Distribution Of IgA Subclasses Research Articles

Expression of Homing Receptors on IgA1 and IgA2 Plasmablasts in Blood Reflects Differential Distribution of IgA1 and IgA2 in Various Body Fluids

Although secretory IgA is the most abundantly produced Ig isotype, the mechanisms underlying the differential distribution of IgA subclasses in various body fluids remain unclear. To explore these mechanisms, we examined the distribution of IgA subclasses, the influence of the nature and sites of encounters with antigens, and the correlation between IgA subclass distribution and homing potentials of circulating IgA plasmablasts. IgA1 predominated in serum, tears, nasal wash fluid, and saliva; the levels of IgA1 and IgA2 were comparable in vaginal wash fluid; and IgA2 predominated in intestinal lavage fluids. Seventy-one percent of circulating IgA plasmablasts secreted IgA1. The intestinal homing receptor (HR), alpha4beta7, was expressed more frequently on IgA2 than on IgA1 plasmablasts, with no differences in the expression of other HRs. IgA subclass distribution among circulating antigen-specific antibody-secreting cells (ASC) was dependent on the nature of the antigen: following vaccination with Salmonella enterica serovar Typhi, unconjugated pneumococcal polysaccharide, or Haemophilus influenzae polysaccharide-diphtheria toxoid conjugate, the proportions of specific IgA1 ASC were 74%, 47%, 56%, and 80%, respectively. HR expression depended on the route of administration: expression of HRs was different after oral than after parenteral vaccination, while no difference was seen between HR expression of antigen-specific IgA1 and IgA2 ASC induced via the same route. The key factors determining IgA subclass distribution in a given secretion are the nature of the antigens encountered at a particular site and the site-specific homing instructions given to lymphocytes at that site. These two factors are reflected as differences in the homing profiles of the total populations of circulating IgA1 and IgA2 plasmablasts.

Human Male Genital Tract Secretions: Both Mucosal and Systemic Immune Compartments Contribute to the Humoral Immunity

In contrast to numerous studies of female genital tract secretions, the molecular properties of Abs and the magnitude of humoral responses in human male genital tract secretions to naturally occurring Ags and to mucosal and systemic immunizations have not been extensively investigated. Therefore, seminal plasma (SP) collected from healthy individuals was analyzed with respect to Ig levels, their isotypes, molecular forms of IgA, and for the presence of Abs to naturally occurring Ags, or induced by systemic or mucosal immunizations with viral and bacterial vaccines. The results indicated that in SP, IgG and not IgA, is the dominant Ig isotype, and that IgM is present at low levels. IgA is represented by secretory IgA, polymeric IgA, and monomeric IgA. In contrast to the female genital tract secretions in which IgA2 occurs in slight excess, the distribution of IgA subclasses in SP resembles that in plasma with a pronounced preponderance of IgA1. The IgG subclass profiles in SP are also similar to those in serum. Thus, SP is an external secretion that shares common features with both typical external secretions and plasma. Specifically, SP contains naturally occurring secretory IgA Abs to environmental Ags of microbial origin and to an orally administered bacterial vaccine, and plasma-derived IgG Abs to systemically injected vaccines. Therefore, both mucosal and systemic immunization with various types of Ags can induce humoral responses in SP. These findings should be considered in immunization strategies to induce humoral responses against sexually transmitted infections, including HIV-1.

IgG and IgA Subclass Distribution of Total Immunoglobulin and Rheumatoid Factors in Rheumatoid Tissue Plasma Cells

The subclass distribution of IgG and IgA plasma cells, and in IgG and IgA rheumatoid factor (RF) producing cells was studied in sections of synovial tissue from seropositive RA and various types of seronegative arthritis, including ankylosing spondylitis, psoriatic arthritis, and Reiter's syndrome. The study was performed with immunofluorescence technique and monoclonal IgG and IgA subclass specific antibodies. IgG RF producing cells were identified by their ability to bind and activate factors both in the early (C3) and late (C5b-9) part of the complement cascade. IgA RF cells were identified by double staining experiments with heat-aggregated IgG and monoclonal antibodies to IgA subclasses. In 23 tissues tested for total IgG, IgG1 cells were usually predominant, while the frequency of IgG3 cells was usually higher than that of IgG2. In 19 tissues also tested for IgA, both IgA subclasses were present in all tissues. IgA1 plasma cells were always predominant, with a mean ratio of IgA1 to IgA2 cells of approximately 10. In the 13 tissues tested for RF-producing cells, the highest frequency of IgG RF cells was found among the IgG3 cells, followed by IgG1 and IgG2. IgA RF cells were found in only one case, all cells being IgA1.

Analysis of human IgG and IgA subclass antibody-secreting cells from localized chronic inflammatory tissue.

The chronic inflammatory diseases in humans have been intensively investigated, however the immune mechanisms underlying diseases such as rheumatoid arthritis (RA), inflammatory bowel disease, and periodontal disease (PD) remain elusive. In this study, we have analyzed the distribution of IgM, IgG, and IgA secreting cells with emphasis on the IgG and IgA subclasses among mononuclear cell populations isolated from gingiva at different stages of PD. Surgically removed tissues were treated with Dispase to gently dissociate cells and the Ficoll-Hypaque gradient centrifugation was used to enrich for viable mononuclear cells rich in lymphocytes, macrophages, and plasma cells. The total numbers of plasma cells increased with the severity of disease. Immunofluorescence analysis showed that most Ig-containing cells were of the IgG isotype; however, significant numbers of IgA-positive cells but few IgM-positive cells were seen. This isolation procedure allowed analysis, at the single cell level, of the distribution of IgG and IgA subclasses of antibody-secreting cells with monoclonal antibodies to human IgG and IgA subclasses. For this, we selected four monoclonal anti-IgG subclass (anti-gamma 1, -gamma 2, -gamma 3, and -gamma 4) antibodies with no subclass cross reactivity for use in the enzyme-linked immunospot assay. Analysis of slight, moderate, and advanced stages of PD showed a progressive increase in spotforming cells (SFC) numbers, and the major isotype of SFC was IgG followed by IgA. The major IgG subclass SFC seen was IgG1 followed by IgG2 whereas similar numbers of IgG3 and IgG4 SFC were observed, a pattern also seen with cells from synovium of RA patients and in mitogen-triggered spleen and PBMC. In terms of the IgA subclass distribution, IgA1 predominated in moderate stages, whereas a selective increase in IgA2 SFC were seen in the more advanced stage of PD. These results show that significant numbers of viable plasma cells/Ig-secreting cells can be isolated from inflamed gingival tissues. Further, careful analysis has shown that IgG subclass responses in gingiva are similar to those found in synovia of RA subjects, and in stimulated PBMC and spleen. However, it should be noted that the number of IgG4- and IgA2-secreting cells increased in the advanced stage of PD.

J-chain expression is more prominent in immunoglobulin A2 than in immunoglobulin A1 colonic immunocytes and is decreased in both subclasses associated with inflammatory bowel disease

Paired immunofluorescence staining demonstrated reduced J-chain positivity of both immunoglobulin A1 (IgA1)- and IgA2-producing cells in colonic mucosa from patients with ulcerative colitis and Crohn's colitis compared with controls (p < 0.002). J-chain expression was generally higher in IgA2 than in IgA1 immunocytes. The median proportion in normal mucosa was 100% for IgA2 vs. 88% for IgA1 (p < 0.005); in ulcerative colitis, 69% vs. 46% (p < 0.004); and in Crohn's colitis, 74% vs. 46% (p < 0.004). Taken together with the overall IgA-subclass distribution, however, these results showed that the proportion of J-chain-positive IgA2 cells in the total IgA-cell population was lower for ulcerative colitis (20%) and Crohn's colitis (32%) than for normal mucosa (63%) (p < 0.002). In relation to the total J-chain-positive IgA-cell population, which contributes to the secretory IgA system, an increased proportion (p < 0.002) belonged to IgA1 in ulcerative colitis (61% vs. normal, 27%), whereas IgA2 was reduced (39% vs. normal, 73%). Similar but smaller trends were noted in Crohn's colitis. The disease-associated reduction of J chain might be compensated by the previously reported twofold numeric increase of IgA cells in colitis. Our study, therefore, did not suggest that the secretory IgA-cell system was quantitatively impaired in inflammatory bowel disease.

IgG and IgA Subclass Distribution of Antitoxin Antibody Responses after Oral Cholera Vaccination or Cholera Disease

The immunoglobulin subclass distribution of cholera antitoxin antibody responses in serum was studied in Swedish volunteers after different routes of immunization with a cholera B subunit-whole cell vaccine (B + WCV) and in Bangladeshi patients convalescing from cholera disease. Both oral and parenteral immunization induced antitoxin antibodies of all the different IgG subclasses (IgG1, IgG2, IgG3 and IgG4) whilst the IgA antibodies were restricted to the IgA1 subclass. A single oral dose of B + WCV induced proportionally higher levels of IgG4 antitoxin in previously cholera-immunized volunteers than in a matched group who had not been cholera-vaccinated before, suggesting that repeated immunization preferentially stimulate formation of IgG4 antibodies. The IgG and IgA subclass distribution of antitoxin antibodies in orally vaccinated Swedes closely resembled that in Bangladeshi cholera patients.

Subclass restriction pattern of antigen-specific antibodies in donors with defective expression of IgG or IgA subclass heavy chain constant region genes

We have developed a method for the measurement of the IgG and IgA subclass distribution of antigen-specific human antibodies. The controls for the specificity of the assay include the use of a number of monoclonal human antibodies and sera from individuals with deletions of particular immunoglobulin heavy chain constant region genes. The system was used to determine the shift in immunoglobulin subclass patterns of specific antibodies against a variety of protein and polysaccharide antigens in individuals with a regulatory deficiency of a given IgG or IgA subclass. Normally, the pattern is quite distinct and antibodies against protein antigens are mainly of the IgG 1 subclass, whereas antibodies against polysaccharide antigens are mainly of the IgG 2 subclass. The results on serum from an IgG 1 deficient donor suggested that IgG 3 and IgG 4 appear to compensate for a lack of IgG 1, whereas isolated deficiencies of IgG 3, IgG 4, or IgA 2 do not markedly influence the expected distribution of specific antibodies. In IgG 2-deficient individuals a more complex pattern was observed where antibodies against protein antigens were retained, whereas levels of antibodies against polysaccharide antigens could vary markedly between donors, which appeared to be dependent on whether the IgG 2 deficiency was an isolated defect or combined with IgG 4 IgA deficiency. However, all the IgG 2-deficient donors had a skewed pattern of anti-polysaccharide antibodies with a shift to IgG 1 to IgG 3.

Immunoglobulin subclass distribution of specific antibodies in allergic patients. Prediction of the IgE inducing capacity of potential allergens.

The IgG and IgA subclass distribution of specific antibodies against a variety of protein and polysaccharide antigens was determined in sera from individuals with high levels of IgE. No shift of the antibody pattern could be observed, suggesting that the aberrant regulation of responses against allergens noted in these patients is limited, encompassing selected antigens only. Antibodies against protein antigens are mainly of the IgG1 subclass. In addition, low levels of specific IgG3 or IgG4 antibodies may be formed. Our data suggest that a given antigen induces either IgG3 or IgG4 and that potential allergens, in addition to IgG1 and IgE, elicit a response restricted to IgG4.

Molecular forms of tear IgA and distribution of IgA subclasses in human lacrimal glands

Tears from subjects without conjunctival inflammation contained 93% polymeric IgA (pIgA) and of secretory IgA standard for tear samples and monomeric IgA standard for serum samples. Tears from subjects without conjunctival inflammation contained 93% polymeric IgA (pIgA) and 7% monomeric IgA; tears from subjects with conjunctival inflammation had less pIgA (79%). Biopsy specimens of 11 lacrimal glands and one accessory lacrimal gland were obtained from 12 additional subjects. Tissues were stained with polyclonal antisera for immunoglobulins, J chain, and secretory component. In addition, tissues were stained for IgA subclasses with monoclonal reagents specific for IgA1 and IgA2. IgA plasma cells predominated over other types of plasma cells, and most were J chain positive. Secretory component, although absent from the interstitium of the gland, was found in most acinar and ductal cells. The average proportion of IgA1 to IgA2 cells was 56% to 44%. We concluded that the lacrimal system of the human resembles other exocrine systems in the production of predominantly pIgA and the nearly equal occurrence of IgA1 and IgA2 plasma cells.

Significance of elevated IgA antibody levels in neonates withgastrointestinal conditions

Four groups of patients, 21 neonates with gastrointestinal anomalies, 8 with necrotizing enterocolitis, 5 with low anorectal anomalies and 15 with spina bifida were studied for the development of IgA and IgA subclass immunoglobulin levels. A highly significant increase in IgA levels (p less than 0.001) was seen in neonates with gastrointestinal anomalies and necrotizing enterocolitis while patients with low anorectal anomalies and spina bifida had normal or undetectable IgA levels. Estimation of IgA sub-class distribution in neonates with raised total serum IgA did not demonstrate any striking correlation between either IgA1 or IgA2 sub-classes and the clinical conditions but rather they reflected the level of total IgA. The levels if IgA, correlated closely with those of total IgA (r = 0.93) in all patients implying that in these patients total serum IgA is composed principally of IgA1 immunoglobulin.

Distribution of IgA subclasses in sera and bone marrow plasma cells of 21 normal individuals.

I would like to report some experiments which Dr. Morell and I have recently performed in Berne, Switzerland. We compared the distribution of the two IgA subclasses in the sera and in the IgA containing cells in the bone marrow of normal individuals. We are indebted to Dr. Hijmans for his help in the immunofluorescence studies of the bone marrow.