IgA Subclasses Research Articles

Primary immunodeficiency in infection-prone children in southern Sweden: occurrence, clinical characteristics and immunological findings.

BackgroundPrimary immunodeficiency diseases (PIDs) comprise a heterogeneous group of disorders mainly characterized by increased susceptibility to infections. The aims of this study were to estimate the occurrence rate of PID in the paediatric (age ≤ 18 years) population of southern Sweden (approx. 265,000 children) and to describe their demographic, clinical and immunological characteristics. During a period of 4 years, in four paediatric speciality clinics in Skåne County in southern Sweden, children being seen for infections and fulfilling specific criteria were evaluated according to a predefined examination schedule. The initial analysis consisted of complete blood counts with analysis of lymphocyte subpopulations (T, B, NK cells), measurement of immunoglobulins (IgG, IgA, IgM, IgE and IgG subclasses), and assessment of the complement system (classical, alternative and lectin pathways). In addition, results of these immunological analyses in other children from the same area and time period were evaluated.ResultsIn total, 259 children (53.6% males) met the criteria and were included. The most common infection was recurrent otitis media. Immunological analyses results for about two thirds of the patients were outside age-related reference intervals. Further examination in this latter group identified 15 children with PID (9 males); 7 (2.7%) had genetically defined PID, representing 4 different diagnoses, and another 8 (3.1%) had a clinically defined PID - common variable immunodeficiency. No additional PID patient was identified from the evaluation of laboratory results in children not included in the study. The median age at diagnosis was 3.5 years (range 1–12 years).ConclusionsThe occurrence rate of PID was about 4 new cases per year in this population. Several different PID diagnoses were found, and the application of specified criteria to identify PID patients was useful. In children who are prone to infection, the use of a predefined set of immunological laboratory analyses at their first examination was beneficial for early identification of patients with PID.

Acute cycling sport causes upper respiratory tract infections and lung function loss among male recreational cyclists of different age groups

Background: Prolonged high intensity aerobic activity could lead to suppression of mucosal immune proteins like salivary IgA subclass, lactoferrin causing temporary immune suppressed state which may lead to Upper respiratory tract infections and loss in lung function. Purpose: This study examined the effect of acute cycling sport on the status of URTI and Forced Expiratory Volume of first second (FEV1) among the recreational cyclists, who participated in their first ever acute cycling sport of minimum 140 kms in a competitive set up. Methods: Seventy five first time recreational volunteer cyclists in four age groups were analysed on their URTI and FEV1. Wisconsin Upper Respiratory Symptom Survey (WURSS-21) score was used to analyse the URTI and Digital spirometer was used to record the FEV1. Fifteen volunteers for each age group were analysed and results compared. Age groups studied were 20-25yrs, 25-30, 30-35 and 35 above. Pre event and post event URTI and FEV1 were analysed with Analysis of Covariance (ANCOVA). Highest scores of the URTI and FEV1 within the first seven days of post event were included. Results: Covariance analysis (P<.05) indicated, for both URTI and FEV1 the four groups showed significant difference among themselves on their post event status. Adjusted post event URTI means were 62.92 (20-25 yrs), 55.03 (25-30), 79.56 (30-35) and 84.63(35 above). Tukey HSD post hoc comparison indicated that 35+ group experienced significant increases (P<.05) in their URTI symptoms, though the 30-35 group also experienced considerable increase in URTI symptoms. Adjusted post event FEV1 were 79.63, 81.51, 78.39 and 78 for the four groups and the Tukey post hoc analysis (P<.05) indicated that there were significant decrements in the FEV1 of both 30-35 and 35+ groups. Discussion: Peripheral airway change and hyper reactivity due to immunologic and inflammatory cell facilitation could be the reason for temporary bronchoconstriction leading to latent loss in lung function. Conclusions: Recreational cyclists of plus 35 years are vulnerable group with respect to the upper respiratory tract infections and loss of lung function considerably due to acute long distance cycling events.

Eosinophilic Esophagitis in Adults Is Associated With IgG4 and Not Mediated by IgE

Eosinophilic esophagitis is usually triggered by foods, by unclear mechanisms. We evaluated the roles of IgE and IgG4 in the development of eosinophilic esophagitis. We performed a prospective, randomized, double-blind, placebo-controlled trial of adults with eosinophilic esophagitis given an antibody against IgE (omalizumab, n= 16) or placebo (n= 14) every 2-4 weeks for 16 weeks, based on weight and serum level of IgE. Endoscopy was performed, esophageal biopsy specimens were collected, and symptoms were assessed at baseline and at 16 weeks. Maximum numbers of eosinophils/high-power field were determined. Homogenates of esophageal biopsy specimens from 11 subjects with eosinophilic esophagitis and 8 without (controls) were assessed for IgM, IgA, and IgG subclasses. In a retrospective analysis, we performed immunofluorescence analysis of IgG4 in fixed esophageal tissues from 2 patients with eosinophilic esophagitis who underwent esophagectomy and 47 consecutive autopsies (controls). We also performed immunofluorescence analysis of IgG4 in esophageal mucosal biopsy specimens from 24 subjects with eosinophilic esophagitis and 9 without (controls). Finally, sera were collected from 15 subjects with eosinophilic esophagitis and from 41 without (controls), and assayed for total and food-reactive IgG4. Omalizumab did not alter symptoms of eosinophilic esophagitis or eosinophil counts in biopsy samples compared with placebo. Homogenates of esophageal tissues from patients with eosinophilic esophagitis had a 45-fold increase in IgG4 compared with controls (P < 3× 10(-5)), but no significant increases in other IgG subclasses, IgM, or IgA. Sparse stromal deposits resembling immune complexes were found in 2 of 5 eosinophilic esophagitis biopsy specimens based on ultrastructural analysis. Esophagectomy samples from 2 patients with eosinophilic esophagitis contained 180 and 300 IgG4 plasma cells/maximal high-power field, mainly in the deep lamina propria; these levels were greater than in tissues from controls. Fibrosis essentially was exclusive to the lamina propria. Granular extracellular IgG4 was detected in biopsy specimens from 21 of 24 patients with eosinophilic esophagitis, butin none of the specimens from 9 controls (P= 6× 10(-6)). The total serum level of IgG4 increased only slightly in patients with eosinophilic esophagitis, compared with controls. Subjects witheosinophilic esophagitis had increased serum levels of IgG4 that reacted with milk, wheat, egg, and nuts-the 4 foods thatmost commonly trigger this condition (P ≤ 3× 10(-4) for each food). In a prospective trial, omalizumab did not reduce symptoms of eosinophilic esophagitis or tissue eosinophil counts compared with placebo. This finding, along with observed granular deposits of IgG4, abundant IgG4-containing plasma cells, and serum levels of IgG4 reactive to specific foods, indicate that, in adults, eosinophilic esophagitis is IgG4-associated, and not an IgE-induced allergy. ClinicalTrials.gov number: NCT00123630.

Perioperative management of an IgA-deficient recipient of a double-lung transplant

When exposed in the perioperative period to blood components containing immunoglobulin (Ig)A IgA-sensitized IgA-deficient patients are at an increased risk of transfusion-associated anaphylaxis. We present the case of an IgA-deficient patient whose candidacy for double-lung transplantation was under review in the preoperative period. A 49-yr-old patient with end-stage chronic obstructive lung disease secondary to deficiencies in IgA and IgG subclasses was being assessed for double-lung transplantation. Early recognition of the ramifications of perioperative transfusion prompted consultation with the transfusion medicine service. This in turn facilitated specialized laboratory testing and the coordinated provision of appropriate blood products for the unpredictable date of transplantation. The theoretical systemic risks of a non-IgA-deficient graft on the sensitized IgA-deficient host were considered. To affirm the patient's candidacy for transplantation, he was ultimately challenged preoperatively with IgA-containing products in a controlled intensive-care setting. Through a multidisciplinary approach [corrected], a successful transplantation outcome was achieved in an IgA-deficient patient undergoing major surgery. Strategies to mitigate risk include the procurement and transfusion of IgA-deficient components, which may be challenging or untenable in emergent perioperative settings.

Human CD19 and CD40L deficiencies impair antibody selection and differentially affect somatic hypermutation

Individuals with genetic defects in CD40 ligand (CD40L) or B-cell antigen receptor coreceptor molecules CD19 and CD81 suffer from an antibody deficiency. Still, these patients carry low levels of memory B cells and serum antibodies. We sought to assess why the remaining memory B cells and antibodies in the blood of these patients do not provide functional immunity. We included CD19-deficient patients (n = 8), CD40L-deficient patients (n = 8), and healthy controls (n = 50) to perform detailed flow cytometry on blood B cells, molecular analysis of IgA and IgG transcripts, as well as functional analysis of B-cell activation. CD19-deficient and CD40L-deficient patients carried reduced numbers of all memory B-cell subsets except CD27(-)IgA(+) B cells. Their immunoglobulin heavy chain class-switched transcripts contained less somatic mutations and reduced usage of IgM-distal IgG2 and IgA2 subclasses. The selection strength of mutations for antigen binding was significantly lower than in controls, whereas selection to maintain superantigen binding was normal. Furthermore, the patients showed impaired selection against inherently autoreactive properties of their immunoglobulins. Somatic hypermutation analysis revealed decreased activation-induced cytidine deaminase and uracil-DNA glycosylase 2 activity in CD40L deficiency and increased uracil-DNA glycosylase 2 but decreased mismatch repair in CD19 deficiency. B-cell activation studies revealed that this was at least in part due to transcriptional regulation of DNA repair genes. This study on CD19 and CD40L deficiencies illustrates that both the B-cell antigen receptor and CD40 signaling pathways are required for the selection of immunoglobulin reactivity. Still, they differentially mediate DNA repair pathways during somatic hypermutation, thereby together shaping the human in vivo antigen-experienced B-cell repertoire.

The EPIC Study: A Clinical Trial To Assess Whether Early Low Dose Prophylaxis In The Absence Of Immunological Danger Signals Reduces Inhibitor Incidence In Previously Untreated Patients (PUPs) With Hemophilia A

▪ BackgroundInhibitor development is a complex, multifactorial immune response involving both patient-specific and treatment-related factors. Of the known risk factors, intensive treatment at an early age has been shown to be significant, and clinical observations have suggested that early prophylaxis (i.e. first exposure to FVIII in the absence of a bleed in the first year of age) may protect patients from inhibitor development by inducing FVIII tolerance. AimThis study aimed to assess prospectively if a once-weekly, low-dose prophylactic regimen started before 1 year of age and before the onset of a severe bleeding phenotype (i.e. joint bleed), together with the minimization of immunological danger signals, could reduce the incidence of inhibitor formation in PUPs with severe and moderately severe hemophilia A to 15% or less. MethodsThe EPIC study was a Phase 3b, prospective, single arm, historically-controlled, international multicenter study to assess the incidence rate of inhibitor formation in PUPs with severe and moderately severe hemophilia A during the first 50 exposure days (EDs) of treatment with ADVATE starting with a once-weekly, low-dose (ADVATE 25 IU/kg once weekly), prophylactic regimen. If clinically indicated, it was permissible to increase the frequency of dosing to 2 or 3 times per week. In addition, infusions during the first 20 EDs had to be given 3 to 4 days before or after any vaccinations, which had to be given subcutaneously, not intramuscularly; infusions had to be avoided if the subject had high fever (above 38°C [100°F]). Main enrolment criteria were: severe and moderately severe hemophilia A (FVIII ≤2%), age <1 year, ≤3 EDs to any FVIII-containing product used for treatment of minor bleeds or for precautionary infusions following injury, adequate venous access (without need for central venous access device), no life-threatening conditions (intracranial hemorrhage, severe trauma) or requirement for surgery at the time of enrollment, no evidence of inhibitor ≥0.6 BU in Nijmegen-modified Bethesda assay at study start, no hemostatic defect other than hemophilia A, no clinically significant chronic disease other than hemophilia A,. Information about type of FVIII gene defect was obtained in 17 subjects. FVIII inhibitor tests were performed at screening, at study infusion #3, 6, 10, 15, 20, 30, 40, and 50, and at any other time point if an inhibitor was suspected. Positive inhibitor testing had to be confirmed by 2 positive inhibitor tests on samples drawn at least 1 week apart. ResultsA total of 22 subjects were enrolled in the study. Of 20 subjects who met all entry criteria, 19 received treatment; of these, all had severe hemophilia A (FVIII<1%). At study entry 11 of these 19 patients were never exposed to FVIII before (PUPs), while the remaining 8 patients had been treated with FVIII concentrates before. FVIII gene mutation analysis revealed intron 22 inversions in 8 out of 17 subjects, hemizygous missense mutations resulting in a stop-codon in 2 subjects, frame-shift mutations in 2 subjects, and hemizygous missense mutations in 5 subjects. A total of 8 subjects developed a confirmed inhibitor: 2 of these 8 subjects had only borderline positivity at inhibitor testing (never above 0.6 BU) with absence of any anti-FVIII antibodies (IgG, IgA, IgM and IgG subclasses) as tested by ELISA. Thus incidence of inhibitors >0.6 BU in PUPs were 27%. A total of 67 major protocol deviations (PD) were reported in 15 patients: 44 PDs of these were reported in 10 subjects and were related to the treatment regimen and therefore have contrasted with the protocol intention, which was to minimize immunological danger signals and low dose prophylactic regimen. As a result of the observed inhibitor incidence the study was terminated based on futility analysis, i.e. the probability to achieve the primary end-point of inhibitor rate reduction to ≤15%. Details on inhibitor patients will be presented. DiscussionThe EPIC study showed no safety issue as confirmed by the Data Safety Monitoring Board. To align treatment decisions in the presence of danger signals (which are not completely avoidable in children around 1 year of age) with a demanding study protocol was found to be challenging. Thus the hypothesis that an early low dose prophylaxis in the absence of immunological danger signals might reduce inhibitor incidence in PUPs with hemophilia A could neither be verified nor disproved within this study. Disclosures:Auerswald:Novo Nordisk: Consultancy, Research Funding; Biotest: Consultancy, Research Funding; CSL-Behring: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; Baxter: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding. Kurnik:Baxter: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; Biotest: Consultancy, Research Funding; CSL-Behring: Consultancy, Research Funding; Novo Nordisk: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding. Blatny:Baxter: speaker fee Other. Reininger:Baxter Innovations GmbH: Employment.

Increased serum concentration of BAFF/APRIL and IgA2 subclass in patients with mixed connective tissue disease complicated by interstitial lung disease

B cell activating factor (BAFF) and a proliferation-inducing ligand (APRIL) are known to be crucial for B cell maturation and survival, and increased expression of these factors in various autoimmune diseases has been reported. Human B cells produce two IgA subclasses: IgA1 and IgA2, the latter being abundant in the distal intestine, saliva, colostrum and bronchial fluid. We investigated these parameters in patients with mixed connective tissue disease (MCTD) complicated by interstitial lung disease (ILD+), and compared them with those in MCTD patients without ILD (ILD−). Sixty-three MCTD patients were divided into two groups: 21 ILD+ patients and 42 ILD− patients. In each patient group we analyzed soluble BAFF/APRIL using ELISA, and IgA1 and IgA2 using double immunodiffusion. Furthermore, we analyzed BAFF–APRIL receptors, BCMA, BAFF-R and TACI, using flow cytometry. The ILD+ patients had significantly higher levels of BAFF/APRIL than the ILD− patients. There were significant correlations between BAFF/APRIL, BAFF/KL-6 and APRIL/KL-6. Although there was no significant inter-group difference in the serum IgA1 level, ILD+ patients had a significantly elevated IgA2 level in comparison with ILD− patients. Moreover, although there were no significant inter-group differences in the expression of BCMA, BAFF-R and TACI on B cells, the expression of BAFF-R was significantly decreased in the ILD+ patients. In recent years, relationships between BAFF/APRIL and IgA subclass have been reported. Our results suggest that an elevated level of BAFF/APRIL drives the maturation of B cells, subsequently leading to IgA2 class switching, and possibly to the development of ILD in patients with MCTD.

Donor-specific HLA antibodies: evaluating the risk for graft loss in renal transplant recipients with isotype switch from complement fixing IgG1/IgG3 to noncomplement fixing IgG2/IgG4 anti-HLA alloantibodies

Human leukocyte antigen alloantibodies have a multitude of damaging effects on the allograft, both complement (C') activation and Fc-independent ones. To date, the clinical significance of non-C' fixing (NCF) HLA donor-specific antibodies (DSA) is still unclear. In this study, we investigated whether renal transplant recipients with NCF-DSA subclasses (IgG2/IgG4, IgA1/IgA2) are at higher risk of graft loss compared to patients with exclusively C' fixing (IgG1/IgG3). Blood samples from 274 patients were analyzed for HLA IgG and IgA subclasses using a modified single-antigen bead assay. We identified 50 (18.2%) patients with circulating NCF antibodies either DSA (n=17) or against third-party HLA (n=33). NCF-DSAs were preferentially of IgG2/IgG4 isotype (11/17) and were mainly directed against HLA class II (13/17). NCF DSA were present as a mixture with strong C' fixing IgG1/IgG3. Graft survival was similar between patients with exclusively C' fixing antibodies and those with a mixture panel (log rang test P=0.162), and also among patients with different immunoglobulin isotype and subclasses (long-rank test, P=0.732). We conclude that expansion of DSA to NCF subclasses postrenal transplantation does not seem to be associated with worse graft survival as compared to the presence of exclusive C' fixing subclasses.

Serum immunoglobulins in the infected and convalescent phases in community-acquired pneumonia

A population-based case-control study was designed to assess changes of serum levels of immunoglobulins and IgG subclasses between infected and convalescent phase in community-acquired pneumonia (CAP). Over a 2-year period, all subjects who were >14 years of age living in the Maresme region (Barcelona, Spain) diagnosed of CAP were registered. Controls were healthy subjects selected from the municipal census. Prognostic factors were assessed and serum levels of total IgG, IgA, IgM, and IgG subclasses were measured at diagnosis and 1 month later (cases). We studied 171 patients with CAP and 90 controls. All immunoglobulins were significantly lower in cases than in controls. At diagnosis, 42.7% of cases showed low levels of some immunologic parameter, mainly total IgG and IgG2. Low immunoglobulin levels at diagnosis were more frequent in patients requiring in-patient care and in those with pneumonia of other etiology than Streptococcus pneumoniae. In the convalescent phase, 26 (23.6%) patients normalized immunological levels. In 27 (24.5%) cases, some parameter with low levels persisted especially in patients with etiology of CAP other than S. pneumoniae. Low serum levels of immunoglobulins particularly total IgG and IgG2 were a common finding in patients with CAP compared to healthy controls. Low immunoglobulin levels may be related to CAP prognosis and persisted in the convalescent phase in one-fourth of cases.

Management of secondary immune deficiencies

Management of secondary immune deficiencies

Myeloid cells as effector cells for monoclonal antibody therapy of cancer

Monoclonal antibodies (mAbs) have become an important addition to chemo- and/or radiotherapy for the treatment of cancer. They have multiple effector functions that can lead to eradication of tumor, including induction of apoptosis, growth inhibition, and initiation of complement-dependent lysis. Furthermore, mAbs can recruit immune effector cells. Traditionally, natural killer cells have been considered as the main effector cell population in mAb-mediated tumor killing. Myeloid cells have potent cytotoxic ability, as well. Monocytes and macrophages have been shown to induce antibody-dependent cytotoxicity and phagocytosis of tumor cells in the presence of IgG anti-tumor mAb. Furthermore, neutrophils are the most abundant population of circulating white blood cells, and as such may constitute a formidable source of effector cells. However, when targeting neutrophils for tumor therapy, antibodies of the IgA subclass may be more effective. This article focuses on enlisting myeloid effector cells for mAb-based immunotherapy of cancer. Additionally, methods to study mAb-dependent phagocytosis of tumor cells by macrophages are compared.

Immunoglobulin values in healthy Thai children aged ≤ 24 months determined by nephelometry

Variation of normal immunoglobulin (Ig) levels between different genetic and environment factors has been studied. Although antibody deficiency diseases can start from infancy, data of Ig reference levels in children aged ≤24 months are still limited, especially in Asian children. The aim of this study was to determine serum IgG, IgA, IgM, and IgG subclasses in healthy Thai children from the newborn period to age 24 months. Serum samples were collected from healthy Thai children age <1-24 months to measured serum IgG, IgA, IgM, and IgG subclasses by nephelometry. Of the 100 infants, 44% were female with a median (range) age of 13 (0.3-24) months. The geometric mean IgG was 803 mg/dL, IgA 36 mg/dL, and IgM 102 mg/dL. The mean IgG1 was 646 mg/dL, IgG2 127 mg/dL, IgG3 45 mg/dL, and IgG4 17 mg/dL. The average ratios of IgG subclass 1:2:3:4 were 77:15:6:2%. No significant differences in each immunoglobulin isotype between genders were found. Our mean IgG level was slightly lower than that in healthy Thai children, measured by radial diffusion method but not significant except 1-3 months (p = 0.016). However, the mean IgG level in our study was higher than that reported by radial diffusion in healthy US children (p <0.001). This study illustrated the importance of having normal Ig values from age- and ethnically-matched controls by high precision nephelometric assay in order to appropriately diagnose immunologic disorders in Asian infants.

Memory B cell compartment constitution and susceptibility to recurrent lower respiratory tract infections in young children

A proportion of children have recurrent LRTIs, mostly as a result of Spn, which persist after 2 years of age. Here, we investigate, by flow cytofluorometry, the constitution of the memory B cell compartment in 90 healthy children and 49 children with recurrent LRTIs to determine if an increased susceptibility to recurrent LRTIs results from a delayed or abnormal ontogeny with poor antibody-mediated protection. Total IgA, IgM, IgG, and IgG subclasses were measured by nephelometry, as well as antipneumococcal antibodies by ELISA. Pneumococcal vaccination status was obtained. We show that the memory B cells increase between birth and 2 years of age (1.6% vs. 21.1%, P<0.001) without further significant increase noted per additional years (3-4 years old: 23.3%; 4-5 years old: 22.2%, P>0.40) to reach adult-like values (31.8±11.8%, P=0.08). Proportions of switched and IgM memory B cells were similar in children and adults. Comparatively, LRTI children had no delay in the constitution of their memory B cell compartment (2-3 years old: 26.9%; 3-4 years old: 18.2%; 4-5 years old: 26.8%, P>0.05). Their switched and IgM memory B cells were similar among age categories, and the distribution was overall similar to that of healthy controls. LRTI children had normal total and pneumococcal serotype-specific antibody values but showed a rapid waning of antipneumococcal antibody levels after vaccination. In summary, our results show that the memory B cell compartment is already similarly constituted at 2 years of age in healthy and LRTI children and thus, cannot explain the increased susceptibility to bacterial pneumonia. However, the waning of antibodies might predispose children to recurrent infections in the absence of revaccination.

Defective immunoglobulin A (IgA) glycosylation and IgA deposits in patients with IgA nephropathy

Defective glycosylation and immune complex (IC) formation may be of primary importance in immunoglobulin A nephropathy (IgAN) pathogenesis. The aim of this study was to determine whether defective IgA1 glycosylation might support renal deposition of IgA and disease activity. IgA was isolated from the serum of 44 IgAN patients and 46 controls and glycosylation analysed by ELISA using glycan-specific lectins. IgA was measured by immunodiffusion and immune complexes by ELISA. IgA subclasses in IC deposits in kidney glomeruli were identified by immunohistochemical methods. A significant increase in N-acetylgalactosamine (GalNAc) in terminal position (p = 0.02) observed in some of the IgAN patients, became more pronounced when sialic acid was removed from IgA1, indicating enhanced expression of α-2,6-sialyltransferase in patients compared with controls (p < 0.0001). Patients with defective galactosylation had lower serum IgA than other IgAN patients (p = 0.003). IgAN patients with both IgA1 and IgA2 glomerular deposits (21.7%) had increased GalNAc in terminal position (p = 0.003). Taken together, our results show that increased IgA glycosylation in IgAN associates with low levels of IgA, concomitant IgA1 and IgA2 glomerular deposits and poor clinical outcome.

Are serum levels of immunoglobulin classes and IgG subclasses involved in delayed pulmonary complications induced by sulfur mustard? Sardasht-Iran Cohort Study

Respiratory complications are the foremost long term debilitating effects after sulfur mustard toxicity. The underlying immunological mechanisms of sulfur mustard induced lung damage are still poorly understood. The question of the involvement of immunoglobulin classes and subclasses in delayed pulmonary complications induced by SM was addressed in this study as a part of Sardasht-Iran Cohort Study (SICS).In SICS, 372 male participants who were exposed to SM 20 years earlier were compared with 128 unexposed age-matched controls. At the time of study (2007), the clinical evaluations and spirometry was performed for all subjects according to the American Thoracic Society Criteria, and at the same time, the sera were isolated, labeled and aliquots were kept frozen in -80°C. Serum immunoglobulin (Ig) levels including IgM, IgA, IgE, IgG, and IgG subclasses (IgG1, IgG2, IgG3 and IgG4) were measured using quantitative Elisa method. It was found that among immunoglobulin classes and IgG subclasses only IgM and IgG4 were significantly decreased in the peripheral blood of exposed cases. IgM level also positively correlated with FEV1 only in the SM exposed group. These results indicated a weak but significant role for IgA in control of the delayed pulmonary complications. There were no strong correlations between other immunoglobulin classes or IgG subclasses with pulmonary disease severity in sulfur mustard intoxicated subjects. The authors proposed that systemic levels of immunoglobulins do not exert essential roles in severity of delayed pulmonary complications following SM toxicity. However, more studies on local and systemic levels of immunoglobulins in more severe groups are suggested.

Multiple antigen peptide consisting of B- and T-cell epitopes of F1 antigen of Y. pestis showed enhanced humoral and mucosal immune response in different strains of mice

Yersinia pestis is a causative agent of plague. F1 and V antigen based vaccines have shown remarkable protection in experimental animals. In order to develop epitope based immunogen, three B and one T-cell epitopes of F1 antigen with palmitate residue at amino terminal were assembled on a lysine backbone as multiple antigen peptide (MAP or F1-MAP). MAP was characterized by SDS-PAGE, immunoblot and immunoreactivity with anti F1 sera. MAP was entrapped in PLGA (polylactide-co-glycolide) microparticles and humoral, mucosal immune responses were studied after intranasal immunization with/without CpG ODN 1826 (CpG)/murabutide in different strains of mice. Serum and mucosal washes were measured for MAP specific IgG, IgA, sIgA and IgG subclasses in three strains of mice. F1-MAP showed high serum antibody and mucosal IgG and IgA peak antibody titers. MAP with CpG showed significantly high (p<0.001) peak antibody titer ranging from 102,400 to 204,800 for IgG and 6400 to 12,800 for IgA. High mucosal sIgA and its secretary component detection confirmed generation of mucosal response in intestinal and lung washes. MAP antisera also showed significant immunoreactivity with individual peptides. Moreover, antibody specific activity (IgG, IgA and sIgA) positively correlates with peak antibody titers. Predominantly IgG2a/IgG2b subclass was observed with CpG formulation but in other formulation a mixed IgG1 and IgG2a response was observed. The present study highlights the importance of multiple antigen peptide approach of F1-antigen with CpG as an alternative approach for subunit vaccine.

A Shorter Time to First Treatment and Worst Overall survival Is Observed in Cll Patients with Reduced Serum IgM As Evaluated with Novel Serum Immunoassays

AbstractAbstract 4585 Background and Aims:Reduced serum IgM levels were proposed as an unfavorable prognostic factor in multiple myeloma1. We therefore investigated the possible impact of decreased serum IgM concentrations on the outcome of Chronic Lymphocytic Leukemia (CLL) patients, a disease that is frequently characterized by severe hypogammaglobulinemia. Serum IgM levels were measured by 2 methods. Patients and methods:188 patients with CLL were studied. Of them 54%, 27%,10%, 7% και 2% where in RAI stage 0, 1, 2, 3, 4 respectively while 65%, 22% and 13% in Binet stages A, B and C. IgM was determined at diagnosis by both classical nephelometry and the new Hevylite™ methods; the first measures total IgM levels while the second one measures the immunoglobulin fractions bound to either kappa or lambda light chains therefore enabling us to determine the ratio of IgM-kappa/lambda (HLCR). Hevylite™ measurements were retrospectively performed in 69/188 patients with available frozen sera aliquots from drawn at the time of diagnosis. Of them the Ig subclasses IgMkappa and IgMlambda, IgGkappa and IgGlambda, IgAkappa and IgAlambda (HLC) were measured by Hevylite™ antibodies nephelometrically. Light chain restriction (kappa or lambda) was determined by flow cytometry or bone marrow/lymph node biopsy immunohistochemistry. The ratios (HLCRs) were calculate with the monoclonal immunoglobulin as numerator, i.e IgMkappa/IgMlambda, IgGkappa/IgGlambda and IgAkappa,/IgAlambda in kappa-restricted patients and IgMlambda/IgMkappa, IgGlambda/IgGkappa, IgAlambda/IgAkappa in lambda-ones. Serum IgM concentration by both methods were related to time to first treatment (TFT) and overall survival (OS). Statistical analysis was performed conventionally; survival curves were drawn by Kaplan-Meyer method and compared by the log-rank test. Results:Total IgM ranged from 12 to 246 mg/dl (median 47.6). The sum of IgMkappa plus IgMlambda from 12 to 369 (median 60). IgMkappa from 7.9 to 349 mg/dL (median 41) in kappa-restricted patients and IgMlambda from 7.6 ωζ 98 mg/dL (median 22) in lambda restricted ones. Low total IgM level (<50 mg/dL) was present in 52% of patients but it did not correlate to TFT or OS. Summated IgMkappa plus IgMlambda lower than 50 mg/dL was present in 41% and correlated with worse OS (p=0.016). Furthermore non-clonal IgM subclass suppression correlated with TFT (p=0.019) and OS (p=0.021). The same was not confirmed for non-clonal IgG and IgA subclasses suppression.Finally IgM-HLCR was abnormal in 35% of patients. Of them, 25% had also abnormal IgG-HLCR and/or IgA-HLCRs leading to the hypothesis of multiple clones coexistence within the neoplastic B-cell lymphocytic population. Conclusions:Low IgM levels determined by novel immunoassays (Hevylite™) correlate to shorter TFT and OS in patients with CLL. Their use may reveal new disease characteristic and are suggestive of a multi-oligoclonal disorder. Disclosures:No relevant conflicts of interest to declare.

The Effect of Red Ginseng Extract on Inflammatory Cytokines after Chemotherapy in Children

Ginseng has been used as an herbal medicine, widely used in Asian countries, for long time. Recently, beneficial effects for immune functions of Korean red ginseng (KRG) have been reported in adults. This study was performed to investigate the effects of ginseng on immune functions in children after cessation of chemotherapy or stem cell transplantation for advanced cancer. Thirty patients, who were diagnosed and treated for leukemia and solid cancer at the department of pediatrics and adolescence of the Yeungnam University Hospital from June 2004 to June 2009, were enrolled for the study. The study group consisted of 19 patients who received KRG extract (60 mg/kg/d) for 1 yr and 11 patients who did not receive KRG extract were the control group. Blood samples were collected every 6 mo. Immune assays included circulating lymphocyte subpopulation, serum cytokines (IL- 2, IL-10, IL-12, TNF-alpha, and IFN-gamma), and total concentrations of serum IgG, IgA, and IgM subclasses. Age at diagnosis ranged from 2 mo to 15 yr (median 5 yr). Nine patients received stem cell transplantation. The cytokines of the KRG treated group were decreasing more rapidly than that of the control group. Lymphocyte subpopulations (T cell, B cell, NK cell, T4, T8, and T4/ T8 ratio) and serum immunoglobulin subclasses (IgG, IgA, and IgM) did not show significant differences between the study and the control groups. This study suggests that KRG extract might have a stabilizing effect on the inflammatory cytokines in children with cancer after chemotherapy.

479 A 3 Year-old Child with Specific Antibody Deficiency and Allergic Rhinitis. A Case Report

BackgroundSpecific antibody deficiency (SAD) is a humoral immunodeficiency characterized by normal levels of IgG, IgA, IgM and IgG subclasses but a failure to polysaccharide antigens, manifested with recurrent bacterial respiratory infections. To establish the SAD diagnosis an inadequate IgG antibody response to more than 50% of pneumococcal serotypes after unconjugated pneumococcal immunization are needed. An adequate response is defined as a post-immunization titre of ≥1.3 μg/mL or ≥4 times the preimmunization value.1,2MethodsThe record of 1 patient was review and relevant clinical data was collected. A review of the literature about SAD was made.ResultsA 4-year old male with family history of atopic disease, esophageal reflux at 3-months, he began with recurrent upper respiratory tract infections at 1-year old, 1 to 2 events per month, fever (39–40° C), persistent cough and hyaline rhinorrhea, nasal itching and sneezing he was treated with multiple antibiotics, inhaled and oral corticosteroids with mild clinical recovery between episodes. A normal blood cell count and normal levels of IgG 1219 mg/dL, IgA 146 mg/dL, IgM 98 mg/dL and IgG subclasses were determined. Allergic rhinitis and asthma were diagnosed at 3-years old, percutaneous prick skin test was positive to Dermatophagoides farinae, Salsola pestifer, Phleum pratense, Heliantus sp. and specific immunotherapy was started. Despite of treatment he continued with recurrent infections so specific antibody response to polysaccharide pneumococcal antigens was evaluated, he responded less than 50% to 14 pneumococcal serotypes after 23-valent unconjugated pneumococcal vaccine, so SAD was diagnosed and treated with prophylactic antibiotic, pneumococcal polysaccharide conjugated vaccine (10-valent) and specific immunotherapy. He showed clinical improvement, with few mild infections, and controlled rhinitis and asthma.ConclusionsThere are several Primary Immunodeficiency Diseases related to allergic diseases as IgA deficiency and SAD. In the atopic patient that does not improve in spite of specific immunotherapy further investigations are needed to exclude them.

Vitamin D3 supplementation in patients with frequent respiratory tract infections: a randomised and double-blind intervention study

BackgroundLow serum levels of 25-hydroxyvitamin D3 are associated with an increased risk of respiratory tract infections (RTIs). Clinical trials with vitamin D3 against various infections have been carried out but...