Macrophages and MHC class II positive cells in the choroid during endotoxin induced uveitis

AIMS/BACKGROUND—Endotoxin induced uveitis has been regarded as a model for acute anterior uveitis and until now little was known about choroidal involvement. The aim of this study was to investigate changes in macrophages and MHC class II positive cells in the choroid of Lewis rats during endotoxin induced uveitis.METHODS—Choroid-sclera wholemounts were isolated from normal Lewis rats and at different time points—4, 8, 16, 24, 48, 72, and 96 hours, and 7, 10, and 14 days after a footpad injection of 200 µg of lipopolysaccharide (LPS). Immunohistochemistry was performed using the monoclonal antibodies ED1 (monocytes, macrophages, dendritic cells), and OX6 (MHC class II antigen).RESULTS—In normal rats, two layers of macrophages were identified in the choroid; a layer located immediately beneath the retinal pigment epithelium (RPE) and a layer bordering the sclera. The density of ED1 positive cells in the layer bordering the RPE cells was 902 (SD 132) cells/mm² whereas the scleral layer had a cell density of 389 (73) cells/mm². Based on morphology, positive cells could be divided into two main categories; pleomorphic/round cells and dendritiform cells with varying appearances, with the latter being predominant in normal eyes. A network of MHC class II positive dendritic cells was found in the choroid, beneath the RPE, with a density of 659 (96) cells/mm². No MHC class II positive cells were found in the macrophage layer bordering the sclera. LPS injection caused a massive influx of ED1 positive macrophages in the area below the RPE cells but did not result in an influx of macrophages at the scleral side of the choroid. The infiltrate reached a maximum at 16 hours following LPS injection and decreased at 96 hours. The morphology of the infiltrating cells was pleomorphic/round at early stages of inflammation and changed into a dendritiform cell population later. The number of MHC class II positive cells on the anterior side of the choroid increased 8 hours after injection and reached a peak at 72-96 hours. MHC class II positive cells were not observed in the vicinity of the sclera at any time after LPS injection. Both resident and MHC class II positive dendritic cell numbers returned to normal values at day 14 following LPS injection.CONCLUSIONS—These results indicate that the choroid is severely inflamed after systemic LPS administration to Lewis rats and suggests that endotoxin induced uveitis may serve as a model for generalised uveitis in humans.     

The influence of penetrating keratoplasty and cyclosporin A therapy on MHC class II (Ia)-positive cells in the rat iris and choroid

Background: The presence of Ia-positive cells (MHC class II equivalent) has been previously reported in the iris and choroid of various species. They have been reported to have both round and dendritic morphologies; the latter may represent classic dendritic cells, potent antigen-presenting cells (APCs). It is possible that the dendritic-like cells play a important role in (auto)immune processes of uveal and other ocular tissues. Using the flat or whole mount technique, the distribution of Ia-positive cells in the rat iris and choroid was investigated following penetrating Keratoplasty (PKP) and following treatment with cyclosporin A (CsA). Methods: Lewis (LW) rats received corneal buttons from Lewis-Brown Norway (LW-BN) donors and were randomly assigned to the following groups: (i) operated, untreated (n=24); (ii) operated, CsA-treated (10 mg/kg i.m.;n=22). Controls were groups (iii) normal LW rats (n=13); (iv) unopcrated, CsA-treated (16 days' treatment;n=8); (v) anterior perforation of the anterior chamber (n=3); (vi) eight corneal sutures only (n=4); (vii) syngeneic operated (LW to LW;n=4). Animals of groups (i) and (ii) were killed on the 5th, 9th and 13th postoperative days and on appearance of the corneal rejection (group i, day 13; group ii, day 16). Both eyes were enucleated, immediately fixed, and iris-choroid flat mounts were examined for Ia-positive cells using APAAP immunohistochemistry. Results: In the normal Lewis rat iris, scattered Ia-positive cells of both nondendritic and dendritic morphology were observed. CsA treatment in the unoperated rat did not result in a significant decrease in the percentage of dendritic cells in the iris or choroid. Anterior chamber perforation, the placement of sutures in the cornea and syngeneic PKP resulted in a moderate increase in iris Ia-positive cells. Allogeneic transplantation resulted in a large increase in both types of Ia-positive cells, particularly on day 13 with corneal rejection. In group ii, an initial decrease in Ia-positive cells until day 13 was observed; upon rejection (day 16), the histological picture was similar to that of untreated animals. Alterations in the operated choroid were also apparent following CsA treatment. Conclusion: Corneal transplantation in the Lewis rat results in an increase in Ia-positive cells in the iris; CsA therapy can delay but not prevent this reaction. Changes in choroidal Ia-positive cells following PKP were not apparent, their numbers being affected only by CsA treatment following grafting.     

Induction of MHC class II antigen in cultured bovine ciliary epithelial cells

The expression of major histocompatibility complex (MHC) class II antigens in cultured bovine ciliary epithelial cells was investigated by means of indirect immunohistochemistry and immunocytofluorometry. Ciliary epithelial cells grown in control tissue-culture medium did not express MHC class II. However, after incubation with bovine gamma-interferon (IFN-G) in concentrations as low as 0.3 units/ml, nearly all cells stained for MHC class II. Tumor necrosis factor increased IFN-G-induced MHC class II expression. A reduction in IFN-G-induced MHC class II expression was observed with dexamethasone, prostaglandin EZ and alpha-interferon. To test whether MHC class II expression in response to IFN-G was specific for the ciliary epithelium, several intraocular tissues were grown in culture and incubated with IFN-G. MHC class II expression was observed in all tissues tested for response to IFN-G, but at different sensitivities. Retinal pigment epithelium and ciliary epithelium exhibited the highest sensitivity, followed by corneal endothelium and lens epithelium; the lowest sensitivity was observed for retinal vascular pericytes. The results are discussed in the context of MHC class II expression on the ciliary epithelium in anterior uveitis.     

Expression of MHC class II antigens and immunoglobulin M by the corneal epithelial cells in herpetic keratitis

The corneal buttons obtained from 4 patients with active epithelial and stromal herpetic keratitis were studied with routine microscopy and immunohistochemistry. We used an immunoperoxidase technique with monoclonal antibodies directed against Langerhans cells, lymphocyte subsets, MHC products and immunoglobulins A, G, M and D. The epithelium and stroma contained an inflammatory infiltrate composed of polymorpho-nuclear leukocytes, dendritic cells, B-lymphocytes and T-lymphocytes (helper/inducer and suppressor/cytotoxic subsets). The epithelial cells of all the corneal buttons expressed MHC class II antigens. IgM was bound to the membrane of the epithelial cells in 3 specimens. HSV antigenic material was localized in the epithelial cells and in the stromal keratocytes by a direct immunofluorescence technique. Our data suggest that cell-mediated as well as antibody-mediated immune responses are involved, with a possible role for an autoimmune mechanism in the pathogenesis of this condition.This work was supported in part by the program of scientific and clinical co-operation in the field of ophthalmic medicine and surgery between the University of Mansoura, Arab Republic of Egypt, and the Catholic University, Leuven, Belgium.     

Nonvascular smooth muscle alpha-actin positive cells in the choroid of higher primates

PURPOSE: As an elaborated network of non-vascular smooth muscle alpha-actin positive cells (NVSMC) is present in the human eye, the purpose of this investigation was to describe the presence of NVSMC in the sclera and choroid of different mammals and primates. METHODS: Whole mounts and tangential serial sections through the sclera and choroid of human donor eyes, of cynomolgus and rhesus monkey, mangabey, owl monkey, tree shrew, pig, cow, rabbit, cat, dog, and rat eyes were stained with a mouse-anti-smooth muscle alpha-actin antibody. The mean number/mm( 2) of NVSMC was calculated in the central region of each quadrant. RESULTS: In cynomolgus and rhesus monkey eyes, numerous NVSMC were located in the suprachoroidal and inner scleral region. In the temporal quadrant, the density of NVSMC was highest. NVSMC were semicircular arranged around the entering long ciliary artery and nerve. In the mangabey and owl monkey choroid, the number of NVSMC was about ten-fold less than in macaque monkeys. Most of the NVSMC were located parallel to large vessels and only some cells were seen in the suprachoroid and inner sclera. In the tree shrew monkey and in all other mammal eyes, no NVSMC could be detected. In the human eyes, in addition to the suprachoroidal and inner scleral NVSMC numerous cells were located in the choroid proper. The regional distribution and the orientation of the NVSMC was comparable in human and macaque monkey eyes. CONCLUSIONS: Comparing these findings with the literature, the presence of NVSMC correlates with the development of a central fovea and an elaborated accommodative system. Its functional significance remains to be determined.     

虹膜脉络膜在闭角型青光眼中的作用机制研究进展

PACG是一种常见的不可逆的致盲性疾病。以往认为其特征性房角关闭的危险因素包括浅前房、短眼轴、厚晶状体等静态解剖因素,还包括虹膜动态变化,脉络膜膨胀等房角动态因素。随着各项眼科影像学技术的发展,人们能更细致地观察到眼球各解剖结构在房角关闭中的作用。其中,虹膜及脉络膜在闭角型青光眼中起到的作用已越来越为人们所重视,借助眼前节OCT（AS-OCT）等各种眼科影像学技术,我们对前房角关闭的机制有了新的认识,可进一步深入阐明虹膜脉络膜与房角关闭之间的联系,为闭角型青光眼早期诊断和个体化治疗做出指导。     

Down-regulation of MHC class II expression on bovine retinal pigment epithelial cells by cytokines.

To determine the effect of interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta) on the down-regulation of MHC class II antigens, bovine retinal pigment epithelial (RPE) cells were incubated with interferon-gamma (IFN-gamma) in different concentrations. Subsequently, the IFN-gamma pretreated RPE cells were cultured with TGF-beta or IL-10 in distinct concentrations and treatment modi. About 10% of native (totally untreated) RPE cells were positive for MHC class II antigens detected with immunocytostaining. Under the influence of IFN-gamma (1,000 U/ml), the number of MHC class II-bearing cells increased to 49.9 +/- 4.5% positive cells after 8 days' incubation. Expression decreased under the most effective TGF-beta treatment (5 ng/ml) to 2.0 +/- 0.9% after 24 h incubation, and under similar IL-10 treatment (200 U/ml) to 3.8 +/- 1.4% after 72 h. This decreasing number of the MHC class II-positive cells may be useful for various eye research studies and, more especially, to RPE cell transplantation in the future.     

Cutaneous melanoma metastasizing to the iris and choroid: a case report

BACKGROUND. Cutaneous melanoma is a primary tumour that rarely produces intraocular metastases. We report on a 47-year-old woman who was referred to our outpatient department because of anisocoria. METHODS. A cutaneous melanoma of the upper left arm had been excised 21 months ago with subsequent polychemotherapy for lymph node involvement. Visual acuity and intraocular tension were normal. The left iris showed a brownish, parapupillary, prominent nodule. In both eyes, multiple circumscribed whitish choroidal sub-pigment epithelial tumours were observed. RESULTS. General metastasis to lymph nodes, pleura, lungs, mucosa and skin was diagnosed. The patient died 2 months later. CONCLUSION. Metastases of cutaneous melanoma to the iris are extremely rare. The ophthalmologist may be helpful in monitoring treatment by quantitative documentation at follow-up.     

Effects of topical carteolol and timolol on tissue circulation in the iris and choroid.

PURPOSE: To evaluate the effects of topical carteolol or timolol on tissue circulation in the iris and posterior choroid. METHODS: After a topical instillation of 20 microl of 2% carteolol, 0.5% timolol, or physiological saline (for control) into one eye, and physiological saline into the other eye of pentobarbital-anesthetized Dutch pigmented rabbits, normalized blur value; a quantitative index of tissue blood velocity in the iris (NB(iris)) and posterior choroid (NB(cho)) was obtained using the laser-speckle method. Intraocular pressure (IOP), blood pressure and pulse rate were serially monitored for 2 hours after instillation. Using separate groups of rabbits, NB(iris) and IOP were measured before and after 20-day twice-daily unilateral treatment of carteolol or timolol. RESULTS: After a single instillation of carteolol, NB(iris) was significantly greater only in the treated eyes than control eyes (P = 0.0050, repeated measures two-way ANOVA), while NB(cho) showed no significant change. IOP in the treated eyes significantly reduced (P = 0.0005). Bilateral reductions of tissue vascular resistance in the iris were found after carteolol instillation (P = 0.0183 approximately 0.0322). After timolol instillation, serial changes in NB(iris) and NB(cho) in the treated eyes were significantly different from those in control eyes (P = 0.0129, 0.0031), while there were no significant differences at any of time points (Mann-Whitney test); IOP in both eyes was significantly reduced (P = 0.0096 approximately 0.0005); tissue vascular resistance in the iris and posterior choroid showed no significant changes. After 20-day treatment, NB(iris) in the both eyes of carteolol-treated rabbits significantly increased from the baseline (P = 0.0280, 0.0425, Wilcoxon signed rank test) and NB(iris) in timolol-treated eyes significantly decreased (P = 0.0280). CONCLUSIONS: A single instillation of topical carteolol significantly increased the iris tissue blood velocity in the treated eye and reduced the tissue vascular resistance in both eyes. Topical timolol tended to decrease tissue blood velocity in the iris and choroid of the treated eye, but showed no significant effects on tissue vascular resistance in the both tissues.     

Mucinous adenocarcinoma metastatic to the iris, ciliary body, and choroid.

A 67-year-old woman presented with signs of severe intraocular inflammation and secondary glaucoma. The initial diagnosis was uveitis, and an anterior chamber paracentesis with cytological study of the aspirate failed to establish an aetiological diagnosis. After three trabeculectomies had failed to control the intraocular pressure, the blind eye was enucleated. On histopathological examination a mucinous adenocarcinoma was found to cover diffusely the iris surface and to involve the ciliary body and peripheral choroid. The patient subsequently developed evidence of widespread metastatic disease and died shortly thereafter. Although a primary tumour was never found, histochemical and immunohistochemical studies of the enucleated eye suggested that the lesion originated in the gastrointestinal tract. In cases of intractable glaucoma and anterior chamber inflammation, metastatic carcinoma should be included in the differential diagnosis, and efforts should be made to substantiate the diagnosis by a systemic examination or a biopsy.     

Density and distribution of dendritiform cells in the peripheral cornea of healthy subjects using in vivo confocal microscopy

PurposeTo establish in a large healthy cohort, dendritiform cell (DC) density and morphological parameters in the central and peripheral cornea using in vivo confocal microscopy (IVCM).MethodsA prospective, cross-sectional, observational study was conducted in 85 healthy volunteers (n = 85 eyes). IVCM images of corneal center and four peripheral zones were analyzed for DC density and morphology to compare means and assess correlations (p < 0.05 being statistically significant).ResultsCentral corneas had lower DC density (40.83 ± 5.14 cells/mm²; mean ± SEM) as compared to peripheral corneas (75.42 ± 2.67 cells/mm², p < 0.0001). Inferior and superior zones demonstrated higher DC density (105.01 ± 7.12 and 90.62 ± 4.62 cells/mm²) compared to the nasal and temporal zones (59.93 ± 3.42 and 51.77 ± 2.98 cells/mm², p < 0.0001). Similarly, lower DC size, field and number of dendrites were observed in the central as compared to the average peripheral cornea (p < 0.0001), with highest values in the inferior zone (p < 0.001 for all, except p < 0.05 for number of dendrites in superior zone). DC parameters did not correlate with age or gender. Inter-observer reliability was 0.987 for DC density and 0.771–0.922 for morphology.ConclusionIn healthy individuals, the peripheral cornea demonstrates higher DC density and larger morphology compared to the center, with highest values in the inferior zone. We provide the largest normative cohort for sub-stratified DC density and morphology, which can be used in future clinical trials to compare differential changes in diseased states. Furthermore, as DC parameters in the peripheral zones are dissimilar, random sampling of peripheral cornea may be inaccurate.