Endothelial Wound Repair of the Organ-Cultured Porcine Corneas

Smeringaiova, Ingrida; Merjava, Stanislava; Stranak, Zbynek; Studený, Pavel; Bednář, Jan; Jirsová, Kateřina

doi:10.1080/02713683.2018.1458883

Cited by 7 publications

(5 citation statements)

References 39 publications

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“…These results were translated to ex vivo human corneas as well. It has been shown that porcine and rabbit corneal endothelial cells have a greater proliferative capacity than humans (23,24,26,52). This baseline difference leads to earlier wound closure of these species compared to the humans.…”

Section: Discussionmentioning

confidence: 99%

Extracellular Vesicles Derived From Human Corneal Endothelial Cells Inhibit Proliferation of Human Corneal Endothelial Cells

et al. 2022

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Corneal endothelial cells (CEnCs) are a monolayer of hexagonal cells that are responsible for maintaining the function and transparency of the cornea. Damage or dysfunction of CEnCs could lead to blindness. Human CEnCs (HCEnCs) have shown limited proliferative capacity in vivo hence, their maintenance is crucial. Extracellular vesicles (EVs) are responsible for inter- and intra-cellular communication, proliferation, cell-differentiation, migration, and many other complex biological processes. Therefore, we investigated the effect of EVs (derived from human corneal endothelial cell line–HCEC-12) on corneal endothelial cells. HCEC-12 cells were starved with serum-depleted media for 72 h. The media was ultracentrifuged at 100,000xg to isolate the EVs. EV counting, characterization, internalization and localization were performed using NanoSight, flow cytometry, Dil labeling and confocal microscopy respectively. HCEC-12 and HCEnCs were cultured with media supplemented with EVs. Extracted EVs showed a homogeneous mixture of exosomes and microvesicles. Cells with EVs decreased the proliferation rate; increased apoptosis and cell size; showed poor wound healing response in vitro and on ex vivo human, porcine, and rabbit CECs. Thirteen miRNAs were found in the EV sample using next generation sequencing. We observed that increased cellular uptake of EVs by CECs limit the proliferative capacity of HCEnCs. These preliminary data may help in understanding the pathology of corneal endothelial dysfunction and provide further insights in the development of future therapeutic treatment options.

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Section: Discussionmentioning

confidence: 99%

Extracellular Vesicles Derived From Human Corneal Endothelial Cells Inhibit Proliferation of Human Corneal Endothelial Cells

et al. 2022

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“…These results were translated to ex vivo human corneas as well. It has been shown that porcine and rabbit corneal endothelial cells have a greater proliferative capacity than humans [23,24,26,40]. Both, porcine and rabbit endothelial cells treated with human EVs showed slow wound healing response.…”

Section: Discussionmentioning

confidence: 99%

Extracellular Vesicles Derived From Human Corneal Endothelial Cells Inhibit Proliferation of Human Corneal Endothelial Cells

Parekh

Rhys

Ramos

et al. 2021

Preprint

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Corneal endothelial cells (CEnCs) are a monolayer of hexagonal cells that are responsible for maintaining the function and transparency of the cornea. Damage or dysfunction of CEnCs could lead to blindness. Human CEnCs (HCEnCs) have shown limited proliferative capacity in vivo hence, their maintenance is crucial. Extracellular vesicles (EVs), are responsible for inter- and intra-cellular communication, proliferation, cell-differentiation, migration, and many other complex biological processes. Therefore, we investigated the effect of EVs (derived from human corneal endothelial cell line – HCEC-12) on corneal endothelial cells. HCEC-12 cells were starved with serum-depleted media for 72 hours. The media was ultracentrifuged at 100,000xg to isolate the EVs. EV counting, characterization, internalization and localization were performed using NanoSight, flow cytometry, Dil labelling and confocal microscopy respectively. HCEC-12 and HCEnCs were cultured with media supplemented with EVs. Extracted EVs showed a homogeneous mixture of exosomes and microvesicles. Cells with EVs decreased the proliferation rate; increased apoptosis and cell size; showed poor wound healing response in vitro and on ex vivo human, porcine, and rabbit CECs. Thirteen miRNAs were found in the EV sample using next generation sequencing. We observed that increased cellular uptake of EVs by CECs limit the proliferative capacity of HCEnCs. These preliminary data may help in understanding the pathology of corneal endothelial dysfunction and provide further insights in the development of future therapeutic treatment options.

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“…Endothelial morphology and cell density are the most commonly used parameters in the assessment of corneal endothelium in humans and animals [1,3,4,6]. Due to the similarities to human eyes, swine eyes have been used extensively in experimental research [9,17,21]. The ocular bulbs and cornea sizes of pigs, their morphological endothelium characteristics, their wide availability and their low prices are some of the largest advantages of this choice in human and veterinary medicine [17].…”

Section: Discussionmentioning

confidence: 99%

Morphology of Endothelial Cells from Different Regions of the Swine Cornea

Clerot

Hünning

Bettio

et al. 2019

Acta Scientiae. Vet.

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Background:The corneal endothelium is a monolayer of polygonal cells which constitute the last layer of the cornea. The integrity of this layer is critical to cornea transparency. The characterization of normal corneal endothelial morphology is important not only to clinical evaluation but also to selection of areas of the cornea with better quality to be employed as donor tissue. The aim of the present study was to evaluate the morphology of endothelial cells from different regions of the swine cornea after alizarin red staining using optical microscopy. Materials, Methods & Results:Twenty-four healthy eyes from 12 swine Large White breed, with 14-monthold, males or females obtained from a licensed Brazilian commercial slaughterhouse were studied. Immediately after humane slaughter, the eyes were enucleated and submitted to ophthalmic examination. Eyes with signs of diseases of the anterior segment were excluded. The cornea, with 3 mm of the sclera, was removed and placed on a glass microscope slide with the endothelial side up. Four radial incisions were made in the periphery of the cornea to better accommodate the cornea on the microscope slide. Alizarin red was diluted in isotonic solution (0.2 g/100 mL) and the pH was adjusted to 4.2 with hydrochloric acid. Three drops of alizarin red were placed on the corneal endothelium. After 90 s, the dye was removed from the cornea with balanced saline solution. The corneal endothelium was examined and photographed using an optical microscope. All evaluations were performed by the same investigator. Photomicrographs were taken of central, superior, inferior, nasal and temporal corneal areas. Parameter studied included endothelial cell morphology. For the statistical analysis, was employed the ANOVA variance test (repeated measures). Differences were considered statistically significant at P < 0.05. Normal endothelium cells were mainly hexagonal (83.7%), pentagonal (7.45%) and heptagonal (8.8%), with a minimal number of cells of other shapes present. There were no significant statistical differences in the proportion of the morphology and the different regions of the cornea (P = 0.31). Discussion: Different techniques are available for the analysis of corneal endothelium, including mainly scanning electron microscopy, specular microscopy and optical microscopy. The analysis of the morphology of corneal endothelium with an optic microscope after staining with alizarin red has been described as an effective, rapid and cost-efficient method, since this dye blends the borated cells, allowing identification. In the present study, using optical microscopy and coloration with alizarin red it was possible to explore and to obtain images of the swine endothelium of all regions of the cornea. The analysis of the cellular morphology or the percentage of hexagonal cells are among the main parameters used to evaluate the health of the corneal endothelium. In this study, the endothelium had the predominance of the hexagonal shape in all regions studied. In swine, there are no studies...

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Endothelial Wound Repair of the Organ-Cultured Porcine Corneas

Cited by 7 publications

References 39 publications

Extracellular Vesicles Derived From Human Corneal Endothelial Cells Inhibit Proliferation of Human Corneal Endothelial Cells

Extracellular Vesicles Derived From Human Corneal Endothelial Cells Inhibit Proliferation of Human Corneal Endothelial Cells

Extracellular Vesicles Derived From Human Corneal Endothelial Cells Inhibit Proliferation of Human Corneal Endothelial Cells

Morphology of Endothelial Cells from Different Regions of the Swine Cornea

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