Cristina M. Kokron

Reduced frequency of CD4+CD25 HIGH FOXP3+ cells and diminished FOXP3 expression in patients with Common Variable Immunodeficiency: A link to autoimmunity?

Common Variable Immunodeficiency (CVID) is a primary immunodeficiency disease characterized by defective immunoglobulin production and often associated with autoimmunity. We used flow cytometry to analyze CD4(+)CD25(HIGH)FOXP3(+) T regulatory (Treg) cells and ask whether perturbations in their frequency in peripheral blood could underlie the high incidence of autoimmune disorders in CVID patients. In this study, we report for the first time that CVID patients with autoimmune disease have a significantly reduced frequency of CD4(+)CD25(HIGH)FOXP3(+) cells in their peripheral blood accompanied by a decreased intensity of FOXP3 expression. Notably, although CVID patients in whom autoimmunity was not diagnosed had a reduced frequency of CD4(+)CD25(HIGH)FOXP3(+) cells, FOXP3 expression levels did not differ from those in healthy controls. In conclusion, these data suggest compromised homeostasis of CD4(+)CD25(HIGH)FOXP3(+) cells in a subset of CVID patients with autoimmunity, and may implicate Treg cells in pathological mechanisms of CVID.

IVIg Immune Reconstitution Treatment Alleviates the State of Persistent Immune Activation and Suppressed CD4 T Cell Counts in CVID

Common variable immunodeficiency (CVID) is characterized by defective B cell function, impaired antibody production, and increased susceptibility to bacterial infections. Here, we addressed the hypothesis that poor antibody-mediated immune control of infections may result in substantial perturbations in the T cell compartment. Newly diagnosed CVID patients were sampled before, and 6–12 months after, initiation of intravenous immunoglobulin (IVIg) therapy. Treatment-naïve CVID patients displayed suppressed CD4 T cell counts and myeloid dendritic cell (mDC) levels, as well as high levels of immune activation in CD8 T cells, CD4 T cells, and invariant natural killer T (iNKT) cells. Expression of co-stimulatory receptors CD80 and CD83 was elevated in mDCs and correlated with T cell activation. Levels of both FoxP3+ T regulatory (Treg) cells and iNKT cells were low, whereas soluble CD14 (sCD14), indicative of monocyte activation, was elevated. Importantly, immune reconstitution treatment with IVIg partially restored the CD4 T cell and mDC compartments. Treatment furthermore reduced the levels of CD8 T cell activation and mDC activation, whereas levels of Treg cells and iNKT cells remained low. Thus, primary deficiency in humoral immunity with impaired control of microbial infections is associated with significant pathological changes in cell-mediated immunity. Furthermore, therapeutic enhancement of humoral immunity with IVIg infusions alleviates several of these defects, indicating a relationship between poor antibody-mediated immune control of infections and the occurrence of abnormalities in the T cell and mDC compartments. These findings help our understanding of the immunopathogenesis of primary immunodeficiency, as well as acquired immunodeficiency caused by HIV-1 infection.

Dysregulated CD1 profile in myeloid dendritic cells in CVID is normalized by IVIg treatment.

To the editor: The CD1 proteins are major histocompatibility complex class I-like molecules specialized in presenting lipid and glycolipid antigens to T cells.[1][1] Group I CD1s include CD1a, CD1b, and CD1c and present bacterial antigens to a diverse repertoire of pathogen-specific T cells.[2][2]

Predictors of Bothrops jararaca venom allergy in snake handlers and snake venom handlers

Since allergic sensitization to snake venom has been reported, anaphylactic reactions to snake venom might be an underestimated factor contributing to fatal snakebites, independently from the toxicity of the venom itself. However, little information is available on the determinants of such reaction. Hence, we studied a group of workers exposed to Bothrops jararaca venom (BJV), in order to clarify the factors related with snake venom allergy. The aim of this work was to investigate the prevalence and predictors of venom allergy among workers exposed to BJV and to confirm the involvement of IgE-mediated mechanisms in this condition. Workers exposed to BJV were assessed for venom allergy using questionnaires and immunological tests. The presence of BJV sensitization was determined through quantification of specific IgE. Allergens were studied using the Western blots and inhibition assays. Of the 67 workers evaluated, 7 (10.4%) presented specific IgE antibodies to BJV. Of those, 6 presented typical symptoms of an IgE-mediated allergic reaction when exposed to BJV. Venom sensitization was associated with length of employment (P=0.042), high levels of total IgE (P=0.034), atopy (P=0.051), and specific tasks, primarily the handling of dried venom (P=0.014). Our observations suggest that exposure to BJV can result in allergic sensitization in snake handlers through IgE-mediated mechanisms. The prevalence rate of this condition appears to be high among these workers, and the handling of dried venom, total IgE level above 100 kU/L, length of employment, and probably history of atopy were predictors of its occurrence.

IL-10-Producing Regulatory B Cells Are Decreased in Patients with Common Variable Immunodeficiency

Common variable immunodeficiency (CVID) is the most prevalent symptomatic primary immunodeficiency in adults. CVID patients often present changes in the frequency and function of B lymphocytes, reduced number of Treg cells, chronic immune activation, recurrent infections, high incidence of autoimmunity and increased risk for malignancies. We hypothesized that the frequency of B10 cells would be diminished in CVID patients because these cells play an important role in the development of Treg cells and in the control of T cell activation and autoimmunity. Therefore, we evaluated the frequency of B10 cells in CVID patients and correlated it with different clinical and immunological characteristics of this disease. Forty-two CVID patients and 17 healthy controls were recruited for this study. Cryopreserved PBMCs were used for analysis of T cell activation, frequency of Treg cells and characterization of B10 cells by flow cytometry. IL-10 production by sorted B cells culture and plasma sCD14 were determined by ELISA. We found that CVID patients presented decreased frequency of IL-10-producing CD24hiCD38hi B cells in different cell culture conditions and decreased frequency of IL-10-producing CD24hiCD27+ B cells stimulated with CpG+PIB. Moreover, we found that CVID patients presented lower secretion of IL-10 by sorting-purified B cells when compared to healthy controls. The frequency of B10 cells had no correlation with autoimmunity, immune activation and Treg cells in CVID patients. This work suggests that CVID patients have a compromised regulatory B cell compartment which is not correlated with clinical and immunological characteristics presented by these individuals.

Establishing a cut-off for the serum levels of specific IgE to milk and its components for cow's milk allergy: Results from a specific population

BACKGROUND Cows milk allergy diagnosis many times requires double-blind placebo-controlled food challenge (DBPCFC), which presents high accuracy but involves risks, specifically in infants and anaphylactic patients. The identification of the cut-off values for specific IgE to milk or its components would contribute to cows milk allergy (CMA) diagnosis. The aim of this study was to compare discriminating concentration of a cows milk specific IgE and its fractions (α-lactoalbumin, β-lactoglobulin, casein) in children for the CMA diagnosis. METHODS this study included 123 patients (M:F=1.3:1) median age at diagnosis=1.91 years, (3.5m to 13.21y) with CMA diagnosis via DBPCFC (n=26), proven anaphylaxis due to cows milk (n=46) or a suggestive clinical history associated with a positive skin prick test (n=51) and open oral food challenge. The control group included 61 patients (1 male:1.1 female) ages ranging from 0.66 to 16.7 years (median=6.83 years). Receiver operator characteristics (ROC) curves were constructed to determine the best cut-offs that guarantees high specificity (>95%) for cows milk and its components. RESULTS considering 98% specificity, cut-off points were: 3.06 kU/L for cows milk, 2.06 kU/L for α-lactalbumin, 1.85 kU/L for β-lactoglobulin and 1.47kU/L for casein. The best ROC curve (area under the curve=0.929) was obtained evaluating cows milk. CONCLUSION this study showed that the cut-off point detected for whole cows milk revealed a better discriminatory capacity for CMA diagnosis without the necessity of the milk components testing.

ANALYSIS OF THE SPUTUM AND INFLAMMATORY ALTERATIONS OF THE AIRWAYS IN PATIENTS WITH COMMON VARIABLE IMMUNODEFICIENCY AND BRONCHIECTASIS

INTRODUCTION: Common variable immunodeficiency is characterized by defective antibody production and recurrent pulmonary infections. Intravenous immunoglobulin is the treatment of choice, but the effects of Intravenous immunoglobulin on pulmonary defense mechanisms are poorly understood. OBJECTIVE: The aim of this study was to verify the impact of intravenous immunoglobulin on the physical properties of the sputum and on inflammatory alterations in the airways of patients with Common variable immunodeficiency associated with bronchiectasis. METHOD: The present study analyzed sputum physical properties, exhaled NO, inflammatory cells in the sputum, and IG titers in 7 patients with Common variable immunodeficiency and bronchiectasis with secretion, immediately before and 15 days after Intravenous immunoglobulin. A group of 6 patients with Common variable immunodeficiency and bronchiectasis but no sputum was also studied for comparison of the basal IgG level and blood count. The 13 patients were young (age=36±17 years) and comprised predominantly of females (n=11). RESULTS: Patients with secretion presented significantly decreased IgG and IgM levels. Intravenous immunoglobulin was associated with a significant decrease in exhaled NO (54.7 vs. 40.1 ppb, p<0.05), sputum inflammatory cell counts (28.7 vs. 14.6 cells/mm3, p<0.05), and a significant increase in respiratory mucus transportability by cough (42.5 vs. 65.0 mm, p < 0.05). CONCLUSION: We concluded that immunoglobulin administration in Common variable immunodeficiency patients results in significant improvement in indexes of inflammation of the airways with improvement in the transportability of the respiratory mucus by cough.

Can patients with common variable immunodeficiency have allergic rhinitis

Background Rhinosinusitis is highly prevalent in patients with common variable immunodeficiency (CVID), and probably allergic rhinitis (AR) may be masked by a history of repeated respiratory infections. The diagnosis of AR is based on the patients symptoms and detection of specific immunoglobulin E (IgE) to aeroallergens. This study was designed to identify rhinitis of probable allergic cause in patients with CVID. Methods This study included 72 adult CVID patients. The patients were divided into three groups according to their history: suggestive of AR, nonallergic rhinitis, and without rhinitis. They were tested for total and specific IgE (in vivo and in vitro). Results The patients’ mean age was 38.2 years. A history of chronic rhinitis was observed in 59 (81.9%) of the cases, 31 of which (43%) had a history suggestive of AR. Patients with a history of rhinitis (whether allergic or nonallergic) presented an earlier onset of symptoms and diagnosis of CVID. Total IgE was undetectable in 86.1% of patients. AR was confirmed by detection of specific IgE to aeroallergens in only 5.6% of the patients. Conclusion In CVID patients, chronic rhinitis may be allergic, because many have personal and family histories suggestive of atopy. However, in this study, allergy was confirmed by specific IgE detection in only 5.6% of cases. CVID patients with a history suggestive of AR commonly present negative results on traditional testing, so additional experiments may be necessary. One suggestion for the investigation of AR in CVID patients would be nasal provocation with the most prevalent allergens.

Slower rescue of ER homeostasis by the unfolded protein response pathway associated with common variable immunodeficiency

Common variable immunodeficiency (CVID) is a primary immunodeficiency characterized by hypogammaglobulinemia and recurrent infections. Herein we addressed the role of unfolded protein response (UPR) in the pathogenesis of the disease. Augmented unspliced X-box binding protein 1 (XBP-1) mRNA concurrent with co-localization of IgM and BiP/GRP78 were found in one CVID patient. At confocal microscopy analysis this patients cells were enlarged and failed to present the typical surface distribution of IgM, which accumulated within an abnormally expanded endoplasmic reticulum. Sequencing did not reveal any mutation on XBP-1, neither on IRE-1alpha that could potentially prevent the splicing to occur. Analysis of spliced XBP-1, IRE-1alpha and BiP messages after LPS or Brefeldin A treatment showed that, unlike healthy controls that respond to these endoplasmic reticulum (ER) stressors by presenting waves of transcription of these three genes, this patients cells presented lower rates of transcription, not reaching the same level of response of healthy subjects even after 48 h of ER stress. Treatment with DMSO rescued IgM and IgG secretion as well as the expression of spliced XBP-1. Our findings associate diminished splicing of XBP-1 mRNA with accumulation of IgM within the ER and lower rates of chaperone transcription, therefore providing a mechanism to explain the observed hypogammaglobulinemia.

Loss of Circulating Mucosal-Associated Invariant T Cells in Common Variable Immunodeficiency Is Associated with Immune Activation and Loss of Eomes and PLZF

Common variable immunodeficiency (CVID) is characterized by low levels of Igs leading to increased risk of infections. Mucosal-associated invariant T (MAIT) cells are a recently identified population of innate T cells with potent antibacterial activity. We hypothesized that CVID is associated with alterations in MAIT cells. Cryopreserved PBMC from CVID patients and healthy controls were used to study the frequency, phenotype, and response to Escherichia coli stimulation of MAIT cells by flow cytometry. MAIT cell frequency and absolute counts were depressed in CVID. Residual MAIT presented elevated coexpression of CD38 and HLA-DR, and reduced expression of CCR6, whereas levels of CD127 (IL-7 receptor) were unchanged. CVID patients also had an accumulation of MAIT cells lacking the critical transcription factors eomesodermin and promyelocytic leukemia zinc finger protein. MAIT cell frequency was inversely associated with levels of soluble CD14, with coexpression of CD38 and HLA-DR, and accumulation of MAIT cells lacking eomesodermin or promyelocytic leukemia zinc finger protein expression. None of these changes were normalized by IgG replacement therapy. Finally, MAIT cells from CVID patients displayed poor IFN-γ responses to E. coli stimulation, in part due to defective Ag presentation, and these responses were increased by pretreatment with IL-7. Defective MAIT cell response may contribute to the increased incidence of microbial infections seen in CVID patients on IgG replacement therapy.