Basic Study
Copyright ©The Author(s) 2019. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Stem Cells. Feb 26, 2019; 11(2): 84-99
Published online Feb 26, 2019. doi: 10.4252/wjsc.v11.i2.84
Anti-inflammatory potential of human corneal stroma-derived stem cells determined by a novel in vitro corneal epithelial injury model
Laura E Sidney, Andrew Hopkinson, Owen D McIntosh, Nagi M Marsit, Mariana Lizeth Orozco Morales
Mariana Lizeth Orozco Morales, Nagi M Marsit, Owen D McIntosh, Andrew Hopkinson, Laura E Sidney, Academic Ophthalmology, Division of Clinical Neuroscience, University of Nottingham, Nottingham, NG7 2UH, United Kingdom
Author contributions: Orozco Morales ML performed the majority of the data acquisition and analysis; Marsit NM processed all amniotic membrane used and helped with acquisition of data; McIntosh OD contributed to conception of the study and acquisition of data; Hopkinson A contributed to conception of the study; Sidney LE contributed to conception of the study, design of experiments, data acquisition, data analysis, and interpretation of data; all authors contributed to drafting the article, making revisions, and had final approval of the manuscript.
Institutional review board statement: Human corneoscleral rims and human AM were used with approval by the Nottingham Research Ethics Committee (Ethics approval numbers: 07/H0403/140 and OY110101, respectively).
Informed consent statement: All corneal tissue was supplied anonymously through Manchester Eye Bank, and all informed consent forms are held at this institution. For amniotic membrane, all donors provided written informed consent prior to tissue collection. During research studies all tissue was anonymised to the researchers.
Conflict-of-interest statement: There are no potential conflicts of interest relevant to this study reported by the authors.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Corresponding author: Laura E Sidney, MSc, PhD, Senior Research Fellow, Academic Ophthalmology, Division of Clinical Neuroscience, University of Nottingham, Queen’s Medical Centre Campus, Nottingham NG7 2UH, United Kingdom.
Telephone: +44-115-8230459
Received: July 24, 2018
Peer-review started: July 24, 2018
First decision: October 5, 2018
Revised: November 1, 2018
Accepted: January 1, 2019
Article in press: January 1, 2019
Published online: February 26, 2019
Research background

The cornea provides two thirds of the eye’s refractive power as well as being the major barrier to the inner content of the eye. At present, the most effective treatment of a diseased or damaged cornea is transplantation of a donor cornea (keratoplasty). However, this is not feasible for 8-10 million individuals. Corneal research has turned to the use of stem cell-based regenerative therapies for corneal tissue regeneration. Recent in vitro studies have shown that mesenchymal stem cell-like cells from the corneal stroma (corneal-derived stromal stem cells, CSSC) contribute to corneal tissue homeostasis, presenting an immunomodulatory response, a non-immunogenic profile, and a regenerative role. Thus, CSSC may be considered an appropriate cell source for the treatment of inflammatory disorders of the cornea.

Research motivation

To investigate whether CSSC seeded on an amniotic membrane (AM) substrate have the ability to modulate an inflamed environment, and therefore whether they were suitable candidates for developing a cell therapy for the front of the eye.

Research objectives

The first objective was to optimise an in vitro injury model mimicking a corneal surface using human corneal epithelial cells (hCEC) stimulated with various injurious agents. Once optimised the second objective was to investigate whether CSSC conditioned media and then CSSC in co-culture could modulate the injury environment. The final objective was to seed CSSC on AM and investigate whether the CSSC-AM construct provided an anti-inflammatory, healing response to the injury.

Research methods

Treatment of hCEC with ethanol and pro-inflammatory cytokines were compared in terms of viability loss, cytotoxicity, and pro-inflammatory cytokine release in order to generate the novel in vitro injury. Co-culture experiments were performed with CSSC alone and with CSSC-AM constructs. The effect of injury and co-culture on viability, cytotoxicity, interleukin (IL)-6 and IL-8 production, and IL1B, TNF, IL6, and CXCL8 mRNA expression were assessed.

Research results

An optimal injury of 20% (v/v) ethanol for 30 s with 1 ng/mL IL-1 beta was developed. Co-culture of the injury model with CSSC inhibited loss of hCEC viability caused by injury. Enzyme linked immunosorbent assay and PCR showed a significant reduction in the production of IL-6 and IL-8 pro-inflammatory cytokines and reduction in pro-inflammatory cytokine mRNA expression during co-culture with CSSC alone and with the AM construct.

Research conclusions

The novel findings of this study confirm the therapeutic potential of the CSSC and the possible use of AM as a cell carrier for application to the ocular surface.

Research perspectives

The novel injury model developed in this study can be adapted for studying the therapeutic effects of many different agents. The findings of this study may lead to the development of practise changing cell therapies for treatment of inflammatory disorders of the cornea in clinic.