ovalbumin and Conjunctivitis

Oral administration of allergens can induce immune tolerance to specific allergens in rodents and hence might be a possibility to prevent and treat allergic diseases in human subjects. However, the gastrointestinal tract of mice is different from that of human subjects. The absorption of specific antigens and subsequent antigen presentation to intestinal T cells is different in both species, making it difficult to extrapolate results.. We investigated primary oral tolerance to ovalbumin (OVA) in an IgE high-responder dog model, which is more predictive for human allergic diseases than corresponding rodent models.. Oral tolerance was induced by means of a 28-day treatment with OVA dissolved in cow's milk.. We observed reduced OVA-specific IgE and IgG production in response to ensuing subcutaneous challenges. Allergic conjunctivitis induced by means of ocular and airway provocation was significantly reduced in tolerized animals compared with that seen in nontolerized control animals. In addition, eosinophilia and neutrophilia in bronchoalveolar lavage fluid and bronchoconstriction after airway allergen challenge were significantly suppressed in tolerized animals. Cytokine analysis by means of real-time PCR on bronchoalveloar fluid cells after allergen challenge revealed a high-level expression of IL-10 and transforming growth factor beta, predominantly in the CD14(+) population.. Feeding infant beagles with OVA for 4 weeks is sufficient to prevent hallmark manifestations of asthma and allergy in adult life. The mechanism of oral tolerance involved an increased expression of IL-10 and transforming growth factor beta cytokines.

Topics: Administration, Oral; Animals; Asthma; Conjunctivitis; Dogs; Hypersensitivity; Immune Tolerance; Immunoglobulin E; Immunoglobulin G; Interleukin-10; Ovalbumin; RNA, Messenger; Transforming Growth Factor beta

To investigate antigen (Ag) specificity, activation, and effector function of the Ag-specific T cells involved in the development of experimental immune-mediated blepharoconjunctivitis (EC), an experimental conjunctivitis.. EC was induced in Brown Norway rats by injection of ovalbumin (OVA)-specific T cells followed by OVA challenge with eye drops. Eyes, including the conjunctivas, were harvested at different time points after challenge. The dependence of EC onset on the challenging Ag was assessed by challenge with an irrelevant Ag or stimulatory OVA peptides. To show the infiltration of transferred T cells into the conjunctiva, T cells were labeled with 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester (CFSE) before transfer. The activation of T cells in the conjunctiva was assessed by measuring phosphorylation of Lck-associated molecules by Western blot analysis. Conjunctivas were also examined by immunohistochemistry and used for reverse transcription-polymerase chain reaction to determine the phenotype of the infiltrating cells and cytokine, chemokine, and chemokine receptor expression. To investigate infiltration of non Ag-specific T cells into the conjunctiva, ragweed (RW)-primed lymphocytes were transferred into OVA-specific T-cell receptor transgenic (DO11.10) mice. The mice were then challenged with RW and the conjunctivas were harvested for immunohistochemistry to detect T cells derived from DO11.10 mice.. EC was induced only when challenged with OVA protein or stimulatory OVA peptides, and CFSE-labeled transferred cells were found in the conjunctiva. Phosphorylation of Lck and an 85-kDa Lck-associated molecule were observed in the conjunctiva 6 hours after challenge. Many cytokines and chemokines began to be expressed at 6 hours, and individual expression patterns over time correlated well with the infiltration patterns of different inflammatory cells. In DO11.10 mice that received RW-primed lymphocytes, T cells derived from the recipient mice infiltrated the conjunctiva after RW challenge.. Ag-specific T cells initiate EC by first infiltrating the conjunctiva, where they become activated by the specific Ag in the conjunctiva.

Topics: Animals; Blepharitis; Blotting, Western; Chemokines; Conjunctiva; Conjunctivitis; Cytokines; Epitopes; Flow Cytometry; Fluoresceins; Immunoenzyme Techniques; Immunophenotyping; Lymphocyte Activation; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Rats; Rats, Inbred BN; Receptors, Chemokine; Reverse Transcriptase Polymerase Chain Reaction; Succinimides; T-Lymphocytes

Experimental immune-mediated blepharoconjunctivitis (EC) in Brown Norway (BN) rats, which is inducible by transfer of antigen-specific T cells, is a model for human allergic conjunctivitis. We investigated the possible inhibition of EC in BN rats by topical application of FK506, which is an immunosuppressive agent that mainly targets T cells.. To induce EC by active immunization, ovalbumin (OVA) adsorbed to alum was injected into the hind footpads of BN rats. Three weeks after the initial immunization, rats were challenged with OVA by eye drops. Twenty-four hours later, lids including conjunctivas, lymph nodes (LNs), and sera were harvested for histology or reverse transcriptase PCR, proliferation assays, and measurement of IgE titer, respectively. For passive immunization, rats were intravenously injected with 10 million of in vitro-stimulated OVA-primed LN cells. Four days after the transfer, rats were challenged with OVA and evaluated as above. The rats were divided into two groups. One group received topical FK506 treatment three times per day from 15 to 21 days after active immunization or from 1 to 4 days after transfer. The other group was treated with vehicle as above.. FK506 treatment suppressed infiltration of both lymphocytes and eosinophils in the conjunctiva either by active or passive immunization (P<0.002). No differences were noted in antigen-specific cellular and humoral immune responses. Concerning cytokine expression in the conjunctiva, a prominent difference was noted only with IL-4, which was more abundantly detected in the vehicle-treated group.. Topical FK506 treatment suppressed EC in BN rats, possibly by inhibition of IL-4 in the conjunctiva.

Topics: Administration, Topical; Animals; Blepharitis; Conjunctivitis; Immunization, Passive; Immunosuppressive Agents; Male; Ophthalmic Solutions; Ovalbumin; Rats; Rats, Inbred BN; Tacrolimus

A study was performed to compare the effects of immunization with ragweed pollen (RW) in two different adjuvants on the characteristics of a previously described model of experimental immune-mediated blepharoconjunctivitis (EC) in rats.. Lewis or Brown Norway (BN) rats were immunized with 100 microg of RW in emulsion with aluminum hydroxide [Al(OH)3] or complete Freund's adjuvant (CFA). Three weeks later, the animals were challenged with eye drops containing RW in PBS. Twenty-four hours after topical challenge, eyes, blood, and lymph nodes were obtained for histology, measurement of antigen-specific antibodies, and proliferation or cytokine assays, respectively. In addition to active immunization, recipients of RW-primed lymph node cells were challenged and evaluated as above.. RW in both adjuvants induced infiltration with predominantly mononuclear cells in Lewis rats and eosinophils in BN rats. As well as active immunization, eosinophils were detected only in BN rats by adoptive transfer of cells. Lymphocyte proliferative responses to RW were high in immunized Lewis rats when CFA was used as an adjuvant. In contrast, proliferative responses in BN rats were higher when Al(OH)3 was used. RW-specific IgE was detected only in BN rats. There were no significant differences in RW-specific IgG1/IgG2a ratio among the four groups. Lewis rats had higher level of RW-specific interferon-gamma in the culture supernatant.. The characteristics of EC are different in Lewis and BN rats, dependent on the genetic background of the rat strains. The response to RW was similar to other previously used antigens, such as ovalbumin.

Topics: Allergens; Aluminum Hydroxide; Animals; Blepharitis; Conjunctivitis; Emulsions; Enzyme-Linked Immunosorbent Assay; Eosinophils; Freund's Adjuvant; Immunization; Immunoglobulin E; Immunoglobulin G; Interferon-gamma; Lymphocyte Activation; Male; Ovalbumin; Passive Cutaneous Anaphylaxis; Plant Proteins; Pollen; Rats; Rats, Inbred BN; Rats, Inbred Lew; Rats, Sprague-Dawley; Th1 Cells; Th2 Cells

Experimental immune-mediated blepharoconjunctivitis (EC) was induced in Lewis rats by immunization with ovalbumin (OVA) in complete Freund's adjuvant (CFA) or aluminum hydroxide [Al(OH)3]. To investigate the affect of genetic factors on the susceptibility of EC, we tested different strains of rats for the development of EC.. Lewis and Brown Norway (BN) rats were immunized once with 100 microg of OVA in CFA or Al(OH)3. Three weeks later they were challenged with OVA in eye drops; 24 hours after the challenge they were sacrificed and their eyes, blood, and lymph nodes were harvested for histological studies, measurement of OVA-specific antibodies (IgG, IgG1, IgG2a, IgE), and proliferation or cytokine assay, respectively. ELISA was used to detect OVA-specific IgG; passive cutaneous anaphylaxis was used for detecting IgE.. EC, OVA-specific IgG, and cellular immunity were induced in Lewis rats by using either adjuvant, whereas IgE was not produced by either adjuvant. In contrast, IgE was produced in BN rats using either adjuvant, whereas cellular immunity was evoked only when CFA was used. Less cellular infiltration as well as cellular proliferation was detected in BN rats immunized with Al(OH)3. In both strains, Al(OH)3 induced a higher IgG1/IgG2a ratio than did CFA. More interferon-gamma by stimulation with OVA was noted in Lewis rats compared to BN rats, whereas interleukin-4 was detected only in BN rats.. The severity of EC evaluated by cellular infiltration was dependent on OVA-specific cellular immunity. Genetic background is more important than adjuvants in determining the nature of EC and immunity.

Topics: Aluminum Hydroxide; Animals; Antibody Formation; Blepharitis; Conjunctivitis; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Freund's Adjuvant; Genetic Predisposition to Disease; Immunity, Cellular; Immunoglobulin E; Immunoglobulin G; Lymph Nodes; Lymphocyte Activation; Male; Ovalbumin; Rats; Rats, Inbred BN; Rats, Inbred Lew; Th1 Cells; Th2 Cells

Covalent conjugates consisting of diverse antigens coupled to optimal numbers of monomethoxypolyethylene glycol (mPEG) molecules have been shown to suppress antigen specific antibody formation. In this study, the possibility was examined that the same conjugates might prevent experimental immune mediated blepharoconjunctivitis (EC, formerly EAC) which had been shown to be caused by CD4(+) T cells-that is, to cell mediated immunity.. 6-8 week old male Lewis rats were used. The test groups of rats received two intravenous injections, each of 300 microg, of a conjugate of ovalbumin mPEG (OVA(mPEG)(11)) in phosphate buffered saline (PBS), 14 and 28 days before the single immunisation with OVA in complete Freund's adjuvant. The rats were challenged 3 weeks later by eye drops containing OVA; 24 hours later they were sacrificed, and their eyes, blood, and lymph nodes were harvested for histological examination and determination of anti-OVA antibody titres and levels of cellular immunity. Two control groups received PBS or OVA in PBS before immunisation. Furthermore, the possibility that OVA(mPEG)(11) may have induced OVA specific suppressor cells was tested by establishing the effects of the co-transfer of splenocytes from OVA(mPEG)(11) treated rats with OVA primed lymph node cells on the manifestations of EC.. Either PBS or OVA pretreated rats, which had not received OVA(mPEG)(11), developed high levels of antibodies and cell mediated immune responses to OVA, and application of eye drops led to blepharoconjunctivitis with massive cellular infiltration. In contrast, pretreatment with OVA(mPEG)(11) prevented cellular infiltration into the lids and conjunctivas, as well as the formation of detectable humoral and cellular immunity against OVA. Co-transfer of splenocytes from OVA(mPEG)(11) treated rats with OVA primed lymph node cells suppressed the cellular infiltration on application of OVA on the conjunctiva.. These data indicate that intravenous injection of OVA(mPEG)(11) conjugates suppressed both humoral and cellular immunity by the effects of antigen specific suppressor cells, thus leading to the inhibition of development of EC.

Topics: Animals; Antibody Formation; Antigens; Blepharitis; Conjunctivitis; Enzyme-Linked Immunosorbent Assay; Immunity, Cellular; Immunosuppression Therapy; Immunosuppressive Agents; Injections, Intravenous; Male; Monocytes; Ovalbumin; Polyethylene Glycols; Rats; Rats, Inbred Lew; Spleen

We recently reported the essential role of cellular immunity on the induction of experimental immune-mediated blepharoconjunctivitis (EC, formerly EAC) by using ovalbumin (OVA) as a model antigen in Lewis rats. The purpose of this study was to investigate the possible induction of EC by immunization with an OVA peptide (OVA 323-339).. Lewis rats were immunized with various doses of OVA or OVA 323-339 in complete Freund's adjuvant. Three weeks later they were challenged with OVA or OVA 323-339 by eye drops; 24 h after challenge, eyes including lids, lymph nodes and blood were harvested after clinical evaluation. An OVA 323-339-specific cell line (S816) was established by periodical stimulation with this peptide. Pathogenicity of S816 was tested by adoptive transfer of S816 into syngeneic recipient rats after challenge with OVA or OVA 323-339.. All rats immunized with OVA 323-339 developed EC after challenge with OVA or OVA 323-339. Rats immunized with OVA 323-339 at doses as low as 0.01 microg had severe clinical scores. OVA-primed rats also developed EC after challenge with OVA 323-339. OVA-primed lymph node cells responded to OVA but not to OVA 323-339. OVA 323-339-primed lymph node cells responded to OVA 323-339 but not to OVA and produced IFN-gamma by stimulation with either OVA or OVA 323-339 (three- to fourfold more than with OVA-primed lymph node cells). Recipient rats of S816 developed severe EC after challenge with either OVA or OVA 323-339.. OVA 323-339 was identified as a potent pathogenic peptide in EC.

Topics: Adoptive Transfer; Animals; Antigens; Blepharitis; CD4-Positive T-Lymphocytes; Cell Line; Conjunctivitis; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Immunity, Cellular; Immunization; Immunoglobulin G; Lymph Nodes; Lymphocyte Activation; Male; Ovalbumin; Peptide Fragments; Rats; Rats, Inbred Lew

Fischer rats were less susceptible to experimental autoimmune uveoretinitis (EAU) than Lewis rats, although both strains have the same MHC molecules. The purpose of this study was to compare the susceptibility of experimental allergic/immune-mediated blepharoconjunctivitis (EAC) between these two strains.. Male Lewis and Fischer rats were immunized with either ovalbumin (OVA) or OVA peptide (OVA323-339) in complete Freund's adjuvant (CFA). Three weeks later, they were challenged with OVA by eye drops. Twenty-four hours later, after clinical evaluation, they were killed and eyes, blood and lymph nodes were harvested for histology, antibody titer and proliferation assay, cytokine production or flow cytometric analysis, respectively.. Fischer rats developed mild EAC compared with Lewis rats. Cellular proliferative responses, IFN-gamma production of culture supernatant and serum IgG specific for OVA were basically the same between the two strains. The same OVA peptides were selected as immunodominant. Flow cytometric analysis demonstrated the same cellular profile of lymph node cells in the two strains.. EAC in Fischer rats was milder than that in Lewis rats, although no apparent differences in immunological parameters between these two strains were detected. These data suggest that factors unrelated to immunological parameters may depend on the susceptibility of EAC.

Topics: Allergens; Animals; Antibody Formation; Blepharitis; Cells, Cultured; Conjunctivitis; Cytokines; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Genetic Predisposition to Disease; Immunity, Cellular; Lymph Nodes; Lymphocyte Activation; Male; Ovalbumin; Peptide Fragments; Rats; Rats, Inbred F344; Rats, Inbred Lew

To analyse the role of stimulation in vitro of lymphocytes on the augmentation of experimental immune mediated blepharoconjunctivitis (EC, formerly EAC) in Lewis rats induced by adoptive transfer.. Two weeks after immunisation with ovalbumin (OVA), rat draining lymph nodes were collected and 50 x 10(6) cells were injected into naive syngeneic recipients either directly or after culture in vitro with OVA, concanavalin A (Con A), or purified protein derivative (PPD) for 3 days. Four days after injection the rats were topically challenged with OVA. 24 hours later, they were sacrificed and eyes and spleens were harvested for histology and proliferation assay. In some experiments, naive recipient rats were irradiated with 7 Gy gamma ray before transfer. The expression of adhesion molecules and cytokine profile of OVA primed lymph node cells were also investigated.. Both infiltrated cell number and splenocyte proliferation in the recipients of stimulated cells were higher than those of unstimulated cells. In vitro stimulation with OVA or Con A induced a severe cellular infiltration, while stimulation with PPD did not. Irradiation markedly diminished cellular infiltration. Stimulation in vitro upregulated the CD4/CD8 ratio by four times and augmented expression of CD25, I-A, ICAM-1 molecules on OVA primed lymph node cells by about five times. IFN-gamma was detected in OVA primed cells by stimulation in vitro, while IL-4 mRNA was extinguished by stimulation in vitro.. Augmentation of EC by stimulation in vitro of transferred lymphocytes might depend on the upregulation of expression of cell surface molecules and cytokine shift as well as augmented antigen specificity.

Topics: Adoptive Transfer; Animals; Blepharitis; Cell Division; Conjunctivitis; Cytokines; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Inguinal Canal; Lymph Nodes; Male; Ovalbumin; Rats; Rats, Inbred Lew; Reverse Transcriptase Polymerase Chain Reaction; Spleen

A novel immunologically provoked inflammatory process was studied in guinea pigs. The animals were immunized by i.p. injections of ovalbumin (OA) suspended in Freund's complete adjuvant and challenged by the application of OA into the conjunctival sac of one eye. An inflammatory reaction was seen a few minutes after provocation and lasted normally for 4-7 days. The process was characterized by early damage to the epithelial layer which was partly detached in small flakes; an intense tearing with the tear fluid soon turning mucous and then purulent; vasodilation in the bulbar conjunctiva, in particular towards the limbal region; margination and emigration of polymorphonuclear, and to a lesser extent, eosinophil, leucocytes which migrated towards and infiltrated the surface epithelial layer. Subsequently, the dominant cell type infiltrating the submucosa was lymphocytes. Later, opacity of the cornea occurred, probably due to oedema and neovascularization of the stroma progressing centrally from the periphery. When the antigenic challenge was repeated, thickening of the conjunctival mucosa, and neoformation of collagen bundles in the submucosa led to the swelling of the upper lids. The facets of this inflammatory trauma may not fit easily into any of the classical types of hypersensitivity. Rather, it may combine features of several of them, at least type 1 and type 4. This syndrome shows several features similar to those of human vernal keratoconjunctivitis.

Topics: Animals; Conjunctiva; Conjunctivitis; Cornea; Disease Models, Animal; Guinea Pigs; Hypersensitivity; Immunization; Leukocyte Count; Male; Microscopy, Electron; Ovalbumin

S-(1,2-Dicarboxyethyl)glutathione (DCE-GS, CAS 1115-52-2) found in rat liver, heart and lens in considerable amounts, showed an anti-inflammatory effect, which was evaluated by testing the inhibition of the experimental conjunctival edema of rats. An intravenous injection (3 mg/kg) prior to the carrageenan injection prevented the conjunctive edema formation by up to 30%. This peptide also inhibited the histamine release from rat mast cells induced by the compound 48/80. The peptide was added to the mast cells before addition of the compound 48/80 and an inhibition of the histamine release up to 96% at a 1 mmol/l concentration occurred. Furthermore, it displayed an antianaphylactic effect in rats using antibody against chicken egg albumin. An injection of the peptide (30 mg/kg) prior to the antigen administration inhibited color deposition up to 43%. Analogues or derivatives of DCE-GS were synthesized and tested for those inhibitory activities. However, there was no other peptide having stronger effects than DCE-GS and little structure-activity relationship among them.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Conjunctivitis; Glutathione; Heart; Histamine Release; In Vitro Techniques; Lens, Crystalline; Liver; Male; Mast Cells; Myocardium; Ovalbumin; p-Methoxy-N-methylphenethylamine; Passive Cutaneous Anaphylaxis; Peptides; Rats; Rats, Sprague-Dawley; Structure-Activity Relationship

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Antitussive Agents; Carrageenan; Conjunctivitis; Cyclohexanes; Cyclohexenes; Edema; Erythema; Inflammation; Mice; Ovalbumin; Rats; Ultraviolet Rays

Repeated topical applications of fluoresceinyl ovalbumin (FL-OA) to the conjunctival sac of guinea pigs sensitized them for conjunctival type 1 hypersensitivity reactions and mast cell degranulation. Guinea pigs infected with Ascaris suum, with high titers of circulating anti-A suum IgE and IgG1 antibody, also produced conjunctival type 1 reactions on topical challenge with A suum antigen. These reactions were no more intense than those of animals topically sensitized and challenged with FL-OA, which in some instances had no detectable serum homocytotropic antibody. Persistently reactive animals that had undergone repeated type 1 conjunctival reactions had histological findings (eg, papillary changes with extensive epithelial eosinophil infiltrates, epithelial thickening or thinning, numerous goblet cells, subepithelial lymphoid cell infiltrates, and new vessel formation) resembling those of human atopic vernal conjunctivitis.

Topics: Animals; Antigens, Helminth; Ascaris; Cell Count; Conjunctivitis; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Fluoresceins; Guinea Pigs; Hypersensitivity, Immediate; Mast Cells; Ovalbumin; Reagins

Topics: Administration, Topical; Animals; Conjunctivitis; Guinea Pigs; Histamine; Leukocytes; Ovalbumin; p-Methoxy-N-methylphenethylamine; Time Factors

Topics: Acacia; Animals; Antibodies; Conjunctiva; Conjunctivitis; Cornea; gamma-Globulins; Germ-Free Life; Guinea Pigs; Hypersensitivity, Immediate; Inflammation; Iris; Iritis; Keratitis; Ophthalmic Solutions; Ovalbumin; Plant Extracts

Topics: Animals; Conjunctivitis; Eosinophilia; Hypersensitivity; In Vitro Techniques; Male; Ovalbumin; Rabbits