinase Pyk2 in normal B cells. After ligation of TLR9, DOCK8 became tyrosine-phosphorylated by Pyk2, bound the Src-family kinase Lyn and linked TLR9 to a Src-kinase Syk-transcription factor STAT3 cascade essential for TLR9-driven B cell proliferation and differentiation. Thus, DOCK8 functions as an adaptor in a TLR9-MyD88 signaling pathway in B cells.

Mycobacterial disease in patients with chronic granulomatous disease: A retrospective analysis of 71 cases.

BACKGROUND: Chronic granulomatous disease (CGD) is a rare primary immunodeficiency caused by inborn errors of the phagocyte nicotinamide adenine dinucleotide phosphate oxidase complex. From the first year of life onward, most affected patients display multiple, severe, and recurrent infections caused by bacteria and fungi. Mycobacterial infections have also been reported in some patients. OBJECTIVE: Our objective was to assess the effect of mycobacterial disease in patients with CGD. METHODS: We analyzed retrospectively the clinical features of mycobacterial disease in 71 patients with CGD. Tuberculosis and BCG disease were diagnosed on the basis of microbiological, pathological, and/or clinical criteria. RESULTS: Thirty-one (44%) patients had tuberculosis, and 53 (75%) presented with adverse effects of BCG vaccination; 13 (18%) had both tuberculosis and BCG infections. None of these patients displayed clinical disease caused by environmental mycobacteria, Mycobacterium leprae, or Mycobacterium ulcerans. Most patients (76%) also had other pyogenic and fungal infections, but 24% presented solely with mycobacterial disease. Most patients presented a single localized episode of mycobacterial disease (37%), but recurrence (18%), disseminated disease (27%), and even death (18%) were also observed. One common feature in these patients was an early age at presentation for BCG disease. Mycobacterial disease was the first clinical manifestation of CGD in 60% of these patients. CONCLUSION: Mycobacterial disease is relatively common in patients with CGD living in countries in which tuberculosis is endemic, BCG vaccine is mandatory, or both. Adverse reactions to BCG and severe forms of tuberculosis should lead to a suspicion of CGD. BCG vaccine is contraindicated in patients with CGD.

Predictive risk factors for relapse after cessation of inhaled corticosteroids in well-controlled childhood asthma.

BACKGROUND: There is limited data for predicting the risk of exacerbations following the cessation of inhaled corticosteroids (ICS) in well controlled childhood asthma. In current study, clinical, functional and inflammatory parameters at the time of ICS withdrawal were investigated in that respect. METHODS: Forty children asymptomatic for at least 3 months on low dose ICS's were enrolled and ICS were discontinued in summer. At enrolment symptom/medication diary, pulmonary function parameters, methacholine provocation testing, peripheral blood eosinophilia, serum total and allergen-specific IgE levels and skin prick testing were performed. In a subgroup of patients, phytohemaglutinin induced secretion of IL-5, IL-13, IFN-γ and IL-10 from blood mononuclear cells were measured. Patients were assessed with symptom/medication diary and pulmonary function test every 2 months for 6 months. RESULTS: Eighteen of 37 patients experienced recurrence of acute asthma symptoms. In patients with acute attack (group I), changes in rhinitis symptom scores at 2nd month vs. baseline were statistically higher. In addition, group I had significantly higher rhinitis symptom scores compared to group II at fourth-month visit. Patients with acute exacerbation revealed a significant decrease in FEV1% at 2nd month compared to baseline. Moreover, group I showed significantly lower FEF 25-75% compared to group II at 2nd month. Baseline bronchial hyper-responsiveness with methacholine was found to be an independent risk factor for asthma exacerbation. CONCLUSIONS: The findings of this study identified baseline bronchial hyperreactivity, higher rhinitis symptom scores and gradual decrease in pulmonary function parameters during follow-up as risk factors for subsequent exacerbation of asthma.

Severe Early-Onset Combined Immunodeficiency due to Heterozygous Gain-of-Function Mutations in STAT1.

PURPOSE: Loss and gain-of-function (GOF) mutations in human signal transducer and activator of transcription 1 (STAT1) lead to distinct phenotypes. Although recurrent infections are common to both types of STAT1 mutations, GOF mutations are distinguished by chronic mucocutaneous candidiasis and autoimmunity. However, the clinical spectra of STAT1 GOF mutations continue to expand. We here describe two patients with STAT1 GOF mutations presenting early in life with combined immunodeficiency (CID). METHODS: Clinical data and laboratory findings including immunophenotyping, level of interferon (IFN)-γ/IL-17(+) T cells, interferon-induced STAT1 phosphorylation, and JAK inhibitor assays were evaluated. Sequencing of STAT1 gene was performed by Sanger sequencer. RESULTS: Patient 1 (P1) had persistent oral candidiasis and cytomegalovirus (CMV) infection since 2 months of age and later developed cavitary lung lesions due to Mycobacterium tuberculosis. Patient 2 (P2) presented with oral candidiasis and recurrent pneumonia at 4 months of age and subsequently developed CMV pneumonitis. Both patients suffered heterozygous missense mutations in STAT1, leading to deleterious amino acid substitutions in the DNA binding domain (P1: c.1154C > T; p.T385M; P2. c.971G > T; p.C324F). Circulating CD4(+) T cells of both patients exhibited increased interferon-γ and decreased IL-17 expression as compared to controls. They also exhibited increased IFN-ß and -γ-induced STAT1 phosphorylation that was reversed upon treatment with the JAK kinase inhibitor ruxolitinib. CONCLUSION: STAT1 GOF mutations may present early in life with CID, consistent with the clinical heterogeneity of the disease. JAK kinase inhibitors may potentially be useful in some patients as adjunct therapy pending definitive treatment with bone marrow transplantation.

The effect of systemic corticosteroids on the innate and adaptive immune system in children with steroid responsive nephrotic syndrome.

UNLABELLED: The severity and duration of immunosuppression caused by corticosteroids (CSs) usage have not been extensively studied. We aimed to investigate the effects of CSs on the various compartments of immune system in relation to timing of initiation and persistence of therapy. Pediatric patients with idiopathic nephrotic syndrome (NS) treated with 2 mg/kg/day prednisolone and healthy control (HC) were enrolled. Blood samples were drawn for immunologic analyses at baseline and at the first and second weeks and first, second, and third months of CS therapy in addition to first and second weeks and first, second, and third months of discontinuation. Fourteen patients (M/F, 7/7) between 1 and 8 years old were evaluated. Untreated NS exhibited high absolute lymphocyte count (ALC)(p = 0.010), absolute CD3(+) T cells (p = 0.020) and absolute CD8(+) T cells (p = 0.006) compared to HC. Suppression in ALC was observed and nadir value was noted at first month of therapy compared to baseline (p = 0.002). The CD4(+) (p = 0.036) and CD8(+) T cell (p = 0.013) counts decreased significantly at the first week of treatment compared to baseline. While baseline B cell counts was indifferent from HC, gradually increased in 2 weeks of CS initiation and decreased during the treatment with a statistical significance compared to HC (p = 0.010). However, after cessation of CS, B cell counts continued to decline and found to be significantly different than baseline at first week (p = 0.008) and at third month (p = 0.040). CONCLUSION: Apart from baseline lymphocyte subset changing observed in untreated NS patients, our data implies that T cells were suppressed very early in the CS treatment. Interestingly, depressed B cell counts were detected later but persisted even after CS cessation. Due to early decrease in T cells, it would be beneficial to assume the patients as immunosuppressed at the very beginning of CS treatment to avoid infections. WHAT IS KNOWN: • Corticosteroids (CSs) are widely used for a variety of diseases including nephrotic syndrome, which is related with complex immune disturbance including T and B cells dysfunctions. • CSs induce neutrophilic leukocytosis concomitant with lymphopenia and eosinopenia leading to immunosupression. What is New: • T cell subsets and proliferation are susceptible to CSs more than B cells; however, the reversibility is faster with dose reduction in CS. • The change of B cells and B cell subtypes (CD27 (+) memory) shows prolonged effect of CSs on B cells which may alter antibody production even after 3 months of CSs cessation.

Potentially Beneficial Effect of Hydroxychloroquine in a Patient with a Novel Mutation in Protein Kinase Cδ Deficiency.

Protein kinase C delta (PRKCD) has essential functions in controlling B-cell proliferation and apoptosis, development of B-cell tolerance and NK-cell cytolitic activity. Human PRKCD deficiency was recently identified to be causative for an autoimmune lymphoproliferative syndrome like disorder with significant B-cell proliferation particularly of immature B cells. Here we report a child with a novel mutation in PRKCD gene who presented with CMV infection and an early onset SLE-like disorder which was successfully treated with hydroxychloroquine.

G6PC3 Deficiency: Primary Immune Deficiency Beyond Just Neutropenia.

Glucose-6-phosphatase catalytic subunit 3 (G6PC3) deficiency was recently defined as a new severe congenital neutropenia subgroup remarkable with congenital heart defects, urogenital malformations, endocrine abnormalities, and prominent superficial veins. Here, we report 3 patients with G6PC3 deficiency presenting with recurrent diarrhea, failure to thrive, and sinopulmonary infections leading to bronchiectasis. In patient I and II, a combined immune deficiency was suspected due to early-onset disease with lymphopenia, neutropenia, and thrombocytopenia, along with variable reductions in lymphocyte subpopulations and favorable response to intravenous γ-globulin therapy. Apart from neutropenia, all 3 patients had intermittent thrombocytopenia, anemia, and lymphopenia. All patients had failure to thrive and some of the classic syndromic features of G6PC3 deficiency, including cardiac abnormalities and visibility of superficial veins in all, endocrinologic problems in PI and PIII, and urogenital abnormalities in PII. Our experience suggests that a diagnosis of congenital neutropenia due to G6PC3 may not be as straightforward in such patients with combined lymphopenia and thrombocytopenia. A high index of suspicion and the other syndromic features of G6PC3 were clues to diagnosis. Screening of all combined immune deficiencies with neutropenia may help to uncover the whole spectra of G6PC3 deficiency.

Dedicator of cytokinesis 8-deficient patients have a breakdown in peripheral B-cell tolerance and defective regulatory T cells.

BACKGROUND: Dedicator of cytokinesis 8 (DOCK8) deficiency is typified by recurrent infections, increased serum IgE levels, eosinophilia, and a high incidence of allergic and autoimmune manifestations. OBJECTIVE: We sought to determine the role of DOCK8 in the establishment and maintenance of human B-cell tolerance. METHODS: Autoantibodies were measured in the plasma of DOCK8-deficient patients. The antibody-coding genes from new emigrant/transitional and mature naive B cells were cloned and assessed for their ability to bind self-antigens. Regulatory T (Treg) cells in the blood were analyzed by means of flow cytometry, and their function was tested by examining their capacity to inhibit the proliferation of CD4(+)CD25(-) effector T cells. RESULTS: DOCK8-deficient patients had increased levels of autoantibodies in their plasma. We determined that central B-cell tolerance did not require DOCK8, as evidenced by the normally low frequency of polyreactive new emigrant/transitional B cells in DOCK8-deficient patients. In contrast, autoreactive B cells were enriched in the mature naive B-cell compartment, revealing a defective peripheral B-cell tolerance checkpoint. In addition, we found that Treg cells were decreased and exhibited impaired suppressive activity in DOCK8-deficient patients. CONCLUSIONS: Our data support a critical role for DOCK8 in Treg cell homeostasis and function and the enforcement of peripheral B-cell tolerance.

Clinical heterogeneity of immunodysregulation, polyendocrinopathy, enteropathy, X-linked: pulmonary involvement as a non-classical disease manifestation.

PURPOSE: IPEX (Immunodysregulation, Polyendocrinopathy, Enteropathy, X-linked) is a rare X-linked recessive life-threatening disorder characterized by autoimmunity and early death. Pulmonary complication related with IPEX has not been elucidated exactly. Here, we report 4 IPEX patients, 3 of which died from severe pulmonary disease. METHODS: Clinical data and laboratory findings including autoantibodies, immunoglobulin levels as well as number of T, B and NK cells were evaluated. FOXP3 expression and T reg activity were analyzed. The FOXP3 gene was sequenced and RNA analysis was performed. RESULTS: Patient I (PI) presented with nephrotic syndrome at 3 years of age and then developed autoimmune hepatitis without eczema, enteropathy or high IgE and died at 9 years of age due to acute respiratory distress syndrome (ARDS). Two cousins of PI had the same hypomorphic splice site mutation leading to a deletion of 27 amino acids, but normal FOXP3 protein expression and normal suppressive capacity of T reg in a proliferation inhibition assay. However, they exhibited typical symptoms such as eczema, diabetes and enteropathy with eosinophilia at early age (PII, PIII) and were transplanted in infancy. One of them had severe respiratory distress right after birth (PIII). Patient IV from another family presented with chronic diarrhea without autoimmune manifestations and died due to ARDS. CONCLUSION: Lung disease related to IPEX syndrome has not been reported before and this entity could be a critical factor in disease outcome.

Clinical, functional, and genetic characterization of chronic granulomatous disease in 89 Turkish patients.

BACKGROUND: Chronic granulomatous disease (CGD) is a rare primary immunodeficiency disorder of phagocytes resulting in impaired killing of bacteria and fungi. A mutation in one of the 4 genes encoding the components p22(phox), p47(phox), p67(phox), and p40(phox) of the leukocyte nicotinamide dinucleotide phosphate reduced (NADPH) oxidase leads to autosomal recessive (AR) CGD. A mutation in the CYBB gene encoding gp91(phox) leads to X-linked recessive CGD. OBJECTIVE: The aim of this study is to show the correlation between clinical, functional, and genetic data of patients with CGD from Turkey. METHODS: We report here the results of 89 patients with CGD from 73 Turkish families in a multicenter study. RESULTS: Most of the families (55%) have an AR genotype, and 38% have an X-linked genotype; patients from 5 families with a suspected AR genotype (7%) were not fully characterized. We compared patients with CGD according to the severity of NADPH oxidase deficiency of neutrophils. Patients with A22(0), A67(0) or X91(0) phenotypes with a stimulation index of 1.5 or less have early clinical presentation and younger age at diagnosis (mean, 3.2 years). However, in p47(phox)-deficient cases and in 5 other AR cases with high residual oxidase activity (stimulation index ≥ 3), later and less severe clinical presentation and older age at diagnosis (mean, 7.1 years) were found. Pulmonary involvement was the most common clinical feature, followed by lymphadenitis and abscesses. CONCLUSION: Later and less severe clinical presentation and older age at diagnosis are related to the residual NADPH oxidase activity of neutrophils and not to the mode of inheritance. CGD caused by A22(0) and A67(0) subtypes manifests as severe as the X91(0) subtype.

Risk factors for persistence of asthma in children: 10-year follow-up.

OBJECTIVE: Risk factors related to the outcome of childhood asthma are not yet well established. We aimed to investigate the long-term outcome for children with asthma to determine the risk factors in predicting persistence of disease. METHODS: Sixty-two children with asthma were evaluated retrospectively at the end of a 10-year follow-up. Patients were asked to complete a questionnaire requesting clinical information, and underwent physical examination, skin prick testing, a pulmonary function test and bronchial provocation testing. Immunologic parameters evaluated were allergen-specific IgE and IgG4 levels, and allergen-induced generation of CD4(+)CD25(+) cells. RESULTS: Mean age at final assessment was 15.9 ± 3.6 years, and duration of follow-up was 10.30 ± 1.27 years. Fifty percent of patients outgrew their asthma during the 10-year follow-up period. All the non-atopic patients outgrew their disease during the study period, whereas 67% of atopic patients did not. We identified two risk factors independently related to the persistence of symptoms: presence of bronchial hyper-responsiveness and presence of rhinitis. Atopic children who were in remission demonstrated significantly higher allergen-induced CD4(+)CD25(+) T cells compared to healthy controls. CONCLUSIONS: Atopy, presence of rhinitis, positive and presence of bronchial hyper-reactivity are important risk factors for the persistence of asthma in children. Allergen-induced CD4(+)CD25(+) T cells were higher in the atopic children who outgrew their disease, implicating an immunological mechanism of asthma remission in children.

A novel approach in allergen-specific immunotherapy: combination of sublingual and subcutaneous routes.

BACKGROUND: Subcutaneous allergen-specific immunotherapy (SIT) has an early onset of action, whereas repeated injections and safety concerns have limited its use in the pediatric age group. Meanwhile, the improved safety profile of the sublingual route has been accepted as an alternative despite its relatively late onset of action. OBJECTIVE: We sought to improve the efficacy and safety of SIT with a combination of the subcutaneous route in the build-up phase and sublingual maintenance in comparison with the sublingual or subcutaneous routes alone. METHODS: Fifty-one house dust mite-sensitized children with mild-to-moderate asthma were randomized into one of 4 groups to receive either (1) subcutaneous immunotherapy (SCIT), (2) sublingual immunotherapy (SLIT), (3) SCIT plus SLIT, or (4) pharmacotherapy. Clinical parameters were evaluated at baseline and months 1, 4, 12, and 18. Allergen-specific immunoglobulin levels and allergen-induced IL-5, IL-10, IL-13, IL-17, TGF-ß, and IFN-γ levels were evaluated as well. RESULTS: In the SCIT and SCIT plus SLIT groups, the number of asthma attacks and inhaled corticosteroid dosage decreased compared with baseline values at the months 4, 12, and 18 but only at month 12 in the SLIT group. The improvement in visual analog scores for rhinitis was significant only in the SCIT plus SLIT group. Increases in the levels of regulatory and T(H)1 cytokines were observed both in the SCIT and SLIT groups, with some differences in dynamics. Antigen-specific IgG(4) levels increased in the SCIT and SCIT plus SLIT groups but not in the SLIT group. Clinical symptom scores were correlated positively with IL-5 levels and negatively with antigen-specific IgG(4), IFN-γ, and TGF-ß levels. CONCLUSION: Our novel regimen of immunotherapy, SCIT plus SLIT, appeared promising in that it successfully combined the advantages of the 2 alternatives: rapid onset and potency in SCIT and safety and avoidance of injections in SLIT.

Serum immunoglobulin levels as a predictive factor for a better outcome of non-atopic childhood asthma.

Childhood asthma is a heterogeneous condition with different phenotypes. Hereby, we aimed to study impact of serum immunoglobulin levels on clinical phenotypes and outcome of asthma. Seventy-eight children (M: 26, F: 52) aged less than 10 yrs (mean = 8.56 ± 3.23 yrs) and diagnosed as mild-moderate persistent asthma, followed up for at least 1 yr were included into the study. Asthmatic children were divided into two groups based on serum immunoglobulin levels at admission and were evaluated with respect to demographic data, allergic sensitization, symptom scores, medication usage, pulmonary functions, and non-specific bronchial hyper-reactivity. The age at onset of symptoms (40.88 ± 32.02 vs. 23.04 ± 26.97 months) was significantly younger in children with hypogammaglobulinemia (n = 28) compared to normogammaglobulinemia group (n = 50) (p = 0.016). Mean follow-up duration was 3.8 ± 2.1 yrs. Atopic sensitization rate was higher in those with normal immunoglobulin levels (81.2% vs. 17.9%), (p < 0.0001). Normal serum immunoglobulin levels were associated with atopic asthma (OR, 4.5; 95% confidence interval (CI): 2.0-10.1). For the prediction of atopic asthma, having normal immunoglobulin levels yielded predictive values of: sensitivity = 88.6%, specificity = 71.8%, positive predictive value = 81.1%, negative predictive value = 82.1%. Furthermore, percentages of atopic dermatitis and allergic conjunctivitis, elevated serum total IgE levels, eosinophilia, and bronchial hyper-reactivity were more common in normogammaglobulinemia with asthma group (p = 0.040, p = 0.003, p = 0.024, p = 0.030, p = 0.040, respectively). Although marked reductions in asthma scores and inhaled corticosteroid usage were observed in both groups over time, the rate of decline was significantly higher and earlier in hypogammaglobulinemia group (p = 0.0001, p = 0.004, respectively). In conclusion, asthmatic children with hypogammaglobulinemia presented at an earlier age, with lower rates of atopy, and earlier clinical improvement accompanied with earlier discontinuation of inhaled corticosteroids than children with normal immunoglobulin levels. Our data demonstrated that in children currently named as early-onset non-atopic asthma, hypogammaglobulinemia might be accompanying, providing evidence for a different phenotype of childhood asthma.

Osteoporosis: an ignored complication of CVID.

BACKGROUND: Multiple factors in common variable immunodeficiency (CVID) might interfere with optimal growth and maturation and potentially compromise bone health. METHODS: We aimed to evaluate bone mineral density (BMD) of patients with CVID using dual energy X-ray absorptiometry (DEXA) and investigate risk factors associated with decreased bone density. RESULTS: Twenty-two patients were included (M: 16, F: 6) with a mean age of 15.6 ± 9.0 yr. DEXA revealed osteopenia in 6/22 (27.3%) and osteoporosis in 9/22 (40.9%) at lumbar spine and osteopenia in 7/19 (37%) and osteoporosis in 3/19 (16%) at femoral neck sites. The age of subjects with osteoporosis was significantly higher than those without (21.6 ± 8.0 vs. 9.0 ± 5.7 yr; p < 0.0001). BMD z-scores were significantly lower in patients with bronchiectasis compared with those without (p = 0.03). Patients with osteoporosis at femoral neck site had lower forced expiratory volume in 1 s (FEV(1) ) (p = 0.024), FEV(1) /forced vital capacity (FVC) (p < 0.0001), PEF (p = 0.008), and FEF 25-75 (p = 0.013) values compared with the patients with normal BMD z-scores. Low serum 25(OH) vitamin D levels were detected in 13/22 patients and low dietary calcium intake in 17/22 patients. BMD z-scores at femoral neck were lower in patients with low B-cell percentage (p = 0.03). BMD z-score at lumbar spine was correlated with folate (r = +0.63, p = 0.004) and serum immunoglobulin G levels (r = +0.430, p = 0.04). CONCLUSION: Osteoporosis appeared as an emerging health problem of patients with CVID, the risk increasing with older age and poorer lung function. Nutritional, biochemical, and immunologic factors appeared to take part in decreased BMD. Insight into the mechanisms of osteoporosis in CVID is crucial to develop preventive strategies.

Vitamin B12 deficiency among asymptomatic healthy infants: its impact on the immune system.

BACKGROUND: The immunomodulatory effects of vitamin B12 deficiency in children have not yet been established in the literature. In the current study, the effects of vitamin B12 on the immune system in asymptomatic and otherwise healthy infants have been studied. METHODS: The study was conducted at Marmara University, "well-child" outpatient clinic. Vitamin B12 level was measured in a cohort of 611 healthy term infants, followed regularly for at least 6 months. Immunoglobulin measurements, lymphocyte subset analysis, cytokine production analysis, lymphocyte proliferation assays and evaluation of lymphocyte apoptosis were performed in a subset of 60 infants. RESULTS: In this cohort, one out of three babies displayed vitamin B12 deficiency. The percentage of CD4+CD25+ regulatory T cells (Tregs) was lower in vitamin B12 deficient babies than in controls. Although the percentage of Tregs increased after treatment, the change was not significant. There was no difference of cytokine levels between vitamin B12 deficient and control groups. However, proinflammatory cytokines were reduced after treatment. No significant difference was observed for immunoglobulins, early apoptosis and lymphocyte proliferation. CONCLUSIONS: Vitamin B12 deficiency is an underestimated health problem among the developing countries. The clinical consequences of the decreased percentage of Tregs associated with vitamin B12 deficiency, and reduction of proinflammatory cytokines after vitamin supplementation needs to be further studied, especially in terms of emerging allergies, autoimmune disorders and anti-inflammatory effects.

Functions of NK and iNKT cells in pediatric and adult CVID, ataxia telangiectasia and agammaglobulinemia patients.

Primary immune deficiencies (PID) are known to be more than 400 genetic defects caused by the impairment in development and/or functions of the immune system. Common Variable Immunodeficiency (CVID), Ataxia Telangiectasia (AT) and Agammaglobulinemia (AG) are examples of the most common immunodeficiency syndrome. Natural killer (NK) cells are a component of innate immune system and play a major role in the host-rejection of both tumors and virally infected cells. iNKT cells have a role in autoimmune and infectious diseases and controlling of tumor rejection. In this study, NK and iNKT cells and their functions, and intracellular cytokine amount are aimed to determine in patients that suffer CVID, AT and AG. NKp30, NKp46, NKG2D, perforin and granzyme mRNA expression levels were analyzed using RT-PCR. Receptors, cytokine amount of NK cell subset and iNKT were analyzed by flow cytometry. Decreased CD3+ T and elevated NK cell subset in pediatric AT were found. Expression of NKp44 was decreased in adult AG, but not in pediatric patients. Low NKp44 expression in CD3-CD16+CD56dim NK cell subset was found in pediatric AT patients. High HLA-DR, perforin and granzyme expression were found in CD3-CD16+CD56dim NK cell subset of pediatric CVID and AT patients. Alteration of the number of NK subsets, NK receptor expression and cytokine production were observed in pediatric patients compared to healthy subjects.

Outcome of hypogammaglobulinemia in children: immunoglobulin levels as predictors.

We evaluated 131 children (M=88, F=43) with hypogammaglobulinemia. Data was analyzed mainly for delineating predictor factors for outcome. The distance from the lower limit of normal (-2SD) for any single measurement of immunoglobulins (Ig) was calculated and transformed into Ig scores. Mean age and duration of follow-up were 5.06 ± 4.05 and 3.7 ± 3.03 years, respectively. The diagnoses were: 22 CVID, 16 IgA deficiency, 33 transient hypogammaglobulinemia of childhood (THC), 3 selective IgM deficiency and 57 unclassified hypogammaglobulinemia (UCH). Low IgA scores (<-0.124) at presentation were indicative of subsequent development of IgA deficiency or CVID, whereas low IgM score (<-0.038) pointed towards more severe and persistent phenotypes. Combination of low IgM score between 2 and 5 years, impaired antibody response and low B cell counts enabled us to predict persistence of hypogammaglobulinemia beyond 5 years (specificity=90.5% and PPV=94.9%) and chronic lung disease (sensitivity=90.4% and specificity=68.3%). The set of criteria including low IgM scores, impaired antibody response and low B cell counts provided a high predictive value in detecting those with persistent hypogammaglobulinemia.

Neonatal BCG vaccination induces IL-10 production by CD4+ CD25+ T cells.

To determine the optimal time of Bacillus Calmette-Guerin (BCG) vaccination for induction of Th1 immunity, we measured the interferon (IFN)-γ and interleukin (IL)-10 secretion in purified protein derivative (PPD)-stimulated peripheral blood mononuclear cell (PBMC) cultures in newborns vaccinated at birth or 2nd month of life. Moreover, role of CD4(+) CD25(+) T cells was studied by depletion assay at 8th month. Nineteen term and healthy newborns were randomized into two groups: Group I composed of 10 newborns vaccinated with BCG at birth and the remaining 9 (group II) at 2nd month of life. PBMCs were isolated at birth, 2nd and 8th months of age, and PPD-stimulated IL-10, 5 and IFN-γ secretion were assessed. The same measurements were repeated for IL-10 and IFN-γ after the depletion of CD4(+) CD25(+) T cells at the 8th month. Children vaccinated at birth demonstrated higher PPD-stimulated IFN-γ and IL-10 levels at 2 months of age when compared to non-vaccinated ones (p = 0.038 and p = 0.022, respectively), whereas at 8 months, no significant differences were detected between the two groups. Moreover, CD4(+) CD25(+)-depleted T-cell cultures resulted in lower PPD-stimulated IL-10 levels in those vaccinated at birth when compared to non-depleted condition at the 8th month (p < 0.001). BCG at birth upregulated PPD-stimulated IFN-γ secretion at the 2nd month and remained still detectable at 8 month after the vaccination, whereas those vaccinated at the 2nd month of life lacked that increase in IFN-γ response at the same time-point. Furthermore, depletion assays suggest that CD4(+) CD25(+) T cells are involved in PPD-stimulated IL-10 secretion in response to BCG vaccination.