RESUMEN

Induction of heterosubtypic immunity to influenza viral antigens is of paramount importance to the prevention of epidemics and potential pandemics. The 1997 incidence of avian influenza infections in humans in Hong Kong heightened the need for pandemic preparedness and a search for vaccines and vaccine delivery systems that can confer broad protection. In this report, we demonstrate that the delivery of H1N1 subtype influenza viral antigens as immunostimulating complexes (ISCOM) induces broad cross-protection in mice against challenge with various influenza virus subtypes, including the avian H9 and the H5 strains that were recently responsible for deaths in humans. The ISCOM delivery system induced high and long-lived serum antiviral antibodies and class I-restricted cytotoxic T-lymphocytes (CTL). Studies with perforin, IFN-gamma, and mu-chain gene knock-out mice demonstrated that the heterosubtypic protection required cross-reactive, functional cytotoxic T cells and nonhemagglutination inhibiting serum antibodies. Interferon-gamma, a major player in viral clearance by nonlytic mechanisms, did not appear to play a role in heterosubtypic immunity. Nonformulated H1N1 influenza antigens failed to induce significant CTL or long-lasting antibody responses or to protect mice against challenge with heterosubtypic viruses. Furthermore, while influenza virus infection induced a dominant nucleoprotein (NP)-specific CTL response in H2 mice, the ISCOM delivery system induced a dominant hemagglutinin-specific CTL response. Moreover, non-neutralizing but cross-reactive antibodies played a role in reducing viral titers by macrophages. These results suggest that exogenous delivery of influenza antigens as ISCOM can influence their antigen processing and presentation, their ability to induce/recall CTL specificities, and their capacity to mediate broad cross-protection against influenza virus variants.

Asunto(s)

ISCOMs/inmunología , Virus de la Influenza A/inmunología , Vacunas contra la Influenza/inmunología , Gripe Humana/prevención & control , Macrófagos/inmunología , Linfocitos T Citotóxicos/inmunología , Animales , Anticuerpos Antivirales/inmunología , Aves , Reacciones Cruzadas , Humanos , Gripe Aviar/prevención & control , Gripe Humana/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Noqueados , Factores de Tiempo

RESUMEN

Profound alterations in humoral and cellular immune responses are a hallmark of aging, and understanding the immunobiology of aging is key to the success of preventive vaccination strategies. With aging, while recall or memory responses to influenza viral antigens for the most part remained unaltered, primary immune responses are severely impaired. The impaired primary responses are partly due to a lack of costimulation, as providing costimulation at the time of induction of primary immune responses against influenza virus vaccine partially reversed aged-related immune dysfunction and conferred enhanced protection. Inclusion of immunomodulators that up-regulate the expression of costimulatory molecules must be considered to improve the efficacy of vaccination in the elderly, particularly to novel immunogens.

Asunto(s)

Envejecimiento/inmunología , Antígenos Virales/inmunología , Memoria Inmunológica , Virus de la Influenza A/inmunología , Vacunas contra la Influenza/farmacología , Infecciones por Orthomyxoviridae/inmunología , Adyuvantes Inmunológicos/biosíntesis , Adyuvantes Inmunológicos/genética , Animales , Anticuerpos Antivirales/biosíntesis , Antígenos CD/biosíntesis , Antígenos CD/genética , Antígeno B7-1/biosíntesis , Antígeno B7-1/genética , Antígeno B7-2 , Células Cultivadas , Femenino , Vectores Genéticos , Activación de Linfocitos , Glicoproteínas de Membrana/biosíntesis , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos DBA , Análisis de Supervivencia , Linfocitos T Citotóxicos/inmunología , Regulación hacia Arriba , Virus Vaccinia/genética

RESUMEN

T560, a mouse B lymphoma that originated in gut-associated lymphoid tissue, expresses receptors that bind dimeric IgA and IgM in a mutually inhibitory manner but have little affinity for monomeric IgA. Evidence presented in this paper indicates that the receptor is poly-Ig receptor (pIgR) known in humans and domestic cattle to bind both IgA and IgM. The evidence includes the demonstration that binding of IgM is J chain dependent, and that pIg-precipitated receptor has an appropriate Mr of 116-120 kDa and can be detected on immunoblots with specific rabbit anti-mouse pIgR. Overlapping RT-PCR performed using template mRNA from T560 cells and oligonucleotide primer pairs designed from the published sequence of mouse liver pIgR indicate that T560 cells express mRNA virtually identical with that of the epithelial cell pIgR throughout its external, transmembrane, and intracytoplasmic coding regions. Studies using mutant IgAs suggest that the Calpha2 domain of dimeric IgA is not involved in high-affinity binding to the T560 pIgR. Inasmuch as this mouse B cell pIgR binds IgM better than IgA, it is similar to human pIgR and differs from rat, mouse, and rabbit epithelial cell pIgRs that bind IgA but not IgM. Possible explanations for this difference are discussed. All clones of T560 contain some cells that spontaneously secrete both IgG2a and IgA, but all of the IgA recoverable from the medium and from cell lysates is monomeric; it cannot be converted to secretory IgA by T560 cells.

Asunto(s)

Inmunoglobulina A/metabolismo , Inmunoglobulina M/metabolismo , Linfoma de Células B/inmunología , Receptores Fc/biosíntesis , Receptores de Inmunoglobulina Polimérica/biosíntesis , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sitios de Unión de Anticuerpos , Unión Competitiva/inmunología , Metabolismo de los Hidratos de Carbono , Carbohidratos/inmunología , Precipitación Química , Reacciones Cruzadas , Activación Enzimática/inmunología , Epítopos de Linfocito B/metabolismo , Epítopos de Linfocito T/metabolismo , Humanos , Immunoblotting , Inmunoglobulina A Secretora/metabolismo , Cadenas J de Inmunoglobulina/fisiología , Linfoma de Células B/metabolismo , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Peso Molecular , Fosfatidilinositol Diacilglicerol-Liasa , Proteína Quinasa C/metabolismo , ARN Mensajero/biosíntesis , Ratas , Receptores Fc/aislamiento & purificación , Receptores de Inmunoglobulina Polimérica/genética , Receptores de Inmunoglobulina Polimérica/aislamiento & purificación , Células Tumorales Cultivadas , Fosfolipasas de Tipo C/metabolismo

RESUMEN

Mice deficient in MHC class II expression (C2d mice) do not make antibody to protein antigens administered systemically, but their ability to produce IgA antibody to antigen administered at mucosal sites has not been described. We investigated IgA production by C2d mice and their IgA antibody response to antigen given orally. Young C2d mice had normal amounts of serum IgA, intestinal-secreted IgA and normal numbers of intestinal IgA plasma cells, compared to control C57BL/6 mice. IgA production by C2d mice increased with age. Following oral immunization with cholera toxin, C57BL/6 mice responded with IgA and IgG antibody, and had increased numbers of IgA plasma cells, but C2d mice gave no response. The Peyer's patch and mesenteric lymph node tissues of C2d mice contained very few CD4-expressing T cells. Thus, C2d mice have no typical mucosal CD4 Th cells and cannot respond to a strong oral immunogen, yet they still produced and secreted IgA. We hypothesized that B-1 lymphocytes could provide a source of IgA independent of antigen-specific T cell help. Young C2d mice had normal numbers of peritoneal B-1a cells and their frequency increased with age. To test the role of these B-1a cells, we bred C2d mice to obtain mice that had no MHC class II expression and expressed the Xid gene that confers deficiency in B-1a cells. These double-deficient mice had 10-fold less serum and secreted IgA than all other F2 littermates. We conclude that B-1a cells are essential for the majority of IgA production in C2d mice. Thus, the C2d mouse may provide a useful tool for analysis of the role of intestinal IgA provided by B-1a cells.

Asunto(s)

Subgrupos de Linfocitos B/inmunología , Antígenos de Histocompatibilidad Clase II/inmunología , Inmunidad Mucosa/inmunología , Inmunoglobulina A/biosíntesis , Intestino Delgado/inmunología , Animales , Toxina del Cólera/administración & dosificación , Toxina del Cólera/inmunología , Antígenos de Histocompatibilidad Clase II/biosíntesis , Inmunoglobulina A/sangre , Inmunoglobulina G/biosíntesis , Ganglios Linfáticos/inmunología , Ratones , Ratones Endogámicos C57BL , Ganglios Linfáticos Agregados/inmunología , Bazo/inmunología , Subgrupos de Linfocitos T/inmunología

RESUMEN

Ageing is associated with a decline in immune function and our primary objective is to 'reverse' age-related decline in protective immune responses to vaccination by formulating vaccines in appropriate delivery systems. In this paper, we demonstrate that influenza vaccine formulated as ISCOMs is highly immunogenic and confers protection in aged mice, when compared to current influenza vaccine. The enhanced protection conferred by Flu-ISCOMs in aged mice correlates with the up-regulation of co-stimulatory molecule, CD86 (B7.2) and to a lesser extent, CD80 (B7.1) expression on antigen presenting cells.

Asunto(s)

Envejecimiento/inmunología , Antígenos CD/biosíntesis , ISCOMs/inmunología , Vacunas contra la Influenza/inmunología , Glicoproteínas de Membrana/biosíntesis , Regulación hacia Arriba , Animales , Antígeno B7-1/biosíntesis , Antígeno B7-2 , Antígenos CD28/inmunología , Ensayo de Inmunoadsorción Enzimática , Pruebas de Inhibición de Hemaglutinación , Inmunidad Innata/inmunología , Ratones , Ratones Endogámicos DBA , Retroelementos

RESUMEN

The mucosal and systemic humoral immune systems can function essentially independent of each other, responding to mucosal and parenteral antigens, respectively. Nevertheless, antigen administered by one route can modify responsiveness to subsequent immunization by an alternate route. Here we demonstrated, in mice, in addition to stimulating rapid and robust sera antibody responses, intragastric (i.g.) immunization with human serum albumin (HSA)-containing starch microparticles (MP) grafted with 3-(triethoxysilyl)-propyl-terminated polydimethylsiloxane (TS-PDMS) primed for enhanced specific sera IgG following a parenteral antigen boost. After as little as one i.g. immunization with microentrapped, but not with soluble, HSA antigen-specific proliferation and antibody secretion were detected in Peyer's patches (PP); this activity peaked after three i.g. MP immunizations. We observed a progressive dissemination of antigen-specific lymphocyte reactivity from PP to splenic tissue following oral MP immunization. Similarly, we observed a shift in HSA-specific antibody-secreting cells from PP and mesenteric lymph nodes to splenic tissue following i.g. MP immunization. We also demonstrated that oral immunization with microentrapped, but not with soluble HSA, resulted in enhanced numbers of spontaneous Th2-cytokine secreting lymphocytes which disseminated from mucosal to systemic lymphoid compartments. This observation coincided with our findings that HSA-specific sera IgG1 responses in animals given HSA in MP were significantly higher than those detected in the sera of mice given soluble HSA i.g., both before and after parenteral antigen challenge. These findings suggest that orally-administered TS-PDMS-grafted MP, by stimulating elements of the mucosal immune system, are a valuable addition to mucosal and systemic vaccine delivery systems.

Asunto(s)

Dimetilpolisiloxanos , Mucosa Gástrica/inmunología , Inmunización/métodos , Linfocitos/inmunología , Tejido Linfoide/inmunología , Albúmina Sérica/administración & dosificación , Siliconas , Almidón , Administración Oral , Animales , Portadores de Fármacos , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Inmunoglobulina A/sangre , Inmunoglobulina G/sangre , Ratones , Ratones Endogámicos BALB C , Microesferas , Albúmina Sérica/inmunología

RESUMEN

Cytotoxic T lymphocytes (CTL) lyse virus-infected target cells by secreting the pore-forming effector molecule, perforin. Perforin-mediated cell death appears to be a major mechanism in viral clearance but its role in regulating immune responses in vivo is unclear. In this report, we show that following immunization with influenza viral antigens, perforin-deficient mice generated about 100-fold greater serum antibody responses than wild-type mice. Further, immune spleen cells from perforin knock-out mice secreted over 10-fold more IFN-gamma following in vitro restimulation than immune spleen cells from control mice. Finally, there were over 10-fold more IFN-gamma-secreting cells in cultures from perforin-deficient mice than those from control mice, suggesting that the enhanced cytokine release by T cells from perforin-deficient mice is due to an increase in the effector cell pool. Collectively, these results suggest that perforin-mediated effector function is required in the down-regulation of the immune response by way of limiting antigen-presenting cell function.

Asunto(s)

Anticuerpos Antivirales/biosíntesis , Antígenos Virales/administración & dosificación , Citocinas/biosíntesis , Virus de la Influenza A/inmunología , Glicoproteínas de Membrana/deficiencia , Animales , Anticuerpos Antivirales/sangre , Células Presentadoras de Antígenos/inmunología , ISCOMs/administración & dosificación , Inmunización , Técnicas In Vitro , Interferón gamma/biosíntesis , Interleucina-5/biosíntesis , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Perforina , Proteínas Citotóxicas Formadoras de Poros , Bazo/citología , Bazo/inmunología , Linfocitos T Citotóxicos/inmunología , Vacunas Virales/administración & dosificación

RESUMEN

Waldeyer's ring is located at the juncture of the respiratory and alimentary tracts, where it is bombarded by inhaled and ingested antigens. However, knowledge of its exact function or consequences of its removal is incomplete. Recently, the murine nasal-associated lymphoid tissue (NALT) has been reported to have functional similarities to Waldeyer's ring and, thus, might be a suitable model to examine the function of oronasopharyngeal lymphoid tissues. To explore the capability of NALT to incite local mucosal and systemic immunity, we immunized mice intranasally (i.n.) with 3-(triethoxysilyl)-propyl-terminated polydimethylsiloxane (TS-PDMS)-grafted microparticles (MP), an inoculant previously shown to induce robust systemic and mucosal humoral immunity following intragastric (i.g.) administration. We demonstrated that i.n. immunization with low doses of microentrapped, but not soluble, human serum albumin (HSA) evoked robust circulating IgG responses (P < 0.05). Additionally, NALT cells isolated from MP-treated mice proliferated in vitro when restimulated with HSA (P < 0.05), suggesting that i.n. immunization with HSA-containing MP incited specific immunity via NALT cell activation. Coinciding with these observations, after i.n. MP administration HSA-specific spot-forming cells (SFC) were observed in NALT, and later posterior cervical lymph nodes (pCLN) and spleen (SPL), suggesting that the observed MP-induced specific systemic antibody responses emanated from the NALT. We also showed that i.n. immunization with HSA-containing TS-PDMS-grafted MP stimulated interleukin-4 (IL-4)-secreting lymphocytes in the NALT. This cytokine microenvironment was probably responsible for driving the IgG1 sera response observed after i.n. MP administration, via the migration of NALT-derived IgG1-committed B cells. Interestingly, unlike i.g. MP administration, i.n. immunization with HSA-containing MP did not evoke detectable specific IgA in any lymphoid tissue examined, or in nasal secretions, probably reflecting differences between NALT and other mucosae-associated lymphoid tissues (MALT).

Asunto(s)

Inmunización , Tejido Linfoide/inmunología , Nasofaringe/inmunología , Albúmina Sérica/inmunología , Administración Intranasal , Animales , Formación de Anticuerpos , Células Productoras de Anticuerpos/inmunología , Técnicas de Cultivo de Célula , División Celular/inmunología , Femenino , Ganglios Linfáticos/inmunología , Linfocitos/inmunología , Ratones , Ratones Endogámicos BALB C , Microesferas , Polímeros

RESUMEN

Influenza immunostimulating complexes (flu-ISCOMs) and a monovalent subvirion vaccine prepared with an H1N1 strain of influenza virus were compared in mice for immunogenicity and protection against challenge with homologous and heterotypic influenza viruses. flu-ISCOMs but not subvirion vaccine fully protected mice against homologous virus challenge after one immunization as assessed by measurement of virus lung titers. The improved protection induced by flu-ISCOMs was associated with a 10-fold higher prechallenge serum hemagglutination inhibition titer. Furthermore, only flu-ISCOMs fully protected mice against mortality and reduced morbidity following challenge with an influenza virus of the serologically distinct H2N2 subtype. This cross-protection correlated with the induction of virus cross-reactive cytotoxic T lymphocytes that recognized a known major histocompatibility class I (H2-Kd)-restricted epitope within the hemagglutinin of influenza virus that is conserved among the H1 and H2 influenza virus subtypes. flu-ISCOMs may offer significant advantages over current commercial formulations as an improved influenza vaccine.

Asunto(s)

ISCOMs , Virus de la Influenza A , Vacunas contra la Influenza , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Linfocitos T Citotóxicos/inmunología , Animales , Reacciones Cruzadas , Relación Dosis-Respuesta a Droga , Femenino , Virus de la Influenza A/clasificación , Virus de la Influenza A/inmunología , Ratones , Ratones Endogámicos BALB C , Infecciones por Orthomyxoviridae/mortalidad , Factores de Tiempo , Virión/inmunología

RESUMEN

Colonization of the nasopharynx by a middle ear pathogen is the first step in the development of otitis media in humans. The establishment of an animal model of nasopharyngeal colonization would therefore be of great utility in assessing the potential protective ability of candidate vaccine antigens (especially adhesins) against otitis media. A chinchilla nasopharyngeal colonization model for nontypeable Haemophilus influenzae (NTHI) was developed with antibiotic-resistant strains. This model does not require coinfection with a virus. There was no significant difference in the efficiency of NTHI colonization between adult (1- to 2-year-old) and young (2- to 3-month-old) animals. However, the incidence of middle ear infection following nasopharyngeal colonization was significantly higher in young animals (83 to 89%) than in adult chinchillas (10 to 30%). Chinchillas that had recovered either from a previous middle ear infection caused by NTHI or from an infection by intranasal inoculation with NTHI were completely protected against nasopharyngeal colonization with a homologous strain and were found to be the best positive controls in protection studies. Systemic immunization of chinchillas with inactivated whole-cell preparations significantly protected animals not only against homologous NTHI colonization but also partially against heterologous NTHI infection. In all protected animals, significant serum anti-P6 and anti-HMW antibody responses were observed. The outer membrane P6 and high-molecular-weight (HMW) proteins appear to be promising candidate vaccine antigens to prevent nasopharyngeal colonization and middle ear infection caused by NTHI.

Asunto(s)

Haemophilus influenzae/inmunología , Nasofaringe/microbiología , Factores de Edad , Animales , Anticuerpos Antibacterianos/análisis , Proteínas de la Membrana Bacteriana Externa/inmunología , Chinchilla , Inmunización , Inmunoglobulina A/análisis , Inmunoglobulina G/análisis , Mucosa Nasal/inmunología , Otitis Media/etiología , Estreptomicina/farmacología

RESUMEN

As the most common cause of sexually transmitted disease in women, chlamydial infections can lead to pelvic inflammatory disease, infertility, and ectopic pregnancy. To better understand the role played by sex hormones in modulating the immune response of the genital tract to microbial infections, we have developed a rat model to study Chlamydia trachomatis infection. Inbred female Lewis rats were primed with progesterone and inoculated by intrauterine instillation of C. trachomatis (mouse pneumonitis strain MoPn) into each uterine horn. When infected animals were examined for the presence of chlamydial antigens 14 days postinfection, both the uterus and vagina were found to be positive compared to those of saline-treated animals, which did not show specific staining. The involvement of local and systemic immune systems following chlamydial infection was determined by analyzing major histocompatibility complex (MHC) class II expression in the reproductive tract and lymphocyte proliferation in response to mitogenic and chlamydia-specific stimulation of cells from the spleen and lymph nodes (LN) draining the reproductive tract. Enhanced proliferation was observed in LN following mitogenic but not antigenic (MOMP [major outer membrane protein]) stimulation. In contrast, spleen cell proliferation was lower in chlamydia-infected rats than in saline-treated controls. MHC class II expression, an indicator of inflammatory responses, was upregulated in the uterus, on glandular epithelial cells, and adjacent to glands in response to chlamydial infection. In other experiments, when rats were infected at estrus and diestrus without prior progesterone priming, chlamydial inclusions were not detected in either the uterus or vagina. However, enhanced lymphocyte proliferation was observed in response to mitogenic and MOMP stimulation in the reproductive tract-draining LN from estrous and diestrous animals. These findings indicate that under appropriate endocrine conditions, the rat uterus is susceptible to C. trachomatis infection and that immune responses to this pathogen can be detected locally and systemically. Further, they suggest that clearance of the infection from the reproductive tract involves immune cells from the LN draining the reproductive tract.

Asunto(s)

Infecciones por Chlamydia/inmunología , Chlamydia trachomatis/inmunología , Porinas , Progesterona/farmacología , Enfermedades Uterinas/inmunología , Animales , Proteínas de la Membrana Bacteriana Externa/inmunología , Chlamydia trachomatis/patogenicidad , Estro , Femenino , Antígenos de Histocompatibilidad Clase II/análisis , Ganglios Linfáticos/patología , Activación de Linfocitos , Ratas , Ratas Endogámicas Lew

RESUMEN

The nasal mucosal is the first site of contact with inhaled antigens. However, the nature of local immune responses and the role of nasal-associated lymphoid tissue (NALT) in those responses have rarely been studied. To characterize the cells involved in mucosally derived immune responses, NALT and Peyer's patch (PP) cells from normal mice, and mice immunized intragastrically or intranasally with cholera toxin (CT), were isolated and analyzed. Compared with PP cells, unstimulated NALT cells contained a higher proportion of T-cells. The CD4:CD8 ratio in NALT cell preparations was less than that observed in PP and more closely resembled that seen in spleen. Additionally, the total B-cell frequency in NALT cell isolates was 20% lower than that observed in PP cell preparations. Although NALT and PP cell isolates contained both mature B-cells and cells undergoing activation to express surface IgA, unlike PP, NALT showed no significant frequency of IgA-switched cells. After intranasal immunization with CT, toxin-specific IgA antibody-forming cells (AFCs) were detected in NALT cell preparations. The numbers of these cells correlated with CT-specific IgA in nasal, but not in gut washes or sera, thus suggesting local nasal production of antigen-specific mucosal antibodies. There was no evidence of anti-CT AFCs in NALT or CT-specific antibody in nasal washes after intragastric CT administration. These results support the notion that nasal mucosal antibody production is best achieved via direct stimulation of IgA-committed, NALT-derived B-cells.

Asunto(s)

Inmunidad Mucosa , Mucosa Nasal/inmunología , Ganglios Linfáticos Agregados/inmunología , Alérgenos/inmunología , Animales , Anticuerpos Antiidiotipos/análisis , Linfocitos B/inmunología , Linfocitos B/ultraestructura , Relación CD4-CD8 , Toxina del Cólera , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Inmunoglobulina A/biosíntesis , Inmunoglobulina A/inmunología , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/inmunología , Tejido Linfoide/inmunología , Tejido Linfoide/ultraestructura , Ratones , Ratones Endogámicos BALB C , Microscopía Electrónica de Rastreo , Mucosa Nasal/ultraestructura , Ganglios Linfáticos Agregados/ultraestructura , Linfocitos T/inmunología , Linfocitos T/ultraestructura

RESUMEN

Aging is associated with a decline in immune function and the elderly are therefore more susceptible to infectious disease and less responsive to vaccination. Influenza antigens complexed as immunostimulatory complexes (ISCOMs) generate more potent protective immune responses compared with non-adjuvanted flu antigens in young adult mice. We report on the protective efficacy of flu-ISCOMs compared with the current split flu vaccine in an aged mouse model. DBA/2 mice aged 2 or 18 months were immunized with flu vaccine, ISCOMs or live virus, prior to challenge with the homologous virus. In aged mice, flu-ISCOMs induced significantly higher serum hemagglutination inhibition (HAI) titers compared to vaccine, similar to the levels obtained in young adult mice that received the split vaccine. Flu-ISCOMs but not vaccine induced cytotoxic T lymphocyte (CTL) responses in young and to a lesser degree in aged mice. In aged mice flu-ISCOMs significantly reduced illness and enhanced recovery from viral infection compared with vaccine. Our data suggests that flu-ISCOMs may offer an improved vaccine strategy for protection of elderly humans against the complications of influenza infection.

Asunto(s)

Envejecimiento/inmunología , Anticuerpos Antivirales/biosíntesis , ISCOMs , Subtipo H1N1 del Virus de la Influenza A , Virus de la Influenza A/inmunología , Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/inmunología , Animales , Antígenos de Superficie/inmunología , Femenino , Pruebas de Inhibición de Hemaglutinación , Ratones , Ratones Endogámicos DBA , Infecciones por Orthomyxoviridae/prevención & control , Linfocitos T Citotóxicos/inmunología

RESUMEN

Covalent conjugates of hen egg lysozyme (HEL) and anti-major histocompatibility complex (MHC) class II monoclonal antibodies (mAb) were used to immunize mice intranasally. Three weeks after intranasal (IN) priming, mice responded rapidly to IN challenge with a mixture of HEL and cholera toxin (CT), by producing large titres of anti-HEL IgA and IgG1 antibody in serum, and IgA antibody in nasal secretions. No secondary response to HEL plus CT occurred in mice that received no priming or mice primed with HEL alone. The secondary serum IgG antibody response was dominated by the IgG1 subclass. HEL combined with CT adjuvant worked much better than HEL alone in producing the secondary response. Control conjugates, containing an IgG isotype-matched mAb without specificity for mouse tissues, provided poor priming. Mice expressing MHC class II molecules, to which the anti-MHC II mAb could not bind, produced a weak antibody response compared with those that expressed the appropriate. MHC class II molecule. Our results demonstrate that immunotargeting to MHC class II molecules via the IN route allows priming of the local IgA and circulating IgG antibody, and indicate that this technique is a feasible approach for delivery of subunit vaccines in the upper respiratory tract.

Asunto(s)

Antígenos/administración & dosificación , Antígenos de Histocompatibilidad Clase II/inmunología , Inmunoconjugados/administración & dosificación , Inmunoglobulina A Secretora/biosíntesis , Inmunoglobulina G/biosíntesis , Administración Intranasal , Animales , Anticuerpos Monoclonales/inmunología , Electroforesis en Gel de Poliacrilamida , Femenino , Inmunoglobulina G/sangre , Memoria Inmunológica , Ratones , Ratones Endogámicos A , Ratones Endogámicos BALB C , Muramidasa/administración & dosificación , Muramidasa/inmunología , Líquido del Lavado Nasal/inmunología

RESUMEN

Recent studies have demonstrated that systemic and mucosal administration of soluble antigens in biodegradable microparticles can potentiate antigen-specific humoral and cellular immune responses. However, current microparticle formulations are not adequate for all vaccine antigens, necessitating the further development of microparticle carrier systems. In this study, we developed a novel microparticle fabrication technique in which human serum albumin (HSA) was entrapped in starch microparticles grafted with 3-(triethoxysilyl)-propyl-terminated polydimethylsiloxane (TS-PDMS), a biocompatible silicone polymer. The immunogenicity of HSA was preserved during the microparticle fabrication process. Following intraperitoneal immunization of mice, TS-PDMS-grafted microparticles (MP) dramatically enhanced serum IgG responses compared with ungrafted MP and soluble HSA alone (P < 0.001). When delivered orally, both TS-PDMS-grafted and ungrafted microparticles elicited HSA-specific IgA responses in gut secretions, in contrast to orally administered soluble antigen. Indeed, TS-PDMS-grafted microparticles stimulated significantly stronger serum IgG (P < 0.005) and IgA (P < 0.001) responses compared with those elicited by ungrafted microparticles. These findings indicate that TS-PDMS-grafted starch microparticles have potential as systemic and mucosal vaccine delivery vehicles.

Asunto(s)

Inmunización/métodos , Mucosa Intestinal/inmunología , Albúmina Sérica/administración & dosificación , Siliconas , Almidón , Administración Oral , Animales , Anticuerpos/análisis , Biodegradación Ambiental , Portadores de Fármacos , Ensayo de Inmunoadsorción Enzimática , Femenino , Inmunoglobulina A/análisis , Inmunoglobulina G/análisis , Inyecciones Intraperitoneales , Ratones , Ratones Endogámicos BALB C , Microesferas , Albúmina Sérica/inmunología

RESUMEN

This case presentation describes a young woman who developed generalized urticaria after receiving the human anti-RhD(D) preparation, WinRho, intravenously. Allergy skin tests and the radioallergosorbent test (RAST) for IgE antibodies to the human anti-D immunoglobulin preparation were positive. Further studies using high-pressure liquid chromatography and protein A column chromatography implicated a nonimmunoglobulin low-molecular-weight contaminant. This case report illustrates an allergic reaction to a highly purified human immunoglobulin preparation, and demonstrates approaches to assessment of such a reaction.

Asunto(s)

Isoanticuerpos/efectos adversos , Globulina Inmune rho(D)/efectos adversos , Urticaria/etiología , Adulto , Canadá , Contaminación de Medicamentos , Femenino , Humanos , Inmunoglobulina E/inmunología , Isoanticuerpos/inmunología , Prueba de Radioalergoadsorción , Sistema del Grupo Sanguíneo Rh-Hr , Globulina Inmune rho(D)/inmunología , Pruebas Cutáneas

RESUMEN

We have explored a new technique for immunization of the intestinal tract of mice, using protein antigens bound to antibodies with specificity for murine MHC class II molecules (MHC-II). Either of two protein antigens, hen avidin (AV) or hen egg lysozyme (HEL) were covalently conjugated to anti-MHC-II antibodies and the purified conjugates were given orally (p.o.) or by direct intraduodenal (i.d.) injection into the intestinal lumen of mice. A secondary immunization p.o. with the same conjugate or with the non-conjugated antigen in the presence of cholera toxin (CTX) resulted in production of both intestinal secretory IgA and serum IgA antibody by those mice. In addition, serum IgG antibodies were produced. Conjugates with appropriate MHC-II specificity targeted the antigen because they induced more IgA and IgG antibody than conjugates with irrelevant antibody specificity or antigen alone, and because they induced antibody in mice that were genetic low responders to antigen. The results indicate the feasibility of oral subunit type vaccines with antibody targeting technology.

Asunto(s)

Anticuerpos Monoclonales/química , Avidina/inmunología , Antígenos de Histocompatibilidad Clase II/inmunología , Mucosa Intestinal/inmunología , Muramidasa/inmunología , Administración Oral , Animales , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/inmunología , Formación de Anticuerpos , Avidina/administración & dosificación , Avidina/química , Sitios de Unión de Anticuerpos , Pollos , Duodeno , Epitelio/inmunología , Femenino , Inyecciones , Mucosa Intestinal/citología , Ratones , Ratones Endogámicos A , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos CBA , Muramidasa/administración & dosificación , Muramidasa/química

RESUMEN

This study was directed to determine the extent of variability in structure or expression of intestinal disaccharidase [gamma-glucoamylase (gamma-GA), sucrase-isomaltase (SI), and lactase] between different strains of mice. Reduced levels of sucrase activity (approximately 20 U/g of protein) were observed in three strains of mice belonging to the CBA/Ca lineage. Four other strains of mice analyzed exhibited higher levels of sucrase activity (approximately 50 U/g of protein). Decreased levels of sucrase in CBA/Ca mice were not associated with decreased levels of activity associated with the isomaltase subunit or with decreased levels of SI mRNA expression. High-performance liquid chromatographic gel filtration, heat inactivation, and kinetic analysis indicated that the differences between strains in sucrase activity might be attributed to structural differences in the sucrase subunit of the SI complex, thus rendering it more susceptible to cleavage and inactivation. However, no differences in kinetic properties of the sucrase subunit were observed between strains. Murine gamma-GA was found to account for a greater proportion of maltase activity (approximately 70%) than that observed in other species (i.e., approximately 20%). In addition, CBA/Ca mice were found to be deficient in intestinal maltase activity (approximately 60 U/g) compared with the other strains studied (approximately 300 U/g).

Asunto(s)

Disacaridasas/metabolismo , Variación Genética , Mucosa Intestinal/enzimología , Intestino Delgado/enzimología , Ratones/metabolismo , Complejo Sacarasa-Isomaltasa/metabolismo , Análisis de Varianza , Animales , Disacaridasas/genética , Expresión Génica , Glucano 1,4-alfa-Glucosidasa/genética , Glucano 1,4-alfa-Glucosidasa/metabolismo , Cinética , Ratones Endogámicos BALB C/metabolismo , Ratones Endogámicos C57BL/metabolismo , Ratones Endogámicos CBA/metabolismo , ARN Mensajero/metabolismo , Especificidad de la Especie , Sacarasa/genética , Sacarasa/metabolismo , Complejo Sacarasa-Isomaltasa/genética , alfa-Glucosidasas/genética , alfa-Glucosidasas/metabolismo

RESUMEN

In previous work, we found that CBA/Ca mice display only 20% of the maltase activity present in other mouse strains. In this study, we characterized more fully the maltase deficiency in CBA/Ca mice. Virtually all of the intestinal maltase activity of CBA/Ca mice was inactivated at 50 degrees C, indicating that it was due only to the sucrase-isomaltase complex. High-performance liquid chromatographic analysis revealed that CBA/Ca mice had undetectable maltase activity displaying the molecular mass characteristic of murine gamma-glucoamylase (gamma-GA) (530 kDa). Gel electrophoretic analysis confirmed that CBA/Ca mice lacked maltase activity with molecular mass of 530 kDa corresponding to gamma-GA. Two-dimensional electrophoretic analysis revealed that the gamma-GA deficiency in CBA/Ca mice was due to the failure to synthesize the enzyme and not to the synthesis of an inactive protein. gamma-GA maltase activity could not be induced in CBA/Ca mice by a diet rich in starch, whereas the activity of other disaccharidases were readily increased. gamma-GA-deficient CBA/Ca mice appear to lack any gross metabolic abnormality resulting from this defect.

Asunto(s)

Disacaridasas/metabolismo , Glucano 1,4-alfa-Glucosidasa/deficiencia , Mucosa Intestinal/enzimología , Ratones Endogámicos CBA/metabolismo , Animales , Cromatografía Líquida de Alta Presión , Disacaridasas/biosíntesis , Electroforesis en Gel de Poliacrilamida , Inducción Enzimática , Glucano 1,4-alfa-Glucosidasa/metabolismo , Íleon/enzimología , Yeyuno/enzimología , Ratones , Especificidad de la Especie , alfa-Glucosidasas/metabolismo

RESUMEN

Immunization of BALB/cJ mice with a peptide corresponding to residues 1 to 23 of glycoprotein D [gD(1-23)] from herpes simplex virus type 2 (HSV-2) elicits antibody responses which correlate with protection against lethal HSV-2 infection. In the present study, we examined the ability of cholera toxin (CTX) to act as an immunogenic carrier for gD(1-23). The number of gD(1-23) residues conjugated to CTX affected its binding to GM1 ganglioside and physiological toxicity, both of which are factors affecting oral immunogenicity. The antibody response elicited after intraperitoneal (i.p.) immunization with the CTX-gD(1-23) conjugate was protective against a lethal i.p. challenge with HSV-2. In other experiments, mice were immunized i.p. on day 0 and subsequent immunizations conducted on days 14 and 28 were administered either intragastrically or intravaginally (i.vag.). Intraperitoneal priming followed by either i.p or intragastric boosting resulted in anti-HSV-2 antibodies in vaginal washings and in protection against a lethal i.vag. challenge with HSV-2. Intraperitoneal priming followed by i.vag. boosting did not elicit anti-HSV-2 antibodies in vaginal washings and did not protect mice against a lethal i.vag. challenge with HSV-2. These results suggest that CTX can act as a systemic and an oral delivery molecule for the covalently linked gD(1-23) peptide and that such conjugates can elicit protective immune responses against systemic and genital HSV-2 infection.

Asunto(s)

Toxina del Cólera/inmunología , Simplexvirus/inmunología , Vacunación/veterinaria , Vacunas Sintéticas/inmunología , Proteínas del Envoltorio Viral/inmunología , Animales , Anticuerpos Antivirales/análisis , Formación de Anticuerpos/inmunología , Sistema Digestivo/inmunología , Sistemas de Liberación de Medicamentos , Epítopos , Femenino , Infusiones Parenterales , Inyecciones Intraperitoneales , Ratones