Clustered ARPE-19 cells distinct in mitochondrial membrane potential may play a pivotal role in cell differentiation.

Age-related macular degeneration (AMD) is associated with the dysfunction and degeneration of retinal pigment epithelium (RPE) cells. Here, we examined how the formation and expansions of cell clusters are regulated by the differentiation of the RPE　cells. In this study, ARPE-19 cells were cultivated in standard or differentiation media, i.

Spatiotemporal Coordination of RPE Cell Quality by Extracellular Vesicle miR-494-3p Via Competitive Interplays With SIRT3 or PTEN.

Purpose: To reveal the molecular mechanism underlying degeneration in human retinal pigment epithelial (hRPE) cells with dysfunctional mitochondrial homeostasis.

Methods: The expression of recently identified miR-494-3p in extracellular vesicles (EV) released from induced-pluripotential-stem-cell-derived human RPE (iPS-hRPE), during coculture with macrophages (Mps) was investigated in iPS-hRPE and ARPE cells differentiated in the presence of nicotinamide (Nic-ARPE). The expression of phosphatase and tensin homolog (PTEN), sirtuin3 (SIRT3), and mitochondrial marker proteins before and after the transfection of miR-494-3p inhibitor and mimic, and the changes in mitochondrial metabolism, membrane potential, and oxidative phosphorylation (OXPHOS) were monitored.

Quiescent innate and adaptive immune responses maintain the long-term integrity of corneal endothelium reconstituted through allogeneic cell injection therapy.

This study aims to clarify the immunogenicity in acquired and innate immune responses of cultured human corneal endothelial cells (hCECs) applied for cell injection therapy, a newly established modality for corneal endothelium failures. Thirty-four patients with corneal endothelial failure received injection of allogeneic hCEC suspension into anterior chamber. No sign of immunological rejection was observed in all 34 patients during the 5-8 years postoperative follow-up period.

Repressed miR-34a Expression Dictates the Cell Fate to Corneal Endothelium Failure.

Purpose: To reveal the mechanism triggering the functional disparity between degenerated and non-degenerated corneal endothelium cells in the water efflux from corneal stroma to the anterior chamber.

Methods: The varied levels of the microRNA (miR)-34, miR-378, and miR-146 family in human corneal endothelium and cultured cells thereof were investigated using 3D-Gene Human miRNA Oligo Chips. Concomitantly, CD44, p53, c-Myc, matrix metalloprotease (MMP)-2 expression, and Ras homolog gene family member A (Rho A) activity was correlated to the expression intensities of these microRNAs, partly complemented with their altered expression levels with the transfection of the corresponding mimics and inhibitors.

Intracellular pH affects mitochondrial homeostasis in cultured human corneal endothelial cells prepared for cell injection therapy.

This study aimed to uncover the mechanism responsible for the clinical efficacy of cell injection therapy with fully differentiated cultured cells. Analysis of polarized expression of ion transporters on cultured human corneal endothelial cells (CECs) subpopulations (SPs) was performed. The intracellular pH (pHi) between two CEC SPs, distinct in the proportion of differentiated cells, was measured, and the association with mitochondrial respiration homeostasis was investigated.

Potential participation of CTRP6, a complement regulator, in the pathology of age related macular degeneration.

Purpose: To investigate the localized expression of C1q/tumor necrosis factor related protein (CTRP) 6 in human age-related macular degeneration (AMD) retinal tissues.

Experimental Study Design: 4 AMD and 3 non-AMD whole eyes of Caucasian donors were used. Eyecups were excised at Eye Bank CorneaGen, Inc.

Superiority of Mature Differentiated Cultured Human Corneal Endothelial Cell Injection Therapy for Corneal Endothelial Failure.

Purpose: To investigate the safety and efficacy of cultured human corneal endothelial cell (hCEC) injection therapy with mature differentiated (mature) cell subpopulations (SPs) for corneal endothelial failure (CEF).

Design: Comparative, interventional case series.

Methods: This study involved 18 eyes with CEF that underwent cultured hCEC injection therapy, categorized into 2 groups: (1) 11 eyes administered a relatively lower proportion (0.

Mitochondrial miRNA494-3p in extracellular vesicles participates in cellular interplay of iPS-Derived human retinal pigment epithelium with macrophages.

To explore new molecular targets for therapy in human model systems by discerning the role of extracellular vesicle (EV) microRNAs (miRs) secreted by human retinal pigment epithelium (hRPE) cells and their cellular interplay with macrophages (Mps). Human Mps differentiated from THP-1 cells stimulated by phorbol myristate acetate were co-cultured with induced pluripotent stem cell-derived differentiated hRPE (iPS-hRPE) cells in Transwell® system separated by 0.40 μm or 0.

CD63 extracellular vesicles from retinal pigment epithelial cells participate in crosstalk with macrophages in the innate inflammatory axis.

The aim of the study is to clarify the participation of extracellular vesicles (EV) secreted by murine primary retinal pigment epithelial (mpRPE) cells in the cell to cell communication with macrophages (Mps), firstly described by the authors in 2016. In ocular inflammation, Mps act as sources of tumor necrosis factor-α (TNF-α), an activator of RPE cells. TNF-α stimulates the production of monocyte chemotactic protein (MCP-1) by RPE cells, thereby causing greater recruitment of Mps to the sub-RPE space.

Epigenetic regulation of the epithelial mesenchymal transition induced by synergistic action of TNF-α and TGF-β in retinal pigment epithelial cells.

To clarify the influence of tumor necrosis factor (TNF)-α on fibrotic phenotypes induced by transforming growth factor (TGF)-β in retinal pigment epithelial cells (RPECs) by epigenetic regulation. Human primary retinal pigment epithelial cells (RPECs including ARPE19) were used in cultures in the presence or absence of TNF-α and/or TGF-β2. RT2 Profiler™ (Qiagen) was used for PCR Array for fibrosis and epithelial mesenchymal transition (EMT).

Mitochondria as a Platform for Dictating the Cell Fate of Cultured Human Corneal Endothelial Cells.

Purpose: Aiming to clarify the role of mitochondria in cell fate decision of cultured human corneal endothelial cell (cHCEC) subpopulations.

Methods: The mitochondrial respiratory ability were examined with Mito stress and Mito fuel flex test assays using an extracellular flux analyzer (XFe24; Agilent Technologies; Santa Clara, CA) for human corneal endothelium tissues, mature cHCECs and a variety of cell state transitioned cHCECs. Tricarboxylic acid cycle and acetyl-coenzyme A-related enzymes was analyzed by proteomics for cell lysates using liquid chromatography-tandem mass spectrometry for cHCEC subpopulations.

Pluripotent epigenetic regulator OBP-801 maintains filtering blebs in glaucoma filtration surgery model.

Inhibition of fibrosis is indispensable for maintaining filtering blebs after glaucoma filtration surgery (GFS). The purpose of this study was to investigate the ability of a pluripotent epigenetic regulator OBP-801 (OBP) to ameliorate extracellular matrix formation in a rabbit model of GFS. Rabbits that underwent GFS were treated with OBP.

Five-Year Follow-up of First 11 Patients Undergoing Injection of Cultured Corneal Endothelial Cells for Corneal Endothelial Failure.

Purpose: To report the safety and efficacy of a novel cell injection therapy using cultured human corneal endothelial cells (hCECs) for endothelial failure conditions via the report of the long-term 5-year postoperative clinical data from a first-in-humans clinical trial group.

Design: Prospective observational study.

Participants: This study involved 11 eyes of 11 patients with pseudophakic endothelial failure conditions who underwent hCEC injection therapy between December 2013 and December 2014.

Polarized Expression of Ion Channels and Solute Carrier Family Transporters on Heterogeneous Cultured Human Corneal Endothelial Cells.

Purpose: To clarify the expression profiles of ion channels and transporters of metabolic substrates among heterogeneous cultured human corneal endothelial cells (cHCECs) distinct in their effectiveness in reconstituting the corneal endothelium.

Methods: Integrated proteomics for cell lysates by liquid chromatography-tandem mass spectrometry was carried out from three aliquots of cHCECs enriched in either cluster of definition (CD)44-/+ (mature) cHCECs or CD44++/+++ cell-state transition (CST) cHCECs. The expression profiles of cations/anions, monocarboxylic acid transporters (MCTs), and solute carrier (SLC) family proteins, as well as carbonic anhydrases (CAs), were investigated.

Metabolites Interrogation in Cell Fate Decision of Cultured Human Corneal Endothelial Cells.

Purpose: Aiming to clarify the metabolic interrogation in cell fate decision of cultured human corneal endothelial cells (cHCECs).

Methods: To analyze the metabolites in the culture supernatants (CS), 34 metabolome measurements were carried out for mature differentiated and a variety of cHCECs with cell state transition through a facility service. Integrated proteomics research for cell lysates by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed for 3 aliquots of each high-quality or low-quality cHCEC subpopulations (SP).

In Vivo Fluorescence Visualization of Anterior Chamber Injected Human Corneal Endothelial Cells Labeled With Quantum Dots.

Purpose: The injection of cultured human corneal endothelial cells (cHCECs) into the anterior chamber (AC) is a newly developed modality for the successful treatment of corneal endothelium dysfunction. Here, we investigated whether or not cHCECs could be labeled using quantum dots (QDs) composed of semiconductor nanoparticle octa-arginine (R8) to trace injected cHCECs and examined the utility of in vivo fluorescence imaging to analyze the dynamics and accumulation of QD-labeled injected cHCECs in a corneal endothelial dysfunction mouse model.

Methods: The cHCECs, either of high quality or with cell-state transition, were labeled by adding a mixture of QDs655 and R8.

A physical biomarker of the quality of cultured corneal endothelial cells and of the long-term prognosis of corneal restoration in patients.

Dysfunction of the corneal endothelium reduces the transparency of the cornea and can cause blindness. Because corneal endothelial cells have an extremely limited proliferative ability in vivo, treatment for corneal endothelial dysfunction involves the transplantation of donor corneal tissue. Corneal endothelium can also be restored via intraocular injection of endothelial cells in suspension after their expansion in vitro.

Distinctly regulated functions and mobilization of CD11c-positive cells elicited by TLR3- and IPS-1 signaling in the cornea.

The human ocular surface epithelium expresses TLR3, which recognizes double-stranded (ds) RNA mimicking polyinosine-polycytidylic acid (polyI:C). Its stimulation induces the secretion of the inflammatory cytokines such as interleukin (IL)-6, IL-8, and type I interferon. The cytoplasmic helicase proteins RIG-I and MDA5 are also expressed on the ocular surface.

Association of Upregulated Angiogenic Cytokines With Choroidal Abnormalities in Chronic Central Serous Chorioretinopathy.

Purpose: To clarify the distinct molecular pathogenesis of central serous chorioretinopathy (CSC) and pachychoroid neovasculopathy (PNV).

Methods: Aqueous humor (AH) was collected from 18 acute CSC, 20 chronic CSC, and 20 PNV patients. Concentrations of 30 cytokines in the AH were analyzed using a multiplex bead immunoassay, and the cytokine profiles were compared among these three groups of patients.

Distinct Aqueous Humour Cytokine Profiles of Patients with Pachychoroid Neovasculopathy and Neovascular Age-related Macular Degeneration.

This study investigated the pathophysiological features of pachychoroid neovasculopathy (PNV) and neovascular age-related macular degeneration (nAMD) by analysing and comparing cytokine profiles in aqueous humour (AH) collected from 18 PNV, 18 nAMD and 11 control patients. Responses to intravitreal injection of aflibercept were also analysed in the PNV and nAMD groups. In the PNV group, vascular endothelial growth factor (VEGF)-A was significantly lower than in the nAMD group (p = 0.