T cell activation within different intraocular compartments during experimental uveitis.
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Animals↗
Biomedical subjects
S W Cousins
Publications and source records attributed to S W Cousins.
Identification of alpha-melanocyte stimulating hormone as a potential immunosuppressive factor in aqueous humor.
The aqueous humor of the eye contains factors that regulate immunological responses within the immunosuppressive ocular microenvironment. Besides TGF-beta, the proteins in the low molecular weight (< 3500 Da) fraction of normal aqueous humor are also immunosuppressive. The low molecular weight fraction of aqueous humor inhibits IFN-gamma production and proliferation of antigen-stimulated lymph node cells. Neuropeptides are one possible family of low molecular weight factors in aqueous humor. Through the utilization of an antigen capturing enzyme-assay, the immunosuppressive neuropeptide alpha-melanocyte stimulating hormone (alpha-MSH) was detected in normal aqueous humor of humans, rabbits, and mice. The mean concentration of alpha-MSH in normal aqueous humor of humans was 20 +/- 3 pM, of rabbits 11 +/- 1 pM, of BALB/c mice 16 +/- 3 pM, and of C57BL/6 mice 14 +/- 3 pM. These physiological concentrations of alpha-MSH inhibited the production of IFN-gamma by antigen-stimulated lymph node cells. In contrast to the low molecular weight fraction, alpha-MSH did not inhibit proliferation. There was a 26% recovery of IFN-gamma production when alpha-MSH was absorbed from the low molecular weight fraction. The results demonstrate neuropeptides to be constitutive components of normal aqueous humor and that factors with the capability of differential regulation of effector T-cell activity may be present within the immunosuppressive ocular microenvironment.
Animals↗
T cell depletion increases susceptibility to murine cytomegalovirus retinitis.
To study the effect of immunosuppression on the development of murine cytomegalovirus (MCMV) retinitis, BALB/c mice were immunosuppressed with methylprednisolone (a corticosteroid) and/or with antibodies against CD4+ and CD8+ T cells and inoculated with low-dose MCMV (5 x 10(2) plaque-forming units) by the supraciliary route. Nonimmunosuppressed mice inoculated with low-dose MCMV by the supraciliary route did not develop necrotizing retinitis. By contrast, 78-100% of immunosuppressed mice developed retinitis after inoculation of low-dose MCMV. To study the effect of depletion of individual T cell subsets, mice were depleted of either CD4+ or CD8+ T cells and inoculated with low-dose MCMV by the supraciliary route. The frequency of retinitis in CD4-depleted mice (30%) was not significantly different from that of nonimmunosuppressed control mice (0%). The frequency of retinitis in the CD8-depleted group (80%) was similar to that observed in mice immunosuppressed with corticosteroid alone (90.9%), with antibodies to both T cell subsets (100%), or with steroid and both T cell subset antibodies (100%). These results support the conclusion that the CD8+ T cell subset is responsible for control of ocular MCMV infection. Furthermore, these results suggest that the CD8+ T cell subset may be important in preventing ocular CMV infection in immunosuppressed patients.
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Quantitative assessment of growth stimulating activity of the vitreous during PVR.
This report postulates that the activity of cellular proliferation along the surface of the retina during proliferative vitreoretinopathy (PVR) is reflected in the aggregate effect of proliferation-inducing growth factors in the vitreous. A method for quantifying the "net" proliferation-inducing capacity of an individual vitreous sample was developed and this assay was used to evaluate the vitreous proliferative activity in a model for experimental PVR. Vitreous was aspirated sequentially after onset of fibroblast-induced PVR in a rabbit model. A simple bioassay for the "aggregate" stimulating activity was developed and each sample was assigned a quantitative value in terms of "proliferation units" (PU). Experimental eyes demonstrated a wide range of stimulating activity (0-1765 PU), but control eyes showed uniformly low levels of activity (0-337 PU). Experimental eyes that ultimately developed retinal detachment displayed higher levels of proliferative activity than did those eyes destined to remain attached. The differences were statistically different by day 3, prior to the onset of clinical retinal changes. We conclude that quantification of vitreous proliferation-stimulating activity is possible and that this method might be useful for screening eyes at high risk for the development of recurrent retinal detachment from PVR.
Animals↗
Fate of the fellow eye after Propionibacterium acnes endophthalmitis.
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Cataract Extraction↗
Immune privilege and suppression of immunogenic inflammation in the anterior chamber of the eye.
Immunologic privilege of the anterior chamber has been associated with the capacity to induce a unique form of deviant systemic immunity after anterior chamber (AC) immunization. However, the capacity of privilege to suppress expression of immunity in the AC has not been examined. We studied the ability of the AC to sustain immunogenic inflammation after direct antigen challenge (delayed hypersensitivity-DH) in C57BL/6 mice primed to M tuberculosis (MT) antigens. Compared to subcutaneous and subconjunctival sites where primed mice demonstrated vigorous and significant DH, the anterior chambers of these mice failed to develop signs of inflammation unless toxic doses of antigen were injected. In an attempt to promote intraocular DH, the AC's of MT-primed mice were pre-treated with subinflammatory intracameral injections of IFN-gamma, a cytokine that antagonizes TGF-beta, recruits antigen presenting cells (APC) from the blood and activates resident APC precursors. It was observed that AC injection of IFN-gamma, followed 3 days later by AC challenge with 200 ng of MT, resulted in severe intraocular inflammation only in primed (but not naive) mice. We conclude that the normal mouse AC resists DH unless its immunosuppressive microenvironment is abolished, as in these experiments by IFN-gamma. We propose that impaired expression of cell-mediated immunity is an important component of immune privilege of the AC.
Animals↗
Retinitis in euthymic mice following inoculation of murine cytomegalovirus (MCMV) via the supraciliary route.
Cytomegalovirus (CMV) retinitis is the most frequent infectious ocular complication of the acquired immune deficiency syndrome (AIDS). Currently, there are few animal models to study the virologic and immunologic factors which contribute to the pathogenesis of CMV retinitis. In these experiments, 1-2 X 10(4) PFU of murine cytomegalovirus (MCMV) was inoculated into the supraciliary space of BALB/c mice. Within three days of inoculation, moderate iridocyclitis was observed which progressed to necrosis of the ciliary body by day 14. Approximately 60% of the mice developed typical retinitis characterized by virus-infected cytomegalic cells in the retina and retinal pigment epithelium, focal retinal infiltrates, transition zones between uninvolved and involved retina, and optic neuritis. The remaining animals exhibited atypical retinitis characterized by non-specific retinal inflammation in the absence of obvious viral infection. This murine model of CMV retinitis shares some features with retinitis observed in AIDS patients with CMV retinitis and may be useful to evaluate the efficacy of immunologic and/or pharmacologic treatment strategies for CMV retinitis.
Animals↗
Studies of tumor-infiltrating lymphocytes from a human choroidal melanoma.
Cell suspensions prepared enzymatically from an ocular choroidal melanoma were cultured in vitro in an effort to generate (1) melanoma tumor cell lines and (2) tumor-infiltrating lymphocytes cytotoxic for ocular melanoma cells. Even though histologic study of the tumor did not show "significant" infiltrating bone marrow-derived cells, lymphocytes were generated readily in cultures to which interleukin-2 was added. Phenotypic analysis of the cultured lymphocytes indicated that T-cells, natural killer (NK) cells, and lymphokine-activated killer (LAK) cells were present. Moreover, functional studies of the cultured lymphocytes revealed NK activity, LAK activity, and most importantly, tumor antigen-specific cytotoxic T-cell activity. It was concluded that it is possible to obtain tumor cell lines and tumor-infiltrating lymphocytes from ocular tumors, both of which would be required if cellular immunotherapy of ocular tumors is contemplated. In addition, these results indicate that ocular melanomas can express unique tumor-specific antigens and that the immune system of a patient with such an ocular tumor can perceive these tumor antigens because antigen-specific precursor cytotoxic T-cells were present in the tumor-containing eye at the time of enucleation. The theoretic and therapeutic implications of these findings are discussed.
Adult↗
Identification of transforming growth factor-beta as an immunosuppressive factor in aqueous humor.
The anterior chamber of the eye is an immunologically privileged site. Over the past 15 years, numerous laboratories documented that the privileged status of this unique site is mediated by multifactorial immunoregulatory processes. Among the participating factors is the aqueous humor that circulates in the anterior chamber and is in contact with most of the tissues in the anterior segment of the eye. Recently, it was found that normal aqueous humor is a powerful inhibitor of antigen-driven T lymphocyte activation, but it spares other important functional properties of T cells. The spectrum of immune inhibitory properties resembles some of the activities of transforming growth factor-beta (TGF-beta), a polypeptide cytokine. Because of this similarity, the authors tried to determine if TGF-beta is present in aqueous humor and whether this cytokine could account for the lymphocyte inhibitory activity of this biologic fluid. They found TGF-beta in aqueous humor by dot-blot analysis. Using the CCL-64 mink lung epithelial cell bioassay for this compound, TGF-beta bioactivity was shown in aqueous humor from several different species, including human. In rabbit and human aqueous humor, most of the biologic activity was due to TGF-beta 2 (80-90%). Dose-response curves generated by using purified porcine TGF-beta showed that aqueous humor contained sufficient concentrations of TGF-beta to account for the observed inhibition in several assays for T cell activation and proliferation. Partial purification of the lymphocyte inhibitor in rabbit aqueous humor by size exclusion in high-performance liquid chromatography demonstrated that several lymphocyte inhibitory fractions contained TGF-beta bioactivity. Finally, neutralizing antisera to TGF-beta 2 were able to reverse most of the lymphocyte inhibitory activity of aqueous humor. It was concluded that TGF-beta was present in high concentration in normal aqueous humor and that this cytokine contributed to the immunosuppressive properties of aqueous humor.
Animals↗
Macroaneurysms associated with retinal branch vein occlusion.
We reviewed the clinical findings, color photographs, and fluorescein angiograms of 147 patients (154 involved eyes) who participated in the Branch Vein Occlusion Study through the Bascom Palmer Eye Institute for the presence of macroaneurysms within or adjacent to the zone of branch vein occlusion. Four distinct categories of macroaneurysms included arterial, venous, capillary, and collateral-associated types. We identified 59 lesions among 25 of the 154 (16.2%) affected eyes. Intraretinal lipid exudation and hemorrhage were commonly observed around the lesions, but these complications usually did not affect visual acuity. Macroaneurysms were associated with capillary nonperfusion on fluorescein angiography. Retinal neovascularization occurred in eight of 25 eyes (32%) with macroaneurysms.
Aged↗
Aqueous humor factors and their effect on the immune response in the anterior chamber.
The immune response to antigens within the anterior chamber is deviant (anterior chamber associated immune deviation - ACAID) in that delayed hypersensitivity is deficient, whereas other immune effector modalities are intact. Experimental evidence indicates that the eye itself is critical to the induction of ACAID. We have examined the antigen processing and presenting potential of cells within the anterior segment of the eye, and have analyzed the potential immunoregulatory properties of these cells, their secretory products, and the aqueous humor itself. Evidence indicates that bone marrow-derived cells within the stroma of the iris and ciliary body inhibit antigen-driven T lymphocyte activation, although they themselves lack the capacity to present antigens to T lymphocytes. The mechanism is in part through secretion of immunosuppressive cytokines. Since aqueous humor contains similar cytokines, it is inferred that these molecules are constitutively secreted. We have determined that a major inhibitory molecule within normal aqueous humor is transforming growth factor-beta (TGFB), which inhibits antigen processing and presentation, and suppresses both T lymphocyte activation and certain aspects of non-specific inflammation. These effects also turn out to be properties of normal aqueous humor. These findings support the hypothesis that local features of the eye modify intraocular antigens such that an ACAID-inducing signal is produced. Experimental evidence suggests that these same properties may play a major role in suppressing efferent immune responses in the eye.
Animals↗
Flow cytometric detection of lymphocyte proliferation in eyes with immunogenic inflammation.
Identification and analysis of antigen-activated lymphocytes from sites of ocular inflammation is important to understanding of the role of infiltrating immune effectors in ocular inflammatory disease. Unfortunately, few assays can distinguish activated lymphocytes from the antigen-irrelevant lymphocytes that randomly migrate to sites of inflammation. The authors describe a method that allows the quantitative recovery and identification of activated lymphocytes from mouse eyes. Recovered subconjunctival lymphocytes were simultaneously stained for specific cell-surface markers (with fluorescein-labeled antibodies) and for DNA content (with propidium iodide), then analyzed by flow cytometry. For any subpopulation of lymphocytes, the relative percentage and absolute number of cells in each phase of the cell cycle was determined. The activated subset of T and B cells was quantitated by determining the number of proliferating cells (ie, the number of cells in the S + G2 + M phases of the cell cycle). Using this method, T cell proliferation was found to be a specific component of the subconjunctival immunogenic inflammatory response induced by a local graft-versus-host reaction and by subconjunctival delayed hypersensitivity reactions. In addition, proliferating T cells could be distinguished from nonproliferating B cells produced by the subconjunctival inoculation of a T cell tumor.
Animals↗
Schisis contributes to necrosis in experimental HSV-1 retinitis.
Injection of the KOS strain of herpes simplex virus type 1 into the anterior chamber of one eye of a BALB/c mouse produces bilateral retinal inflammation which progresses in the contralateral eye to total retinal necrosis by day 10 post-inoculation. Although both immune and direct viral cytopathic effects have been proposed as causes of the initial infiltration, the precise mechanism(s) of the contralateral retinal necrosis remains uncertain. In this study, histopathological examination of the uninoculated eyes of HSV-1 injected BALB/c mice revealed the presence of vitreous traction against the internal limiting membrane of the retina which was associated with separation of the nerve fiber layer from the underlying ganglion cell layer. This separation resulted in avulsion of Mueller cell footplates and retinal vessels, hemorrhage into the schisis cavity and disruption of the underlying retinal architecture. The schisis began focally on day 8 or 9 post-inoculation, and the entire retina of most animals was involved by day 10 or 11. These results suggest that retinal schisis involving separation of the nerve fiber layer from the ganglion cell layer contributes to necrosis of the retina in the uninoculated contralateral eye.
Animals↗
Herpes simplex retinitis in the mouse. Clinicopathologic correlations.
BALB/c mice inoculated with herpes simplex virus type 1 in one anterior chamber develop retinitis with subsequent retinal necrosis in the contralateral eye. The clinical features of this disease were characterized with indirect ophthalmoscopy and fundus photography. Three phases of the disease were delineated: (1) acute retinitis; (2) retinal necrosis; and (3) resolution; the clinical findings in each phase were correlated with the microscopic features. In addition, we compared the virus titers from individual animals with and without clinical evidence of retinitis to demonstrate that retinal inflammation correlated with the titer of virus in the uninoculated eye. This animal model is useful for the analysis of the clinical, virologic and immunologic manifestations of herpetic retinitis.
Animals↗
HSV-1 retinitis and delayed hypersensitivity in DBA/2 and C57BL/6 mice.
Following uniocular anterior chamber (a.c.) inoculation of HSV-1 into BALB/c mice, the architecture of the uninoculated eye is destroyed by a pan-retinal inflammatory process within 10 days. BALB/c inoculated with HSV-1 via the a.c. route also develop anterior chamber associated immune deviation (ACAID), one characteristic of which is impairment of virus-specific delayed hypersensitivity (DH) responsiveness. To determine whether other strains of mice develop ACAID and contralateral retinitis, DBA/2 and C57BL/6 mice were inoculated with HSV-1 (KOS strain) via the a.c. route. At intervals after inoculation, animals were examined clinically for evidence of retinitis in the uninoculated eye and were either (1) sacrificed for virus recovery studies or (2) ear-challenged for DH assays. Seven of 15 DBA/2 mice had clinical evidence of retinitis in the uninoculated eye by day 10 post-inoculation (p.i.). At this time, the average titer of virus in the uninoculated eyes was 3.37 log10 PFU/ml. Animals of this strain also had significantly impaired HSV-1 virus-specific DH responses consistent with ACAID. No (0/12) C57BL/6 developed contralateral retinitis; at day 10 p.i., the average virus titer in the uninoculated eyes of C57BL/6 mice was 1.10 log10 PFU/ml. Following a.c. inoculation of HSV-1, C57BL/6 mice developed vigorous DH responses comparable to those of subcutaneously immunized, DH-positive animals. The results of these experiments suggest that different strains of mice vary in their susceptibility to HSV-1 retinitis and that the development of retinitis is linked to the amount of virus in the uninoculated eye.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Endotoxin-induced uveitis in the rat: observations on altered vascular permeability, clinical findings, and histology.
A single intraperitoneal dose of endotoxin (lipopolysaccharide or LPS) induces an acute inflammatory response in the uveal tract of rats. This inflammation is characterized by a breakdown of the blood/aqueous barrier within 3 hr after the LPS and the subsequent development of clinical disease and a cellular infiltrate. Early change in vascular permeability, clinical, and pathological changes were dose dependent with the two highest doses (100 micrograms or 500 micrograms) producing more severe pathology. Clinical and histopathologic abnormalities peaked at 24 hr and were resolving by 48 hr. Although clinical and histologic changes correlated well, the degree of breakdown of the blood/aqueous barrier at 3 hr failed to predict the extent of the cellular exudate measured by either clinical or histologic criteria. In addition, pharmacologic suppression of the early vascular permeability changes with indomethacin, cyproheptadine, or both agents failed to protect the animals consistently from subsequently developing significant clinical disease or cellular infiltrates on histopathology. LPS-induced uveitis in the rat provides a simple, reproducible model for ocular inflammation without requiring direct eye manipulation. The mediators responsible for the early vascular permeability in this model appear to be distinct from the mediators primarily responsible for the subsequent cellular exudate.
Animals↗
Uveitis and arthritis: experimental models and clinical correlates.
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Animals↗
Ocular albumin fluorophotometric quantitation of endotoxin-induced vascular permeability.
Bacterial endotoxin (lipopolysaccharide; LPS) is known to alter systemic vascular permeability, but this effect is difficult to monitor and quantitate in vivo. The ocular vessels of the rabbit are particularly sensitive to LPS. Using a slit lamp equipped with a fluorophotometer, we have adapted a method to quantitate endotoxin-induced ocular vascular permeability by measuring the accumulation of fluorescein isothiocyanate-conjugated albumin into the anterior chamber of the eye. After intravenous administration of Salmonella typhimurim LPS, the anterior chamber fluorescence and blood fluorescence were measured at intervals of 15 min and 1 h, respectively, over 4 h. In controls, maximal fluorescence in the anterior chamber was 3.1 +/- 0.8% of blood fluorescence. Doses of LPS as low as 0.25 mug/kg produced an ocular/serum fluorescence ratio of 17.6 +/- 4.9. A dose of 2.5 mug of LPS per kg tended to produce a higher ratio (68.0 +/- 7.1) than a larger dose of 50 mug/kg (30.5 +/- 16.6). Permeability changes began within 30 min after LPS, and the rate of dye accumulation varied over time, with maximal leakage usually occurring 90 min after LPS, but occasionally occurring much later. Repeated doses produced tolerance. By conjugating albumin to rhodamine and utilizing a second filter with the slit lamp to measure accumulation of this dye, we demonstrated the persistence of marked permeability during a period when intraocular fluorescein isothiocyanate and albumin levels were relatively constant. This methodology indicates that extremely low doses of LPS induce ocular permeability changes and that neither the time course nor the dose response of this effect is linear. Ocular fluorophotometry is a sensitive, noninvasive technique to study the dynamics and pharmacology of LPS-induced permeability changes.
Animals↗