The localization of IgA and IgE globulins in the palatinal tonsils,nasal mucous membrane and nasal polypi

Using the immunofluorescence method, the localization of IgA and IgE globulins was studied in tonsillar tissue, nasal and pharyngeal mucous membranes and nasal polypi of patients hospitalised at the Clinic of Otorhinolaryngology. No difference in either localization and number of cells containing blood and/or secretory IgA globulin could be found in individual patients. Great amounts of IgE globulin were found in nasal mucous membrane and nasal polypi in subepithelial plasma cells even in patients without demonstrable allergy.     

Human peripheral blood mononuclear cells produce IgA anti-influenza virus antibody in a secondary in vitro antibody response

The function and immunoregulation of human IgA memory B cells producing anti-influenza virus antibody was analyzed in vitro in antigen-stimulated cultures. Peripheral blood mononuclear cells (PBMC) from seven of eight normal adult volunteers naturally immunized to influenza virus produced IgA anti-influenza virus antibody when stimulated in vitro with inactivated A/Aichi/68 [H3N2] influenza virus. This IgA antibody response was approximately one-eighth the IgG antibody response. PBMC from each of five patients with selective IgA deficiency failed to produce any measurable IgA antibody. When tonsillar mononuclear cells (TMC) were studied in a similar manner, a relatively higher IgA antibody response was obtained (one-third the IgG antibody) than with PBMC. Additional studies were undertaken to investigate the immunoregulation of this IgA antibody production and the relatively lower amount produced by PBMC than by TMC. Co-cultures of peripheral blood B cells with irradiated peripheral blood T cells (to possibly inactivate a radiosensitive IgA suppressor cell) did not result in a relative increase in IgA antibody production. Also, co-cultures of B cells with increasing numbers of T cells produced parallel increases of IgG and IgA antibody when plotted on a log scale with slopes of approximately 1, suggesting that a single helper T cell was limiting for both isotypes. Finally, pokeweed mitogen-stimulated co-cultures of peripheral blood and tonsillar B and T cells revealed that the B cell population, but not the T cell population, determined the amount of IgA anti-influenza virus antibody produced. Precursor frequency analyses of tonsillar and peripheral blood B cells in antigen-stimulated cultures confirmed that tonsils contained a higher precursor frequency of B cells for IgA anti-influenza virus antibody production (3.95/10(6) B cells) than did peripheral blood B cells (0.65/10(6) B cells). Thus, IgA memory cells are preferentially found in tonsillar tissue as compared with the peripheral blood, consistent with the role of the tonsils as a mucosal immune organ.     

A novel human IgA monoclonal antibody protects against tuberculosis

Abs have been shown to be protective in passive immunotherapy of tuberculous infection using mouse experimental models. In this study, we report on the properties of a novel human IgA1, constructed using a single-chain variable fragment clone (2E9), selected from an Ab phage library. The purified Ab monomer revealed high binding affinities for the mycobacterial α-crystallin Ag and for the human FcαRI (CD89) IgA receptor. Intranasal inoculations with 2E9IgA1 and recombinant mouse IFN-γ significantly inhibited pulmonary H37Rv infection in mice transgenic for human CD89 but not in CD89-negative littermate controls, suggesting that binding to CD89 was necessary for the IgA-imparted passive protection. 2E9IgA1 added to human whole-blood or monocyte cultures inhibited luciferase-tagged H37Rv infection although not for all tested blood donors. Inhibition by 2E9IgA1 was synergistic with human rIFN-γ in cultures of purified human monocytes but not in whole-blood cultures. The demonstration of the mandatory role of FcαRI (CD89) for human IgA-mediated protection is important for understanding of the mechanisms involved and also for translation of this approach toward development of passive immunotherapy of tuberculosis.     

Cloning the arterial IgA antibody response during acute Kawasaki disease

Kawasaki disease (KD) is the most common acquired cardiac disease in children in developed nations. The etiology of KD is unknown but likely to be a ubiquitous microbial agent. Previously, we showed that oligoclonal IgA plasma cells infiltrate coronary arteries and other inflamed tissues in acute KD. We demonstrated that a synthetic Ab made using an alpha H chain sequence prevalent in acute KD arterial tissue detected Ag in acute KD coronary arteries, lung, and other inflamed tissues and that Ag localized to cytoplasmic inclusion bodies in the acute KD ciliated bronchial epithelium. In this study, we synthesized a panel of mAbs from alpha and kappa chain sequences present in the KD arterial wall and tested the Abs for binding to acute KD tissues. We report that all of the synthetic mAbs that bind to acute KD tissues detect Ag in cytoplasmic inclusion bodies in the acute KD ciliated bronchial epithelium. Abs made from alpha sequences that were prevalent in KD arterial tissue show stronger binding to acute KD tissues than Abs made from less prevalent sequences. These findings highlight the likely importance of the inclusion bodies in the etiopathogenesis of acute KD, confirm that the IgA Ab response in acute KD is Ag driven, and demonstrate the usefulness of cloning the Ab response in diseased tissues to identify disease-relevant Ags.     

Naturally occurring plasmodium-specific IgA antibody in humans from a malaria endemic area

Blood samples collected from individuals belonging to an endemic area in Uttar Pradesh, were tested for plasmodial antigen specific immunoglobulin A (IgA) by enzyme immuno assay using soluble extract ofPlasmodium falciparum from culture. Among 773 (20.18%,P < 0.0001) samples 156 sera demonstrated a detectable seropositivity for antigen specific IgA. IgA levels were higher among individuals who experienced repeated attacks of malaria compared to acute infected patients. Among seropositive individuals the IgA titers were found increased with the age. Immunoglobulin isolated from sera having high level of IgA showed growth inhibitory effect inPlasmodium falciparum in vitro. A group of sera with high IgA antibody againstPlasmodium falciparum crude antigen showed seronegativity with specific peptides. Statistically, no positive or negative correlations were observed between antigen specific IgG and IgA. However, there was a tendency towards negative correlation between IgA and IgM. Mechanisms for the parasite specific IgA production remain to be established.     

Matrix protein-specific IgA antibody inhibits measles virus replication by intracellular neutralization

Measles virus (MV) is still an imposing threat to public health. The matrix (M) protein has been shown not only to function as a structure block in the assembled MV virions, but also to regulate viral RNA synthesis, playing an important role in MV's replication and assembly. In the present study, we generated a panel of IgG monoclonal antibodies (MAbs) against M protein and successfully obtained one IgA MAb (5H7) from the IgG panel. Employing the polarized Vero cells grown in the two-chamber transwell model, we investigated whether M-specific 5H7 IgA MAb could suppress MV's replication and assembly. The data presented indicate that, while failing to show the activities of traditional neutralization and immune exclusion, M-specific IgA MAb was able to effectively inhibit viral replication by intracellular neutralization (78%), supporting the notion that the M protein is important for MV assembly and replication and implying that the M protein was an effective target antigen. The data also showed that MV had a long entry and assembly phase during viral replication, providing an extended window for IgA intervention. The colocalization of M proteins and M-specific 5H7 IgA MAbs demonstrated that the intracellular neutralization was due to the direct binding of the M-specific 5H7 IgA MAbs to the M proteins. In summary, the present study has added another example showing that IgA antibodies targeting internal viral antigens could proactively participate in mucosal immune protection by intracellular neutralization and has provided evidence that M protein might be included as a target antigen in future MV vaccine design.     

Diagnostic value of dengue virus-specific IgA and IgM serum antibody detection

The diagnostic value of dengue virus (DV)-specific immunoglobulin A (IgA) serum antibody detection, by an indirect immunofluorescence assay (IFA) was evaluated. For this study, the kinetics of DV-specific IgA serum antibodies was analysed in two experimentally immunised macaques, paired samples from 35 patients suspected of a primary or secondary DV infection, paired sera from patients with high levels of IgA specific antibodies against influenza virus (n = 15), sera from patients with other viral infections (n = 40) and healthy blood donors (n = 10), which served as controls. The presence of DV-specific IgA serum antibodies in humans and in monkeys was compared with that of DV-specific IgM demonstrated in a capture enzyme-linked immunosorbent assay (ELISA). The development of DV-specific IgA and IgM antibodies in macaques proved to be similar to that observed in humans with a DV infection. In sera obtained from suspected primary DV patients during the acute phase and convalescent phase, DV-specific IgA was detected in 1/6 (17%) and 6/6 (100%), whereas IgM was detected in 4/6 (67%) and 5/6 (83%), respectively. In sera from suspected secondary DV patients during the acute phase and convalescent phase, DV-specific IgA was detected in 18/29 (62%) and 28/29 (97%), whereas IgM was detected in 20/29 (69%) and 28/29 (97%), respectively. The control group consisted of five paired serum samples from yellow fever vaccinated individuals and a patient with acute tick-borne encephalitis, 15 paired serum samples from patients with high levels of IgA antibodies specific for influenza virus and 40 serum samples from patients with specific IgM antibodies against other viruses. Ten serum samples from healthy blood donors were included. Among the control serum samples, in one patient, both DV-specific IgA and IgM antibodies were present, and in three sera DV-specific IgM antibodies could be demonstrated. These data suggest that detection of DV-specific IgA serum antibodies by IFA may have additional value for the diagnosis of DV infection.     

Inhibitory effect of intestinal anti-Gb3 IgA antibody on verotoxin-induced cytotoxicity

The effect of intestinal IgA antibody against the receptor for verotoxin (VT), globotriaosylceramide (Gb3), on VT-mediated cytotoxicity was examined. Intestinal IgA antibodies against Gb3 were prepared by oral immunization of mice with Gb3 and adjuvant monophosphoryl lipid A (MPL)-containing liposomes composed of dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylserine and cholesterol (1 : 1 : 2, molar ratio) (PS-liposome). Oral administration with Gb3 and MPL-containing PS-liposome induced significant IgA responses to Gb3 in the intestinal lavage fluid in all mice tested. Furthermore, anti-Gb3 IgA antibodies in the lavage fluid effectively inhibited the cytotoxicity of VT2 to Vero cells in a dose-dependent manner. These results suggest that anti-Gb3 IgA antibodies produced in the intestinal tract, upon oral immunization with Gb3-containing liposome, function as inhibitors against VT and also indicate the potential usefulness of oral PS-liposome vaccines containing MPL for the induction of a protective mucosal immune response against intestinal diseases.     

Fecal IgA antibody responses after oral poliovirus vaccination in infants and elder children

We investigated fecal IgA antibody responses after oral polyvalent poliovirus vaccination. Infants were given vaccines twice with an interval of 6 weeks. Specific IgA antibodies in the feces were determined by enzyme-linked immunosorbent assay, and viruses were isolated in tissue cultures. We found that, after the first vaccination, antibody responses seemed to be elicited only against the serotypes of isolated viruses. After the second vaccination, however, antibodies were detected to all three serotypes with higher titers, suggesting that the first vaccination induced the immunologic memory. The IgA antibodies had virus-neutralizing activity, and existed in the feces as both intact 11S and fragmented 4S molecules. Next, children were given the third vaccination 3 or 9 years later. Fecal IgA antibody responses were found to be poorer in elder children, while they responded with high serum neutralization titers. The secretory IgA memory seemed to last much shorter the serum IgG memory.     

Passive transfer of local immunity to influenza virus infection by IgA antibody

Secretory IgA is presumed to be the mediator of mucosal immunity based on many studies that show a correlation between protection and secretory IgA titers; however, a causal relationship has not yet been established. Classically, passive transfer of antibody has been used to demonstrate causality, but the passive transfer of local immunity with physiologically transported IgA has not been previously reported. In this study mice were injected intravenously with polymeric IgA (pIgA), monomeric IgA (mIgA), or IgG1 mAb specific for the H1 hemaglutinin of PR8 influenza virus. pIgA was shown to be specifically transported into nasal secretions relative to the mIg. The transported pIgA was functional, as evidenced by its ability to bind to virus in an ELISA assay and to protect nonimmune mice against intranasal infection with H1N1 but not H3N2 influenza virus. Intravenous injection of similar virus-neutralizing doses of anti-influenza IgG1 mAb did not protect against nasal viral challenge. IgA-mediated protection could be abrogated by the intranasal administration of antiserum against the alpha chain of IgA. These data demonstrate the passive transfer of local immunity by the i.v. administration of pIgA antibody and show that the IgA in secretions can protect against influenza virus infection. This general approach could provide a model for the evaluation of the role of local IgA in host defense against other pathogens.     

Serodiagnostics of chlamydial infections--significance of positivity in IgA and/or IgM antibody classes only

There was followed the development of serological findings in patients with proved positivity only in classes IgA and/or IgM of chlamydial antibodies (without IgG), which can be suspected of showing "false" positivity. 184 patients were repeatedly examined for chlamydial antibodies in their sera (interval between collections up to three months) using a genus specific rELISA. Sera were also tested for the evidence of IgM antibodies against capside antigen of Epstein-Barr virus (EBV) and against cytomegalovirus (CMV) using ELISA methods. In 75 (40.8%) of patients, IgA/IgM individual positivities were demonstrated even during the following sample test(s). In 28 (15.2%) of them, IgG evidence preceded and in 29 (15.7%) other patients positive seroconversion followed in this class. In 13 (7.1%) patients, IgG antibodies disappeared and subsequently reappeared. Only in 39 (21.2%) of these probands, antibodies IgA/IgM were not demonstrated at another examination. Active EBV, resp. CMV infection was proved in 24 (13.0%), resp. in 18 (9.8%) of patients. It is concluded that the evidence of positivities only in classes IgA and/or IgM mostly signal the onset of a primary infection (reinfection) or an active infection in patients with IgG production failures respectively. In these cases, a "false" positivity can be supposed to occur only in a minor extent.     

Vaccination-induced changes in human B-cell repertoire and pneumococcal IgM and IgA antibody at different ages

It is well known that older people are more susceptible to morbidity and mortality from infectious diseases, particularly from pulmonary diseases such as pneumococcal pneumonia where vaccines do not provide efficient protection as in younger populations. We have previously shown that the B-cell repertoire in the old is reduced and hypothesise that this may contribute to the impaired humoral responses of the elderly. Here, we investigated the repertoire and antibody responses to winter vaccination in two age groups, aged 18-49 and 65-89. We found that the serum IgM and IgA pneumococcal responses were significantly impaired in the older group, with no difference in IgG levels. IGHM spectratype analysis seems to be the most promising in terms of its predictive ability for vaccine responses. Spectratypes showed a clear change in the repertoire at day 7 after vaccination, with a return to the baseline levels at day 28. The changes at day 7 reflected expansion of IGH sequences that have smaller, more hydrophilic, CDR3 regions, and these changes were attenuated in the older group. The older group was more likely to have spectratypes indicative of a reduced diversity at day 0 and day 28. On average, the baseline repertoire in the older group was comprised of larger CDR3 regions than in the younger group. In conclusion, IgA and IgM responses are significantly impaired in the elderly pneumococcal response and are likely key mediators of protection. Hydrophilicity and/or small size of the IGH CDR3 appear to be important in these responses.