Intranasal priming induces local lung-resident B cell populations that secrete protective mucosal antiviral IgA.

Antibodies secreted at the mucosal surface play an integral role in immune defense by serving to neutralize the pathogen and promote its elimination at the site of entry. Secretory immunoglobulin A (IgA) is a predominant Ig isotype at mucosal surfaces whose epithelial cells express polymeric Ig receptor capable of transporting dimeric IgA to the lumen. Although the role of IgA in intestinal mucosa has been extensively studied, the cell types responsible for secreting the IgA that protects the host against pathogens in the lower respiratory tract are less clear. Here, using a mouse model of influenza virus infection, we demonstrate that intranasal, but not systemic, immunization induces local IgA secretion in the bronchoalveolar space. Using single-cell RNA sequencing, we found a heterogeneous population of IgA-expressing cells within the respiratory mucosa, including tissue-resident memory B cells, plasmablasts, and plasma cells. IgA-secreting cell establishment within the lung required CXCR3. An intranasally administered protein-based vaccine also led to the establishment of IgA-secreting cells in the lung, but not when given intramuscularly or intraperitoneally. Last, local IgA secretion correlated with superior protection against secondary challenge with homologous and heterologous virus infection than circulating antibodies alone. These results provide key insights into establishment of protective immunity in the lung based on tissue-resident IgA-secreting B cells and inform vaccine strategies designed to elicit highly effective immune protection against respiratory virus infections.

Survival of Recombinant Monoclonal Antibodies (IgG, IgA and sIgA) Versus Naturally-Occurring Antibodies (IgG and sIgA/IgA) in an Ex Vivo Infant Digestion Model.

To prevent infectious diarrhea in infants, orally-supplemented enteric pathogen-specific recombinant antibodies would need to resist degradation in the gastrointestinal tract. Palivizumab, a recombinant antibody specific to respiratory syncytial virus (RSV), was used as a model to assess the digestion of neutralizing antibodies in infant digestion. The aim was to determine the remaining binding activity of RSV F protein-specific monoclonal and naturally-occurring immunoglobulins (Ig) in different isoforms (IgG, IgA, and sIgA) across an ex vivo model of infant digestion. RSV F protein-specific monoclonal immunoglobulins (IgG, IgA, and sIgA) and milk-derived naturally-occurring Ig (IgG and sIgA/IgA) were exposed to an ex vivo model of digestion using digestive samples from infants (gastric and intestinal samples). The survival of each antibody was tested via an RSV F protein-specific ELISA. Ex vivo gastric and intestinal digestion degraded palivizumab IgG, IgA, and sIgA (p < 0.05). However, the naturally-occurring RSV F protein-specific IgG and sIgA/IgA found in human milk were stable across gastric and intestinal ex vivo digestion. The structural differences between recombinant and naturally-occurring antibodies need to be closely examined to guide future design of recombinant antibodies with increased stability for use in the gastrointestinal tract.

Oral administration of an anti-CfaE secretory IgA antibody protects against Enterotoxigenic Escherichia coli diarrheal disease in a nonhuman primate model.

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of diarrhea-associated illness in developing countries. There is currently no vaccine licensed to prevent ETEC and the development of an efficacious prophylaxis would provide an intervention with significant impact. Recent studies suggested that effective protection could be achieved by inducing immunity to block colonization of ETEC. Here, we evaluated the efficacy of secretory (s) IgA2 and dimeric (d) IgA2 of an anti-colonization factor antigen antibody, 68-61, in the Aotus nancymaae nonhuman primate (NHP) ETEC challenge model via oral and parental delivery. Thirty-nine animals were distributed across 3 groups of 13, and challenged with 5.0x1011 colony forming unit (CFU) of H10407 on Day 0. Group 1 received a dIgA2 68-61 subcutaneously on day 0. Group 2 received a SIgA2 68-61 orally on days -1, 0, and +1, and Group 3 received an irrelevant SIgA2 antibody orally on days -1, 0, and +1. All animals were observed for symptoms of diarrhea, and stools were collected for ETEC colony counts. Anti-CfaE SIgA2 treatment significantly lowered the attack rate, resulting in a protective efficacy of 74.1% (p = 0.025) in Group 2 as compared to Group 3. The anti-CfaE dIgA2 treatment group had reduced diarrheal attack rate, although the reduction did not reach significance (57.1%; p = 0.072) as compared to the irrelevant SIgA2 Group 3. Our results demonstrated the feasibility of oral administration of SIgA as a potential immunoprophylaxis against enteric infections. To our knowledge, this is the first study to demonstrate the efficacy of administrated SIgA in a nonhuman primate model.

Oral Passive Immunization With Plasma-Derived Polyreactive Secretory-Like IgA/M Partially Protects Mice Against Experimental Salmonellosis.

Secretory immunoglobulins have a critical role in defense of the gastrointestinal tract and are known to act by preventing bacterial acquisition. A stringent murine model of bacterial infection with Salmonella enterica Typhimurium was used to examine protection mediated by oral passive immunization with human plasma-derived polyreactive IgA and IgM antibodies (Abs) reconstituted as secretory-like immunoglobulins (SCIgA/M). This reagent has been shown to trigger Salmonella agglutination and to limit the entry of bacterium into intestinal Peyer's patches via immune exclusion. We now demonstrate that upon administration into ligated intestinal loops, SCIgA/M properly anchors in the mucus and is protected from degradation to a better extent that IgA/M or IgG. Moreover, prophylactic oral administration of SCIgA/M before intragastric infection of mice with a virulent strain of S. enterica Typhimurium allows to protect infected animals, as reflected by reduced colonization of both mucosal and systemic compartments, and conserved integrity of intestinal tissues. In comparison with IgA/M or IgG administration, SCIgA/M provided the highest degree of protection. Moreover, such protective efficacy is also observed after therapeutic oral delivery of SCIgA/M. Either prophylactic or therapeutic treatment with passively delivered SCIgA/M ensured survival of up to 50% of infected mice, while untreated animals all died. Our findings unravel the potential of oral passive immunization with plasma-derived polyreactive SCIgA/M Abs to fight gastrointestinal infections.

Systemic administration of an HIV-1 broadly neutralizing dimeric IgA yields mucosal secretory IgA and virus neutralization.

We investigated the mucosal distribution and neutralization potency of rhesus recombinant versions of the HIV-specific, broadly neutralizing antibody b12 (RhB12) following intravenous administration to lactating rhesus monkeys. IgG and dimeric IgA (dIgA) administration resulted in high plasma concentrations of broadly neutralizing antibody (bnAb), but the monomeric IgA (mIgA) was rapidly cleared from the systemic compartment. Interestingly, differences in the distribution of the RhB12 isoform were observed between the mucosal compartments. The peak concentration of RhB12 IgG was higher than dIgA in saliva, rectal, and vaginal secretions, but the bnAb concentration in milk was one to two logs higher after dIgA administration than with IgG or mIgA infusion. Neutralization was observed in plasma of all animals, but only those infused with RhB12 dIgA showed moderate levels of virus neutralization in milk. Remarkably, virus-specific secretory IgA was detected in mucosal compartments following dIgA administration. The high milk RhB12 dIgA concentration suggests that passive immunization with dIgA could be more effective than IgG to inhibit virus in breast milk.

Monoclonal Antibodies Against Porcine sIgA and Their Use in Immunohistochemistry.

Secretory IgA (sIgA) is known as the predominant immunoglobulin in the mucosal system. It prevents pathogens from invading an animal's body through mucosa, making homeostasis. However, few studies examining the secretion of sIgA in mucosal-associated tissues of porcines based on immunohistochemistry methods have been done. In this study, BALB/c mice were immunized with porcine sIgA and the splenocytes were then fused with myeloma cells. Finally, three hybridoma cell lines secreting monoclonal antibody (MAb) against porcine sIgA were obtained. All three MAbs had no cross-reaction with porcine IgG confirmed by Western blot analysis. Furthermore, lungs, tracheas, and intestines were collected from healthy porcines to prepare tissue slices, followed by incubation with the MAb produced in this study. The results showed that sIgA existing in respiratory and digestive systems could be detected by this newly produced MAb. These generated MAbs against porcine sIgA might have a potential use in mucosal research of porcines.

Inhibition of verotoxin (VT) 2 absorption into systemic blood from intestine by repeated administration of bovine immune colostral antibody against VT2 in mice.

BACKGROUND/PURPOSE: Whether absorption of verotoxin (VT) 2 from the intestine in mice is inhibited by administration bovine immune colostral antibody against VT2 was investigated. METHODS: Three-week-old mice were administered VT2 solution at 477.8 ng/mL or 955.6 ng/mL, and bovine immune colostral antibody against VT2 was then administered three times. Whey without antibody against VT2 was administered to control mice. Serum levels of VT2 were measured by fluorescence enzyme immunoassay. RESULTS: Serum levels of VT2 in mice administered VT2 solution at 477.8 ng/mL and bovine immune colostral antibody against VT2 scarcely changed. By contrast, serum levels of VT2 in control mice increased and peaked 12 hours after administration. Peak values were 15.4 ± 5.04 ng/mL. Furthermore, serum levels of VT2 at 12 hours and 16 hours in control mice were significantly higher than in mice administered bovine colostral antibody against VT2. Serum levels of VT2 in mice administered antibody at 955.6 ng/mL showed no significant differences between repeated administration of bovine immune colostral antibody and controls. CONCLUSION: These results suggest that absorption of VT2 from the intestine was inhibited by repeated administration of bovine immune colostral antibody against VT2 at early stages of Escherichia coli O157:H7 infection, whereas VT2 in the intestine remained at low levels.

Passive administration of purified secretory IgA from human colostrum induces protection against Mycobacterium tuberculosis in a murine model of progressive pulmonary infection.

BACKGROUND: Immunoglobulin A is the most abundant isotype in secretions from mucosal surfaces of the gastrointestinal, respiratory and genitourinary tracts and in external secretions such as colostrum, breast milk, tears and saliva. The high concentration of human secretory IgA (hsIgA) in human colostrum strongly suggests that it should play an important role in the passive immune protection against gastrointestinal and respiratory infections. MATERIALS AND METHODS: Human secretory IgA was purified from colostrum. The reactivity of hsIgA against mycobacterial antigens and its protective capacity against mycobacterial infection was evaluated. RESULTS: The passive administration of hsIgA reduces the pneumonic area before challenge with M. tuberculosis. The intratracheal administration of M. tuberculosis preincubated with hsIgA to mice greatly reduced the bacterial load in the lungs and diminished lung tissue injury. CONCLUSIONS: HsIgA purified from colostrum protects against M. tuberculosis infection in an experimental mouse model.

Alteration of DSS-mediated immune cell redistribution in murine colitis by oral colostral immunoglobulin.

BACKGROUND: Oral bovine colostrum prophylaxis accelerates the recovery of dextran sulfate sodium (DSS)-induced colitis. In the present study the beneficial effects on acute intestinal inflammation of two major colostral components, secretory immunoglobulin A and lactoferrin, were investigated. Outbred NMRI mice received whole bovine colostrum (BC, 20 mg/kg body weight), colostral bovine lactoferrin (bLf, 150 mg/kg), or secretory immunoglobulin A (sIgA, 1-2 mg/kg body weight) daily by oral gavage, either two weeks before induction of colitis (prophylaxis) or after disease establishment (therapy). Bovine serum albumin (BSA, 150 mg/kg body weight) and immunoglobulin G (IgG, 1 and 2 mg/kg body weight) served as protein controls. Colitis was induced by providing 5% DSS solution ad libitum for seven days. RESULTS: Compared to BSA, BC therapy improved occult blood, stool consistency, and clinical recovery from colitis but did not prevent initial weight loss. In contrast, administration of bLf did not influence the course of colitis in either the prophylactic or the therapeutic setting. Therapeutic application of sIgA promoted weight gain in the recovery phase of colitis but failed to improve other clinical parameters. Prophylactically-fed sIgA influenced immune cell redistribution, normalized peripheral blood CD11c⁺CD83⁺ mature dendritic cells, modulated colonic immune cell infiltration, and altered the numbers of both DSS-induced regulatory γδ TCR⁺ T cells and CD11b⁺Gr-1⁺ myeloid suppressor cells in the lymph nodes and spleens of mice. CONCLUSIONS: These data demonstrated the potential of colostrum in disease recovery and epithelial homeostasis following intestinal injury. Colostral sIgA failed to improve acute disease activity but promoted weight gain and modulated immune cell responses that are involved in the genesis of colitis.

Topical curcumin can inhibit deleterious effects of upper respiratory tract bacteria on human oropharyngeal cells in vitro: potential role for patients with cancer therapy induced mucositis?

PURPOSE: Curcumin exerts its anti-inflammatory activity via inhibition of nuclear factor κB. Oropharyngeal epithelia and residing bacteria closely interact in inflammation and infection. This in vitro model investigated the effects of curcumin on bacterial survival, adherence to, and invasion of upper respiratory tract epithelia, and studied its anti-inflammatory effect. We aimed to establish a model, which could offer insights into the host-pathogen interaction in cancer therapy induced mucositis. METHODS: Moraxella catarrhalis (Mcat) and the oropharyngeal epithelial cell line Detroit 562 were used. Time-kill curves assessed the inhibition of bacterial growth and adherence assays and gentamicin protection assays determined the effect of curcumin-preincubated cells on bacterial adherence and invasion. Curcumin-mediated inhibition of pro-inflammatory activation by Mcat was determined via interleukin-8 concentrations in the supernatants. The synergistic role of secretory IgA (sIgA) on adherence was investigated. RESULTS: Curcumin was bactericidal at concentrations >50 µM. Preincubation of Detroit cells for 60 min demonstrated that concentrations >100 µM inhibited bacterial adherence. Together with sIgA, curcumin inhibited adherence at concentrations ≥50 µM. Both 100 and 200 µM curcumin significantly inhibited Mcat cell invasion. Finally, curcumin inhibited Mcat-induced pro-inflammatory activation by strongly suppressing IL-8 release. At a concentration of 200 µM, 10 min of curcumin exposure inhibited IL-8 release significantly, and complete suppression required a pre-exposure time of ≥45 min. CONCLUSION: Curcumin, in clinically relevant concentrations for topical use, displayed strong antibacterial effect against a facultative upper respiratory tract pathogen by inhibiting bacterial growth, adherence, invasion, and pro-inflammatory activation of upper respiratory tract epithelial cells in vitro.

[Clinical laboratory approaches to parodontitis treatment optimization].

In order to determine cycloferon liniment clinical-pathogenetic efficacy in comprehensive parodontitis therapy examination and treatment of 80 patients was done. It was determined that the cycloferon liniment use in comprehensive treatment of patients with parodontitis let to reduce infectious load in parodontal pockets and local inflammation intensity, to normalize the secretory immunoglobulin level and immune status indices that provided speed up of healing process and reduction relapse frequency.

Hiv-specific secretory IgA in breast milk of HIV-positive mothers is not associated with protection against HIV transmission among breast-fed infants.

OBJECTIVES: To test whether secretory immunoglobulin A (sIgA) to human immunodeficiency virus (HIV) antigens in breast milk of HIV-positive women is associated with protection against HIV transmission among breast-fed infants. STUDY DESIGN: Nested, case-control design in which HIV-specific sIgA was measured in breast milk collected from 90 HIV-positive women enrolled in a study in Lusaka, Zambia. Milk samples were selected to include 26 HIV-positive mothers with infected infants (transmitters) and 64 mothers with uninfected infants (nontransmitters). RESULTS: HIV-specific sIgA was detected more often in breast milk of transmitting mothers (76.9%) than in breast milk of nontransmitting mothers (46.9%, P = .009). There were no significant associations between HIV-specific sIgA in breast milk and other maternal factors, including HIV RNA quantities in breast milk, CD4 count, and plasma RNA quantities. CONCLUSIONS: HIV-specific sIgA in breast milk does not appear to be a protective factor against HIV transmission among breast-fed infants.

Secretory IgA induces antigen-independent eosinophil survival and cytokine production without inducing effector functions.

BACKGROUND: Eosinophils in human beings reside in tissues, especially the mucosal tissues of the gastrointestinal tract and inflamed airways. Secretory IgA (S-IgA) is the predominant antibody secreted by these tissues and likely plays a role in the innate immune response. OBJECTIVE: Because eosinophils and S-IgA are often colocalized in mucosal tissues, we examined the potential regulatory effects of S-IgA without antigens on survival, gene expression, and effector functions of human eosinophils. METHODS: Eosinophils were incubated with S-IgA in solution without antigens (soluble S-IgA) or with S-IgA immobilized to mimic multivalent antigen cross-linking. Eosinophil activation was monitored by superoxide anion generation and degranulation. Survival was assessed between 24 and 96 hours. Gene and protein expression were examined by microarray and ELISA. Eosinophil lysates were examined by immunoblot for extracellular signal-regulated kinase (ERK) phosphorylation. RESULTS: Immobilized S-IgA stimulated eosinophil superoxide production and degranulation; soluble S-IgA did not. Although immobilized S-IgA inhibited eosinophil survival in vitro, soluble S-IgA enhanced survival; this involved autocrine production of GM-CSF. Soluble S-IgA without antigens induced increases in mRNA levels of various cytokines, chemokines, signal transduction molecules, antiapoptotic factors, and cell surface markers. By using ELISA, we confirmed protein expression of selected mediators. Eosinophil interaction with soluble S-IgA likely involves FcalphaRI (CD89) and ERK pathway activation. CONCLUSION: Secretory IgA without multivalent antigens may regulate survival and gene expression of eosinophils. Eosinophils in mucosal tissues can be either primed for action (cytokine production and survival) or fully activated (degranulation and superoxide release) by different forms of S-IgA.

Targeting of secretory IgA to Peyer's patch dendritic and T cells after transport by intestinal M cells.

In addition to being instrumental to the protection of mucosal epithelia, secretory IgA (SIgA) adheres to and is transported by intestinal Peyer's patch (PP) M cells. The possible functional reason for this transport is unknown. We have thus examined in mice the outcome of SIgA delivered from the intestinal lumen to the cells present in the underlying organized mucosa-associated lymphoreticular tissue. We show selective association of SIgA with dendritic cells and CD4(+) T and B lymphocytes recovered from PP in vitro. In vivo, exogenously delivered SIgA is able to enter into multiple PP lining the intestine. In PP, SIgA associates with and is internalized by dendritic cells in the subepithelial dome region, whereas the interaction with CD4(+) T cells is limited to surface binding. Interaction between cells and SIgA is mediated by the IgA moiety and occurs for polymeric and monomeric molecular forms. Thus, although immune exclusion represents the main function of SIgA, transport of the Ab by M cells might promote Ag sampling under neutralizing conditions essential to the homeostasis of mucosal surfaces.

Monoclonal IgA antibody-mediated expulsion of Trichinella from the intestine of mice.

To assess the potential role of IgA antibody in expulsion of the nematode of the genus Trichinella from the intestine, a panel of IgA monoclonal antibodies (mAbs) were produced from the mesenteric lymph node cells from BALB/c mice orally vaccinated with irradiated muscle larvae of Trichinella britovi. One IgA mAb, HUSM-Tb1, formed immunoprecipitates on the surface of live muscle larvae, and by immunohistochemistry reacted with their stichocytes and cuticular surface, but not with those tissues of the adult stage or newborn larvae. Intraperitoneal injection of BALB/c mice with this mAb 5 h before challenge conferred a high level of protection (more than 95%) against T. britovi infection, when 2.0 mg of specific IgA/20 g body weight was given to a mouse. The same treatment produced a similar effect in SCID mice lacking functional T- and B-cells, indicating no requirement of synergistc T-cell factors for the effect. Passive transfer of the mAb at the time of challenge or later showed less or no effect upon worm expulsion. It is concluded that the mucosal IgA response, when adequately induced, can impede the establishment of infective Trichinella parasites in the mouse intestine.

Inmunoglobulina a Secretora en niños internados el Hospital Materno Infantil y Hospitaldel Niño con diagnostico de Infeccion Respiratoria e Infeccion Gastrointestinal / Secretory Inmunoglobulina to in children committed in the Infantile Maternal Hospital and Hospital of the Boy with I diagnose of Respiratory Infection and Gastrointestinal Infection

La presente investigación tuvo como objetivo estudiar los niveles de IgA secretora en pacientes con diagnostico de infección respiratoria o infección gastrointestinal. Participaron del estudio 129 niños de ambos sexos, 43 correspondían a niños aparentemente sanos (grupo control) y 86 a niños con infección gastrointestinal o respiratoria (grupo de estudio) a los mismos que se les tomó las muestras de saliva de la cavidad sublingual. Posterioremente se procedió a dosificar IgA secretora a través del método ELISA de captura. Se completó un cuestionario en el que se incluyeron datos personales, diagnóstico clínico, antecedentes patológicos y exámenes laboratoriales, permitiendo así, la clasificación de la población. Los resultados muestran niveles de IgA s normales o ligeramente incrementados en los niños con infección respiratoria e infección gastrointestinal, pese a tratarse de niños internados y en su mayoría con estados deficientes nutricionalmente, sugiriéndose la posibilidad de una hiperestimulación constante del medio sobre el sistema imnunológico

Theophylline inhibits the release of eosinophil survival cytokines--is Raf-1 the protein kinase A target?

Increased numbers of activated eosinophils in bronchial tissue is a feature of asthma and may, in part, be attributed to the prolonged cytokine-dependent survival of eosinophils within the inflamed microenvironment. Low-dose oral theophylline was previously shown to reduce the number of activated eosinophils within the sub-mucosa following allergen exposure. A number of inhibitory actions of theophylline have been described which relate to eosinophil recruitment and activation, including inhibition of cell migration and release of granule basic proteins. In this study we investigated the ability of theophylline to inhibit the release of preformed GM-CSF and IL-8 from eosinophils in vitro, as these cytokines may serve an autocrine function in eosinophil survival in vivo. Eosinophils rapidly released GM-CSF and IL-8 spontaneously, and release was further enhanced in response to sIgA-coated beads. Theophylline inhibited the stimulated, but not the spontaneous, release of both cytokines. We previously reported the role of protein kinase A in inhibition of arachidonic acid mobilization and LTC4 synthesis. Therefore we speculate that cAMP-dependent activation of protein kinase A following theophylline treatment of eosinophils resulted in inhibition of Raf-1 and MAPK/MAPKK dependent activation of phospholipase A2 and consequently inhibition of degranulation and cytokine release.

Human milk proteins including secretory IgA fail to elicit tolerance after feeding.

Oral administration of large doses of protein antigen generally induces a state of systemic unresponsiveness currently termed mucosally induced tolerance. In this study, we used human milk protein (HMP) without casein as a multi-protein antigen for the study of mucosally induced tolerance. The HMP utilized in this study mainly contained secretory (S) IgA, lactoferrin (Lf) and alpha-lactalbumin (Lact). When mice were given 1 or 25 mg of HMP orally 3 times or 25 mg orally four consecutive weeks prior to systemic immunization, antigen-specific serum IgG responses to HMP were induced by subsequent parenteral immunization with 100 microg of HMP. Analysis of IgG subclasses revealed that IgG1 followed by IgG2b accounted for the IgG responses noted. When both HMP and ovalbumin (OVA) were fed to mice, tolerance developed to OVA but not to HMP. To further investigate the nature of immune responses seen following oral gavage of HMP, we examined responses to individual protein of HMP. Brisk serum IgG1 and IgG2b responses to both S-IgA and Lf were induced by oral followed by systemic immunization with HMP. Analysis of splenic CD4+ T cells from mice given oral HMP revealed production of Th2- but not Th1-type cytokines. These results show that oral administration of HMP preferentially induces exclusive Th2-type immune responses, which may prevent the development of HMP (S-IgA and Lf)-specific mucosally induced tolerance.