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J Liversidge

Publications and source records attributed to J Liversidge.

At least 37 records · Page 2Linked to original sources
PubMed1994

Rat retinal pigment epithelial cells express an inducible form of nitric oxide synthase and produce nitric oxide in response to inflammatory cytokines and activated T cells.

In this report we show that rat retinal pigment epithelial (RPE) cells express an inducible form of nitric oxide synthase (iNOS) and secrete high levels of nitric oxide (NO.) when co-cultured with activated lymphocytes. We have previously shown that cultured rat RPE cells suppress syngeneic lymphocyte proliferation, an effect attributed to prostaglandin E2 (PGE2) secretion by the RPE cells. However supernatants from such co-cultures were also found to contain high levels of nitrite (NO2-), the stable end-product of NO. synthesis. RPE cell secretion of NO. was stimulated by the cytokines interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha), an effect enhanced by endotoxin [lipopolysaccharide (LPS)], reduced by the competitive inhibitor of L-arginine metabolism, NG-monomethyl-L-arginine (L-NMMA) and inhibited by cycloheximide. These effects were dose dependent. Using reverse transcription (RT)/PCR a product of 1398 bp was amplified which showed sequence identity with iNOS cloned from rat vascular smooth muscle. Northern blot analysis of total RNA extracted from rat RPE before and after cytokine stimulation showed induction of a 4.5 kb (kilobase) transcript which hybridized with a 1398 bp (base pair) polymerase chain reaction (PCR)-generated cDNA probe derived from the sequence of rat RPE cell iNOS. These results indicate RPE cells express an inducible form of nitric oxide synthase (NOS) and that high levels of NO. may be produced locally in the eye by the RPE in the presence of activated lymphocytes. Given the cytostatic and cytotoxic properties of this molecule, NO. may play an important role as an inducible mediator of immunosuppressive mechanisms within the microenvironment of the eye at the site of lymphocyte activation.

Amino Acid Oxidoreductases↗
PubMed1994

ICAM-1/LFA-1 interactions in T-lymphocyte activation and adhesion to cells of the blood-retina barrier in the rat.

To identify the signals involved in the adhesion and subsequent migration of lymphocytes across the endothelium (REC) and pigment epithelium (RPE) of the blood-retina barrier we have studied the effects of monoclonal antibodies (mAb) to rat adhesion/accessory molecules on the binding of normal and concanavalin A (Con A)-activated rat spleen lymphocytes to cultured unstimulated and interferon-gamma (IFN-gamma)-stimulated RPE and REC. Forty to 48% of unactivated T cells were found to bind to normal REC or RPE by leucocyte function-associated antigen-1/intercellular adhesion molecule-1 (LFA-1/ICAM-1)-independent mechanisms, despite constitutive expression of ICAM-1 by the RPE cells and LFA-1 by the T cells. Con A-activated lymphocytes showed an enhanced adhesion to both RPE and REC. However, IFN-gamma-stimulated RPE and REC did not demonstrate a significant increase in adhesiveness for normal lymphocytes highlighting the importance of lymphocyte integrin activation from low-affinity to high-affinity state. Activated lymphocyte adhesion to unstimulated RPE and REC was significantly blocked by LFA-1 mAb (35%, P < 0.0001) and ICAM-1 mAb (20%, P < 0.001). Inhibition of adhesion by antibody to CD2 was not significant. Both ICAM-1 and LFA-1 mAb also significantly (P < 0.05) blocked antigen presentation following retinal extract stimulation of lymphocytes from immunized rats in proliferation assay. These data suggest that the ICAM-1/LFA-1 system is important in lymphocyte trafficking into the eye only after lymphocyte activation.

Animals↗
PubMed1994

Intranasal administration of retinal antigens suppresses retinal antigen-induced experimental autoimmune uveoretinitis.

Bovine retinal extract (RE) is a heterologous mixture of highly uveitogenic proteins including S-Antigen (S-Ag), interphotoreceptor retinol binding protein (IRBP) and rhodopsin, and is a potent inducer of experimental autoimmune uveoretinitis (EAU). Intranasal inoculation of Lewis rats with RE performed daily for 10 days prior to immunization with RE suppresses both the severity and the incidence of the clinical response and histopathological changes in EAU. Significant suppression of the disease in treated animals could be achieved with a total (cumulative) intranasal inoculum of 42 micrograms of antigen. Animals which were treated with extract exhibited a normal total antibody response to S-Ag, IRBP and retinal extract when compared with controls [phosphate-buffered saline (PBS) treated] animals. The antibody response in tolerized animals was predominantly anti-S-Ag IgG2a with suppression of anti-S-Ag IgM response. Treated animals had a significantly suppressed delayed-type hypersensitivity (DTH) response to retinal extract but normal response to purified protein derivative (PPD) compared to control animals. Adoptive transfer of splenocytes from treated animals also demonstrated some protection against RE-induced EAU. These results demonstrate that tolerance induction impairs the onset and severity of EAU by inhibiting the DTH response to heterologous mixture of retinal antigens.

Administration, Intranasal↗
PubMed1994

Localization and characterization of major histocompatibility complex class II-positive cells in the posterior segment of the eye: implications for induction of autoimmune uveoretinitis.

PURPOSE: To identify potential antigen-presenting cells in the choroid and retina of the normal rat eye, with a view to proposing a role for such cells in the induction and perpetuation of experimental autoimmune uveoretinitis, a model of human uveoretinal inflammation. METHODS: Immunohistochemical and electron microscopic studies using a panel of monoclonal antibodies were performed on frozen sections of the perfused-fixed normal Lewis rat eye, choroid whole mounts, and cytospin preparations of cells harvested from choroid/ciliary body explant cultures. In addition, time-lapse video recordings of migratory uveal tract cells in culture were taken. RESULTS: No major histocompatibility complex class II-positive cells were found in the normal Lewis rat retina. However, at least three populations of potential antigen-presenting cells were found in the uveal tissues of the eye: classical dendritic cells expressing high levels of major histocompatibility complex class II antigen; resident dendritiform macrophages, which were negative for major histocompatibility complex class II antigen, but expressed specific macrophage markers (ED2); and blood-borne macrophages (ED1) that had emigrated from the vasculature into the tissue compartment. In addition there were small numbers of cells expressing novel markers such as markers usually found only on macrophage subsets in splenic tissue (ED3) and a recently described marker for veiled dendritic cells (OX62). Dendritic cells and resident dendritiform macrophages closely interacted with each other and with tissue cells, particularly retinal pigment epithelial cells. CONCLUSIONS: The posterior uveal tract is richly populated with classical dendritic cells expressing constitutive high levels of major histocompatibility complex class II antigen. There are also several types of macrophages with the potential to modulate immune responses in the posterior segment. Interactions among these cells and with resident tissue cells such as retinal pigment epithelial cells are probably central to the initiation of (auto)immune responses in the posterior segment of the eye.

Animals↗
PubMed1993

Retinal pigment epithelial cells modulate lymphocyte function at the blood-retina barrier by autocrine PGE2 and membrane-bound mechanisms.

Retinal pigment epithelial cells (RPE) in vitro display many characteristics typical of antigen-presenting cells (APC) yet are poor inducers of lymphocyte proliferation and in vivo appear to remain MHC class II negative within the acute inflammatory chorioretinal lesions associated with experimental autoimmune uveoretinitis. In an in vitro assay system designed to mimic the in vivo situation as closely as possible, RPE cells profoundly suppressed lymphocyte proliferation to antigen, mitogen, and IL-2 even in the presence of exogenous APC. RPE immunosuppression was found to comprise both soluble and membrane-bound components and to be reversible as lymphocytes could be restimulated by thymocytes after coculture on RPE cells. Indomethacin restored proliferative responses of cocultured lymphocytes to specific antigen and mitogen indicating that the soluble component of suppression was mediated in part by PGE2 production by the RPE cells. Constitutive production of PGE2 by RPE cells was found to be low (approximately 10(-9) M), but increased to high levels (approximately 10(-6) M) in coculture with activated lymphocytes. In addition, antigen-specific primed lymphocyte responses during coculture with RPE were enhanced in the presence of indomethacin compared to similar splenocyte responses in the absence of RPE cells. However, indomethacin did not reverse RPE inhibition of IL-2-driven lymphocyte proliferation, and the failure of lymphocyte proliferation could not be attributed to TGF beta production by RPE cells, indicating that other RPE cell-immunosuppressive mechanisms were involved. Trypsin pretreatment of RPE cell monolayers removed an as yet unidentified inhibitory factor present on the RPE cell membrane and in combination with indomethacin enabled RPE cells to function as antigen-presenting cells.

Animals↗
PubMed1993

Nasal administration of retinal antigens suppresses the inflammatory response in experimental allergic uveoretinitis. A preliminary report of intranasal induction of tolerance with retinal antigens.

Current immunotherapy of posterior uveitis is non-specific and limited by drug toxicity and unpredictable relapses on therapy. Alternative modes of therapy being investigated using the rat model of experimental autoimmune uveoretinitis (EAU) have included the induction of tolerance with oral administration of milligram quantities of retinal antigens. In this preliminary report we demonstrate that tolerance to retinal antigens can be induced via the upper respiratory tract with microgram doses of antigen, preventing subsequent induction of EAU.

Animals↗
PubMed1993

Adhesion molecule expression in acute and fibrotic sympathetic ophthalmia.

Samples of iris ciliary body, choroid and retina from normal eyes and from 2 cases of sympathetic ophthalmitis (one acute and one late stage fibrosis) were examined for the expression of the VLA integrins beta 1 and alpha 1-6, and the integrin beta 3, in addition to ICAM-1, VCAM-1, ELAM-1 and CD44 using an APAAP staining technique. The expression of VLA-4, VLA-5, VCAM-1, ICAM-1, and CD44 was significantly increased and ELAM-1 was slightly increased in acute sympathetic ophthalmitis in comparison to fibrotic and normal eyes. VLA-6 was moderately increased in acute and fibrotic cases and VLA-2 VLA-3 and beta 3 were moderately expressed on all tissues examined. The differential expression of molecules known to be involved in lymphocyte activation and adhesion in acute sympathetic ophthalmitis suggests that certain adhesion molecules play a role in the pathogenesis of intraocular inflammation and may be suitable targets for immunotherapy.

Acute Disease↗
PubMed1993

Retinal antigen specific lymphocytes, TCR-gamma delta T cells and CD5+ B cells cultured from the vitreous in acute sympathetic ophthalmitis.

CD5+ B lymphocytes and TCR gamma-delta T lymphocytes, phenotypes implicated in the pathogenesis of autoimmune disease, were isolated from the vitreous in a case of acute sympathetic ophthalmitis. These cells were obtained using a method which allows the selective maintainance in vitro of in vivo activated T lymphocytes. Dual colour flow cytometry showed that after 3 days culture in IL-2 containing medium 61% of cells were CD5/CD19 + ve and 41% CD3/TCR gamma delta + ve. Of the total CD3 + ve population, 15% were gamma/delta negative. These cells formed a population which also responded in a proliferation assay to retinal antigens. Histologically the eye showed a marked mononuclear cell infiltration of the retina, ciliary body and choroid. Granulomatous lesions within the choroid contained lymphocytes, plasma cells and multinucleate giant cells. Immunocytochemistry showed lymphocyte populations to be predominantly CD2 + ve CD3 + ve T lymphocytes of the CD4 sub-set. Distribution of monocytes/macrophages throughout the lesions and restriction of B-lymphocytes to granulomata were all consistent with a DTH type reaction. Despite immunosuppressive therapy, the expression of activation antigens HLA-DR and ICAM-1 on infiltrating and resident ocular tissue cells was high, although IL-2 receptor (CD25) expression was virtually absent. Flow cytometric analysis of peripheral blood cells prior to treatment with Cyclosporin-A showed systemic activation of lymphocytes, with high levels of HLA-DR and CD25 expression and a raised CD4/CD8 ratio.

Antigens↗
PubMed1992

Immunocytochemical analysis of blood lymphocytes in uveitis.

We studied the surface expression of activation markers IL2-R, HLA-DR and CD45-RO on peripheral T-lymphocytes in two groups of patients (n = 26) with idiopathic uveoretinitis, compared with controls. Thirteen patients were analysed by alkaline phosphatase anti-alkaline phosphatase (APAAP) immunocytochemistry, which demonstrated a significant rise in expression of HLA-DR and IL2-R surface markers. Flow cytometric analysis was performed on a further 13 patients, which confirmed a significant rise in IL2-R expression in uveitis patients. Within this group systemic activation was confined to patients with idiopathic retinal vasculitis. Dual flow cytometry confirmed a CD4+,IL2--R+ T--lymphocyte phenotype. A further 4 patients with retinal vasculitis who had been treated with cyclosporin A demonstrated a 32% reduction in IL2-R expression over a 3-month period. Analysis of CD45-RO and CD5+ cells was found to be uninformative in this study. We have demonstrated activated peripheral lymphocytes in patients predominantly with retinal vasculitis, the significance of which is discussed.

Biomarkers↗
PubMed1992

Experimental autoimmune uveoretinitis: a model system for immunointervention: a review.

Experimental autoimmune uveoretinitis (EAU) is a useful model of human posterior uveitis and as such, permits the analysis of strategies for immuno-intervention. Modulation of the autoimmune response may be attempted at the stages of induction of EAU, during homing of autoreactive lymphocytes to the target organ, the retina, or during the effector stage of the disease. This paper presents a brief overview of current immuno-therapeutic modalities and assesses the usefulness for extrapolation to human disease.

Animals↗
PubMed1992

Antigen processing and presentation in the eye: a review.

The recognition event between self-antigen, the MHC and the T cell receptor has become the target for potential immunotherapy of T cell mediated autoimmune disease. For this approach to succeed in uveoretinitis, uniformity in T cell receptor usage, restricted MHC usage and limited epitope recognition by individuals would be required. In this study we have shown that despite clonal heterogeneity of response to multideterminate retinal extract antigens, proliferation to the antigens tested was restricted by the IA MHC class II molecule. Different patterns of reactivity to retinal antigens in the presence of various protease inhibitors was observed. Natural processing of IRBP appears to be complex, requiring a number of enzymes to generate immunogenic fragments, in contrast, for S-antigen, plasmin alone may suffice. The RPE cells which are potential processors and presenters of retinal antigens produce PGE2 and may act as suppressors of ocular inflammation.

Animals↗
PubMed1992

Interphotoreceptor retinoid binding protein induced experimental autoimmune uveitis: an immunophenotypic analysis using alkaline phosphatase anti-alkaline phosphatase staining, dual immunofluorescence and confocal microscopy.

Using a Lewis rat model of IRBP induced EAU, we have examined the progress of leucocytes infiltrating the uveitic eye. APAAP and dual immunofluorescence were used to show that ED7 and 8 (CD11b/CD18) positive monocytes, W3/25 and OX8 (CD4 and CD8) lymphocytes were prominent in the initial inflammatory exudate around the retinal vessels and in the choroid. ED1 positive monocytes were also observed in the choroid. As disease progressed, these cells moved into the inner retina, vitreous and ROS. ED8 positive cells were the first to appear in the ROS. This was followed by the later appearance of ED2 tissue macrophages in the vitreous and ED3 inflammatory macrophages in the vitreous and ROS.

Alkaline Phosphatase↗
PubMed1990

Comparison of clinical and experimental uveitis.

Uveitis is a term which encompasses many clinical syndromes which would appear to be discrete entities. Both clinically and experimentally, the separation of uveitis affecting only the anterior segment from that affecting the posterior segment has a sound pathogenetic basis. However, clear distinctions among the various forms of endogenous posterior uveitis are more difficult to maintain in the light of evidence from experimental models of autoimmune uveitis (EAU). EAU can be induced by a variety of retinal antigens and each antigen has been shown to induce somewhat different forms of EAU, depending on such factors as dose of antigen, species and strains of animal model, and the type(s) of adjuvant used. However, within each model a similar spectrum of uveoretinal responses can be induced by each antigen suggesting that the pathogenetic mechanisms are probably similar also. In addition, if these models are analogous to human disease, then each clinical entity within this apparently heterogeneous group of clinical posterior uveitis syndromes may represent one aspect of a general organ-specific uveoretinal response to autoantigens.

Adjuvants, Immunologic↗
PubMed1990

Are accessory molecules implicated in activation of T cells at the blood-retinal barrier?

Co-culture of rat retinal pigment epithelial (RPE) cells or retinal capillary endothelial (EC) cells with T lymphocytes from immunised syngeneic rats indicated that RPE cells but not EC could process retinal S-antigen and induce antigen specific T cell activation. Activation of lymphocytes by EC however was also antigen independent, and may be due to T cell (CD2) endothelial cell (LFA-3) ligand interactions which are functional on EC but not RPE.

Animals↗
PubMed1990

Regulation of the local immune response by retinal cells.

Experimental autoimmune uveoretinitis (EAU) induced by retinal antigens is a CD4+ (Th) lymphocyte mediated disease. Generation of autoreactive CD4+ cells requires the processing and presentation of autoantigen by antigen presenting cells (APC) in combination with MHC Class II antigen. Efficient presentation of antigen to T cells has also been shown to depend on accessory molecules of adhesion such as intercellular adhesion molecule-1 (ICAM-1) and leukocyte function-associated antigen-1 (LFA-1). Aberrant expression of Class II antigens by local tissue cells has been suggested as a possible mechanism in autoimmune processes. Several ocular cells express Class II antigens during inflammation, while other cells such as Muller cells inhibit antigen presentation in vitro. We have also shown that retinal pigment epithelial cells (constitutively) and endothelial cells (after induction) express ICAM-1 and that CD4+ lymphocyte adhesion to these cells is inhibited by antibodies to ICAM-1. Accessory molecules may therefore be important, not only in local presentation of antigen but in recruitment of circulating autoreactive cells to the eye since these cells represent the site of the blood-retinal barrier. Regulation of the local immune response in the eye therefore, may occur at several levels.

Animals↗
PubMed1990

Interactions between lymphocytes and cells of the blood-retina barrier: mechanisms of T lymphocyte adhesion to human retinal capillary endothelial cells and retinal pigment epithelial cells in vitro.

We have studied the adhesion of human CD4+ lymphocytes to cultured human retinal vascular endothelial cells (EC) and human retinal pigment epithelial cells (RPE), both of which comprise the cellular components of the blood-retina barrier. We have observed differences in the lymphocyte-RPE and the lymphocyte-EC interactions. Firstly, RPE cells were found to express high levels of the adhesion molecule ICAM-1 constitutively, whereas EC expressed ICAM-1 only after induction with IFN-gamma. In addition, lymphocyte binding to normal and minimally stimulated RPE (5 U/ml, 4 hr) was predominantly ICAM-1 dependent, but after maximal stimulation (500 U/ml, 4 days), increased lymphocyte adhesiveness included an ICAM-1-independent component, which was apparently not due to involvement of MHC class II or CD2 molecules. In contrast, binding of lymphocytes to unstimulated EC involved both an ICAM-1-dependent and an ICAM-1-independent mechanism, the latter being subject to inhibition by monoclonal antibody to CD2. Studies of adhesion at 4 indicated that no binding occurred to normal or stimulated RPE, but binding to EC was observed, albeit reduced to 50% of the 37 binding level, and this implies that the LFA-3/CD2 adhesion pathway may also be involved in lymphocyte binding to EC. Overall, the results indicate a functional difference between RPE and EC affecting T-cell adhesion, migration and activation at the blood-retinal barrier, which must be considered when devising therapies to prevent lymphocyte infiltration of the eye.

Antibodies, Monoclonal↗