Claudins regulate gene and protein expression of the retinal pigment epithelium independent of their association with tight junctions

Liu, Fanfei; Xu, Tao; Peng, Shaomin; Adelman, Ron A.; Rizzolo, Lawrence J.

doi:10.1016/j.exer.2020.108157

Cited by 10 publications

(2 citation statements)

References 53 publications

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“…To further support the importance of semi-permeable membrane qualities on cell extrusions, we cultured retina pigment epithelial cells (hTERT RPE-1, ATCC) known to have less developed cell junctions ( 25 ) on our wave substrates. And, as shown in Video 15, no cell extrusions can be detected from confluent monolayers of these cells over our wave substrates.…”

Section: Resultsmentioning

confidence: 99%

Surface curvature and basal hydraulic stress induce spatial bias in cell extrusion

Huang

Yong

She

et al. 2022

Preprint

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Cell extrusion is a coordinated biological process by which epithelial cells are removed from the epithelium without compromising the tissue's natural barrier function. Cell extrusions not only occur when injuries or genetic mutations arise, they also serve to maintain homeostatic cell numbers and have been found to drive morphogenesis. However, the effects of unique environmental settings---e.g., the complex geometric landscape and varying solute gradients in intestinal epithelia---on cell extrusions are less explored. Here we show, using epithelial monolayers cultured on periodic wave substrates, that surface curvature can induce a spatial bias in cell extrusions, with cells more frequently extruding from the negatively curved valleys. Surprisingly, increasing media osmolarity significantly reduced this bias, and also overall extrusion numbers. Furthermore, basal fluid spaces were found smaller under monolayers in hyper-osmotic media compared to iso-osmotic conditions, and similarly smaller on the positively curved hills as compared to valleys. The latter corresponded with the measured inward pointing normal forces on the hills. Remarkably, higher focal adhesion kinase (FAK) and Akt activation were observed in hyper-osmotically treated monolayers. Likewise, higher FAK activation was also observed in wave-hill residing cells. Our results show that surface curvature can play a modulating role in osmotically induced hydraulic stress levels, which act to disrupt focal adhesions and affect cell extrusions via FAK and the downstream Akt survival pathway. These findings highlight the fact that multiple environmental cues can synergistically influence cell extrusions. And an awareness of this will benefit our understanding of spatiotemporal preferences in cell extrusions seen in homeostasis, disease and development.

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Section: Resultsmentioning

confidence: 99%

Surface curvature and basal hydraulic stress induce spatial bias in cell extrusion

Huang

Yong

She

et al. 2022

Preprint

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“…Tight junctions form in the apical periphery of contacting cells mediating the diffusion from the choroid to the subretinal space and vice versa. ZO1 and especially CL19 are responsible for the formation of the tight junction barrier in RPE and thereby take part in the modulation of transepithelial diffusion [ 38 ]. Disruption of this barrier enables an uncontrollable leakage of molecules and nutrients [ 37 ].…”

Section: Discussionmentioning

confidence: 99%

Cell maturation influences the ability of hESC-RPE to tolerate cellular stress

et al. 2022

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Background Transplantation of human pluripotent stem cell-derived retinal pigment epithelium (RPE) is an urgently needed treatment for the cure of degenerative diseases of the retina. The transplanted cells must tolerate cellular stress caused by various sources such as retinal inflammation and regain their functions rapidly after the transplantation. We have previously shown the maturation level of the cultured human embryonic stem cell-derived RPE (hESC-RPE) cells to influence for example their calcium (Ca2+) signaling properties. Yet, no comparison of the ability of hESC-RPE at different maturity levels to tolerate cellular stress has been reported. Methods Here, we analyzed the ability of the hESC-RPE populations with early (3 weeks) and late (12 weeks) maturation status to tolerate cellular stress caused by chemical cell stressors protease inhibitor (MG132) or hydrogen peroxide (H2O2). After the treatments, the functionality of the RPE cells was studied by transepithelial resistance, immunostainings of key RPE proteins, phagocytosis, mitochondrial membrane potential, Ca2+ signaling, and cytokine secretion. Results The hESC-RPE population with late maturation status consistently showed improved tolerance to cellular stress in comparison to the population with early maturity. After the treatments, the early maturation status of hESC-RPE monolayer showed impaired barrier properties. The hESC-RPE with early maturity status also exhibited reduced phagocytic and Ca2+ signaling properties, especially after MG132 treatment. Conclusions Our results suggest that due to better tolerance to cellular stress, the late maturation status of hESC-RPE population is superior compared to monolayers with early maturation status in the transplantation therapy settings.

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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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Claudins regulate gene and protein expression of the retinal pigment epithelium independent of their association with tight junctions

Cited by 10 publications

References 53 publications

Surface curvature and basal hydraulic stress induce spatial bias in cell extrusion

Surface curvature and basal hydraulic stress induce spatial bias in cell extrusion

Cell maturation influences the ability of hESC-RPE to tolerate cellular stress

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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