• Clinical and Experimental Reproductive Medicine
• /
• v.38 no.4
• /
• pp.216-221
• /
• 2011

Objective: To differentiate the human embryonic stem cells (hESCs) into the retinal pigment epithelium (RPE) in the defined culture condition and determine its therapeutic potential for the treatment of retinal degenerative diseases. Methods: The embryoid bodies were formed from hESCs and attached on the matrigel coated culture dishes. The neural structures consisting neural precursors were selected and expanded to form rosette structures. The mechanically isolated neural rosettes were differentiated into pigmented cells in the media comprised of N2 and B27. Expression profiles of markers related to RPE development were analyzed by reverse transcription-polymerase chain reaction and immunostaining. Dissociated putative RPE cells ($10^5$ cells/5 ${\mu}L$) were transplanted into the subretinal space of rat retinal degeneration model induced by intravenous sodium iodate injection. Animals were sacrificed at 1, 2, and 4 weeks after transplantation, and immnohistochemistry study was performed to verify the survival of the transplanted cells. Results: The putative RPE cells derived from hESC showed characteristics of the human RPE cells morphologically and expressed molecular markers and associated with RPE fate. Grafted RPE cells were found to survive in the subretinal space up to 4 weeks after transplantation, and the expression of RPE markers was confirmed with immunohistochemistry. Conclusion: Transplanted RPE cells derived from hESC in the defined culture condition successfully survived and migrated within subretinal space of rat retinal degeneration model. These results support the feasibility of the hESC derived RPE cells for cell-based therapies for retinal degenerative disease.

• Molecules and Cells
• /
• v.44 no.8
• /
• pp.613-622
• /
• 2021

In vertebrate eyes, the retinal pigment epithelium (RPE) provides structural and functional homeostasis to the retina. The RPE takes up retinol (ROL) to be dehydrogenated and isomerized to 11-cis-retinaldehyde (11-cis-RAL), which is a functional photopigment in mammalian photoreceptors. As excessive ROL is toxic, the RPE must also establish mechanisms to protect against ROL toxicity. Here, we found that the levels of retinol dehydrogenases (RDHs) are commonly decreased in phosphatase tensin homolog (Pten)-deficient mouse RPE, which degenerates due to elevated ROL and that can be rescued by feeding a ROL-free diet. We also identified that RDH gene expression is regulated by forkhead box O (FOXO) transcription factors, which are inactivated by hyperactive Akt in the Pten-deficient mouse RPE. Together, our findings suggest that a homeostatic pathway comprising PTEN, FOXO, and RDH can protect the RPE from ROL toxicity.

• Molecules and Cells
• /
• v.46 no.7
• /
• pp.441-450
• /
• 2023

β-Catenin (Ctnnb1) has been shown to play critical roles in the development and maintenance of epithelial cells, including the retinal pigment epithelium (RPE). Ctnnb1 is not only a component of intercellular junctions in the epithelium, it also functions as a transcriptional regulator in the Wnt signaling pathway. To identify which of its functional modalities is critically involved in mouse RPE development and maintenance, we varied Ctnnb1 gene content and activity in mouse RPE lineage cells and tested their impacts on mouse eye development. We found that a Ctnnb1 double mutant (Ctnnb1^dm), which exhibits impaired transcriptional activity, could not replace Ctnnb1 in the RPE, whereas Ctnnb1^Y654E, which has reduced affinity for the junctions, could do so. Expression of the constitutively active Ctnnb1^∆ex3 mutant also suppressed the development of RPE, instead facilitating a ciliary cell fate. However, the post-mitotic or mature RPE was insensitive to the loss, inactivation, or constitutive activation of Ctnnb1. Collectively, our results suggest that Ctnnb1 should be maintained within an optimal range to specify RPE through transcriptional regulation of Wnt target genes in the optic neuroepithelium.

• Molecules and Cells
• /
• v.44 no.3
• /
• pp.168-178
• /
• 2021

The retinal pigment epithelium (RPE) forms a monolayer sheet separating the retina and choroid in vertebrate eyes. The polarized nature of RPE is maintained by distributing membrane proteins differentially along apico-basal axis. We found the distributions of these proteins differ in embryonic, post-natal, and mature mouse RPE, suggesting developmental regulation of protein trafficking. Thus, we deleted tumor susceptibility gene 101 (Tsg101), a key component of endosomal sorting complexes required for transport (ESCRT), in embryonic and mature RPE to determine whether ESCRT-mediated endocytic protein trafficking correlated with the establishment and maintenance of RPE polarity. Loss of Tsg101 severely disturbed the polarity of RPE, which forms irregular aggregates exhibiting non-polarized distribution of cell adhesion proteins and activation of epidermal growth factor receptor signaling. These findings suggest that ESCRT-mediated protein trafficking is essential for the development and maintenance of RPE cell polarity.

• Biomolecules & Therapeutics
• /
• v.30 no.3
• /
• pp.291-297
• /
• 2022

Dry age-related macular degeneration (AMD) is a type of progressive blindness that is primarily due to dysfunction and the loss of retinal pigment epithelium (RPE). The accumulation of N-retinylidene-N-retinylethanolamine (A2E), a by-product of the visual cycle, causes RPE and photoreceptor degeneration that impairs vision. Genes associated with dry AMD have been identified using a blue light model of A2E accumulation in the retinal pigment epithelium and transcriptomic studies of retinal tissue from patients with AMD. However, dry macular degeneration progresses slowly, and current approaches cannot reveal changes in gene transcription according to stages of AMD progression. Thus, they are limited in terms of identifying genes responsible for pathogenesis. Here, we created a model of long-term exposure to identify temporally-dependent changes in gene expression induced in human retinal pigment epithelial cells (ARPE-19) exposed to blue light and a non-cytotoxic dose of A2E for 120 days. We identified stage-specific genes at 40, 100, and 120 days, respectively. The expression of genes corresponding to epithelial-mesenchymal transition (EMT) during the early stage, glycolysis and angiogenesis during the middle stage, and apoptosis and inflammation pathways during the late stage was significantly altered by A2E and blue light. Changes in the expression of genes at the late stages of the EMT were similar to those found in human eyes with late-stage AMD. Our results provide further insight into the pathogenesis of dry AMD induced by blue light and a novel model in vitro with which relevant genes can be identified in the future.

• Advances in nano research
• /
• v.13 no.1
• /
• pp.87-96
• /
• 2022

Drug self-delivery systems can easily realize combination drug therapy and avoid carrier-induced toxicity and immunogenicity because they do not need non-therapeutic carrier materials. So, designing appropriate drug self-delivery systems for specific diseases can settle most of the problems existing in traditional drug delivery systems. Retinal pigment epithelium is very important for the homeostasis of retina. However, it is vulnerable to oxidative damage and difficult to repair. Worse still, the antioxidants can hardly reach the retina by non-invasive administration routes due to the ocular barriers. Herein, the targeted group (folic acid) and antioxidant (melatonin) have been grafted on the surface of ZnO quantum dots to fabricate a new kind of drug self-delivery systems as a protectant via eyedrops. In this study, the negative nanoparticles with size ranging in 4~6 nm were successfully synthesized. They could easily and precisely deliver drugs to retinal pigment epithelium via eyedrops. And they realized acid degradation to controlled release of melatonin and zinc in retinal pigment epithelium cells. Consequently, the structure of retinal pigment epithelium cells were stabilized according to the expression of ZO-1 and β-catenin. Moreover, the antioxidant capacity of retinal pigment epithelium were enhanced both in health mice and photic injury mice. Therefore, such new drug self-delivery systems have great potential both in prevention and treatment of oxidative damage induced retinal diseases.

• BMB Reports
• /
• v.56 no.2
• /
• pp.84-89
• /
• 2023

The implications of nutrient starvation due to aging on the degeneration of the retinal pigment epithelium (RPE) is yet to be fully explored. We examined the involvement of AMPK activation in mitochondrial homeostasis and its relationship with the maintenance of a healthy mitochondrial population and epithelial characteristics of RPE cells under nutrient starvation. Nutrient starvation induced mitochondrial senescence, which led to the accumulation of reactive oxygen species (ROS) in RPE cells. As nutrient starvation persisted, RPE cells underwent pathological epithelial-mesenchymal transition (EMT) via the upregulation of TWIST1, a transcription regulator which is activated by ROS-induced NF-κB signaling. Enhanced activation of AMPK with metformin decelerated mitochondrial senescence and EMT progression through mitochondrial biogenesis, primed by activation of PGC1-α. Thus, by facilitating mitochondrial biogenesis, AMPK protects RPE cells from the loss of epithelial integrity due to the accumulation of ROS in senescent mitochondria under nutrient starvation.

• Polymer(Korea)
• /
• v.38 no.1
• /
• pp.24-30
• /
• 2014

Retinal pigment epithelium (RPE) plays an important role in maintaining the visual function and the degeneration of the RPE causes several retinal degeneration disease. In order to fabricate the suitable carrier for RPE transplantation, the hybrid poly(lactide-co-glycolide) (PLGA) film with hesperidin was prepared. Hesperidin has an anti-inflammatory and antioxidant characteristics. ARPE-19 was seeded on hesperidin/PLGA film and then, cell proliferation was determined by the MTT assay, and cell adhesion and cell morphology were confirmed by SEM. Also, RT-PCR was performed to confirm the expression of the specific genes, and AEC immunohistochemical staining was performed to determine the expression of RPE65. As a result, we confirmed that attachment, proliferation and phenotype maintenance of RPE cells were more excellent on hesperidin/PLGA film than PLGA film, thereby we were able to confirm the potential applications of hesperidin/PLGA film as tissue engineering carrier for regeneration of retina.

• BMB Reports
• /
• v.53 no.5
• /
• pp.284-289
• /
• 2020

Tamoxifen, a nonsteroidal estrogen receptor (ER) antagonist, is used routinely as a chemotherapeutic agent for ER-positive breast cancer. However, it is also causes side effects, including retinotoxicity. The retinal pigment epithelium (RPE) has been recognized as the primary target of tamoxifen-induced retinotoxicity. The RPE plays an essential physiological role in the normal functioning of the retina. Nonetheless, potential therapeutic agents to prevent tamoxifen-induced retinotoxicity in breast cancer patients have not been investigated. Here, we evaluated the action mechanisms of sulfasalazine against tamoxifen-induced RPE cell death. Tamoxifen induced reactive oxygen species (ROS)-mediated autophagic cell death and caspase-1-mediated pyroptosis in RPE cells. However, sulfasalazine reduced tamoxifen-induced total ROS and ROS-mediated autophagic RPE cell death. Also, mRNA levels of tamoxifen-induced pyroptosis-related genes, IL-1β, NLRP3, and procaspase-1, also decreased in the presence of sulfasalazine in RPE cells. Additionally, the mRNA levels of tamoxifen-induced AMD-related genes, such as complement factor I (CFI), complement factor H (CFH), apolipoprotein E (APOE), apolipoprotein J (APOJ), toll-like receptor 2 (TLR2) and toll-like receptor 4 (TLR4), were downregulated in RPE cells. Together, these data provide novel insight into the therapeutic effects of sulfasalazine against tamoxifen-induced RPE cell death.

• Journal of Microbiology and Biotechnology
• /
• v.17 no.6
• /
• pp.1024-1030
• /
• 2007

We investigated the human apolipoprotein E2 (apoE2) transgenic mouse as an animal model system for age-related macular degeneration (AMD). Transgenic mice expressing human apoE2 and C57BL/6J mice were fed normal chow or a high-fat diet for 4 weeks. Eyes were collected from the mice and lipid deposits in retinal pigment epithelium (RPE) were assessed using electron microscopy. The expressions of apoE, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and pigment-epithelium derived factor (PEDF), which are molecular markers for angiogenesis, were assessed with immunohistochemistry. Eyes from apoE2 mice, regardless of diet, contained lipid accumulation in RPE under electron microscopy, whereas control C57BL/6J eyes did not. Lipid accumulation was found predominantly in the RPE and the Bruch's membrane and increased in the eyes of apoE2 mice after one month of a high-fat diet ($8{\pm}2\;per\;50{\mu}m^2$ for normal chow and $11{\pm}2\;per\;50\;{\mu}m^2,\;p<0.05)$. ApoE expression was similar in the apoE2 and control mice; however, VEGF and bFGF were overexpressed in the retinal pigment epithelium of apoE2 eyes compared with control eyes, and PEDF expression was slightly decreased. These expression patterns of VEGF, bFGF, and PEDF suggest angiogenesis is progressing in apoE2 eyes. In conclusion, the eyes of apoE2 mice develop typical lipid accumulations, a common characteristic of AMD, making them a suitable animal model for AMD. The expression profile of VEGF and bFGF on the retinal pigment epithelium suggests that apoE2 may induce neovascularization by altering angiogenic cytokines.