The cell biology of the retinal pigment epithelium

Lakkaraju, Aparna; Umapathy, Ankita; Tan, Li Xuan; Daniele, Lauren L.; Philp, Nancy J.; Boesze‐Battaglia, Kathleen; Williams, David S.

doi:10.1016/j.preteyeres.2020.100846

Cited by 256 publications

(232 citation statements)

References 354 publications

(533 reference statements)

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“…Additionally, as mentioned, the αVβ5 integrin is essential for RPE cell function, i.e. binding of POS and subsequent phagocytosis through CD36 and MERTK [3,[6][7][8][9]. Our results also fit with gene expression profiles that have been published for ARPE19 cells [101], fetal and adult human RPE cells [102,103] as well as stem cell-derived RPE cells [104,105].…”

Section: Discussionsupporting

confidence: 87%

“…For instance, on RPE cells, they mediate the interaction with Bruch's membrane and the phagocytosis of shed POS [1,3,5]. In this study we analyzed and compared the expression of integrin subunits in different RPE cells using immunostaining.…”

Section: Resultsmentioning

confidence: 99%

“…RPE cells are critical for the development and integrity of the outer retina. They transport nutrients from blood vessels to photoreceptors and phagocytose shed photoreceptor outer segments (POS) as well as playing a role in maintaining the blood-retina barrier [1][2][3]. Integrins are crucial for the proper functioning of the RPE cells.…”

Section: Introductionmentioning

confidence: 99%

“…The adhesion of the RPE monolayer to its normal substrate, Bruch's membrane, is mediated via integrins [4,5]. Additionally, αVβ5 integrin is essential for POS binding and subsequent phagocytosis through CD36 and the protooncogene tyrosine-protein kinase MER (MERTK) [3,[6][7][8][9]. Integrins are transmembrane signaling molecules that are formed through the hetero-dimerization of an α and a β subunit [10,11].…”

Section: Introductionmentioning

confidence: 99%

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Immunochemical analysis of the integrin expression in retinal pigment epithelial cells

Heller

Heintz

Kwok

et al. 2020

Preprint

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Retinal pigment epithelial (RPE) cells have been used in disease modelling and transplantation studies in retinal diseases. Several types of RPE cells have been trialed, ranging from primary cells and immortalized cell lines to stem cell-derived RPE cells. During aging and in disease, the extracellular environment of the RPE cells changes, interfering with RPE cell adhesion. We hypothesize that this could be a key problem in transplantation studies that have reported lack of adhesion and survival of the transplanted RPE cells. Integrins are essential for the proper function of the RPE, mediating adhesion to Bruch's membrane, and the binding and subsequent phagocytosis of photoreceptor outer segments. Variability that has been found in clinical trials might be due to the variability of cell types used and their expression profiles of surface molecules. Here, we set out to analyze integrin expression in primary rat RPE cells and in the human cell line ARPE-19 using immunochemistry. We found that both cell types express integrins to varying degrees. After long-term culturing, ARPE-19 cells resemble mature RPE cells, and increase integrin expression. We hence argue that it is important to test the properties of these cells prior to transplantation to avoid failure of adhesion and to facilitate correct function.

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Section: Discussionsupporting

confidence: 87%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Immunochemical analysis of the integrin expression in retinal pigment epithelial cells

Heller

Heintz

Kwok

et al. 2020

Preprint

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“…Autophagy is an essential clearance mechanism for the post-mitotic RPE, where debris cannot be diluted by cell division (4). RPE from AMD donors exhibit several autophagic defects including decreased LC3B lipidation (a marker for autophagosome biogenesis) and accumulation of p62/SQSTM1 (a measure of completion of autophagy) (55,56).…”

Section: Biochemical Measurements Of Total Cell Cholesterol Showed Thmentioning

confidence: 99%

Mitochondria-dependent phase separation of disease-relevant proteins drives pathological features of age-related macular degeneration

Cunza

Tan

Thamban

et al. 2020

Preprint

Self Cite

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The retinal pigment epithelium (RPE) is the site of initial damage leading to photoreceptor degeneration and vision loss in age-related macular degeneration (AMD). Genetic and histopathological studies implicate cholesterol dysregulation in AMD; yet mechanisms linking cholesterol to RPE injury and drusen formation remain poorly understood. Especially enigmatic are allelic variants of the cholesterol transporter APOE, major risk modifiers in Alzheimer’s disease that show reversed risk associations with AMD. Here, we investigated how ApoE isoforms modulate RPE health using live-cell imaging of primary RPE cultures and high-resolution imaging of human donor tissue. We show that the AMD-protective ApoE4 efficiently transports cholesterol and safeguards RPE homeostasis despite cellular stress. In contrast, ApoE2-expressing RPE accumulate cholesterol, which promotes autophagic deficits and complement-mediated mitochondrial fragmentation. Redox-related order-disorder phase transitions in ApoE2 drive the formation of intracellular biomolecular condensates as potential drusen precursors. Drugs that restore mitochondrial function limit condensate formation in ApoE2-RPE. Autophagic and mitochondrial defects correlate with intracellular ApoE aggregates in AMD donor RPE. Our study elucidates how AMD risk variants act as tipping points to divert the RPE from normal aging towards AMD by disrupting critical metabolic functions, and identifies mitochondrial stress-mediated aberrant phase transitions as a novel mechanism of drusen biogenesis.

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Cytocompatibility of electrospun poly‐L‐lactic acid membranes for Bruch's membrane regeneration using human embryonic stem cell‐derived retinal pigment epithelial cells

Abbasi,

O'Neill

2024

J Biomedical Materials Res

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Cell replacement therapy is under development for dry age‐related macular degeneration (AMD). A thin membrane resembling the Bruch's membrane is required to form a cell‐on‐membrane construct with retinal pigment epithelial (RPE) cells. These cells have been differentiated from human embryonic stem cells (hESCs) in vitro. A carrier membrane is required for cell implantation, which is biocompatible for cell growth and has dimensions and physical properties resembling the Bruch's membrane. Here a nanofiber electrospun poly‐L‐lactic acid (PLLA) membrane is tested for capacity to support cell growth and maturation. The requirements for laminin coating of the membrane are identified here. A porous electrospun nanofibrous PLLA membrane of ∼50 nm fiber diameter was developed as a prototype support for functional RPE cells grown as a monolayer. The need for laminin coating applied to the membrane following treatment with poly‐L‐ornithine (PLO), was identified in terms of cell growth and survival. Test membranes were compared in terms of hydrophilicity after laminin coating, mechanical properties of surface roughness and Young's modulus, porosity and ability to promote the attachment and proliferation of hESC‐RPE cells in culture for up to 8 weeks. Over this time, RPE cell proliferation, morphology, and marker and gene expression, were monitored. The functional capacity of cell monolayers was identified in terms of transepithelial electrical resistance (TEER), phagocytosis of cells, as well as expression of the cytokines, vascular endothelial growth factor (VEGF) and pigment epithelium‐derived factor (PEDF). PLLA polymer fibers are naturally hydrophobic, so their hydrophilicity was improved by pretreatment with PLO for subsequent coating with the bioactive protein laminin. They were then assessed for amount of laminin adsorbed, contact angle and uniformity of coating using scanning electron microscopy (SEM). Pretreatment with 100% PLO gave the best result over 10% PLO treatment or no treatment prior to laminin adsorption with significantly greater surface stiffness and modulus. By 6 weeks after cell plating, the coated membranes could support a mature RPE monolayer showing a dense apical microvillus structure and pigmented 3D polygonal cell morphology. After 8 weeks, PLO (100%)‐Lam coated membranes exhibited the highest cell number, cell proliferation, and RPE barrier function measured as TEER. RPE cells showed the higher levels of specific surface marker and gene expression. Microphthalmia‐associated transcription factor expression was highly upregulated indicating maturation of cells. Functionality of cells was indicated by expression of VEGF and PEDF genes as well as phagocytic capacity. In conclusion, electrospun PLLA membranes coated with PLO‐Lam have the physical and biological properties to support the distribution and migration of hESC‐RPE cells throughout the whole structure. They represent a good membrane candidate for preparation of hESC‐RPE cells as a monolayer for implantation into the subretinal space of AMD patients.

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The cell biology of the retinal pigment epithelium

Cited by 256 publications

References 354 publications

Immunochemical analysis of the integrin expression in retinal pigment epithelial cells

Immunochemical analysis of the integrin expression in retinal pigment epithelial cells

Mitochondria-dependent phase separation of disease-relevant proteins drives pathological features of age-related macular degeneration

Cytocompatibility of electrospun poly‐L‐lactic acid membranes for Bruch's membrane regeneration using human embryonic stem cell‐derived retinal pigment epithelial cells

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