Biomedical subjects

Peter Wiedemann

Publications and source records attributed to Peter Wiedemann.

At least 55 records · Page 3Linked to original sources

PubMed2005

Triamcinolone does not alter glial cell activation in the experimentally detached rabbit retina.

AIMS: Retinal detachment induces neural and photoreceptor cell degeneration and fast activation of micro- (immune) and macroglial cells. Hypoxia caused by increased distance between the choriocapillaris and the neural retina, and retinal oedema during detachment, are factors causing gliotic responses and cell degeneration. Triamcinolone may inhibit some cellular responses that accompany hypoxia. Therefore, we investigated whether triamcinolone acetonide may be effective to reduce the gliotic alterations in the detached retina. METHODS: Local retinal detachment in rabbit eyes was created by subretinal injection of sodium hyaluronate, and triamcinolone acetonide (8 mg) was applied intravitreally. Wholecell patch-clamp records from Muller cells and Ca2+ imaging from retinal wholemounts were carried out. Microglial/immune cells in the nerve-fiber layer of retinal wholemounts were labeled with Griffonia simplicifolia agglutinin (GSA) isolectin. Additionally, two morphological parameters which characterize microglial activation/immune cell infiltration were estimated: the cross-sectional area of the somata of the cells in the nerve-fiber layer and the number of cell processes which evolve from the soma. RESULTS: Three days after detachment, gliotic alterations were apparent in Muller cells isolated from both detached and nondetached retinal areas, as indicated by the cellular hypertrophy, by the downregulation of the plasma membrane K+ conductance, and by the upregulation of intracellular Ca2+ responsiveness to stimulation of purinergic P2Y receptors. Intravitreal triamcinolone did not alter these gliotic alterations of Muller cells. Furthermore, triamcinolone could not inhibit the immune cell activation present in detached and attached retinal areas. However, intravitreal triamcinolone led to a strong decrease in the process number of GSA lectin-positive cells from detached retinas. CONCLUSIONS: The results suggest that triamcinolone is ineffective to inhibit gliotic responses in the detached retina. However, the immune cell activation after detachment was partially influenced by triamcinolone.

Animals↗

PubMed2005

Neuronal versus glial cell swelling in the ischaemic retina.

Under normal conditions, the pigment epithelium dehydrates the outer retina while Müller glial cells mediate the rapid water transport within the inner retina. Gliotic alterations of Müller cells may be implicated in the development of oedema in the post-ischaemic retina. Here, we suggest a mechanism of Müller cell-supported neuronal cell swelling and apoptosis in the ischaemic retina. During ischaemia, over-excitation of ionotropic glutamate receptors leads to neuronal cell depolarization that causes excess Ca(2+) influx into the cells, and to activation of the apoptosis machinery. The ion fluxes into the retinal neurons are associated with water movements that are mediated by aquaporin-4 water channels expressed by Müller cells and result in neuronal cell swelling. After reperfusion, the glial cells may swell due to the down-regulation of their K(+) conductance, which results in intracellular K(+) overload and water movements from the blood and vitreous into the cells. An inhibition of the glial cell-mediated water movements during ischaemic episodes should reduce the ion shifts at the neuronal synapses, resulting in decreased neuronal cell swelling and apoptosis. An inhibition of the water movements in the post-ischaemic phase may prevent cytotoxic Müller cell swelling but may impair the fluid clearance from retinal tissue in the presence of vasogenic oedema. Thus, pharmacological modification of the ion and fluid clearance functions of Müller cells may become a novel way to resolve both cytotoxic and vasogenic oedema in the retina.

Animals↗

PubMed2005

Resensitization of P2Y receptors by growth factor-mediated activation of the phosphatidylinositol-3 kinase in retinal glial cells.

PURPOSE: To determine whether activation of receptor tyrosine kinases enhances the responsiveness of purinergic P2Y receptors in Muller glial cells, known to induce Muller cell proliferation. METHODS: The P2Y receptors of primary cultured Muller cells of the guinea pig were desensitized by short (30 seconds to 10 minutes)- and long (24 or 48 hours)-term application of adenosine 5'-triphosphate (ATP). Recordings of ATP-evoked intracellular calcium responses showed whether short-term application of different growth factors evoke a resensitization of the receptors. Coapplication of pharmacologic inhibitors showed whether activation of protein kinases is involved in receptor resensitization. RESULTS: Both short- and long-term incubation with ATP induced a significant P2Y receptor desensitization, which was indicated by a strongly reduced intracellular calcium mobilization and which lasted for at least 48 hours. However, the receptors were significantly resensitized after short-term application of platelet-derived, epidermal, or nerve growth factors. The growth factor-mediated resensitization was dependent on an intact cytoskeleton and on the activation of protein phosphatases and of the phosphatidylinositol-3 kinase (PI3K), but was independent of the activation of protein kinase C, src kinases, or extracellular signal-regulated kinases. CONCLUSIONS: The results show that activation of receptor tyrosine kinases causes, via activation of PI3K and protein phosphatases, a resensitization of P2Y receptors formerly desensitized by agonist application. The growth factor-mediated resensitization may underlie the previously observed enhanced responsiveness of P2Y receptors in retinal glial cells in experimental retinal detachment and proliferative vitreoretinopathy and may contribute to the induction of reactive gliosis and Muller cell proliferation.

Adenosine Triphosphate↗

PubMed2005

Ischemia-reperfusion causes exudative detachment of the rabbit retina.

PURPOSE: To characterize the activation of macroglial (Müller) and microglial cells, as well as neuronal cell degeneration, during ischemia-reperfusion in rabbit retina and to test the possible effect of triamcinolone acetonide on gliosis. METHODS: Transient retinal ischemia was produced by increasing intraocular pressure for 60 minutes. Triamcinolone (8 mg) was intravitreally applied immediately after the cessation of ischemia. At 3 and 8 days after reperfusion, the K+ currents of acutely isolated Müller cells were recorded, and the Ca2+ responses of Müller cells on stimulation of P2Y receptors were recorded fluorometrically in retinal wholemounts. Microglial/immune cells in the nerve fiber layer of retinal wholemounts were labeled with isolectin. To evaluate neuronal and Müller cell loss, the numbers of cells were counted in retinal slices. RESULTS: Transient ischemia caused exudative detachment of the central retina that was characterized by disruption of the pigment epithelial monolayer, the presence of scattered pigment epithelial and immune cells in the expanded subretinal space, and retinal folds. A significant loss of photoreceptor cells was observed at 8 days after reperfusion. At 3 and 8 days after reperfusion, Müller cell gliosis was apparent, as indicated by cellular hypertrophy, downregulation of K+ channel expression, and an increased number of cells that displayed P2Y receptor-mediated Ca2+ responses. The number of microglial/immune cells increased strongly after reperfusion. Intravitreal triamcinolone did not affect the parameters of Müller cell gliosis but decreased the number of microglial/immune cells. CONCLUSIONS: Ischemia-reperfusion of the rabbit retina causes exudative retinal detachment that is characterized by a loss of photoreceptor cells, whereas the inner retina remains largely preserved. Micro- and macroglial cells are activated early during reperfusion, even before dropout of the photoreceptor cells. Intravitreal triamcinolone may decrease the degree of microglial/immune cell activation.

Animals↗

PubMed2005

Identification of P2Y receptor subtypes in human muller glial cells by physiology, single cell RT-PCR, and immunohistochemistry.

PURPOSE: Retinal Müller glial cells are known to express metabotropic P2Y receptors. The present study was conducted to identify certain subtypes of P2Y receptors in human Müller cells. METHODS: The patch-clamp technique was used to measure increases of Ca(2+)-dependent K+ currents mediated by the activation of P2Y receptors in freshly isolated human Müller cells. Several P2 agonists were used. Subsequently, the cells were harvested into the patch pipette and a single cell RT-PCR was performed. Moreover, retinal tissue from organ donors was used for immunohistochemistry. RESULTS: The electrophysiological data were consistent with the expression of P2Y1, P2Y2, P2Y4, and P2Y6 receptor subtypes. RT-PCR revealed that mRNA for all these subtypes was present in Müller cells. However, the incidence of P2Y2 receptor mRNA was significantly lower than that of the other subtypes. Immunoreactivity for all four subtypes was found in retinal tissue, partly colocalized with immunoreactivity for vimentin. CONCLUSIONS: The presented data obtained by different techniques revealed that human Müller cells express P2Y1, P2Y2, P2Y4, and P2Y6 receptors. The specific roles of these receptor subtypes in retinal physiology and/or pathophysiology remain to be investigated in future studies.

Fluorescent Antibody Technique, Indirect↗

PubMed2004

Glutamate-evoked alterations of glial and neuronal cell morphology in the guinea pig retina.

Neuronal activity is accompanied by transmembranous ion fluxes that cause cell volume changes. In whole mounts of the guinea pig retina, application of glutamate resulted in fast swelling of neuronal cell bodies in the ganglion cell layer (GCL) and the inner nuclear layer (INL) (by approximately 40%) and a concomitant decrease of the thickness of glial cell processes in the inner plexiform layer (IPL) (by approximately 40%) that was accompanied by an elongation of the glial cells, by a thickening of the whole retinal tissue, and by a shrinkage of the extracellular space (by approximately 18%). The half-maximal effect of glutamate was observed at approximately 250 mum, after approximately 4 min. The swelling was caused predominantly by AMPA-kainate receptor-mediated influx of Na+ into retinal neurons. Similar but transient morphological alterations were induced by high K+ and dopamine, which caused release of endogenous glutamate and subsequent activation of AMPA-kainate receptors. Apparently, retinal glutamatergic transmission is accompanied by neuronal cell swelling that causes compensatory morphological alterations of glial cells. The effect of dopamine was elicitable only during light adaptation but not in the dark, and glutamate and high K+ induced strong ereffects in the dark than in the light. This suggests that not only the endogenous release of dopamine but also the responsiveness of glutamatergic neurons to dopamine is regulated by light-dark adaptation. Similar morphological alterations (neuronal swelling and decreased glial process thickness) were observed in whole mounts isolated immediately after experimental retinal ischemia, suggesting an involvement of AMPA-kainate receptor activation in putative neurotoxic cell swelling in the postischemic retina.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed2004

PEDF derived from glial Müller cells: a possible regulator of retinal angiogenesis.

A precise balance between stimulators and inhibitors of angiogenesis, such as vascular endothelial growth factor (VEGF) and pigment epithelium-derived factor (PEDF), respectively, is essential for angiogenic homeostasis in ocular tissues. Retinal hypoxia is accompanied by some pathological conditions that may promote intraocular neovascularization. Here we demonstrate that retinal glial (Müller) cells express and release pigment epithelium-derived factor (PEDF). Decreasing oxygen concentrations cause strong attenuation of PEDF release resulting in enhanced VEGF/PEDF ratios. Exposure of Müller cells to VEGF suppressed PEDF release in a dose-dependent manner. This may represent a novel mechanism of ocular angiogenic homeostasis sufficient in the control of PEDF levels during normoxia or mild hypoxia but supplemented by other (hitherto unknown) mechanisms in cases of strong hypoxia. In spite of the enhanced VEGF/PEDF ratios resulting from hypoxia, conditioned media of Müller cells failed to stimulate additional proliferation of retinal endothelial cells. These findings suggest that in the ischemic retina, Müller cells generate a permissive condition for angiogenesis by secreting more VEGF and less PEDF, but the onset of retinal endothelial cell proliferation requires another triggering signal that remains to be identified.

Animals↗

PubMed2004

Neuropeptide Y-evoked proliferation of retinal glial (Muller) cells.

BACKGROUND: Glial cells in human retinas and in fibrocellular membranes from patients with proliferative vitreoretinopathy (PVR) have been described to upregulate their expression of Y1 receptors for neuropeptide Y (NPY) (Soler et al.: Glia 39:320, 2002). However, it is unknown whether Y1 receptor activation causes proliferation of retinal glial cells. We investigated whether NPY exerts a proliferation-stimulating effect on retinal glial cells, and compared the NPY-evoked signaling with the signaling of purinergic P2Y receptors. METHODS: Proliferation assays using bromodeoxyuridine were carried out on primarily cultured Muller glial cells of the guinea pig, in the absence and presence of blockers of Y1 receptors, of receptor tyrosine kinases (RTKs), of mitogen-activated protein kinases (MAPKs) and of phosphatidylinositol-3 kinase (PI3K). RESULTS: NPY exerted a biphasic effect on Muller cell proliferation. At low concentrations (0.1 ng/ml and 1 ng/ml) it decreased the proliferation rate of the cells, while at higher concentration (100 ng/ml) it increased Muller cell proliferation. The NPY-evoked proliferation was mediated by Y1 receptor stimulation and by activation of the p44/p42 MAPKs and partially of the p38 MAPK. Moreover, Y1 receptor-induced activation of PI3K as well as transactivations of the platelet-derived and the epidermal growth factor RTKs were necessary for full mitogenic effect of NPY. Y1 and P2Y receptors share partially common signal transduction pathways in Muller cells. CONCLUSION: It is suggested that NPY may be involved in stimulation of retinal glial cell proliferation during PVR when it is released at higher amounts into the injured retina.

Adenosine Triphosphate↗

PubMed2004

Angiogenesis-related factors derived from retinal glial (Müller) cells in hypoxia.

Retinal glial (Müller) cells may play a major role in vascular eye diseases as they secrete vascular endothelial growth factor (VEGF), a hypoxia-induced angiogenic cytokine. They also release significant amounts of the anti-angiogenic factors, transforming growth factor (TGF)-beta2, pigment epithelium derived factor (PEDF), and thrombospondin-1 (TSP-1). Exposure of human (MIO-M1) and guinea-pig Müller cells to hypoxia resulted in a decreased release of TGF-beta2 and PEDF but in an elevated secretion of TSP-1. When retinal endothelial cells were exposed to VEGF/anti-angiogenic factor ratios mimicking those found in culture media of Müller cells under normoxia or hypoxia, their proliferation was significantly inhibited by TGF-beta2, PEDF or TSP-1. Thus Müller cells may provide a permanent anti-proliferative condition for retinal endothelial cells.

Analysis of Variance↗

PubMed2004

Characterization of the basic fibroblast growth factor-evoked proliferation of the human Müller cell line, MIO-M1.

BACKGROUND: Basic fibroblast growth factor (bFGF) has been suggested to mediate activation of Müller glial cells in the ischemic-hypoxic retina. However, the intracellular pathways activated by bFGF in human Müller cells have been little explored. We characterized the signaling transduction pathways which are involved in the control and growth factor-evoked proliferation of a recently described human Müller cell line, MIO-M1. In addition, we investigated whether bFGF evoked the release of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) from the cells. METHODS: The growth factor-evoked proliferation of cultured MIO-M1 cells was estimated by means of a bromodeoxyuridine immunoassay, in the absence and presence of blockers of mitogen-activated protein kinases (MAPKs) and of the phosphatidylinositol-3 kinase (PI3K). The activation state of the p44/p42 MAPK was determined by Western blotting, and the bFGF-evoked release of VEGF and HGF was evaluated by ELISA. RESULTS: bFGF evoked a concentration-dependent increase of the cell proliferation, with an EC50 of approximately 1 ng/ml, via activation of both the p44/p42 MAPK and the p38 MAPK. In contrast, the mitogenic effects of the platelet-derived and the heparin-binding epidermal growth factors were dependent on p44/p42 MAPK activation and independent of activation of p38 MAPK. The transforming growth factors beta1 and beta2 also evoked cell proliferation which was independent of activation of the MAPKs investigated. bFGF evoked a release of VEGF and of HGF by the cells; these effects were independent of MAPK activation and were possibly mediated by activation of the PI3K signaling pathway. CONCLUSION: bFGF evokes multiple intracellular signaling pathways in human Müller cells which underlie the gliotic cell responses upon ischemic-hypoxic insults in the retina. Beside the stimulation of cell proliferation, which is dependent on activation of p44/p42 and p38 MAPKs, bFGF induces the secretion of VEGF and HGF by Müller cells.

Blotting, Western↗

PubMed2004

Selective staining by vital dyes of Müller glial cells in retinal wholemounts.

Müller glial cells within the retina may respond to different signaling molecules with an elevation of their intracellular free calcium. To prove the localization of the recorded calcium responses in Müller cells within acutely isolated retinal wholemounts, retinal pieces from adult animals and humans were exposed to different vital dyes just after the calcium imaging records were finished. The dyes, Mitotracker Orange, Mitotracker Green, Celltracker Orange, Celltracker Green, and monochlorobimane, are all selectively taken up by Müller glial cells, while neuronal cells remain largely devoid of the dyes. By using this method, it can be demonstrated that the free calcium alterations within the wholemounts indeed occur within Müller cells. Moreover, the cross-sectional areas of (dye-filled) Müller glial cell bodies, as well as of (dye-free) neuronal cell bodies, can be measured in retinal wholemounts, and the spatial densities of both types of cells can be determined. The vital dye loading of Müller cells may facilitate investigations of stimulus-induced alterations of retinal glial cell physiology and morphology.

Animals↗

PubMed2004

GABA(A) receptors in Müller glial cells of the human retina.

The present study was aimed at characterizing the GABA(A) receptor-mediated currents in acutely isolated glial (Müller) cells of the human retina and investigating their subcellular localization across the Müller cell membrane. Extracellular application of GABA evoked two current responses in human Müller cells: a fast transient GABA(A) receptor-mediated current that inactivated within 10 s and that was independent of extracellular Na(+), and a sustained current that was dependent on extracellular Na(+) and that was mediated by high-affinity GABA transporters. The receptor current was half-maximally activated at a GABA concentration of 32 microM, while the transporter current showed an affinity constant of 7.9 microM GABA. The receptor currents were blocked by bicuculline and picrotoxin and were also activated by muscimol or by other amino acids. The receptor currents are Cl(-) currents, as indicated by the close relationship between the reversal potential of these currents and the Cl(-) equilibrium potential. Using perforated-patch recordings, a mean intracellular Cl(-) concentration of 37 +/- 12 mM was determined in human Müller cells. Using electrophysiological and fluorescence imaging methods, it was revealed that GABA(A) receptors are unevenly distributed across the Müller cell membrane, with higher densities at the endfoot, at the soma, and at the distal sclerad end of the cells. It is concluded that GABA(A) receptor expression may allow a sensing of retinal GABAergic neuronal signal transmission by Müller cells.

Action Potentials↗

PubMed2004

A potassium channel-linked mechanism of glial cell swelling in the postischemic retina.

The cellular mechanisms underlying glial cell swelling, a central cause of edema formation in the brain and retina, are not yet known. Here, we show that glial cells in the postischemic rat retina, but not in control retina, swell upon hypotonic stress. Swelling of control cells could be evoked when their K(+) channels were blocked. After transient ischemia, glial cells strongly downregulated their K(+) conductance and their prominent Kir4.1 protein expression at blood vessels and the vitreous body. In contrast, the expression of the aquaporin-4 (AQP4) (water channel) protein was only slightly altered after ischemia. Activation of D(2) dopaminergic receptors prevents the hypotonic glial cell swelling. The present results elucidate the coupling of transmembraneous water fluxes to K(+) currents in glial cells and reveal the role of altered K(+) channel expression in the development of cytotoxic edema. We propose a mechanism of postischemic glial cell swelling where a downregulation of their K(+) conductance prevents the emission of intracellularly accumulated K(+) ions, resulting in osmotically driven water fluxes from the blood into the glial cells via aquaporins. Inhibition of these water fluxes may be beneficial to prevent ischemia-evoked glial cell swelling.

Animals↗

PubMed2004

Peeling of the basal membrane in the human retina: ultrastructural effects.

PURPOSE: To study the effects of internal limiting membrane (ILM) peeling, usually performed in cases of macular hole formation, on the structural integrity of the human retina. DESIGN: Experimental study. MATERIAL: Two human donor eyes. METHODS: A human donor retina was subjected to ILM peeling shortly after enucleation, fixed, and studied by electron microscopy. MAIN OUTCOME MEASURES: Electron microscopic findings. RESULTS: Within the peeled area, and even at its margins, a substantial number of Müller (glial) cell endfeet and vitreal Müller cell processes was severely damaged. However, many glial cells remained intact, and the cell bodies of the injured cells showed no signs of destruction. CONCLUSIONS: We have shown that ILM peeling results in minor, but demonstrable, damage of the adjacent retina. Clinical studies are required to demonstrate unequivocally that ILM peeling does not induce functional deficits rather than improvement of visual function.

Basement Membrane↗

PubMed2004

Drug delivery systems for vitreoretinal diseases.

The eye has an environment that is specific unto itself in terms of pharmacokinetics: the inner and outer blood-retinal barriers separate the retina and the vitreous from the systemic circulation and vitreous body, which physiologically has no cellular components, occupies the vitreous cavity, an inner space of the eye, and reduces practical convection of molecules. Considering this, development of a drug delivery system (DDS) is becoming increasingly important in the treatment of vitreoretinal diseases not only to facilitate drug efficacy but also to attenuate adverse effects. The DDS has three major goals: enhances drug permeation (e.g., iontophoresis and transscleral DDS), controls release of drugs (e.g., microspheres, liposomes, and intraocular implants), and targets drugs (e.g., prodrugs with high molecular weight and immunoconjugates). Comprehensive knowledge of these should lead to development of innovative treatment modalities.

Absorbable Implants↗

PubMed2004

Vitreoretinal endoscope for the assessment of the peripheral retina and the ciliary body after large retinectomies in severe anterior PVR.

PURPOSE: We evaluated endoscopically the changes of the peripheral retina and the ciliary body after large retinectomies. METHODS: The peripheral retina and the ciliary body of 5 patients with anterior proliferative vitreoretinopathy after large retinectomies (> 180 degrees) were visualized endoscopically. RESULTS: The endoscope allowed a complete assessment of the peripheral retina and the ciliary body. Even the 12 o'clock part of the eye could be evaluated with the 30 degrees optic. The peripheral retina, a fibrosis and a detachment of the ciliary body can be visualized and assessed. The cause of the postoperative hypotony after large retinectomies is mainly related to fibrosis and detachment of the ciliary body. Surgeons can expect a postoperative hypotony if fibrosis and a large detachment of the ciliary body is seen during surgery with the help of an endoscope. CONCLUSION: Endoscopic vitreoretinal visualization of the pars plana and ciliary body is a useful additional technique after large retinectomies in the treatment of anterior PVR.

Ciliary Body↗

PubMed2004

Glial cell expression of hepatocyte growth factor in vitreoretinal proliferative disease.

The hepatocyte growth factor (HGF) has been crucially implicated in the development of proliferative retinal diseases; however, it is unclear whether retinal glial cells express or respond to HGF. Therefore, we examined the expression of HGF and of the receptor for HGF, c-Met, by immunohistochemical costaining with glial fibrillary acidic protein (GFAP) in epiretinal membranes of patients with proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR), respectively. Furthermore, it was determined whether cells of the human retinal glial cell line, MIO-M1, secrete HGF protein, and whether HGF stimulates proliferation and chemotaxis, and secretion of the vascular endothelial growth factor (VEGF). Neuroretinas of patients with PVR express elevated mRNA level for HGF in comparison to control retinas. In epiretinal membranes of patients with PVR or PDR, immunoreactivity for HGF and for c-Met, respectively, partially colocalized with immunoreactivity for GFAP. Fetal bovine serum and basic fibroblast growth factor, but not heparin-binding epidermal or platelet-derived growth factors, evoked HGF secretion by cultured retinal glial cells. HGF displayed only a marginal effect on cell proliferation while it stimulated chemotaxis. HGF promoted the secretion of VEGF, via activation of the phosphatidylinositol-3 kinase. It is concluded that glial cells in epiretinal membranes express both HGF protein and c-Met receptors. The results suggest an autocrine/paracrine role of HGF in glial cell responses during proliferative vitreoretinal disorders as well as in retinal neovascularization, by stimulating of VEGF release.

Adolescent↗

PubMed2004

Contrary effects of cytokines on mRNAs of cell cycle- and ECM-related proteins in hRPE cells in vitro.

PURPOSE: Retinal pigment epithelial (RPE) cells play a pivotal role in the pathogenesis of proliferative vitreoretinopathy (PVR). As a result of a breakdown of the blood-retina barrier, growth factors obtain access to the subretinal space and stimulate several retinal cell types. The aim of the present study was to analyze the effect of several growth factors on the proliferation of human (h)RPE cells, and on the mRNA expression of transcription factors, cell cycle proteins, and extracellular matrix (ECM) proteins. METHODS: hRPE cells were incubated in the presence of TGF-beta1, TGF-beta2, PDGF, VEGF, or bFGF for 24-72 h. Cell proliferation was assessed by determinating BrdU incorporation. Changes in mRNA expression of c-fos, c-myc, PCNA, FEN1, Ki67, collagen III, and collagen IV were investigated by ribonuclease protection assay (RPA). RESULTS: RPE cell proliferation was significantly increased by exposure to PDGF and bFGF for 48 h, and was decreased by application of TGF-beta1 and TGF-beta2 for 48 and 72 h. All the tested growth factors significantly elevated the amounts of c-fos mRNA (after 1 h) and of c-myc mRNA (after 24 h). PDGF and bFGF up-regulated the expression of Ki67 mRNA, and down-regulated that of collagen III and collagen IV mRNA after 24 h. TGF-beta1 and TGF-beta2 decreased the expression of Ki67 mRNA, and increased that of collagen III and collagen IV mRNA. CONCLUSION: Our results show that distinct cytokines may induce contrary effects with respect to proliferation of, and ECM formation by, hRPE cells in vitro. This knowledge may be useful for the development of improved therapeutic approaches.

Cell Cycle Proteins↗