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Title: A novel approach of harvesting viable single cells from donor corneal endothelium for cell-injection therapy
Authors: Ong, Hon Shing
Peh, Gary
Neo, Dawn Jin Hui
Ang, Heng-Pei
Adnan, Khadijah
Nyein, Chan Lwin
Morales-Wong, Fernando
Bhogal, Maninder
Kocaba, Viridiana
Mehta, Jodhbir Singh
Keywords: Science::Medicine
Issue Date: 2020
Source: Ong, H. S., Peh, G., Neo, D. J. H., Ang, H.-P., Adnan, K., Nyein, C. L., . . . Mehta, J. S. (2020). A novel approach of harvesting viable single cells from donor corneal endothelium for cell-injection therapy. Cells, 9(6), 1428-. doi:10.3390/cells9061428
Project: JRNMRR163801 
Journal: Cells 
Abstract: Donor corneas with low endothelial cell densities (ECD) are deemed unsuitable for corneal endothelial transplantation. This study evaluated a two-step incubation and dissociation harvesting approach to isolate single corneal endothelial cells (CECs) from donor corneas for corneal endothelial cell-injection (CE-CI) therapy. To isolate CECs directly from donor corneas, optimization studies were performed where donor Descemet’s membrane/corneal endothelium (DM/CE) were peeled and incubated in either M4-F99 or M5-Endo media before enzymatic digestion. Morphometric analyses were performed on the isolated single cells. The functional capacities of these cells, isolated using the optimized simple non-cultured endothelial cells (SNEC) harvesting technique, for CE-CI therapy were investigated using a rabbit bullous keratopathy model. The two control groups were the positive controls, where rabbits received cultured CECs, and the negative controls, where rabbits received no CECs. Whilst it took longer for CECs to dislodge as single cells following donor DM/CE incubation in M5-Endo medium, CECs harvested were morphologically more homogenous and smaller compared to CECs obtained from DM/CE incubated in M4-F99 medium (p < 0.05). M5-Endo medium was hence selected as the DM/CE incubation medium prior to enzymatic digestion to harvest CECs for the in vivo cell-injection studies. Following SNEC injection, mean central corneal thickness (CCT) of rabbits increased to 802.9 ± 147.8 μm on day 1, gradually thinned, and remained clear with a CCT of 385.5 ± 38.6 μm at week 3. Recovery of corneas was comparable to rabbits receiving cultured CE-CI (p = 0.40, p = 0.17, and p = 0.08 at weeks 1, 2, and 3, respectively). Corneas that did not receive any cells remained significantly thicker compared to both SNEC injection and cultured CE-CI groups (p < 0.05). This study concluded that direct harvesting of single CECs from donor corneas for SNEC injection allows the utilization of donor corneas unsuitable for conventional endothelial transplantation.
ISSN: 2073-4409
DOI: 10.3390/cells9061428
Schools: School of Materials Science and Engineering 
Rights: © 2020 The Authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (
Fulltext Permission: open
Fulltext Availability: With Fulltext
Appears in Collections:MSE Journal Articles

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