Uveitis In Lewis Rats Research Articles

Attenuation of Experimental Autoimmune Uveitis in Lewis Rats by Betaine.

Experimental autoimmune uveitis (EAU) is an animal model of human autoimmune uveitis that is characterized by the infiltration of autoimmune T cells with concurrent increases in pro-inflammatory cytokines and reactive oxygen species. This study aimed to assess whether betaine regulates the progression of EAU in Lewis rats. EAU was induced via immunization with the interphotoreceptor retinoid-binding protein (IRBP) and oral administration of either a vehicle or betaine (100 mg/kg) for 9 consecutive days. Spleens, blood, and retinas were sampled from the experimental rats at the time of sacrifice and used for the T cell proliferation assay, serological analysis, real-time polymerase chain reaction, and immunohistochemistry. The T cell proliferation assay revealed that betaine had little effect on the proliferation of splenic T cells against the IRBP antigen in an in vitro assay on day 9 post-immunization. The serological analysis showed that the level of serum superoxide dismutase increased in the betaine-treated group compared with that in the vehicle-treated group. The anti-inflammatory effect of betaine was confirmed by the downregulation of pro-inflammation-related molecules, including vascular cell adhesion molecule 1 and interleukin-1β in the retinas of rats with EAU. The histopathological findings agreed with those of ionized calcium-binding adaptor molecule 1 immunohistochemistry, further verifying that inflammation in the retina and ciliary bodies was significantly suppressed in the betaine-treated group compared with the vehicle-treated group. Results of the present study suggest that betaine is involved in mitigating EAU through anti-oxidation and anti-inflammatory activities.

Anti-inflammatory Effects of Sinomenium Acutum Extract On Endotoxin-induced Uveitis in Lewis Rats.

Purpose: Recent studies have reported the anti-inflammatory effect of Sinomenium acutum. We investigated the anti-inflammatory effect of sinomenine on endotoxin-induced uveitis in a rat model.Methods: Endotoxin-induced uveitis was induced in rat by lipopolysaccharide (LPS) immunization. Sinomenine (50mg/kg and 100mg/kg) was administered at 30 minutes before, 6 hours and 12 hours after LPS immunization. Clinical and histological severity was evaluated. Protein concentration and levels of tumor necrosis factor (TNF)-α and prostaglandin (PG)-E2 in aqueous humor were measured. Expression of activated Nuclear factor (NF)-κB p65 in ciliary body was also observed.Results: Clinical and histological severities were significantly milder in sinomenine-treated rat than in controls (P < 0.001). Sinomenine suppressed protein leakage and down-regulated the production of TNF-α and PG-E2 in a dose-dependent manner. Sinomenine treatment suppressed the translocation of the NF-κB p65 subunit into the nuclei.Conclusion: Systemic administration of sinomenine suppressed the inflammation of ocular tissues. These findings suggest that sinomenine could be a novel therapeutic agent for the control of endogenous ocular inflammatory disease.

Therapeutic anti-inflammatory effects of luteolin on endotoxin-induced uveitis in Lewis rats.

The present study aimed to investigate the therapeutic efficacy of post-inflammatory treatment with luteolin on endotoxin-induced uveitis (EIU) in rats. Intraperitoneal injection of 10 mg/kg luteolin or 1 mg/kg prednisolone (Pred) at 4 hr post-lipopolysaccharide (LPS) injection (200 µg) was associated at 24 hr post-LPS injection with decreased clinical severity scores, number of inflammatory cells, protein levels and levels of tumor necrosis factor (TNF)-α, nitric oxide (NO) and prostaglandin (PG) E2 in the aqueous humor (AqH) and degrees of histological ocular tissue injury. The anti-inflammatory potency of luteolin was comparable to that of Pred. Luteolin exhibited robust efficacy in the treatment of EIU in rats, indicating its potential clinical utility in treating uveitis.

Luteolin attenuates endotoxin-induced uveitis in Lewis rats.

The aim of the present study was to investigate the efficacy of luteolin on endotoxin-induced uveitis (EIU) in rats. EIU was induced in Lewis rats by subcutaneous injections of lipopolysaccharide (LPS). One hr before the LPS injection, 0.1, 1 or 10 mg/kg luteolin or 1 mg/kg prednisolone was intraperitoneally injected. We investigated its effect upon clinical scores, cellular infiltration and protein leakage, as well as on the level of tumor necrosis factor (TNF)-α, nitric oxide (NO) and prostaglandin (PG) E2 in the aqueous humor (AqH). Histologic examination and immunohistochemical analysis in the iris-ciliary body (ICB) were performed to determine the expressions of cyclooxygenase (COX)-2 and inducible NO synthase (iNOS), and then the activated nuclear factor (NF)-κB p65, I kappa B (IκB)-α degradation, phosphorylated (p)-IκB kinase (IKK) α/β and activator protein (AP)-1 c-Jun. Luteolin suppressed, in a dose-dependent manner, the clinical scores, number of inflammatory cells, the protein concentration, and the TNF-α, NO and PGE2 levels in the AqH and improved the histiologic status of the ocular tissue. Luteolin suppressed the expression of iNOS and COX-2 and the activated NF-κB p65, IκB-α degradation, p-IKKα/β and AP-1 p-c-Jun in the ICB. The anti-inflammatory potency of 10 mg/kg luteolin was as strong as that observed with 1 mg/kg prednisolone. These results demonstrate that luteolin attenuates ocular inflammation by inhibiting expression and release of inflammatory markers, along with the inhibition of the activated NF-κB pathway and at least partly AP-1 activity in the ICB.

Grouping annotations on the subcellular layered interactome demonstrates enhanced autophagy activity in a recurrent experimental autoimmune uveitis T cell line.

Human uveitis is a type of T cell-mediated autoimmune disease that often shows relapse–remitting courses affecting multiple biological processes. As a cytoplasmic process, autophagy has been seen as an adaptive response to cell death and survival, yet the link between autophagy and T cell-mediated autoimmunity is not certain. In this study, based on the differentially expressed genes (GSE19652) between the recurrent versus monophasic T cell lines, whose adoptive transfer to susceptible animals may result in respective recurrent or monophasic uveitis, we proposed grouping annotations on a subcellular layered interactome framework to analyze the specific bioprocesses that are linked to the recurrence of T cell autoimmunity. That is, the subcellular layered interactome was established by the Cytoscape and Cerebral plugin based on differential expression, global interactome, and subcellular localization information. Then, the layered interactomes were grouping annotated by the ClueGO plugin based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases. The analysis showed that significant bioprocesses with autophagy were orchestrated in the cytoplasmic layered interactome and that mTOR may have a regulatory role in it. Furthermore, by setting up recurrent and monophasic uveitis in Lewis rats, we confirmed by transmission electron microscopy that, in comparison to the monophasic disease, recurrent uveitis in vivo showed significantly increased autophagy activity and extended lymphocyte infiltration to the affected retina. In summary, our framework methodology is a useful tool to disclose specific bioprocesses and molecular targets that can be attributed to a certain disease. Our results indicated that targeted inhibition of autophagy pathways may perturb the recurrence of uveitis.

Effects of the immunomodulator, VGX‐1027, in endotoxin‐induced uveitis in Lewis rats

VGX-1027 is a novel, low molecular weight, immunomodulatory compound that has shown efficacy against a variety of immuno-inflammatory disease models in animals including autoimmune diabetes in NOD mice, collagen-induced arthritis and chemically induced inflammatory colitis. Here, we have studied the effects of VGX-1027 on the development of endotoxin-induced uveitis (EIU) in male Lewis rats, as a model of inflammatory ocular diseases in humans. EIU was induced by a single footpad injection of 200 microg lipopolysaccharide (LPS). Groups of rats were treated with either VGX-1027 (25 mg kg(-1)) or its vehicle at different time points (30 min, 6 h or 12 h) after the challenge with LPS or, as positive control, with dexamethasone. The rats were killed within 16 h after LPS challenge, and the eyes and aqueous humor were collected to study serological, immunological and histological signs of EIU. The rats treated with VGX-1027 within 6 h after LPS challenge exhibited milder clinical, histological and laboratory signs of EIU than those treated with vehicle. This study provides the first evidence that systemic treatment with VGX-1027 counteracts the uveitis-inducing effect of LPS in rats and suggests that this drug may have potential in the treatment of immuno-inflammatory conditions of the eye in humans.

Inhibitory Effects of Glucosamine on Endotoxin-Induced Uveitis in Lewis Rats

Glucosamine sulfate (GS) is a naturally occurring sugar that exerts immunosuppressive effects in vitro and in vivo. The authors investigated whether GS modulates the inflammatory reaction in endotoxin-induced uveitis (EIU) of rats and the mechanisms by which it exerts its effects. Two-hundred micrograms of lipopolysaccharide (LPS) was injected subcutaneously into Lewis rats to induce EIU. Doses of GS (10, 100, or 1000 mg/kg) were divided into three aliquots and administered intraperitoneally 30 minutes before LPS injection, concurrently with LPS injection, and 30 minutes after LPS injection. Twenty-four hours after LPS injection, aqueous humor was collected for cell counting and measurement of protein concentration. Immunohistochemical staining of the iris-ciliary body was performed to evaluate the effects of GS on intercellular adhesion molecule (ICAM)-1 and nuclear factor (NF)-kappaB activation and to demonstrate macrophage infiltration. The effects of various doses of GS pretreatment were also examined using a mouse macrophage cell line (RAW264.7 cells) and LPS stimulation. Levels of prostaglandin (PG)-E2 and nitric oxide (NO) were determined. Expression of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 were measured using Western blot analysis. The effect of GS on LPS-induced NF-kappaB activation in RAW cells was also examined. Cell counting and analysis of protein concentration in aqueous humor revealed that GS suppressed EIU in rats treated with a high dose of GS (1000 mg/kg). Immunohistochemistry showed that treatment with GS reduced ICAM-1 expression and suppressed activation of NF-kappaB in the iris-ciliary body. The main inflammatory cells in the iris-ciliary body during EIU were macrophages. In LPS-stimulated macrophage RAW cell culture, GS inhibited the production of NO and PG-E2, the expression of iNOS and COX-2, and the activation of NF-kappaB. GS suppresses EIU in rats by blockading the NF-kappaB-dependent signaling pathway and the subsequent production of ICAM-1 and proinflammatory mediators. This study has extended the authors' previous observation that GS is a potentially important compound for reducing ICAM-1-mediated inflammatory effects in the eye.

Inhibitory Effects of Lutein on Endotoxin-Induced Uveitis in Lewis Rats

Lutein deposits in the macula and lens of human eyes with high concentration and is well known as an eye-protective nutrient for its beneficial effects on eye disease such as age-related macular degeneration and cataract. The purpose of the present study was to investigate the effects of lutein on endotoxin-induced uveitis (EIU) in rats. EIU was induced in male Lewis rats by subcutaneous injection of 200 microg lipopolysaccharide. Lutein or dexamethasone was administered intravenously at 30 minutes before, at the same time as, and at 30 minutes after LPS treatment. The aqueous humor was collected at 24 hours after LPS injection, the number of infiltrating cells, the protein concentration, and the levels of nitric oxide (NO), tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, prostaglandin (PG)-E2, monocyte chemoattractant protein-1 (MCP-1), and macrophage inflammatory protein (MIP)-2 in the aqueous humor were determined. Immunohistochemical staining with a monoclonal antibody against activated nuclear factor (NF)-kappaB was performed to evaluate the effect of lutein on NF-kappaB activation in the iris-ciliary body (ICB) of rats. A mouse macrophage cell line (RAW264.7 cells) was stimulated with LPS in the presence or absence of lutein. Expression of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and degradation of inhibitor kappaB (IkappaB) were analyzed by Western blot analysis. Lutein suppressed the development of EIU in a dose-dependent fashion. The anti-inflammatory effect of 100 mg/kg lutein was as strong as that of 1 mg/kg dexamethasone. Treatment with lutein reduced the concentrations of NO, TNF-alpha, IL-6, PGE2, MCP-1, and MIP-2 in aqueous humor. Lutein also suppressed the activation of NF-kappaB in the ICB as well as iNOS and COX-2 expression and IkappaB degradation in RAW cells. These findings indicate that lutein has anti-inflammatory effects on EIU by inhibiting the NF-kappaB dependent signaling pathway and the subsequent production of proinflammatory mediators.

Immunostimulatory and immunomodulatory peptides derived from the α1 domain of HLA-B27 in experimental autoimmune diseases in Lewis rats

Peptides derived from amino acid sequence 60–80 of HLA-B27 (B27PA, aa 60–72 and B2702PA, aa 60–80) mimic cytokeratin and are able to induce in vitro proliferation of human peripheral blood lymphocytes as well as arthritis in Lewis rats. Here we show that the pathogenic epitope recognized by autoaggressive rat T cells is located at the N-terminus of the sequence, between aa 60 and 72. A C-terminally elongated 25mer peptide (B2702.60–84) showed increased pathogenicity, indicating either a second arthritogenic epitope or an immunomodulatory region within this peptide. B2702.60–84 has been described to inhibit murine and human CD8 + cytotoxic T cells (CTL) and was even successfully used for the treatment of allograft rejection. In addition to pathogenicity we have investigated the immunomodulatory effect of peptide B2702.60–84 in our rat model of experimental autoimmune uveitis (EAU), induced with retinal S-Antigen peptide PDSAg. We found that disease exacerbated following coimmunization of PDSAg with B2702.60–84. In vitro, the B27-peptide enhanced the proliferation of CD4 + T cell lines specific for retinal autoantigen peptides during coincubation of B2702.60–84 with the respective antigen. Oral tolerance induction, an effective mechanism to prevent uveitis in Lewis rats, is abrogated by cofeeding peptide B2702.60–84 with the tolerogen PDSAg. In rat EAU, naturally occurring regulatory T cells and orally induced γδTCR + suppressor cells are CD8 +, which might be impeded by peptide B2702.60–84. As a consequence of their abrogated suppressive capacity disease was exacerbated. We propose a similar role of HLA-B27 in man: disturbing the mechanisms down-regulating self-responses might lead to autoimmune diseases. This could explain the high association of HLA-B27 with a variety of autoimmune diseases targeting different organs or tissues.

Autoimmune uveitis induced by molecular mimicry of peptides from rotavirus, bovine casein and retinal S-antigen.

Antigenic mimicry of infectious agents and autoantigens is a proposed pathomechanism for autoimmune diseases. Here, we describe antigenic mimicry of a peptide from rotavirus, a nutritional protein from bovine milk (alphas2-casein) and a peptide thereof as well as a highly pathogenic peptide from retinal S-antigen (PDSAg), a major autoantigen in experimental autoimmune uveitis in Lewis rats. Immunization of rats with the peptides and the casein protein induced uveitis, an intraocular inflammation leading to decreased vision and even blindness. The peptides elicited cross-reactive T cell responses and uveitis in rats and were also recognized by lymphocytes and sera from uveitis patients. Oral tolerization with PDSAg, but not with rotavirus- and casein-derived peptides or casein protein, prevented PDSAg-induced uveitis in rats. Cofeeding of casein with cholera toxin induced uveitis in rats, suggesting that breaking oral tolerance to casein during gastrointestinal infections might also be able to initiate uveitis in humans.

Effects of cloricromene, a coumarin derivative, on endotoxin-induced uveitis in Lewis rats.

To investigate the effects of cloricromene, a coumarin derivative, in rats subjected to endotoxin-induced uveitis (EIU). Endotoxin uveitis was induced in male Lewis rats by a single footpad injection of 200 microg lipopolysaccharide (LPS). Cloricromene was topically applied to the rat eye twice at 1 hour before and 7 hours after injection of LPS. A separate group of animals was treated with vehicle. Rats were killed 16 hours after injection and the eyes enucleated for histologic examination and immunohistochemical analysis. The effect of treatment was also evaluated by slit lamp examination, by the number of intraocular inflammatory cells on histologic sections, and by measuring the protein and TNFalpha levels in the aqueous humor. Nitrite and nitrate production was also measured in the aqueous humor. The histopathology of the iris-ciliary body included inflammatory cell infiltration and nuclear modification of vessel endothelial cells. Cloricromene treatment reduced the inflammatory cell infiltration and improved histologic status of the ocular tissue. Immunohistochemical analysis for P-selectin, intracellular adhesion molecule (ICAM)-1, nitrotyrosine, and poly(ADP-ribose) synthetase (PARS) revealed a positive staining in inflammatory cell infiltration from LPS-treated rats. The degree of staining for P-selectin, ICAM-1, nitrotyrosine, and PARS was markedly reduced in tissue sections obtained from LPS-recipient rats that had received cloricromene. Cloricromene strongly inhibited cell infiltration, protein exudation, TNFalpha production, and nitrite-nitrate formation. This study provides the first evidence that cloricromene, a coumarin derivative, attenuates the degree of inflammation and tissue damage associated with EIU in rats.

Immunopathogenesis and prevention of uveitis with the Behçet's disease-specific peptide linked to cholera toxin B.

Behcet’s disease (BD) is a multisystemic disease affecting mucocutaneous, ocular, central nervous system, joint and vascular tissues. The aetiology of BD has been associated with a variety of microorganisms, especially Herpes simplex virus and some strains of Streptococcus sanguis. A common microbial agent may be involved, such as heat shock protein (HSP) which has significant homology with human cellular HSP. The evidence in favour of HSP as an aetiological factor in BD is based on T cell proliferative responses and B cell antibodies to the defined HSP60 epitope p336-351 in patients with BD in Britain, Japan and Turkey. The proportion of γδ T cells is significantly increased in BD and HSP65 or HSP70 upregulates γδ T cells. The peptide determinant defined in patients with BD elicits uveitis in Lewis rats when administered by the oral mucosal or systemic route. It is noteworthy that the MICA gene is a cell stress response gene and shares nucleotide sequences with the human HSP70 promoter. The cytokine and chemokine networks stimulated by HSP induce polarisation towards the TH1 cytokines during active disease. The evidence from these studies has converged towards the concept that the multi-system immunopathology of BD is generated by an over-reaction to microbial stress proteins. We now demonstrate that p336-351 induced uveitis in rats can be prevented by oral tolerization with the peptide linked to recombinant cholera toxin B subunit. Preliminary evidence in an open clinical trial suggests that this novel tolerizing strategy may be effective in preventing relapse of uveitis in BD.

Induction of arthritis and uveitis in Lewis rats by antigenic mimicry of peptides from HLA-B27 and cytokeratin.

Ankylosing spondylitis (AS) is highly associated with HLA-B27. We have previously shown that peripheral blood lymphocytes from AS patients respond to stimulation with a peptide from the sequence of HLA-B27. Here we report on molecular mimicry of peptides from HLA-B27 and cytokeratin, the latter being specifically expressed in synovial membranes and eyes, the main targets of the autoaggressive immune response in AS patients. Immunization of rats with these peptides induced an inflammatory response in joints, spine and eyes, resembling the symptoms in AS. Furthermore, both HLA-B27- and cytokeratin-derived peptides, are effective oral tolerogens: feeding these peptides ameliorated arthritis and uveitis induced with the cytokeratin peptide. Our model might elucidate the role of peptides from the sequence of HLA-B27 as an antigen of the immune response in AS, introducing a new aspect of antigenic mimicry between HLA-B27 and tissue-specific antigens. We propose this as a mechanism directing a systemic autoimmune response to specific target organs by antigenic mimicry of T cell epitopes.

Epitope recognition and T cell receptors in recurrent autoimmune anterior uveitis in Lewis rats immunized with myelin basic protein

Lewis rats immunized with myelin basic protein (MBP) develop experimental autoimmune encephalomyelitis (EAE) and associated anterior uveitis (AU). Rats recover and become resistant to further reinduction of EAE. We investigated whether the resistance to reinduction of EAE was associated with the resistance to AU in LEW rats reinjected with MBP. We demonstrated that while rats remained resistant to EAE, they become susceptible to uveitis after recovery, and suffered a second episode of disease. The susceptibility to reinduced disease was associated with the recognition of new MBP epitopes. In contrast to the initial episode of AU, TCR Vβ8.2 predominance was not observed in the iris/ciliary body. Our results suggest that T cells specific for MBP, which are rapidly reactivated when re-exposed to antigen, are sufficient to induce clinical uveitis in LEW rats. This process may involve a shifting of T cell specificity from the major encephalitogenic peptide utilizing the Vβ8.2 receptor to a more diverse cell repertoire.

Effect of chloroquine on experimental melanin-protein-induced uveitis in Lewis rats.

To evaluate the effect of chloroquine on experimental recurrent autoimmune melanin-protein-induced uveitis in Lewis rats. Five Lewis rats were immunized with melanin to induce uveitis (group I); another six were immunized and treated with 5 mg/kg/day of chloroquine (group II). To reinduce uveitis, they received two booster injections of melanin. They were killed when uveitis was present after the last booster injection; every eye was evaluated by means of clinical and histopathological examination. A control group of four rats received the same chloroquine dose to evaluate retinal toxicity (group III). Rats in group I showed more severe uveitis episodes than rats in group II. The second uveitis episode was more severe in group I than in group I. Rats in group III showed no clinical or histopathological abnormalities. Chloroquine decreased the severity and duration of uveitis, and in two rats it prevented recurrences. Thus, chloroquine can be considered an effective treatment for chronic, recurrent experimental uveitis.

Corticotropin-releasing hormone and inflammation.

Corticotropin-releasing hormone (CRH) is a major regulator of the hypothalamic-pituitary-adrenal axis (HPA) and principal coordinator of the stress response. As in stress, intracerebroventricular administration of CRH suppresses the immune system indirectly, via glucocorticoid and/or sympathetic system-mediated mechanisms. Also, during inflammatory stress, the cytokines TNF alpha, IL-1, and IL-6 stimulate hypothalamic CRH and/or vasopressin secretion as a way of preventing the inflammatory reaction from overreacting. Recently, CRH receptors were described in peripheral sites of the immune system, and CRH was found to promote several immune functions in vitro. We demonstrated a direct role of CRH in the inflammatory immune process in vivo, by first studying the effect of systemic CRH immunoneutralization in an experimental model of carrageenin-induced aseptic inflammation in Spague-Dawley rats. We extended these observations to other forms of experimental inflammation, including streptococcal cell wall polysaccharide- and adjuvant-induced arthritides and peptide R16 (epitope of the interphotoreceptor retinoid-binding protein)-induced uveitis in Lewis rats. We also studied human disease states, including rheumatoid arthritis, Hashimoto thyroiditis, and ulcerative colitis. Inflamed tissues contained large amounts of IR CRH, reaching levels similar to those observed in the hypophyseal portal system. We also demonstrated the presence of CRH mRNA and CRH receptors in inflammatory cells and identified the mast cells as a major immune target for CRH. In addition to production by immune cells, the peripheral nervous system, including the postganglionic sympathetic neurons and the sensory fibers type C, appears to contribute to IR CRH production in inflammatory sites. The production of CRH from the postganglionic sympathetic neurons may be responsible for the stress-induced activation of allergic/autoimmune phenomena, such as asthma and eczema, via mast cell degranulation. Antalarmin, a novel nonpeptide CRH receptor antagonist, displaced 125I-labeled ovine CRH binding in rat pituitary, frontal cortex, and cerebellum, but not heart, consistent with antagonism at the CRHR1 receptor. In vivo antalarmin significantly inhibited CRH-stimulated ACTH release and carrageenin-induced subcutaneous inflammation in rats. Thus, antalarmin and other related compounds that antagonize CRH at the level of its own receptor have therapeutic potential in some forms of inflammation directly mediated by type 1 CRH receptors and promise to enhance our understanding of the many roles of CRH in immune/inflammatory reactions.

Recognition of B‐Cell Epitopes of the 65 kDa HSP in Behçet's Disease

B-cell epitopes of the mycobacterial 65 kDa heat shock protein (HSP) were mapped in sera from patients with Behçet's Disease (BD). A series of 47 overlapping synthetic peptides (15ers) derived from the sequence of the Mycobacterium tuberculosis 65 kDa HSP was used in ELISA. Significant increases in IgA and IgG antibody levels were observed with peptides 111-125, 154-172 and 311-326 in sera from BD, compared with those from controls. Homologous peptides derived from the sequence of the human mitochondrial 60 kDa HSP were then examined. Peptides 136-150 and 336-351 showed comparable results to the homologous mycobacterial peptides 111-125 and 311-326, respectively. The B-cell epitopes defined in this investigation overlap with the T-cell epitopes the authors have previously reported in BD. Inhibition studies are consistent with the view that antibodies to each of the three B-cell epitope peptides represent a small proportion of the total B-cell epitope repertoire elicited by the 65 or 60 kD HSP. Sequential antibody studies suggest that IgA and IgG antibody titres to one or all three peptides tested may increase during exacerbation of ocular disease. The functional role of these antibodies needs to be determined, but the peptides may be involved in the immunopathogenesis of BD as they can induce experimental uveitis in Lewis rats, which is a principal manifestation of BD.

The effect of chlorpromazine on endotoxin-induced uveitis in the Lewis rat.

Chlorpromazine (CPZ) has been used extensively in the treatment of psychiatric disorders, and has recently been shown to possess systemic anti-inflammatory properties as well. To investigate the potential effects of CPZ on ocular inflammation, we evaluated its action on endotoxin-induced uveitis (EIU) in Lewis rats. At three different dosage levels, CPZ produced highly significant reductions in the mean aqueous aspirate inflammatory cell counts and histological inflammatory scores as compared to controls treated with vehicle only. Analysis of aqueous fluid demonstrated a similar decrease in protein concentration and phospholipase A2 (PLA-2) activity in the treated animals. The ability of CPZ to inhibit the development of EIU may be related to its properties as a calcium channel blocker and inhibitor of the enzyme phospholipase A2.

Generation of chemiluminescence in experimental autoimmune uveitis.

Using experimental model of uveitis, the inflammation-associated chemiluminescence was measured by luminol amplification method. The induction of uveitis in Lewis rats was carried out by immunizing bovine S-antigen in complete Freund's adjuvant on hind foot-pad. The chemiluminescence activity from six sets of retina and choroid measured at the peak of inflammation was found to be in the range of 300,000-400,000 counts, amounting to nearly 20-fold increase from non-immunizated control animals under the same conditions. The phagocyte-derived superoxide anion and hydroxyl radical appear to be the two major radicals in the chemiluminescent species. This was revealed by in vitro suppression of chemiluminescence by 5,400 units of superoxide dismutase and 10 mM of D-mannitol. The decreases in counts were 21% and 35% for superoxide dismutase and D-mannitol respectively. The presence of hydrogen peroxide was not established since catalase with dose up to 20,000 units did not cause any significant suppression in counts. In the time sequence studies covering the entire course of uveitis development, the level of chemiluminescent species at day 9 postimmunization increased to 1.5-fold that of day 0, continued to increase to a maximum of 18-fold on day 12 and decreased slowly to about 3-fold in day 19. The pattern of increase appears to coincide with our previous findings in the number of polymorphonuclear leukocytes, the extent of membrane lipid peroxidation and the degree of retinal degeneration. Thus, these radicals play an important role in initiation as well as perpetuation of the membrane oxidative processes that lead to retinal degeneration.

Effect of iron chelation on severity of ocular inflammation in an animal model.

Metabolites of oxygen-free radicals generated by polymorphonuclear leukocytes and macrophages are believed to inflict the initial tissue damage in acute inflammations. Of the various oxygen products, hydroxyl radicals are known to be potent toxic agents, and their generation depends largely on the presence of free iron. Treatment of experimental uveitis in Lewis rats with an iron chelator, deferoxamine mesylate, resulted in marked reduction in choroidal inflammation and suppression of retinal damage. These findings suggest that in experimental uveitis the severity of ocular inflammation and tissue damage may be mediated by the iron-catalyzed generation of hydroxyl radicals, and deferoxamine may thus serve as an anti-inflammatory agent.