Retinal Pigment Epithelium Migration Research Articles

Tricetin suppresses the cell migration and BMP-6 expression through p38 signaling pathways in human retinal pigment epithelium cells.

Proliferative vitreoretinopathy (PVR) is a visual-threatening disease, which cause from the migration of retinal pigment epithelium (RPE). Tricetin, a family of flavonoids, can inhibit the metastasis of several cancers. Herein, we aim to evaluate the possible effect of tricetin on inhibiting ARPE-19 cells migration. The Boyden chamber assay, wound healing assay, RNA sequencing, and Western blot analysis were applied in our experiment. The results revealed that tricetin inhibited the cell migration abilities of ARPE-19 cells. Moreover, using RNA sequencing technology, we revealed that tricetin repressed bone morphogenetic protein-6 (BMP-6) gene expressions in ARPE-19 cells. Overexpression of BMP-6 resulted in significant restoration of cell migration capabilities of tricetin-treated ARPE-19 cells. Furthermore, tricetin suppressed the phosphorylation of the p38 signaling pathway. Moreover, blocking the p38 pathway also inhibits BMP-6 expression and migration in the ARPE-19 cells. In conclusion, this study revealed that tricetin inhibits the ARPE-19 cell migration mainly via the suppression of BMP-6 expression and p38 signaling pathway.

Complete Resolution of Central Serous Chorioretinopathy Cases Treated with Low Power Mode Laser Technology : A Novel Approach

Introduction : Central serous chorioretinopathy (CSC) is an idiopathic condition mostly affecting men in their 20s to 50s. It is characterized by serous fluid accumulation behind the neurosensory retina resulting in a localized macular detachment. Aldosterone-receptor antagonist and photodynamic laser therapy are the current mainstay of treatments, although most CSC cases can resolves spontaneously within 2 to 3 months. Low power mode (LPM) laser has emerged as a new effective technology to treat CSC.
 Case Illustration : Case 1: A 50-year-old male was diagnosed with recurrent chronic CSC since 2 months ago. His visual acuity declined even after given oral aldosterone receptor antagonist for two weeks, so he was scheduled to undergo LPM. After three weeks of laser treatment, CSC resolved and the visual acuity improved (Figure1). Case 2 : A 47-year-old male complained of decreased vision and metamorphopsia on his right eye since two weeks ago. He was diagnosed with CSC and underwent LPM. Three weeks after LPM, his vision improved and the metamorphopsia resolved (Figure2). LPM helps the migration of retinal pigment epithelium (RPE) cells to restore the outer blood-retinal barrier through expression of heat shock proteins. We used MC-500ViXi laser system (Nidek Co., Gamagori, Japan) and the laser parameters used in this study were 100?m spot size, 0.01s exposure time, 0.25-0.5 spacing, and 30% LPM ratio.
 Discussion : 
 Conclusion : LPM is a new yet promising treatment modalities to treat acute and chronic CSC. Its low power nature makes LPM a safe choice of treatment with less damage to the surrounding retina.

Transdifferentiation and Intrachoroidal Migration of Melanotic Retinal Pigment Epithelium in Punctate Inner Choroiditis

ABSTRACT Purpose: To report the multimodal imaging features of hyperpigmented chorioretinal lesions originating from the retinal pigment epithelium (RPE) within punched-out lesions of punctate inner choroidopathy (PIC). Methods: Retrospective case report. Multimodal imaging findings including fundus photography, optical coherence tomography (OCT), and OCT-angiography (OCTA) were analyzed. Results: A 49-year-old female with myopic degeneration developed progressive lesions of PIC requiring immunosuppressive therapy with adalimumab. Within areas of punched-out chorioretinal atrophic lesions, the occurrence of hyperpigmented lesions were observed which enlarged and extended into the choroid over a multiyear follow-up. Conclusion: This case illustrates the development of pigmented choroidal lesions appearing to originate from the RPE through transdifferentiation following previous chorioretinal inflammatory lesions. The introduction of adalimumab treatment may have activated the cellular migration of the RPE. To the best of our knowledge, this is the first report of intrachoroidal RPE migration in PIC.

Extracellular vesicles from retinal pigment epithelial cells expressing R345W-Fibulin-3 induce epithelial-mesenchymal transition in recipient cells.

We have shown previously that expression of R345W-Fibulin-3 induces epithelial-mesenchymal transition (EMT) in retinal pigment epithelial (RPE) cells. The purpose of the current study was to determine if extracellular vesicles (EVs) derived from RPE cells expressing R345W-Fibulin-3 mutation are sufficient to induce EMT in recipient cells. ARPE-19 cells were infected with luciferase-tagged wild-type (WT)- Fibulin-3 or luciferase-tagged R345W-Fibulin-3 (R345W) using lentiviruses. EVs were isolated from the media by ultracentrifugation or density gradient ultracentrifugation. Transmission electron microscopy and cryogenic electron microscopy were performed to study the morphology of the EVs. The size distribution of EVs were determined by nanoparticle tracking analysis (NTA). EV cargo was analysed using LC-MS/MS based proteomics. EV-associated transforming growth factor beta 1 (TGFβ1) protein was measured by enzyme-linked immunosorbent assay. The capacity of EVs to stimulate RPE migration was evaluated by treating recipient cells with WT- or R345W-EVs. The role of EV-bound TGFβ was determined by pre-incubation of EVs with a pan-TGFβ blocking antibody or IgG control. EM imaging revealed spherical vesicles with two subpopulations of EVs: a group with diameters around 30nm and a group with diameters over 100nm, confirmed by NTA analysis. Pathway analysis revealed that members of the sonic hedgehog pathway were less abundant in R345W- EVs, while EMT drivers were enriched. Additionally, R345W-EVs had higher concentrations of TGFβ1 compared to control. Critically, treatment with R345W-EVs was sufficient to increase EMT marker expression, as well as cell migration in recipient cells. This EV-increased cell migration was significantly inhibited by pre-incubation of EVs with pan-TGFβ-neutralising antibody. In conclusion, the expression of R345W-Fibulin-3 alters the size and cargo of EVs, which are sufficient to enhance the rate of cell migration in a TGFβ dependent manner. These results suggest that EV-bound TGFβ plays a critical role in the induction of EMT in RPE cells.

Prolonged Lifetimes of Histologic Autofluorescence in Ectopic Retinal Pigment Epithelium in Age-Related Macular Degeneration.

The purpose of this study was to investigate histologic autofluorescence lifetimes and spectra of retinal pigment epithelium (RPE) on the transition from normal aging to RPE activation and migration in age-related macular degeneration (AMD). Autofluorescence lifetimes and spectra of 9 donor eyes were analyzed in cryosections by means of 2-photon excited fluorescence at 960 nm. Spectra were detected at 483 to 665 nm. Lifetimes were measured using time-correlated single photon counting in 2 spectral channels: 500 to 550 nm (short-wavelength spectral channel [SSC]) and 550 to 700 nm (long-wavelength spectral channel [LSC]). Fluorescence decays over time were approximated by a series of three exponential functions. The amplitude-weighted mean fluorescence lifetime was determined. Markers for retinoid activity (RPE65) and immune function (CD68) were immunolocalized in selected neighboring sections. We identified 9 RPE morphology phenotypes resulting in 399 regions of interest (ROIs) for spectral and 497 ROIs for lifetime measurements. RPE dysmorphia results in a shorter wavelength peak of spectral emission: normal aging versus RPE migrated into the retina (intraELM) = 601.7 (9.5) nm versus 581.6 (7.3) nm, P < 0.001, whereas autofluorescence lifetimes increase: normal aging versus intraELM: SSC 180 (44) picosecond (ps) versus 320 (86) ps, P < 0.001; and LSC 250 (55) ps versus 441 (76) ps, P < 0.001. Ectopic RPE within the neurosensory retina is strongly CD68 positive and RPE65 negative. In the process of RPE degeneration, comprising different steps of dysmorphia and migration, lengthening of autofluorescence lifetimes and a hypsochromic shift of emission spectra can be observed. These autofluorescence changes might provide early biomarkers for AMD progression and contribute to our understanding of RPE-driven pathology.

Biocompatibility and Efficacy of a Linearly Cross-Linked Sodium Hyaluronic Acid Hydrogel as a Retinal Patch in Rhegmatogenous Retinal Detachment Repairment.

To prevent the migration of retinal pigment epithelium (RPE) cells into the vitreous cavity through retinal breaks after the pars plana vitrectomy for the repair of rhegmatogenous retinal detachment (RRD), sealing retinal breaks with an appropriate material appears to be a logical approach. According to a review of ocular experiments or clinical trials, the procedure for covering retinal breaks with adhesives is complex. A commercially available cross-linked sodium hyaluronic acid (HA) hydrogel (Healaflow®) with the injectable property was demonstrated to be a perfect retinal patch in RRD clinical trials by our team. Based on the properties of Healaflow®, a linearly cross-linked sodium HA hydrogel (HA-engineered hydrogel) (Qisheng Biological Preparation Co. Ltd. Shanghai, China) with the injectable property was designed, whose cross-linker and cross-linking method was improved. The purpose of this study is to report the characteristics of an HA-engineered hydrogel using Healaflow® as a reference, and the biocompatibility and efficacy of the HA-engineered hydrogel as a retinal patch in the rabbit RRD model. The HA-engineered hydrogel exhibited similar dynamic viscosity and cohesiveness and G′ compared with Healaflow®. The G′ of the HA-engineered hydrogel varied from 80 to 160 Pa at 2% strain under 25°C, and remained constantly higher than G″ over the range of frequency from 0.1 to 10 Hz. In the animal experiment, clinical examinations, electroretinograms, and histology suggested no adverse effects of the HA-engineered hydrogel on retinal function and morphology, confirming its favorable biocompatibility. Simultaneously, our results demonstrated the efficacy of the HA-engineered hydrogel as a retinal patch in the RRD model of rabbit eyes, which can aid in the complete reattachment of the retina without the need for expansile gas or silicone oil endotamponade. The HA-engineered hydrogel could play the role of an ophthalmologic sealant due to its high viscosity and cohesiveness. This pilot study of a small series of RRD models with a short-term follow-up provides preliminary evidence to support the favorable biocompatibility and efficacy of the HA-engineered hydrogel as a promising retinal patch for sealing retinal breaks in retinal detachment repair. More cases and longer follow-up studies are needed to assess its safety and long-term effects.

Overexpression of HTRA1 increases the proliferation and migration of retinal pigment epithelium cells.

Age-related macular degeneration (AMD) mainly affects the central region of retina and has many late-stage manifestations. Age-related macular degeneration is a leading cause of irreversible blindness in older people. The main feature of AMD is retinal pigment epithelium (RPE) degeneration. In this study, we aimed to explore the influence of HTRA1 expression on the proliferation and migration of RPE cells. Human ARPE-19 cells were transfected with an HTRA1 overexpression lentivirus or HTRA1 siRNA to silence HtrA1 expression. Quantitave reverse-transcription polymerase chain reaction (qRT-PCR) and western blotting were used to verify the relative level of HTRA1 mRNA and expression of HTRA1 protein of transfected human ARPE-19 cells. The MTT clone formation and transwell assays were used to confirm the effect of HTRA1 expression on the proliferation, colony forming ability and migration of ARPE-19 cells. The proliferation capacity (shown as optical density value) of ARPE-19 cells in the HTRA1-overexpressing group at culture times of 24 h and 48 h were 0.595 ±0.032 and 0.867 ±0.037 respectively, which were much higher than in the mock group. However, the proliferative capacity of cells in the HTRA1-silenced group decreased with increasing time of culture, compared with the mock group. The number of cloned and migrating cells in the HTRA1-overexpressing group were much higher than in the mock group, whereas the numbers in the HTRA1-silenced group were significantly lower. Overexpression of HTRA1 promotes proliferation and migration of RPE cells, which can help maintain the function of sensory neurons in the retina. Therefore, HTRA1 may be a suitable target for AMD treatments.

Choroideremia Gene Therapy.

Choroideremia Gene Therapy.

Bone Morphogenetic Protein (BMP)4 But Not BMP2 Disrupts the Barrier Integrity of Retinal Pigment Epithelia and Induces Their Migration: A Potential Role in Neovascular Age-Related Macular Degeneration.

Disruption of retinal pigment epithelial (RPE) barrier integrity and RPE migration are hallmark features in neovascular age-related macular degeneration (nAMD), but the underlying causes and pathophysiology are not completely well-defined. Herein, we aimed to evaluate the effect of bone morphogenetic proteins (BMPs) on the barrier function and migration of RPE. In particular, we investigated the role of BMP2 and BMP4 in these processes as our analysis of RNA-sequencing (seq) data from human donor eyes demonstrated that they are highly differentially expressed BMP members in macular RPE/choroid versus macular retina. We used electrical cell-substrate impedance sensing (ECIS) system to monitor precisely in real time the barrier integrity and migration of ARPE-19 after treatment with various concentrations of BMP2 or BMP4. Immunofluorescence was also used to assess the changes in the expression and the organization of the key tight junction protein, zona occludens (ZO)-1, in ARPE-19 cells under BMP2 or BMP4 treatment. This was followed by measuring the activity of matrix metalloproteinases (MMPs). Finally, RNA-seq and ELISA were used to determine the local and circulating levels of BMP2 and BMP4 in retinas and serum samples from nAMD donors. Our ECIS results showed that BMP4 but not BMP2 decreased the transcellular electrical resistance (TER) of ARPE-19 and increased their migration in comparison with control (vehicle-treated cells). Furthermore, immunofluorescence showed a disorganization of ZO-1 in BMP4-treated ARPE-19 not in BMP2-treated cells or vehicle-treated controls. This effect of BMP4 was associated with significant increases in the activity of MMPs, specifically MMP2. Lastly, these results were corroborated by additional findings that circulating but not local BMP4 levels were significantly higher in nAMD donor samples compared to controls. Collectively, our results demonstrated unreported effects of BMP4 on inducing RPE dysfunction and suggest that BMP4 but not BMP2 may represent a potential therapeutic target in nAMD.

Long-term Evolution and Remodeling of Soft Drusen in Rhesus Macaques

PurposeTo characterize the evolution and structure of soft drusen in aged rhesus macaques using in vivo multimodal retinal imaging and ex vivo histologic and ultrastructural analyses as a nonhuman primate model of early age-related macular degeneration (AMD).MethodsMultimodal imaging including fundus photography, spectral domain optical coherence tomography (SD-OCT), and fundus autofluorescence (FAF) were used to characterize and track individual drusen lesions in 20 aged rhesus macaques (mean age 23.3 ± 2.7 years) with drusenoid lesions over 2 years, followed by semithin histologic analysis and transmission electron microscopy (TEM).ResultsAlthough most drusen gradually increased in size, a portion spontaneously regressed or collapsed over 2 years. Histologic analyses showed that soft drusen exhibit hypertrophy and dysmorphia of overlying retinal pigment epithelium (RPE), as seen in early and intermediate AMD, but do not exhibit RPE atrophy, RPE migration, or photoreceptor degeneration characteristic of advanced AMD. Ultrastructure of soft drusen showed abundant lipid particles within Bruch's membrane and AMD-related basal linear deposits (BlinD) resembling those in human drusen.ConclusionsThe dynamic remodeling, histologic findings, and ultrastructural features of soft drusen in aged rhesus macaques support nonhuman primates as an animal model of early AMD and reveal important insights into drusen biogenesis and AMD development.

Phosphoinositide 3-kinase δ inactivation prevents vitreous-induced activation of AKT/MDM2/p53 and migration of retinal pigment epithelial cells

Phosphoinositide 3-kinases (PI3Ks) are a family of lipid kinases that play a critical role in transmitting signals from cell-surface molecules to intracellular protein effectors. Key PI3Ks include PI3Kα, PI3Kβ, and PI3Kδ, which are regulated by receptors. The signaling pathway comprising the PI3Ks, along with a Ser/Thr kinase (AKT), a proto-oncogene product (mouse double minute (MDM)2), and a tumor suppressor protein (p53), plays an essential role in experimental proliferative vitreoretinopathy (PVR), which is a fibrotic blinding eye disorder. However, which PI3K isoforms are involved in PVR is unknown. A major characteristic of PVR is the formation of epi (or sub)-retinal membranes that consist of extracellular matrix and cells, including retinal pigment epithelium (RPE) cells, glial cells, and macrophages. RPE cells are considered key players in PVR pathogenesis. Using immunoblotting and immunofluorescence analyses, we herein provide the evidence that PI3Kδ is highly expressed in human RPEs when it is primarily expressed in leukocytes. We also found that PI3Kδ inactivation through two approaches, CRISPR/Cas9-mediated depletion and a PI3Kδ-specific inhibitor (idelalisib), not only blocks vitreous-induced activation of AKT and MDM2 but also abrogates a vitreous-stimulated decrease in p53. Furthermore, we demonstrate that PI3Kδ inactivation prevents vitreous-induced proliferation, migration, and contraction of human RPEs. These results suggest that PI3Kδ may represent a potential therapeutic target for RPE-related eye diseases, including PVR.

Microglia Regulate Neuroglia Remodeling in Various Ocular and Retinal Injuries.

Reactive microglia and infiltrating peripheral monocytes have been implicated in many neurodegenerative diseases of the retina and CNS. However, their specific contribution in retinal degeneration remains unclear. We recently showed that peripheral monocytes that infiltrate the retina after ocular injury in mice become permanently engrafted into the tissue, establishing a proinflammatory phenotype that promotes neurodegeneration. In this study, we show that microglia regulate the process of neuroglia remodeling during ocular injury, and their depletion results in marked upregulation of inflammatory markers, such as Il17f, Tnfsf11, Ccl4, Il1a, Ccr2, Il4, Il5, and Csf2 in the retina, and abnormal engraftment of peripheral CCR2+ CX3CR1+ monocytes into the retina, which is associated with increased retinal ganglion cell loss, retinal nerve fiber layer thinning, and pigmentation onto the retinal surface. Furthermore, we show that other types of ocular injuries, such as penetrating corneal trauma and ocular hypertension also cause similar changes. However, optic nerve crush injury-mediated retinal ganglion cell loss evokes neither peripheral monocyte response in the retina nor pigmentation, although peripheral CX3CR1+ and CCR2+ monocytes infiltrate the optic nerve injury site and remain present for months. Our study suggests that microglia are key regulators of peripheral monocyte infiltration and retinal pigment epithelium migration, and their depletion results in abnormal neuroglia remodeling that exacerbates neuroretinal tissue damage. This mechanism of retinal damage through neuroglia remodeling may be clinically important for the treatment of patients with ocular injuries, including surgical traumas.

Down-regulation of microRNA-27b promotes retinal pigment epithelial cell proliferation and migration by targeting Nox2

Aberrant proliferation and migration of retinal pigment epithelium (RPE) cells contributes to the pathology of various ocular diseases. miR-27b has been reported to be crucial in the regulation of cell differentiation, proliferation, apoptosis, and migration. However, the role of miR-27b on RPE proliferation and migration remains largely unknown. Here the effect of miR-27b on ARPE-19 cells under platelet-derived growth factor (PDGF)-BB stimulation was explored. In this study, we found that the expression level of miR-27b was significantly reduced in ARPE-19 cells under PDGF-BB stimulation. Ectopic expression of miR-27b remarkably inhibited PDGF-BB-induced proliferation and migration in ARPE-19 cells. Furthermore, bioinformatic analysis and luciferase reporter assay showed that NADPH oxidase 2 (Nox2) was a direct target for miR-27b, and that knockdown of Nox2 expression mimicked the inhibitory effect of miR-27b on PDGF-BB −induced proliferation and migration in ARPE-19 cells, whereas, restoration of Nox2 expression showed an opposite effect. In addition, the ROS production and the activation of P13K/AKT/mTOR signaling induced by PDGF-BB were also suppressed by miR-27b overexpression or Nox2 silencing. Thus, these findings indicated that miR-27b exerted its protective role in RPE cells under PDGF-BB stimulation was partially through regulation of Nox2 and its downstream P13K/AKT/mTOR signaling, which might be a potential therapeutic approach for treatment of diseases caused by RPE proliferation, and migration.

C-Jun-mediated microRNA-302d-3p induces RPE dedifferentiation by targeting p21Waf1/Cip1

Dedifferentiation of retinal pigment epithelium (RPE) cells and choroidal neovascularization (CNV) contributes to the pathogenesis of age-related macular degeneration (AMD). MicroRNAs (miRNAs) have crucial roles in AMD onset and progression. We thus aim to investigate the effects of miRNAs on RPE dedifferentiation and endothelium cell (EC) behavior, and analyze its downstream pathways. We have previously identified miR-302d-3p as the most downregulated miRNA signature along with RPE differentiation. Herein, in vitro study supported that miR-302d-3p induces RPE dedifferentiation typified by reduction of RPE characteristic markers, interrupts its phagocytosis, and promotes its migration, proliferation, and cell-cycle progression. c-Jun was identified as a potential upstream transcript factor for MIR302D, which might modulate RPE function by regulating miR-302d-3p expression. P21Waf1/Cip1, a cyclin-dependent kinase inhibitor encoded by the CDKN1A gene, was identified as a downstream target of miR-302d-3p. Our data suggested that p21Waf1/Cip1 could promote RPE differentiation, and inhibit its proliferation, migration, and cell-cycle progression. We also demonstrated that miR-302d-3p suppresses RPE differentiation through directly targeting p21Waf1/Cip1. In addition, the miR-302d-3p/CDKN1A axis was also involved in regulating tube formation of ECs, indicating its potential involvement in CNV formation. Taken together, our study implies that miR-302d-3p, regulated by c-Jun, contributes to the pathogenesis of both atrophic and exudative AMD. MiR-302d-3p promotes RPE dedifferentiation, migration, proliferation and cell-cycle progression, inhibits RPE phagocytosis, and induces abnormal EC behavior by targeting p21Waf1/Cip1. Pharmacological miR-302d-3p inhibitors are prospective therapeutic options for prevention and treatment of AMD.

Novel NADPH oxidase inhibitor VAS2870 suppresses TGF‑β‑dependent epithelial‑to‑mesenchymal transition in retinal pigment epithelial cells

NADPH oxidases (NOXs) are important in the pathophysiology of fibrotic diseases. The expression and activity of NOXs are regulated by growth factors, including transforming growth factor (TGF-β). The proliferation of retinal pigment epithelial (RPE) cells following epithelial- to-mesenchymal transition (EMT) is a major pathological change involved in proliferative vitreoretinopathy (PVR). The aim of the present study was to determine the effects of the novel NOX inhibitor VAS2870 on the TGF-β-dependent expression of NOX4 and associated cellular events in RPE cells. Cell viability was examined using a Cell Counting Kit-8 assay and cell cycle progression was detected by flow cytometric analysis. Immunofluorescence analysis and western blot analysis were performed to assess EMT. It was found that TGF-β increased the expression of NOX4 and that pre-incubation with VAS2870 eliminated this effect. Additionally, TGF-β promoted RPE migration and increased EMT. Pre-incubation with VAS2870 significantly prevented TGF-β2-induced EMT by decreasing the levels of α-smooth muscle actin and E-cadherin, and also inhibited the migratory ability of the RPE cells, as demonstrated by scratch assays. Finally, VAS2870 suppressed the proliferation of RPE cells, and led to G1-phase cell cycle arrest and a significant downregulation of the expression of cyclin D1. In conclusion, the pharmacological inhibition of NOX may be a promising tool for the treatment of PVR.

IL-10 inhibits retinal pigment epithelium cell proliferation and migration through regulation of VEGF in rhegmatogenous retinal detachment.

Rhegmatogenous retinal detachment (RRD) is a disorder of the eye that affects physical and mental health. Retinal pigment epithelium (RPE) is closely associated with RRD, and it is hypothesized that RPE-secreted inflammatory cytokines may induce early pathological changes of RRD and may participate in RPE proliferation and migration. The present study determined a role for interleukin (IL)‑10 as an RPE‑secreted immune regulatory factor that contributes to RRD. A rat RRD model was established and RPE cells were isolated and cultivated. RPE cells were randomly divided into four groups, three treated with different concentrations of IL‑10 (100, 50, and 20mM) and one untreated. RPE cell proliferation was evaluated by MTT assay and the activity of caspase‑3 was also measured. RPE cell invasion was determined by Transwell assay. Vascular endothelial growth factor A (VEGF) expression was examined by reverse transcription‑quantitative polymerase chain reaction and western blotting; IL‑1 and IL‑6 levels were measured by ELISA. IL‑10 treatment suppressed RPE cell proliferation and migration, promoted caspase‑3 activity, inhibited VEGF mRNA and protein expression, and downregulated the secretion of inflammatory cytokines IL‑1 and IL‑6 in RRD group compared with the untreated Model group. The aforementioned effects of IL‑10 became more evident with increasing IL‑10 concentration. IL‑10 suppressed inflammation, facilitated RPE cell apoptosis and inhibited cell proliferation and migration through the regulation of VEGF expression.

Evaluation of intraretinal migration of retinal pigment epithelial cells in age-related macular degeneration using polarimetric imaging

The purpose of the present study was to evaluate the intraretinal migration of the retinal pigment epithelium (RPE) cells in age-related macular degeneration (AMD) using polarimetry. We evaluated 155 eyes at various AMD stages. Depolarized light images were computed using a polarization-sensitive scanning laser ophthalmoscope (PS-SLO), and the degree of polarization uniformity was calculated using polarization-sensitive optical coherence tomography (OCT). Each polarimetry image was compared with the corresponding autofluorescence (AF) images at 488 nm (SW-AF) and at 787 nm (NIR-AF). Intraretinal RPE migration was defined by the presence of depolarization at intraretinal hyperreflective foci on PS-SLO and PS-OCT images, and by the presence of hyper-AF on both NIR-AF and SW-AF images. RPE migration was detected in 52 of 155 eyes (33.5%) and was observed in drusenoid pigment epithelial detachment (PED) and serous PED with significantly higher frequencies than in other groups (P = 0.015). The volume of the migrated RPE cluster in serous PED was significantly correlated with the volume of the PED (R2 = 0.26; P = 0.011). Overall, our results showed that intraretinal RPE migrations occurred in various AMD stages, and that they occurred more commonly in eyes with serous and drusenoid PED.

Activated Retinal Pigment Epithelium, an Optical Coherence Tomography Biomarker for Progression in Age-Related Macular Degeneration.

PurposeTo summarize and contextualize recent histology and clinical imaging publications on retinal pigment epithelium (RPE) fate in advanced age-related macular degeneration (AMD); to support RPE activation and migration as important precursors to atrophy, manifest as intraretinal hyperreflective foci in spectral-domain optical coherence tomography (SDOCT).MethodsThe Project MACULA online resource for AMD histopathology was surveyed systematically to form a catalog of 15 phenotypes of RPE and RPE-derived cells and layer thicknesses in advanced disease. Phenotypes were also sought in correlations with clinical longitudinal eye-tracked SDOCT and with ex vivo imaging–histopathology correlations in geographic atrophy (GA) and pigment epithelium detachments (PED).ResultsThe morphology catalog suggested two main pathways of RPE fate: basolateral shedding of intracellular organelles (apparent apoptosis in situ) and activation with anterior migration. Acquired vitelliform lesions may represent a third pathway. Migrated cells are packed with RPE organelles and confirmed as hyperreflective on SDOCT. RPE layer thickening due to cellular dysmorphia and thick basal laminar deposit is observed near the border of GA. Drusenoid PED show a life cycle of slow growth and rapid collapse preceded by RPE layer disruption and anterior migration.ConclusionsRPE activation and migration comprise an important precursor to atrophy that can be observed at the cellular level in vivo via validated SDOCT. Collapse of large drusen and drusenoid PED appears to occur when RPE death and migration prevent continued production of druse components. Data implicate excessive diffusion distance from choriocapillaris in RPE death as well as support a potential benefit in targeting drusen in GA.

FAK phosphorylation plays a central role in thrombin-induced RPE cell migration

The migration of retinal pigment epithelial (RPE) cells is an important step in various pathologic conditions including subretinal neovascularization (SRN), proliferative vitreoretinopathy (PVR) and, importantly, as a consequence of retinal surgery. Therefore, the elucidation of the mechanisms underlying RPE trans-differentiation and migration is essential for devising effective treatments aimed to the prevention of these disorders. A common event in these pathologies is the alteration of the blood-retina barrier (BRB), which allows the interaction of RPE cells with thrombin, a pro-inflammatory protease contained in serum. Our previous work has demonstrated that thrombin induces RPE cell cytoskeletal remodeling and migration, hallmark processes in the development of PVR; however, the molecular mechanisms involved are still unclear. Cell migration requires the disassembly of focal adhesions induced by Focal Adhesion Kinase (FAK) phosphorylation, together with the formation of actin stress fibers. The aim of the present work was to identify thrombin-activated signaling pathways leading to FAK phosphorylation and to determine FAK participation in thrombin-induced RPE cell migration. Results demonstrate that the activation of PAR1 by thrombin induces FAK autophosphorylation at Y397 and the subsequent phosphorylation of Y576/577 within the activation loop. FAK phosphorylation was shown to be under the control of c/nPKC and PI3K/PKC-ζ, as well as by Rho/ROCK, since the inhibition of these pathways prevented thrombin-induced FAK phosphorylation and the consequent disassembly of focal adhesions, in parallel to FAK-dependent actin stress fiber formation and RPE cell migration. These findings demonstrate, for the first time, that thrombin stimulation of RPE cell transformation and migration are regulated by FAK tyrosine phosphorylation. Thus, targeting FAK phosphorylation may provide a strategical basis for PVR treatment.

Retinal pigment epithelial features indicative of neovascular progression in age-related macular degeneration

Background/aimsTo identify characteristic retinal pigment epithelium (RPE) changes in fellow eyes of patients with neovascular age-related macular degeneration (nAMD) using polarisation-sensitive optical coherence tomography (PS-OCT).MethodsThirty-one fellow eyes of 31 patients...