March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Vitronectin and its Receptor αvβ5 in Limbal Epithelial Stem Cell Maintenance
Author Affiliations & Notes
  • Nick Di Girolamo
    School of Medical Sciences - Pathology, University of New South Wales, Sydney, Australia
  • Paula Ordonez
    School of Medical Sciences - Pathology, University of New South Wales, Sydney, Australia
  • Sharron Chow
    School of Medical Sciences - Pathology, University of New South Wales, Sydney, Australia
  • Denis Wakefield
    School of Medical Sciences - Pathology, University of New South Wales, Sydney, Australia
  • Footnotes
    Commercial Relationships  Nick Di Girolamo, None; Paula Ordonez, None; Sharron Chow, None; Denis Wakefield, None
  • Footnotes
    Support  University of New South Wales Gold Star Award
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 1709. doi:
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      Nick Di Girolamo, Paula Ordonez, Sharron Chow, Denis Wakefield; Vitronectin and its Receptor αvβ5 in Limbal Epithelial Stem Cell Maintenance. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1709.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : Limbal stem cell deficiency (LSCD) is a blinding corneal condition treatable through expensive, time consuming, and complicated autologous or allogeneic stem cell (SC) transplantation. If corneo-limbal SCs (LSCs) can be more readily identified, isolated, and maintained ex vivo, better quality and more stable grafts could be prepared for patients suffering this debilitating disease. We postulate that the extracellular matrix (ECM) glycoprotein, vitronectin (VN) and its integrin receptor αvβ5 play an integral role in supporting LSCs within their niche microenvironment.

Methods: : Histological and immunolocalization studies were conducted on cadaveric human fetal and adult ocular tissues. Limbal epithelial cells were expanded from donor human corneal rims using a tissue explant technique or enzymatic dissociation and αvβ5 expression determined by flow cytometry. Integrin αvβ5+/- cells were further selected using either magnetic microbeads or FACS sorting. Functional colony forming assays, and immunocytological and RT-PCR assays for SC marker expression were conducted on fractionated cells.

Results: : VN was identified as an ECM component on the limbal basement membrane, and was restricted to SC-like clusters within the limbal region, but was not expressed in the differentiated compartment of the central cornea. Furthermore, its receptor αvβ5-integrin co-localized to N-cadherin+/CK-15+ putative LSCs. αvβ5 expression was restricted to less than 5% of total isolated limbal epithelial cells and was used to enrich for a population of presumed LSCs based on functional colony forming capacity and enhanced expression of LSC-associated genes.

Conclusions: : Our tissue-based studies indicate that a VN/αvβ5 axis plays an important role in maintaining LSCs within their niche. Our in vitro data supports this concept; however until the precise proteomic profile of the niche is disclosed, it may be difficult to establish the ideal culture conditions to nurture these cells long-term. Due to the unique, transparent and accessible nature of the human limbus and its SC reservoir, information gained from these studies may inform researchers developing treatment strategies for other human organ systems.

Keywords: cornea: epithelium • extracellular matrix • cell adhesions/cell junctions 
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