Control of the Corneal Epithelial Stem/Progenitor Cell Niche

Lead Research Organisation: Cardiff University
Department Name: Optometry and Vision Sciences

Abstract

The corneal epithelium is vital for healthy vision and its malfunction owing to injury or disease leads to irreversible blindness. The epithelium is maintained by a population of slow-cycling adult stem/progenitor cells located at the edge of the cornea. These exist basally and within radial infoldings of the epithelial basement membrane. We hypothesise that cellular cross-talk and signals from the matrix help maintain the stem/progenitor cell niche. This is based on our discovery that specific sulphation motifs of chondroitin sulphate are putative biomarkers for articular cartilage progenitor cells (J Histochem Cytochem 2008;56:125-131). Pilot investigations of rabbit cornea support this concept and indicate that chondroitin sulphate is preferentially located next to the epithelial basal cells at the corneal periphery.
This studentship will comprise an in-depth investigation of the stem/progenitor cell niche at the edge of the cornea and in corneal cells in culture. Immunohistochemistry will be conducted using a battery of monoclonal antibodies developed in the Caterson laboratory against variously sulphated chondroitin sulphate epitopes. This will allow identification of matrix markers of the corneal stem/progenitor cell niche. Co-localisation with putative identifiers of stem/progenitor cells (CD-90, CD-105, p63 and Beta-catenin) will clarify specific co-associations between cells and their immediate environment in vivo and in vitro. Higher resolution imaging by immunoelectron microscopy with nanogold particles will distinguish cell surface-associated, or matrix-associated, localisation of the chondroitin sulphate biomarkers. Pilot studies of rabbit cornea have also provided tantalising evidence of what appear to be direct connections between fibroblasts and epithelial cells at the cornea's edge (Fig 2). This will be investigated at high magnification and in three-dimensions using the emerging technique of serial block face scanning electron microscopy to ascertainthe nature and extent of these proposed intercellular connections in, and away from, the niche. Finally, to identify the role of the stem/progenitor cell niche in corneal epithelial dysfunction, changes in the presence, distribution and sulphation specificity of chondroitin sulphate -- and of the pattern of cell-cell connections across the epithelial basement membrane -- will be investigated in tissue obtained postoperatively from eyes with corneal epithelial stem cell deficiencies, caused either by disease or chemical injury.
It is also intended that in the second year of study, the student will have a summer placement in the lab of the research collaborators in Kyoto, Japan.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M009122/1 01/10/2015 30/09/2023
1803680 Studentship BB/M009122/1 01/10/2016 31/03/2021 Greg Hammond
 
Description The human cornea is comprised of five layers; the pig cornea has historically been stated to only have four layers- missing Bowman's layer between the surface epithelial cell layer and the stroma. I have demonstrated that the pig cornea does include this layer and have acquired numerical data to further support the presence of this layer.
A specific form of the carbohydrate chondroitin sulphate has been identified as part of the stem cell niche of corneal epithelium of the pig, in close association with cells positive for potential stem cell markers. This is now the third mammalian species to demonstrate this and strongly suggests this form of chondroitin sulphate could have a role in maintaining stem cells.
I have shown that X-ray micro-computed tomography can be used for visualising and creating 3D reconstructions of the corneal epithelium.
I have also shown, with 3D tissue reconstruction, that direct cell-to-cell contact occurs between stromal cells of the corneal epithelial stem cell niche and basal epithelial cells in the pig eye. This is again the third mammalian species to show this.
Exploitation Route As the chondroitin sulphate form identified by antibody 6C3 has now been identified as part of the pig, rabbit, and human limbal stem cell niches, work needs to be done to characterise this form and its effect on limbal stem cells. This could lead to better methods of culturing stem cells and incorporation into niche reconstruction therapies that could lead to better health outcomes for sufferers of limbal stem cell deficiency.
There is also further work required to examine Bowman's layer and the use of microCT with ocular tissue which could be done by researchers at multiple levels.
Sectors Healthcare,Manufacturing, including Industrial Biotechology,Other

 
Description Contribution to Welsh eye care policy during Professional Internship Placement
Geographic Reach Local/Municipal/Regional 
Policy Influence Type Participation in a advisory committee
 
Description Continuing collaboration with KPUM 
Organisation Kyoto Prefectural University of Medicine
Country Japan 
Sector Academic/University 
PI Contribution The School of Optometry and Vision Sciences at Cardiff University has enjoyed strong links with the Ophthalmology Department of Kyoto Prefectural University of Medicine and this project has continued these links. We contributed to this collaboration by sending researchers to KPUM for short research placements and to present the Research Group's work to the Ophthalmology department.
Collaborator Contribution KPUM hosted me for three weeks to carry out research on their banked human tissue, including limbal stem cell deficient tissue, which we would not have access to in the UK.
Impact Added support to the Research Group's application to the Japanese Society for the Promotion of Science (JSPS) for continued funding to support the collaboration.