ovalbumin and Leukopenia

Leukopenia is a severe condition resulting from both pathological processes and some treatments, like chemotherapy in cancer patients. However, the activation of the patient immune system is required for the success of immunotherapeutic strategies, as cancer vaccines. In this regard, leukopenia constitutes a major hurdle to overcome, mainly due to the impairment of cytotoxic T lymphocyte (CTL) responses. Adjuvants are basic components of vaccine formulations, which might be useful to stimulate immunity under this immunosuppressed condition. To this aim, we tested the capacity of a novel nanoparticulated complex, very small size proteoliposomes (VSSP), to promote CTL even in a leukopenic scenario. Noteworthy, we observed that a VSSP-based OVA vaccine induced a normal antigen-specific CTL response in mice rendered leukopenia by the administration of high doses of the chemotherapeutic agent cyclophosphamide (CY), while under the same conditions the OVA antigen formulated in the TLR-3 agonist polyinosinic-polycytidylic acid (P(I:C)) was ineffective. Moreover, an appropriate combination of VSSP with the P(I:C) vaccine was able to restore the CD8(+) T cell effector function in leukopenic mice. VSSP induced not only a faster repopulation of immune cells in CY-receiving animals, but also enhanced the recovery of memory T lymphocytes and myeloid dendritic cells (DCs) while simultaneously abrogated the immunosuppressive capacity of myeloid-derived suppressor cells (MDSCs). Our results suggest that VSSP could be a particularly suitable immunomodulator to be used in CTL-promoting active immunotherapy strategies operating in severe immune compromised scenarios.

Topics: Adjuvants, Immunologic; Animals; Female; Leukopenia; Mice; Mice, Inbred C57BL; Neisseria meningitidis; Ovalbumin; Proteolipids; T-Lymphocytes, Cytotoxic

In certain models of allergic airway disease, mast cells facilitate the development of inflammation and airway hyper-responsiveness (AHR). To define the role of the high affinity IgE receptor (FcepsilonRI) in the development of AHR, mice with a disruption of the alpha subunit of the high affinity IgE receptor (FcepsilonRI(-/-)) were exposed on 10 consecutive days to nebulized OVA. Forty-eight hours after the last nebulization, airway responsiveness was monitored by the contractile response of tracheal smooth muscle to electrical field stimulation (EFS). After the 10-day OVA challenge protocol, wild-type mice demonstrated increased responsiveness to EFS, whereas similarly challenged FcepsilonRI(-/-) mice showed a low response to EFS, similar to nonexposed animals. Further, allergen-challenged FcepsilonRI(-/-) mice showed less airway inflammation, goblet cell hyperplasia, and lower levels of IL-13 in lung homogenates compared with the controls. IL-13-deficient mice failed to develop an increased response to EFS or goblet cell hyperplasia after the 10-day OVA challenge. We transferred bone marrow-derived mast cells from wild-type mice to FcepsilonRI(-/-) mice 1 day before initiating the challenge protocol. After the 10-day OVA challenge, recipient FcepsilonRI(-/-) mice demonstrated EFS-induced responses similar to those of challenged wild-type mice. Transferred mast cells could be detected in tracheal preparations. These results show that FcepsilonRI is important for the development of AHR after an aerosolized allergen sensitization protocol and that this effect is mediated through FcepsilonRI on mast cells and production of IL-13 in the lung.

Topics: Adjuvants, Immunologic; Adoptive Transfer; Aerosols; Allergens; Animals; Bone Marrow Transplantation; Bronchial Hyperreactivity; Electric Stimulation; Female; Goblet Cells; Hyperplasia; Immunoglobulin E; Immunoglobulin G; Inflammation; Interleukin-13; Leukopenia; Lung; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Mutant Strains; Mice, Transgenic; Nebulizers and Vaporizers; Ovalbumin; Receptors, IgE; Trachea

Activin A, a homodimeric protein (betaAbetaA) and a member of the TGF-beta superfamily, is involved in the inflammatory repair process. Using cDNA microarray analysis, we discovered strong induction of the activin betaA gene in human mast cells (MC) on stimulation with PMA and calcium ionophore (A23187). Activin betaA mRNA was also highly induced in primary cultured murine bone marrow MC (BMMC) after stimulation by IgE receptor cross-linking. Secretion of activin A was evident in human mast cell-1 line cells 3 h after stimulation and progressively increased over time. Activin A was present in the cytoplasm of activated but not unstimulated murine bone marrow MC as demonstrated by immunofluorescence studies, suggesting that secretion of activin A by MC was due to de novo synthesis rather than secretion of preformed protein. Activin A also colocalized with human lung MC from patients with asthma by double-immunofluorescence staining. Furthermore, secretion of activin A was significantly increased in the airway of wild-type mice after OVA sensitization followed by intranasal challenge. Secretion of activin A, however, was greatly reduced in MC-deficient WBB6F(1)-W/W(v) mice as compared with wild-type mice, indicating that MC are an important contributor of activin A in the airways of a murine asthma model. Additionally, activin A promoted the proliferation of human airway smooth muscle cells. Taken together, these data suggest that MC-derived activin A may play an important role in the process of airway remodeling by promoting the proliferation of airway smooth muscle.

Topics: Activins; Animals; Asthma; Blotting, Northern; Bone Marrow Cells; Bronchoalveolar Lavage Fluid; Cell Division; Cell Line; Cells, Cultured; Cytoplasm; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Humans; Inhibin-beta Subunits; Leukopenia; Lung; Mast Cells; Mice; Mice, Mutant Strains; Microscopy, Confocal; Muscle, Smooth; Oligonucleotide Array Sequence Analysis; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction

In the mucosal immune system, resident dendritic cells are specialized for priming Th2-polarized immunity, whereas the Ag-presenting activity of macrophages has been linked with the development of Th1 phenotype. As an immune switch toward Th1 can protect against Th2-mediated allergic response, this study investigated the capacity of lung macrophages to stimulate Th1 responses during the secondary exposure to inhaled allergen, thereby suppressing Th2-mediated allergic airway inflammation in a murine model of allergic asthma. Following airway macrophage depletion in OVA-sensitized mice, lung T cells defaulted to a phenotype that produced less Th1 (IFN-gamma) and more Th2 (IL-4 and IL-5) cytokines, leading to more severe airway hyperreactivity and inflammation after intranasal Ag challenge. After OVA pulsing and adoptive transfer, lung macrophages selectively promoted a Th1 response in Ag-sensitized recipients and did not induce pulmonary eosinophilia. By contrast, OVA pulsing and adoptive transfer of a lung cell preparation, consisting of dendritic cells, B cells, and macrophages, promoted a Th2 response with an associated inflammatory response that was suppressed when macrophages were present and pretreated with IFN-gamma, but exacerbated when macrophages were depleted before IFN-gamma treatment. In addition, Th1-promoting activity of lung macrophages was not related to the autocrine production of IL-12p40. These results suggest that the Th1-promoting APC activity may be an inherent property of the lung macrophage population, and may play an important role, upon stimulation by IFN-gamma, in antagonizing an ongoing Th2 immunity and Th2-dependent allergic responses.

Topics: Administration, Intranasal; Adoptive Transfer; Allergens; Animals; Antigen Presentation; Antigen-Presenting Cells; Clodronic Acid; Cytokines; Eosinophilia; Female; Immunophenotyping; Inflammation; Interferon-gamma; Interleukin-12; Leukopenia; Liposomes; Lung; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Hypersensitivity; Th1 Cells; Th2 Cells; Up-Regulation

1. In guinea-pigs previously sensitized with ovalbumin, the intra-plantar administration of the antigen induced dose-dependent and sustained oedema. An intense infiltrate of neutrophils and eosinophils was observed at the peak of the oedema (4 h). 2. Oedema induced by ovalbumin at the doses of 50 or 200 micrograms/paw was not inhibited by antihistamines (meclizine and cetirizine), a PAF antagonist (BN 50730), a cyclo-oxygenase inhibitor (indomethacin), a lipoxygenase inhibitor (MK-886), a dual type lipo- and cyclo-oxygenase inhibitor (NDGA), a bradykinin antagonist (Hoe 140) or the combination of cetirizine, MK-886, indomethacin and BN 50730. These drugs did inhibit paw oedema induced by their specific agonists or by carrageenin. These results suggest that histamine, PAF, prostaglandins, leukotrienes or bradykinin are not important in the development of immune paw oedema in guinea-pigs. 3. Dexamethasone (10 mg kg-1) inhibited oedema induced by ovalbumin (50 or 200 micrograms/paw, P < 0.05). This effect apparently does not result from inhibition of arachidonate metabolism, since indomethacin, MK-886 and NDGA were without effect. 4. Oedema induced by ovalbumin (50 or 200 micrograms/paw) was also inhibited by azelastine. This effect was not due to the anti-histaminic property of azelastine since two other potent-antihistamines, meclizine and cetirizine, were ineffective. 5. Intravenous injection of lipopolysaccharide (LPS) dose-dependently inhibited the oedema induced by ovalbumin (200 micrograms/paw). This effect could not be attributed to hypotension or leucopenia since the maximal dose applied (81 micrograms kg-1) did not induce significant changes in the blood pressure or in the white blood cell levels of the animals. It is suggested that the effect of LPS is mediated by the endogenous release of cytokines, including tumour necrosis factor (TNF alpha). Murine TNF alpha dose dependently(9-81 microg kg-1) inhibited the paw oedema induced by ovalbumin.7. The anti-oedematogenic effects of LPS and/or TNF alpha are possibly associated with their capacity to inhibit leucocyte emigration. Accordingly, guinea-pigs rendered leucopenic with vinblastine exhibited less intense oedema after ovalbumin. Vinblastine did not affect oedema induced by PAF or bradykinin,indicating that vascular responsiveness was not involved.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Blood Pressure; Dexamethasone; Edema; Enzyme-Linked Immunosorbent Assay; Foot; Guinea Pigs; Hypersensitivity; Immunoglobulin G; Leukocyte Count; Leukocytes; Leukopenia; Lipopolysaccharides; Male; Mice; Ovalbumin; Phthalazines; Platelet Aggregation Inhibitors; Tumor Necrosis Factor-alpha; Vinblastine

Bronchoconstriction (BC) is the main feature of anaphylaxis in the guinea pig. Since LPS induces lung inflammation and antigen-induced BC depends on the endogenous formation of histamine and arachidonate metabolites, we studied whether LPS might modulate antigen-induced BC. Guinea pigs were sensitized subcutaneously with 10 micrograms ovalbumin (OA) on days 0 and 14. LPS (100 micrograms/kg) was injected intravenously on day 21, and daily injections of LPS were continued before the antigenic challenge on day 22, 23, 24, or 25. Intratracheal injection of 100 micrograms OA induced an abrupt and reversible BC. Single or repetitive injections of LPS reduced BC. LPS is likely to reduce the OA-induced BC by affecting the histamine-dependent component of BC, since (a) LPS induced a partial degranulation of lung mast cells; (b) BC is reduced by mepyramine, an histamine receptor antagonist; (c) LPS did not affect BC in mepyramine-treated guinea pigs; (d) LPS reduced histamine release by OA-stimulated guinea pig lungs in vitro. Moreover, the in vitro OA-induced production of arachidonate metabolites was also reduced by LPS. The decreased formation of TXB2 was not only secondary to a reduced release of histamine, since LPS inhibited TXB2 formation in the presence of mepyramine. Finally, the FMLP-induced BC and mediator release were inhibited by LPS, whereas the platelet activating factor-induced pulmonary responses were not. Thus, the protective effect of LPS is not antigen-specific and does not result from a general desensitization. These studies indicate that a single dose of LPS reduces the antigen-induced BC by reducing histamine release from lung mast cells, although a decreased formation of eicosanoids may contribute to the protective effect of LPS.

Topics: Animals; Antigens; Aspirin; Bronchoconstriction; Cell Degranulation; Endotoxins; Escherichia coli; Guinea Pigs; Hemodynamics; Histamine; Hypersensitivity; Leukopenia; Lipopolysaccharides; Mast Cells; Microscopy, Electron; N-Formylmethionine Leucyl-Phenylalanine; Ovalbumin; Platelet Activating Factor; Pyrilamine; Thromboxane B2

1. Ovalbumin administration to animals injected with 0.5-5 x 10(6) washed platelets microliter-1 from actively sensitized animals induced a marked decrease (maximum of around 50% at 60 min) in the number of circulating leucocytes, whereas platelet counts were unaffected. The intensity of the decrease in leucocyte counts was dependent upon the concentration of the injected platelets. 2. No drop in leucocyte counts was measured upon antigen challenge of guinea-pigs injected with platelets from non-sensitized animals. 3. This phenomenon was unaffected by pretreatment of the recipient animals with a platelet-activating factor (PAF) antagonist WEB 2086 (3 mg kg-1 i.v.) but was markedly reduced (around 50% inhibition) by the anti-allergic drug nedocromil sodium (100 mg kg-1 i.v.). By contrast, indomethacin (5 mg kg-1 i.v.) caused a significant (P less than 0.01) enhancement of the antigen-induced leucopenia, whereas the mixed cyclooxygenase and lipoxygenase inhibitor, BW 755C (10 mg kg-1 i.v.) suppressed the drop in leucocyte counts evoked by ovalbumin administration. 4. These results indicate that platelets from actively sensitized guinea-pigs transferred to normal animals still responds to the specific antigen with the activation of circulating leucocytes. This phenomenon appears to be independent of the production of PAF and of cyclo-oxygenase metabolites. By contrast, the production of the metabolites of the lipoxygenase pathway by platelets could account for the marked leucopenia observed following the immunological stimulation.

Topics: 4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; Animals; Antigens; Azepines; Blood Platelets; Female; Guinea Pigs; Immunization; In Vitro Techniques; Indicators and Reagents; Indium Radioisotopes; Leukocyte Count; Leukopenia; Male; Nedocromil; Ovalbumin; Quinolones; Triazoles

We examined the kinetics of proteoglycan (PG) depletion in rabbits with antigen-induced arthritis. There was a rapid loss of PG from arthritic cartilage, reaching 35-40% at day 7. Thereafter, the rate of PG depletion declined, and by day 42, the maximum loss was 55-60%. The initial loss of PG was accompanied by the appearance of large amounts of sulfated glycosaminoglycans (GAGs) in the joint fluid (measured as total sulfated GAGs by dye binding and as keratan sulfate by radioimmunoassay). However, by day 14, the levels of sulfated GAGs in arthritic joint fluid declined to control levels, even though the cartilage demonstrated a sustained depletion of PG. The cartilage PG degradation observed in antigen-induced arthritis could also be produced in normal animals by a single intraarticular injection of recombinant interleukin-1. The acute loss of cartilage PG occurred independently of neutrophil accumulation, both in the case of antigen-induced arthritis and after injection of interleukin-1.

Topics: Animals; Antigens; Arthritis; Arthritis, Experimental; Cartilage, Articular; Glycosaminoglycans; Immunization; Injections, Intra-Articular; Interleukin-1; Joints; Keratan Sulfate; Knee Joint; Leukopenia; Male; Mechlorethamine; Neutrophils; Ovalbumin; Proteoglycans; Rabbits

One characteristic of experimental allergic encephalomyelitis (EAE) in all species is the presence of a considerable leukocyte infiltrate in the central nervous system (CNS). By adoptive transfer of EAE into irradiated or nonirradiated Lewis strain rats we now show that the bulk (greater than 90%) of infiltrating cells in the CNS are superfluous to the induction of disease, as lethally irradiated recipients, despite having very few infiltrating cells in the CNS, acquire severe paralytic EAE. The reduction in the level of infiltration in irradiated recipients is selective, however, as both irradiated and nonirradiated diseased animals have very similar numbers of cells expressing IL-2-R. Disease in irradiated recipient animals is associated with substantial submeningeal hemorrhage in the spinal cord and brain stem and similar hemorrhages are found in recipients rendered leukopenic with cytotoxic drugs. Clinical signs of disease and hemorrhage are preventable, however, by administration to the recipient rats of mAbs specific for the CD4 antigen. Classic delayed-type hypersensitivity (DTH) reactions are transferable with the same cells that produce EAE in both irradiated and nonirradiated recipient rats, but such transfer of DTH is observed only in nonirradiated recipient animals and not in irradiated rats. Collectively, the findings reported herein support the conclusion that the paralysis characteristic of acute EAE is mediated by the direct action of very small numbers of activated CD4+ lymphocytes that infiltrate the CNS and produce their effects by inducing vascular damage. The findings are not consistent with reports that the lesions in EAE are produced by a classic DTH reaction.

Topics: Animals; Busulfan; Cells, Cultured; Central Nervous System; Cerebral Hemorrhage; Chlorambucil; Encephalomyelitis, Autoimmune, Experimental; Female; Hemorrhage; Hypersensitivity, Delayed; Immunization, Passive; Leukopenia; Male; Myelin Basic Protein; Ovalbumin; Rats; Rats, Inbred Lew; Receptors, Immunologic; Receptors, Interleukin-2; Spinal Cord Diseases; Spleen; T-Lymphocytes, Helper-Inducer; Whole-Body Irradiation

Topics: Acute Disease; Anaphylaxis; Animals; Apnea; Carotid Arteries; Erythrocyte Count; Hemodynamics; Histamine; Hypersensitivity, Immediate; Hypertension; Hypertension, Pulmonary; Leukocyte Count; Leukopenia; Ovalbumin; Swine; Swine Diseases; Thrombocytopenia

Topics: Animals; Blood Proteins; Cattle; Enterobacter; Female; Immunodiffusion; Leukocyte Count; Leukopenia; Mastitis, Bovine; Ovalbumin; Staphylococcal Infections