bromochloroacetic-acid and Retinal-Detachment

To develop a large animal model of proliferative vitreoretinopathy (PVR) in the swine to eventually study disease pathophysiology, as well as novel therapies.. PVR was induced in domestic swine by creation of a posterior vitreous detachment, creation of a retinal detachment by the injection of subretinal fluid, and intravitreal injection of green fluorescent protein-positive retinal pigment epithelial (GFP+ RPE) cells. Control eyes had the same surgical procedures without RPE cell injection. PVR was clinically graded on days 3, 7, and 14. Animals were euthanized on day 14, and enucleated eyes were analyzed by light microscopy and immunohistochemistry.. Injection of GFP+ RPE cells into the vitreous cavity produced localized, traction retinal detachments by day 14 in all eyes (14 of 14); in contrast, the retina spontaneously reattached by day 3 and remained attached in all control eyes (10 of 10). Contractile epiretinal membranes on the inner retinal surface that caused the traction retinal detachments consisted predominantly of GFP+ RPE cells. These cells stained positive for cytokeratin, confirming their epithelial origin, and also expressed α-SMA and fibronectin, markers for myofibroblasts and fibrosis, respectively.. We established a swine PVR model that recapitulates key clinical features found in humans and, thus, can be used to study the pathophysiology of PVR, as well as new novel therapies. GFP+ RPE cells injected into the vitreous cavity formed contractile membranes on the inner retinal surface and caused localized traction retinal detachments.

Topics: Actins; Animals; Disease Models, Animal; Epiretinal Membrane; Female; Fibronectins; Fibrosis; Green Fluorescent Proteins; Keratins; Luminescent Agents; Myofibroblasts; Retina; Retinal Detachment; Retinal Pigment Epithelium; Swine; Vitreoretinopathy, Proliferative

To observe the changes in N-cadherin expression in the retina after experimental retinal detachment (RD) and reattachment in the rat and to explore the role N-cadherin might play after RD.. Forty rat retinas were detached by transscleral injection of 1.4% sodium hyaluronate into the subretinal space. The eyes were enucleated at different time intervals (n = 5), followed by fixation, embedding, and sectioning. The differences in N-cadherin expression in the normal retina, detached retina, and spontaneously reattached retina were determined. Furthermore, an N-cadherin antagonist was injected in combination with 1.4% sodium hyaluronate into the subretinal space in another 10 eyes, in an attempt to demonstrate the role N-cadherin plays after RD.. N-cadherin was not expressed in the RPE layer of the normal rat retina. After RD, intense immunolabeling of N-cadherin was seen in the RPE cells, the photoreceptors, and the outer limiting membrane (OLM). An increasing number of cytokeratin (CK)-positive cells likely to be RPE cells was found attached to the outer surface of the detached neural retina. Where the retina was reattached, the N-cadherin immunolabeling rapidly decreased. In eyes treated with an N-cadherin antagonist, the retinas appeared thinner than that in eyes without treatment, and the photoreceptor nuclei showed significantly loss. Moreover, CK-positive cells attached to the outer surface of the detached retina were markedly fewer in number.. Increased expression of N-cadherin in the RPE cells, the photoreceptor cells, and the OLM of the retina after RD may contribute to RPE cell migration and photoreceptor survival. These changes could be reversed by retinal reattachment.

Topics: Animals; Bruch Membrane; Cadherins; Disease Models, Animal; Female; Fluorescent Antibody Technique, Indirect; Keratins; Male; Photoreceptor Cells, Vertebrate; Pigment Epithelium of Eye; Rats; Rats, Sprague-Dawley; Retinal Detachment

Endothelin one (ET-1) is a vasomodulator peptide that plays a role on ocular blood flow, glial proliferation, and collagen matrix contraction by retinal pigmented epithelial (RPE) cells. Both glial and RPE cells have been involved in the formation of epiretinal membranes (ERMs). This investigation was conducted to determine whether ET-1 may be associated with ERMs, either idiopathic (IERMs) or from proliferative vitreoretinopathy (PVR).. Plasma and vitreous samples were collected from patients classified by the presence of PVR membranes, retinal detachment (RD), and other ocular conditions, such as IERMs, that made the patients candidates for vitrectomy. Immunoreactive endothelin one (IR-ET-1) was tested in plasma and vitreous by radioimmunoassay. Immunoreactive-ET-1 was localized in IERMs and PVR membranes immunohistochemically. Expression of endothelin receptors A (ETA) and B (ETB) was confirmed by reverse transcription-polymerase chain reaction.. IR-ET-1 levels in plasma and vitreous were higher in patients with PVR and in patients with RD than in those of the control group. Eyes with IERMs also showed higher IR-ET-1 levels than the control group cases. IR-ET-1 levels in eyes with PVR were higher than those in eyes with IERMs. IR-ET-1 levels in eyes with RD were also higher than those of eyes with IERMs. Immunoreactive ET-1 was localized in the cellular and stromal components of both IERMs and PVR membranes. Furthermore, ETA and ETB receptors were expressed in both IERMs and PVR membranes.. IR-ET-1 in human vitreous is elevated in PVR, RD, and IERMs. ET-1 and its receptors ETA and ETB are present in epiretinal tissue of both idiopathic and PVR membranes. These data suggest an involvement of ET-1 in retinal disease.

Topics: Adult; Aged; Aged, 80 and over; Endothelin-1; Epiretinal Membrane; Female; Glial Fibrillary Acidic Protein; Humans; Keratins; Male; Middle Aged; Radioimmunoassay; Receptor, Endothelin A; Receptor, Endothelin B; Retinal Detachment; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Vitreoretinopathy, Proliferative; Vitreous Body

To characterize the extracellular matrix and cellular components of a 'snowbank' removed during vitreous surgery for treatment of retinal detachment complicating pars planitis.. The 'snowbank' was examined using immunohistochemical techniques and a panel of monoclonal and polyclonal antibodies directed against glial fibrillary acidic protein (GFAP), cytokeratin, alpha smooth muscle actin, tenascin, laminin, fibronectin, and collagen types I, II, and III.. The 'snowbank' was acellular except on the uveal side where there were cytokeratin-positive retinal pigment epithelial cells. There were no cells positive for the glial cell marker GFAP and the myofibroblast cell marker alpha smooth muscle actin. The extracellular matrix of the 'snowbank' contained tenascin and collagen types I, II, and III. There was no immunoreactivity for laminin and fibronectin.. These results on the immunohistochemical components of the 'snowbank' may be useful in clarifying the nature of the chronic inflammatory process in pars planitis. They indicate extensive tissue repair and remodeling, leading to major loss of function.

Topics: Actins; Antibodies, Monoclonal; Child; Collagen; Extracellular Matrix; Extracellular Matrix Proteins; Female; Fibronectins; Glial Fibrillary Acidic Protein; Humans; Immunoenzyme Techniques; Keratins; Laminin; Lens, Crystalline; Pars Planitis; Retinal Detachment; Scleral Buckling; Tenascin; Vitrectomy

To report an unusual case of intravitreal inflammation in a human eye caused by the presence of residual perfluorodecalin in a case of giant retinal tear and retinal detachment.. The posterior capsule of the lens, which was infiltrated with deposits, was collected during surgery. The specimen was stained with hematoxylin and eosin, with periodic acid-Schiff, and for melanin. Part of it was examined with electron microscopy. Immunohistochemical staining was performed to demonstrate CD68 antigens, cytokeratin, and glial fibrillary acid protein.. Vacuolated macrophages and retinal pigment epithelial cells infiltrated the posterior capsule. Electron microscopy showed the presence of membrane-lined vacuoles within the macrophages. A monolayer of epithelial cells covered the cellular infiltration.. Residual perfluorodecalin can induce an intraocular chronic macrophage response.

Topics: Adult; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Cryosurgery; Endophthalmitis; Eye Diseases; Fluorocarbons; Glial Fibrillary Acidic Protein; Humans; Immunoenzyme Techniques; Keratins; Lens Capsule, Crystalline; Macrophages; Male; Phacoemulsification; Reoperation; Retinal Detachment; Retinal Perforations; Scleral Buckling; Vitrectomy; Vitreous Body

Hepatocyte growth factor/scatter factor (HGF/SF) possesses mitogenic, motogenic, and morphogenic properties and has recently been implicated in various retinal diseases. The role of HGF/SF in proliferative vitreoretinal disease was investigated.. Sections of epiretinal membranes were stained immunohistochemically for cytokeratins, to identify HRPE cells, and for HGF/SF receptor (c-Met). Cultured HRPE cells were stained for c-Met and investigated for shape change in response to HGF/SF, by using image analysis. The dose-response relationship for HRPE cells to HGF/SF was investigated by a cell migration assay and the specificity of this response evaluated by a neutralization experiment. Subretinal fluid (SRF) and vitreous from patients with retinal detachment and proliferative vitreoretinopathy (PVR) plus vitreous from eyes obtained after death, eyes with macular hole, and eyes with proliferative diabetic retinopathy (PDR) were investigated for the presence of HGF/SF using an enzyme-linked immunosorbent assay (ELISA). HGF/SF activity was measured using an MDCK cell scatter assay.. HRPE cells in epiretinal membranes and in culture expressed c-Met. Cultured HRPE cells responded to HGF/SF by an epithelial-to-mesenchymal shape change and by cell migration, a response that increased with increasing concentrations of HGF/SF. This response was reduced in the presence of neutralizing antibody. There was evidence of HGF/SF in increasing concentrations in more severe PVR and in PDR when measured by ELISA, and, conversely, there was evidence of correspondingly decreasing HGF/SF activity when measured by MDCK cell scatter assay in these diseases.. HGF/SF is present in normal and pathologic vitreous. HRPE cells respond by shape change and cell migration to HGF/SF. Concentrations of HGF/SF increase in proliferative vitreoretinal disease and increase in turn with increased severity of the disease, but HGF/SF bioactivity decreases (consistent with activator depletion). These findings are consistent with the hypothesis that HGF/SF may play a role in the HRPE mesenchymal transformation that typifies PVR.

Topics: Adult; Aged; Aged, 80 and over; Animals; Cell Line; Cell Movement; Cell Size; Dogs; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Epiretinal Membrane; Hepatocyte Growth Factor; Humans; Immunoenzyme Techniques; Keratins; Kidney; Middle Aged; Pigment Epithelium of Eye; Proto-Oncogene Proteins c-met; Retinal Detachment; Vitreoretinopathy, Proliferative; Vitreous Body

To determine whether syngeneic retinal cells injected in the vitreous cavity of the rat are able to initiate a proliferative process and whether the ocular inflammation induced in rats by lipopolysaccharide (LPS) promotes this proliferative vitreoretinopathy (PVR).. Primary cultured differentiated retinal Müller glial (RMG) and retinal pigmented epithelial (RPE) cells isolated from 8 to 12 postnatal Lewis rats were injected into the vitreous cavity of 8- to 10-week-old Lewis rats (10(5) cells/eye in 2 microlieter sterile saline), with or without the systemic injection of 150 microgram LPS to cause endotoxin-induced uveitis (EIU). Control groups received an intravitreal injection of 2 microliter saline. At 5, 15, and 28 days after cell injections, PVR was clinically quantified, and immunohistochemistry for OX42, ED1, vimentin (VIM), glial fibrillary acidic protein (GFAP), and cytokeratin was performed.. The injection of RMG cells, alone or in combination with RPE cells, induced the preretinal proliferation of a GFAP-positive tissue, that was enhanced by the systemic injection of LPS. Indeed, when EIU was induced at the time of RMG cell injection into the vitreous cavity, the proliferation led to retinal folds and localized tractional detachments. In contrast, PVR enhanced the infiltration of inflammatory cells in the anterior segment of the eye.. In the rat, syngeneic retinal cells of glial origin induce PVR that is enhanced by the coinduction of EIU. In return, vitreoretinal glial proliferation enhanced the intensity and duration of EIU.

Topics: Animals; Cell Transplantation; Cells, Cultured; Fluorescent Antibody Technique, Indirect; Glial Fibrillary Acidic Protein; Injections; Keratins; Lipopolysaccharides; Neuroglia; Pigment Epithelium of Eye; Rats; Rats, Inbred Lew; Receptors, Complement 3b; Retina; Retinal Detachment; Salmonella typhimurium; Transplantation, Isogeneic; Uveitis; Vimentin; Vitreoretinopathy, Proliferative; Vitreous Body

In the late stages of proliferative diabetic retinopathy (PDR) the vascularized posterior cortical gel (PCG) contracts leading to a partial posterior hyaloidal separation, hemorrhage, and traction retinal detachment (TRD). "Additional epiretinal membranes" have been described previously. These are thin, usually transparent epiretinal membranes which extend anteriorly from the point of attachment of the elevated posterior cortical gel to the edge of the TRD. The origin and frequency of the occurrence as well as the clinical significance of such additional epiretinal membranes are the subjects of controversy.. To quantitate the authors' clinical impression that additional epiretinal membranes are common in advanced PDR, to characterize them immunohistochemically, and to demonstrate the rationale for the authors' surgical approach.. Intraoperative observations for all patients undergoing diabetic vitrectomy and delamination over the last 2 years were reviewed retrospectively. The presence of additional epiretinal membrane was searched for in the initial stages of vitrectomy. When identified, their apparent continuity with the elevated portion of the PCG was confirmed. Surgical specimens were obtained from nine patients for immunohistochemical study.. Additional epiretinal membranes were observes in 145 (81%) of 179 consecutive eyes with PDR that underwent surgery for macular TRD. Immunohistochemical staining with type II collagen antibody was positive in all specimens, suggesting that these membranes were of vitreous origin.. It is likely that the additional epiretinal membranes represent the posterior leaf of a split PCG, the anterior leaf being the elevated portion of the PCG. The two leaves remain fused in the main, fibrovascular portion of the epiretinal membrane. These findings help explain the clinical experience that once the posterior leaf of the PCG is identified and elevated, it provides an accurate point of entry into the surgical plane facilitating delamination of the fused (vascularized) portion of the PCG from the detached retina.

Topics: Aged; Collagen; Diabetic Retinopathy; Eye Diseases; Female; Glial Fibrillary Acidic Protein; Humans; Immunoenzyme Techniques; Keratins; Male; Middle Aged; Retinal Detachment; Retrospective Studies; Vitrectomy; Vitreoretinopathy, Proliferative; Vitreous Body

The standard treatment for retinal breaks is thermal adhesion. Breaks in the posterior pole (i.e., macular holes) recently have been treated using vitrectomy and the recombinant cytokine transforming growth factor-beta. This has been shown to achieve closure of the retinal breaks by stimulating localized fibrocellular proliferation. Serum has been shown to contain chemoattractants and mitogens for many types of cells. The authors studied the clinical and histologic effect of autologous serum application to retinal breaks in an experimental model.. Twenty-four rabbits underwent pars plana lensectomy, vitrectomy, retinectomy, fluid-air exchange, application of test solution (12 with Hank's buffered salt solution and 12 with autologous serum), and air-gas exchange. Clinical examination with indirect ophthalmoscopy was performed, and animals were killed 5, 14, and 28 days after treatment. Tissue sections through the retinectomy were studied by light microscopy, electron microscopy, and immunocytochemistry.. None of the serum-treated eyes showed retinal detachment at the site of the retinectomy by evaluation with indirect ophthalmoscopy at each of the time points. In contrast, in control eyes retinal detachment developed at the retinectomy site from 0% at day 5 to 50% at day 14 and 75% at day 28. By light microscopy, serum-treated eyes contained multilayers of fibroblast-like cells adhering the retinectomy edges to the underlying retinal pigment epithelium and choroid. The control eyes had nonadherent retinal edges at the retinectomy site with little sign of fibrocellular response. Results were confirmed by electron microscopy. The fibroblast-like cells by immunocytochemistry contained vimentin, cytokeratin 18, and/or glial fibrillary acidic protein.. This study suggests that serum induces a localized fibrocellular response at the retinectomy edges compared with control eyes. This response, characterized by light microscopy, electron microscopy, and immunocytochemistry, appears to involve a mixed population of glial, retinal pigment epithelial, and/or fibroblastic cells. These cells seem to enhance adhesion and subsequent reattachment of the edges of the retinectomies at the time points studied when compared with controls.

Topics: Animals; Blood; Cell Movement; Disease Models, Animal; Fibroblasts; Glial Fibrillary Acidic Protein; Immunoenzyme Techniques; Keratins; Male; Neuroglia; Pigment Epithelium of Eye; Rabbits; Retina; Retinal Detachment; Retinal Perforations; Vimentin

Immunohistochemical techniques were used to investigate the relation between retinal pigment epithelial cells (RPE), traction retinal detachment (TRD) membranes, and combined traction rhegmatogenous retinal detachment (CTR) membranes in proliferative diabetic retinopathy. Seven CTR and five TRD membranes were obtained during closed microsurgery. Six of the seven CTR membranes and one of the five TRD membranes contained RPE. Eleven of the 12 diabetic membranes incorporated glial cells. The findings emphasise that the intravitreal membranes of proliferative diabetic retinopathy contain a diversity of cell types and indicate that RPE tend to contribute to CTR, rather than TRD, membranes. The histopathological appearance of CTR membranes is that of a hybrid between TRD and proliferative vitreo-retinopathy membranes.

Topics: Diabetic Retinopathy; Humans; Immunohistochemistry; Keratins; Pigment Epithelium of Eye; Retinal Detachment

Topics: Animals; Blood Platelets; Cell Membrane; Disease Models, Animal; Drug Administration Schedule; Free Radical Scavengers; Fundus Oculi; Ginkgo biloba; Humans; Injections; Keratins; Plant Extracts; Rabbits; Retinal Detachment; Retinal Neovascularization; Vitreous Body

Immunotoxins directed against a membrane marker of cell proliferation, transferrin receptor, were investigated to inhibit the growth of retinal pigment epithelial (RPE) cells in proliferative vitreoretinopathy (PVR). We undertook an immunocytological study in specimens of vitreous, subretinal fluid, and epiretinal membranes from patients with PVR to address the expression of transferrin receptor by proliferating pigment epithelial cells during the course of PVR and in normal human ocular structures. Thirty four specimens of vitreous and subretinal fluid, as well as seven epiretinal membranes, were immunocytologically examined using monoclonal antibodies to transferrin receptor. They showed a strong expression of this marker by a large majority of the cells in these two periretinal fluids (mean percentages 80 and 91% in vitreous and subretinal fluid, respectively). In contrast, only a few cells within epiretinal membranes were found to express transferrin receptor. In normal human eye sections conjunctival and corneal epithelial cells, subcapsular epithelium of the lens strongly expressed transferrin receptor, whereas RPE cells remained negative to antitransferrin receptor antibodies. A few iris or ciliary pigment epithelial cells reacted weakly. Thus, this study shows that most intravitreal and subretinal fluid proliferating cells strongly express transferrin receptor on their surface. Also confirmed is that immunotoxins to this membrane antigen could constitute potentially useful therapeutic agents in PVR.

Topics: Adult; Aged; Aged, 80 and over; Antibodies, Monoclonal; Cell Membrane; Conjunctiva; Epithelium; Exudates and Transudates; Eye Diseases; Fluorescent Antibody Technique; Humans; Keratins; Lens, Crystalline; Middle Aged; Pigment Epithelium of Eye; Receptors, Transferrin; Retinal Detachment; Retinal Diseases; Vitreous Body

The identification of cells comprising epiretinal membranes is difficult because of the phenotypic changes that occur. Examination of intermediate filament protein content by immunocytochemical analysis can help to identify some cells with altered ultrastructure but is not always definitive because altered expression of intermediate filament proteins can also occur. To examine this issue further, the authors utilized a postembedding immunocytochemical technique with epiretinal membranes in which they were able to double label for keratin, a useful marker for identifying retinal pigment epithelial cells, and glial fibrillary acidic protein (GFAP), a useful marker for identifying glial cells. Nine of ten idiopathic epiretinal membranes contained cells that labeled for GFAP and not keratin. Two of these membranes also contained cells that labeled only for keratin and one membrane contained cells that simultaneously labeled for both GFAP and keratin. Other types of epiretinal membranes had an equal participation by cells that expressed only GFAP or keratin (12 of 17 membranes contained cells positive for keratin; 13 of 17 contained cells positive for GFAP). Ten of 17 nonidiopathic membranes contained cells simultaneously expressing GFAP and keratin, although they comprised only a minor subpopulation of the total number of cells present. These findings demonstrate that keratin and GFAP are not mutually exclusive intermediate filament proteins in cells of epiretinal membranes and that, although each may provide a helpful adjunct for cell type identification, neither is an absolutely specific marker.

Topics: Eye Diseases; Glial Fibrillary Acidic Protein; Humans; Keratins; Membranes; Retina; Retinal Detachment; Retinal Diseases; Staining and Labeling; Vitreous Body

During vitreoretinal surgery, 23 epiretinal membranes from eyes treated with silicone oil were removed. They were examined by immunohistochemical methods and compared with 15 membranes from eyes affected by proliferative vitreoretinopathy (PVR) and not treated with silicone oil and with 4 membranes from eyes with intermediate uveitis. PVR membranes from eyes treated with silicone oil showed a high level of macrophages and a strong expression of HLA-DR. Additionally, lymphocytes were found in PVR membranes, a finding that has not been described before. Similar changes were seen in proliferations removed from eyes affected by uveitis, but these membranes were found to have smaller amounts of extracellular substance. In contrast to this, most cells in the PVR membranes from eyes not treated with silicone oil react with vimentin, GFAP and cytokeratin. In none of these membranes were we able to find T-lymphocytes. It is not possible to say whether or not the different findings are attributable ot the silicone oil.

Topics: Extracellular Matrix; Follow-Up Studies; Glial Fibrillary Acidic Protein; HLA-DR Antigens; Humans; Immunoenzyme Techniques; Keratins; Macrophages; Pigment Epithelium of Eye; Retina; Retinal Detachment; Retinal Neovascularization; Silicone Oils; T-Lymphocytes; Uveitis; Vimentin; Vitrectomy

Culture of cells from subretinal fluid (SRF) was performed using 29 SRF samples obtained at retinal reattachment surgery. Proliferating cells were found in 58.6% of the samples studied. The cells were of retinal pigment epithelial (RPE) origin, as evidenced by their brown pigmentation in primary culture and their positive immunostaining for cytokeratins 8/18. The age of the patients did not affect the proliferative capacity of the cells. Proliferating cells were present in all samples from eyes with proliferative vitreoretinopathy (PVR) of grade C1 or more. In primary culture the cells had a fibroblast-like morphology, resembling that of ordinary RPE cells exposed to the vitreous. We conclude that the SRF of many patients with PVR contains viable proliferating RPE cells and that SRF offers a new source of RPE cells for studies on the pathogenesis of PVR.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Cell Division; Cell Separation; Cells, Cultured; Extracellular Space; Female; Fibronectins; Humans; Keratins; Male; Microscopy, Fluorescence; Middle Aged; Pigment Epithelium of Eye; Retinal Detachment; Retinal Diseases

Topics: Adult; Aged; Cells, Cultured; Desmin; Eye Diseases; Female; Fluorescent Antibody Technique; Glial Fibrillary Acidic Protein; Histocytochemistry; Humans; Keratins; Male; Middle Aged; Retinal Detachment; Vimentin; Vitreous Body

Immunohistochemical techniques were used to identify cells containing cytokeratins in sections or tissue-culture monolayers from ocular (reference) tissues and also from 22 epiretinal membranes obtained during closed microsurgery for macular pucker or massive preretinal retraction. Results of cytokeratin immunostaining in reference tissues indicated that this is a valuable means of determining the contribution and distribution of epithelial cells in epiretinal membranes, and that the epithelial cells in the membranes were probably derived from the retinal pigment epithelium. Epithelial cells were identified in 17 of the 22 epiretinal membranes, but they did not usually constitute the predominant cell type. We concluded that the fibroblasts or fibroblast-like cells thought to be responsible for the contraction of epiretinal membranes are seldom of retinal pigment epithelial origin. Biomicroscopic pigmentation of a membrane was shown to be a poor guide to its epithelial cell population.

Topics: Animals; Aotus trivirgatus; Cattle; Culture Techniques; Fluorescent Antibody Technique; Humans; Immunoenzyme Techniques; Keratins; Membranes; Pigment Epithelium of Eye; Protein Precursors; Rabbits; Reference Standards; Retina; Retinal Detachment