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Results for “Retinal pigment epithelium”

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At least 19 recordsLinked to original sources

PubMed2000

Creatine kinase in human retinal pigment epithelium.

Retinal pigment epithelial ion transport activity, and consequent ATP consumption vary significantly as a function of photoreceptor activity. In a variety of cell types, ATP levels are maintained during high-energy usage by phosphocreatine hydrolysis, catalysed by the enzyme creatine kinase. The present work was designed to assess the importance of creatine kinase in retinal pigment epithelial cell metabolism. To this end, activity measurements, non-denaturing gel electrophoresis, Western blot analysis and immunohistochemistry were used to characterize creatine kinase in retinal pigment epithelium. Total creatine kinase activity in the retinal pigment epithelium is approximately 0.05 micromol ATP mg protein(-1) min(-1). The bulk of this activity was mediated by the B-CK isoform. However, by immunoblotting, non-denaturing gel electrophoresis and immunohistochemistry, the presence of the M-CK isoform of creatine kinase was also detected. The M-CK isoform was plasma membrane associated and predominately localized to the apical surface. Creatine kinase in the retinal pigment epithelium could function in a spatial energy shuttle that helps to sustain apical plasma membrane ion transport activity.

Blotting, Western↗

PubMed1990

Computerised visual field deficits in tears of the retinal pigment epithelium.

Retinal Pigment epithelial tears have been well documented as a complication of pigment epithelial detachment in patients with age related macular degeneration. Spontaneous and iatrogenic separation of detached retinal pigment epithelium, with subsequent retraction and exposure of the underlying choriocapillaris and Bruch's membrane, usually results in poor visual function in the affected areas. However, exact characterisation of the resultant scotomas has not been previously described. We present two patients with spontaneous pigment epithelial tears who underwent Octopus computerised visual field analysis. The density and characteristics of their associated field loss is compared with their clinical and fluorescein angiographic appearance.

Aged↗

PubMed2006

Retinal pigment epithelium.

Retinal pigment epithelium (RPE) arises from neuroectoderm and plays a key role in support of photoreceptor functions. Several degenerative eye diseases, such as macular degeneration or retinitis pigmentosa, are associated with impaired RPE function that may lead to photoreceptor loss and blindness. RPE derived from human embryonic stem (hES) cells can be an important source of this tissue for transplantation to cure such degenerative diseases. This chapter describes differentiation of hES cells to RPE, its subsequent isolation, maintenance in culture, and characterization.

Cell Culture Techniques↗

PubMed1985

Isolation of plasma membranes from the bovine retinal pigment epithelium.

Retinal pigment epithelium plasma membranes have been isolated by differential and density gradient centrifugation of glass-bead-bound, collagenase-treated cells. Electron microscopic evidence indicates that the glass-bead-bound cells were devoid of red blood cells, rod outer segments and other ocular cell contaminants. The plasma membranes were recovered in 4-6 micrograms/eye yields and purified 10-fold by 5'-nucleotidase and alkaline phosphodiesterase I, and 6.5-fold by (Na+ + K+)-ATPase. Plasma membrane purity as measured by covalent labeling of the epithelial cell plasma membrane proteins with p-(diazonium) benzene[32S]sulfonic acid was 8-19-fold. In purified plasma membranes contamination by mitochondria was undetectable and lysosomal contamination reduced 100-fold, while endoplasmic reticulum was 2-fold enriched. SDS-polyacrylamide gel electrophoresis of the plasma membrane proteins revealed 23-26 major bands by Coomassie blue staining and 12-16 major bands by radioactive labeling. The plasma membranes exhibited a 3-fold lower concentration of docosahexaenoic acid, a 3-fold higher cholesterol/phosphate ratio, and were 10-fold enriched in cholesterol per micrograms protein when compared to the whole cell fraction. Retinal epithelial plasma membranes contain an average of 1 mol cholesterol per mol of lipid phosphorus, a high palmitic acid concentration (39 mol%) and a low concentration of docosahexaenoic acid (2 mol%). The lipid profile of the retinal pigment epithelial plasma membranes indicates that they are typical of plasma membranes from many other cell types and that they appear to be less fluid than total rod outer segment membranes.

5'-Nucleotidase↗

PubMed1983

Furosemide-sensitive Cl transport in embryonic chicken retinal pigment epithelium.

Retinal pigment epithelium- (RPE) choroid-sclera preparations from embryonic chickens were mounted in an Ussing chamber. A spontaneous transepithelial voltage (Ve) of 5.1 mV (retinal side positive) and a resistance of 114 omega . cm2 can be attributed to the RPE. Furosemide and ouabain inhibited the Ve without affecting the resistance when applied to the retinal surface of the preparation but had no effect when applied to the scleral surface. Acetazolamide and amiloride were without effect when applied to either surface. The Ve fell by 100% in Na-free medium, by 70% in Cl-free medium, and was unchanged in HCO3-free medium. When Na and Cl were returned to the medium, the Ve recovered. Dilution potentials and unidirectional flux studies indicated that the RPE is more permeable to Na than to Cl. Isotope flux studies performed under open-circuit conditions showed a net retina-to-choroid flux for both Na and Cl with the net Cl flux abolished by furosemide. Analysis of the Na+ and Cl- electrochemical gradients and the Na permeability across the RPE suggests that the net retina-to-choroid Na+ flux is largely passive, whereas the net retina-to-choroid Cl- flux results from active transport. These results indicate that embryonic chicken RPE possesses furosemide-sensitive Cl transport function. A model of embryonic chicken RPE transepithelial transport is presented.

Acetazolamide↗

PubMed1997

Expression of 17 beta-hydroxysteroid dehydrogenase type IV in chick retinal pigment epithelium.

Retinal pigment epithelium (RPE) has essential roles that maintain functions of the neural retina. Previously, we described that the 3b5 monoclonal antibody (3b5 mAb) recognized the RPE cells in the chick embryonic eye. In the present study, a gene coding for the 3b5 antigen has been cloned and sequenced. A 10-day-old embryonic RPE cDNA library was constructed in lambda Uni-ZAP XR, and screened with the 3b5 mAb as a probe. The complete nucleotide sequence of the antigen recognized by the 3b5 mAb was obtained. The cDNA encodes a 735 amino acid protein with a calculated molecular mass of 80,147 Da. The deduced amino acid sequence shares 72% identity with the human 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) type IV. Northern blot analysis detected a 2.9 kb transcript. Immunohistochemical and in-situ hybridization studies revealed that 17 beta-HSD type IV was preferentially expressed in the RPE cells. The expression of steroid metabolizing enzymes in the RPE cells is able to create estrogenic environment, which may have a role of maintenance of the neural retina.

17-Hydroxysteroid Dehydrogenases↗

PubMed1995

Reorganization of actin microfilaments and microtubules in regenerating retinal pigment epithelium.

Retinal pigment epithelium (RPE) regenerating after experimental damage in rabbits exhibits major changes in cell shape, polarity and junctions--features that depend on the cytoskeleton. This report correlates these changes with the redistribution of actin microfilaments and microtubules, using electron microscopy and confocal laser scanning microscopy. We compare immature cells with the more mature cells that form the new epithelial monolayer. Two populations of immature RPE cells are interspersed at the edge of the regenerating RPE sheet. One population of immature cells makes few junctions with their neighbors or the basement membrane. They form pseudopodia and exhibit a prominent network of actin microfilaments beneath the plasma membrane. These cells are probably motile and advance the epithelial sheet. Another population of immature cells contains numerous stress fibers that insert into large basement membrane attachments. The cells make focal adhesions with their neighbors, rather than the junctional complexes characteristic of mature RPE cells. These cells are probably not motile and mature into the cells forming the new monolayer--cuboidal cells with numerous basal folds and apical villi and a complete belt of intercellular junctions. Stress fibers are lost as the circumferential bundle associated with the zonula adherens re-forms. Microtubules, which form prominent longitudinal bundles running through the processes of immature cells, take on the meshwork organization characteristic of mature RPE as the immature cells differentiate.

Actin Cytoskeleton↗

PubMed2002

Oxidant and antioxidant modulation of chloride channels expressed in human retinal pigment epithelium.

Retinal pigment epithelium (RPE) possesses regulated chloride channels that are crucial for transepithelial fluid and ion transport. At present, little is known about the molecular nature of chloride channels in human adult RPE (haRPE) or the effects of oxidative stress on membrane conductance properties. In the present study, we assessed ClC channel and cystic fibrosis transmembrane conductance regulator (CFTR) expression and membrane chloride conductance properties in haRPE cells. ClC-5, ClC-3, ClC-2, and CFTR mRNA expression was confirmed with RT-PCR analysis, and protein expression was detected with Western blot analysis and immunofluorescence microscopy. Whole cell recordings of primary cultures of haRPE showed an outwardly rectifying chloride current that was inhibited by the oxidant H(2)O(2). The inhibitory effects of H(2)O(2) were reduced in cultured human RPE cells that were incubated with precursors of glutathione synthesis or that were stably transfected to overexpress glutathione S-transferase. These findings indicate a possible role for ClC channels in haRPE cells and suggest possible redox modulation of human RPE chloride conductances.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1988

Alpha-1 adrenergic receptors on rabbit retinal pigment epithelium.

Retinal pigment epithelium (RPE)-choroid-sclera preparations from black, dutch-belted rabbits were sealed in an Ussing chamber. The RPE-generated trans-RPE voltage (Ve) and electrical resistance (R) were monitored. Epinephrine (an alpha and beta adrenergic agonist) reduced Ve by as much as 39% without affecting R. A response to epinephrine was noted at concentrations as low as 1.4 X 10(-7) M. Phenylephrine (an alpha adrenergic agonist) had essentially the same effect as epinephrine at identical concentrations. Clonidine (an alpha-2 adrenergic agonist) had a very slight effect but only at 10(-4) M. Isoproterenol (a beta adrenergic agonist) had no apparent effect upon Ve or R. The RPE response to epinephrine and phenylephrine was blocked by the alpha adrenergic antagonist phentolamine and by the alpha-1 adrenergic antagonist prazosin but not by the alpha-2 adrenergic antagonist yohimbine or by the beta adrenergic antagonist propranolol. Dibutyryl cyclic AMP (in the presence and absence of IBMX) and cyclic GMP (as cGMP and in the dibutyryl form) had no apparent effect upon Ve or R. These results indicate that rabbit RPE possesses an alpha-1 adrenoreceptor which, when stimulated, substantially reduces the RPE generated trans-RPE electrical current.

Animals↗

PubMed1992

Preserved para-arteriolar retinal pigment epithelium retinitis pigmentosa.

In two patients, we studied retinitis pigmentosa with preservation of the retinal pigment epithelium adjacent to and under the retinal arterioles (despite panretinal degeneration). Both patients with preserved para-arteriolar retinal pigment epithelium also exhibited a peculiarly strong hyperopia. In addition to previously reported features, these patients also had sheathing of the major vascular arcades, which suggested a vascular involvement in this uncommon form of retinitis pigmentosa.

Adult↗

PubMed2001

Differential roles of CD36 and alphavbeta5 integrin in photoreceptor phagocytosis by the retinal pigment epithelium.

Retinal pigment epithelial (RPE) cells employ alphavbeta5 integrin and CD36 receptors to phagocytose photoreceptor outer segment fragments (OS). We explored special properties of RPE phagocytosis to identify the contribution of CD36 to RPE phagocytosis measuring effects of CD36 antibodies on OS binding and internalization kinetics. Early, CD36 antibodies had no effect on OS binding or internalization. Both control and CD36 antibody treated RPE initiated internalization approximately 2 hours after OS challenge. Later, bivalent CD36 IgG accelerated OS engulfment while monovalent Fab fragments inhibited engulfment. Cross-linking Fab fragments restored the accelerating activity of intact IgG. Strikingly, antibodies were effective even if added to OS already bound by RPE. alphavbeta5 blocking antibody reduced OS binding equally well in the presence of CD36 antibodies but CD36 antibodies accelerated internalization of remaining bound OS. Furthermore, CD36 ligation at either apical or basal RPE surface partially substituted for soluble factors that are required for internalization but not for binding of OS at the RPE apical surface. Our results demonstrate that CD36 ligation is necessary and sufficient to activate the OS internalization mechanism of RPE. They suggest that CD36 acts as a signaling molecule in postbinding steps of RPE phagocytosis independently of the OS binding receptor alphavbeta5 integrin.

Animals↗

PubMed1992

Experimental autoimmune anterior uveitis (EAAU). II. Dose-dependent induction and adoptive transfer using a melanin-bound antigen of the retinal pigment epithelium.

Retinal pigment epithelial cell fractions have been investigated for their capacity to induce experimental uveitis. Cells of the dark (melanotic) and light areas of the bovine RPE have subsequently been extracted by buffer, Triton X-100, sodium dodecyl sulfate (SDS), and treated with various reagents in order to study some characteristics of the antigen. The SDS-insoluble melanotic fraction, consisting of spindle-shaped, mature melanin granules, proved to be the most uveitogenic preparation. Using pertussis toxin as coadjuvant, 1 microgram of melanin-protein (3.4 x 10(6) granules) was able to induce experimental autoimmune anterior uveitis (EAAU) in Lewis rats. The pathogenic activity of the responsible pathogen (PEP-X) was not diminished by SDS, nor eliminated by mildly alkaline SDS or formic acid treatment. However, HCl-deproteinized granules were not uveitogenic. The results show that PEP-X is a highly stable melano-antigen that is probably covalently bound to the granule surface. This is the first time that a melanin-bound antigen has been demonstrated to evoke specific autoaggressive activity. EAAU could adoptively be transferred by sensitized and in vitro stimulated CD4 T-lymphocytes. The evoked inflammation started 3-4 days after injection, was similar to those induced by immunization, and consisted mainly of severe iridocyclitis accompanied by dense flare and cells in the anterior chamber. Choroiditis developed in severe cases of EAAU but no inflammation was detected in the retina, pineal gland or other organs of these rats. EAAU could not be transferred by serum. Immunized PVG rats and guinea-pigs did not develop ocular inflammation. In monkeys a high dose of antigen evoked a very mild EAAU accompanied by choroiditis. In view of its characteristics, EAAU may be a new model for human anterior uveitis.

Animals↗

PubMed1983

Co-cultivation of retinoblastoma with fibroblasts, iris pigment epithelium, and retinal pigment epithelium in tissue culture.

Retinoblastoma cells of the Y79 line were co-cultivated with human fibroblasts and bovine iris and retinal pigment epithelium in tissue culture. The Y79 cells, which characteristically grow as a suspension culture, were found to attach directly to the fibroblasts and pigment epithelium on the flask surface. Electron microscopic examination of the fibroblast-retinoblastoma co-cultures revealed numerous pinocytotic vesicles lining the fibroblast cell borders that were in contact with the tumor cells. The retinoblastoma cells contained increased numbers of ribosomes, endoplasmic reticulum, and-mitochondria. Fibroblastic processes appeared to wrap around and engulf tumor cells. In both the iris and retinal pigment epithelium co-cultures with retinoblastoma cells, there were increased numbers of mitochondria in the tumor cells in areas adjacent to pigment epithelium but no pinocytotic vesicles were seen. The pigment epithelium attached to Y79 cells showed fewer processes than did the fibroblasts in co-culture. In summary, both fibroblast and pigment epithelium functioned as an effective carrier cell layer for retinoblastoma cells. In addition, we believe that the fibroblast layer removed substances secreted by the tumor cells via pinocytotic vesicles.

Animals↗

PubMed1993

Expression of the rasT24 oncogene in the ciliary body pigment epithelium and retinal pigment epithelium results in hyperplasia, adenoma, and adenocarcinoma.

We examined eye lesions in five lines of transgenic mice carrying the human rasT24 oncogene driven by the type I gamma glutamyl transferase (gamma GT) promoter. In three lines, hyperplasia developed as early as 11.5 days postconception in the outer neuroectodermal layer, which gives rise to ciliary body and retinal pigment epithelium. At birth, the eyes from many animals contained adenomas, and by day 27, mice developed invasive adenocarcinomas originating in the region of the ciliary body. Microphthalmia, cataracts, and chronic nongranulomatous inflammation involving the anterior and/or posterior segments of the eye were also found. gamma GT is detectable histochemically as early as 11.5 gestational days in the outer neuroectodermal layer and after birth is more abundant in the ciliary body than in the retinal pigment epithelium. Using a reverse transcriptase-polymerase chain reaction, we found that type I (but not types II or III) gamma GT RNA is made by the mouse eye; the gamma GT(I)rasT24 transgene transcription product was detected in the eyes of all five transgenic lines. The sequential progression of hyperplasia to invasive neoplasms in the ciliary body in response to gamma GT(I)rasT24 expression differs from the process in the kidney of these animals in which tubular hyperplasia and microadenomas with little evidence of progression are the major lesions.

Adenocarcinoma↗

PubMed1990

Ultrastructural and immunocytochemical changes in retinal pigment epithelium, retinal glia, and fibroblasts in vitreous culture.

Retinal pigment epithelial (RPE) cells, retinal glial, and fibroblasts, three cell types believed to play a role in the pathogenesis of epiretinal membrane formation, were maintained in vitreous culture to determine the influence of vitreous on their ultrastructure and expression of cytokeratin, glial fibrillary acidic protein (GFAP), vimentin, and glutamine synthetase (GS). Using a highly sensitive, preembedding technique for the immunolocalization of these antigens at the ultrastructural level, most RPE cells were found to lose cytokeratin and vimentin within 1 day after seeding on irradiated vitreous. The percentage of keratin-positive cells then increased with time in culture. If the vitreous was placed on RPE cells cultured in monolayer instead of placing the cells on the vitreous, keratin and vimentin were expressed these intermediate filament proteins diminished with time. Glutamine synthetase was found in RPE cells grown in monolayer with or without a vitreous overlay, but not in RPE cells grown on the surface of vitreous. Retinal glial grown on vitreous showed a time-dependent decrease in the number of cells expressing GFAP and a corresponding increase in cells expressing vimentin or GS. Some fibroblasts in vitreous culture expressed vimentin but not the other antigens evaluated. A substantial number of cells in each culture did not stain positively for cytokeratin, GFAP, vimentin, or GS. All three cell types showed phenotypic diversity at the ultrastructural level with each cell type being capable of assuming the same morphologic appearance under certain conditions. These results demonstrate the phenotypic plasticity of RPE cells, retinal glia, and fibroblasts when grown in contact with vitreous and provide further evidence that neither ultrastructure, intermediate filament protein expression, nor the presence of GS is sufficient to determine the cell type of origin of cells in epiretinal membranes.

Animals↗

PubMed1997

Comparison of tight junction permeability for albumin in iris pigment epithelium and retinal pigment epithelium in vitro.

BACKGROUND: The degenerative retinal diseases are one of the major causes of visual loss in the western world. Although heterologous RPE transplants rescue the photoreceptors in the dystrophic rat model, rejection remains a major limiting factor. Given the common embryonic origin, iris pigment epithelial (IPE) cells might be able to take over the functions of retinal pigment epithelial (RPE) cells, serving as an autologous graft for transplantation and thereby preventing rejection. One of the main functions of RPE cells is the generation of tight junctions which form the outer blood-retinal barrier. In this study we compared the tight junction permeabilities of IPE and RPE cells isolated from Long Evans rats by measuring their albumin clearances. METHODS: IPE and RPE cells were cultured on semipermeable filter supports with and without the addition of 0.02% ethylenediaminetetraacetic acid (EDTA). At selected intervals, the albumin clearances of the IPE and RPE cells were measured spectrophotometrically and compared. The morphology of the cells was compared using electron microscopy and fluorescent labeling. RESULTS: IPE and RPE cells both restricted the passage of albumin in vitro. After the modulation of tight junctions with 0.02% EDTA, the clearance increased in both types of cells in a similar fashion. The morphology of tight junctions was visualized with electron microscopy. CONCLUSION: These results indicate that the functional barrier for macromolecules is similar in IPE and RPE cells in vitro. This raises the possibility that IPE cells would form tight junctions in the subretinal space, thereby substituting for the blood-retinal barrier normally formed by RPE cells.

Actins↗

PubMed1994

Autosomal recessive retinitis pigmentosa with preserved para-arteriolar retinal pigment epithelium.

Retinitis pigmentosa with preserved para-arteriolar retinal pigment epithelium is a rare form of retinitis pigmentosa that starts early in life with preservation of retinal pigment epithelium adjacent to and under the retinal arterioles and that has hitherto been described as an isolated form. We examined 22 patients from one large family, together with two isolated patients, and confirmed the presumed autosomal recessive mode of inheritance in this type of retinitis pigmentosa. New findings associated with retinitis pigmentosa with preserved para-arteriolar retinal pigment epithelium were asteroid hyalosis in four (17%) of 24 patients, tortuosity of retinal arterioles in 11 (46%) of 24 patients, peripheral regions of opacified vessels in eight (33%) of 24 patients, and preservation not only of the para-arteriolar pigment epithelium, but also of the peripheral retinal pigment epithelium in 13 (54%) of 24 patients. Previously reported signs present in these patients were nystagmus in six (25%) of 24 patients, hypermetropia in 23 (96%) of 24 patients, optic nerve head drusen in nine (38%) of 24 patients, vascular sheathing in 11 (46%) of 24 patients, maculopathy in all 24 patients (100%), yellow round deposits in the posterior pole in nine (38%) of 24 patients, exudates resembling those in Coats' disease in two (8%) of 24 patients, visual field defects in all 24 patients (100%), and nondeductible electroretinograms in 21 (91%) of 23 patients. Linkage analysis carried out in the large family resulted in the assignment of a gene for retinitis pigmentosa with preserved para-arteriolar retinal pigment epithelium to chromosome 1q31-q32.1.

Adolescent↗

PubMed2006

Use of iris pigment epithelium to replace retinal pigment epithelium in age-related macular degeneration: a gene expression analysis.

OBJECTIVE: To determine the gene expression profiles of primary retinal pigment epithelium (RPE) and iris pigment epithelium (IPE) using microarrays. METHODS: Primary RPE and IPE from 6 human donor eyes were collected, and total RNA was isolated. Differences in gene expression were determined using a human genechip (human U95Av2 [12 600 probes]; Affymetrix Inc, Santa Clara, Calif). RESULTS: Hierarchical cluster analysis differentiated the gene expression profiles of RPE and IPE clusters into 2 distinct groups. A mean +/- SD of 5308 +/- 416 gene probes were expressed in RPE vs 6130 +/- 205 in IPE. Sixty-eight genes were expressed only in RPE; 154 genes were expressed only in IPE. Twenty-two additional genes had greater than 3-fold increased expression in RPE vs IPE, and 147 genes had greater than 3-fold decreased expression in RPE vs IPE. CONCLUSION: There are major differences in the gene expression profiles of primary RPE vs IPE. Clinical Relevance The different gene expression profiles of primary RPE vs IPE harvested from the same donor eyes infer that it may be difficult for IPE to replace all aspects of damaged RPE function in transplantation studies.

Aged↗