May 2006
Volume 47, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2006
Optimization of Human Endothelial Cell Culture and Donor Stromal Button for the Reconstruction of a Partial Thickness Posterior Cornea
Author Affiliations & Notes
  • P. Carrier
    Departments of Ophthalmology and Surgery, LOEX/Laval University, Quebec, PQ, Canada
  • S. Proulx
    Departments of Ophthalmology and Surgery, LOEX/Laval University, Quebec, PQ, Canada
  • J.–M. Bourget
    Departments of Ophthalmology and Surgery, LOEX/Laval University, Quebec, PQ, Canada
  • A. Deschambeault
    Departments of Ophthalmology and Surgery, LOEX/Laval University, Quebec, PQ, Canada
  • I. Brunette
    Department of Ophthalmology, Montreal University, Quebec, PQ, Canada
  • L. Germain
    Departments of Ophthalmology and Surgery, LOEX/Laval University, Quebec, PQ, Canada
  • Footnotes
    Commercial Relationships  P. Carrier, None; S. Proulx, None; J. Bourget, None; A. Deschambeault, None; I. Brunette, None; L. Germain, None.
  • Footnotes
    Support  Canadian Institutes of Health Research (CIHR) Grant, Réseau de Recherche en Santé de la Vision from the Fonds de la Recherche en Santé du Québec (FRSQ)
Investigative Ophthalmology & Visual Science May 2006, Vol.47, 4389. doi:https://doi.org/
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      P. Carrier, S. Proulx, J.–M. Bourget, A. Deschambeault, I. Brunette, L. Germain; Optimization of Human Endothelial Cell Culture and Donor Stromal Button for the Reconstruction of a Partial Thickness Posterior Cornea . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4389. doi: https://doi.org/.

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

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Abstract

Purpose: : Bullous keratopathy and Fuchs' corneal dystrophy are responsible for more than 40% of corneal transplantations in Canada and the US. However, complications related to endothelial cells attrition and endothelial rejection reduce the success rate of this procedure. This study was conducted in order to improve the outcome of corneal transplantation. Our purpose was to evaluate the feasibility of an autologous endothelial graft by transplanting the host’s cultured endothelial cell on the posterior surface of a donor cornea.

Methods: : Our first goal was to optimize culture of human endothelial cells. We analyzed the morphology and proliferation of cells cultured on either a fibronectin and collagen matrix (FNC), plastic or a feeder layer of 3T3. We then studied the growth behavior of central and peripheral corneal cells. Next, endothelial cells were isolated, seeded and cultured for 24h or 14d on the posterior surface of allogeneic lamellar stromal buttons and processed for histology. We also evaluated whether Descemet’s membrane should be peeled off or left intact for a better adhesion of the cells. Finally, the effect of presence of living stromal keratocytes within the donor button was assessed.

Results: : Human endothelial cells cultured on FNC proliferated faster then those cultured on plastic or with a feeder layer. However, cells cultured on this matrix presented a fibroblast–like morphology. Cells isolated from the peripheral cornea were characterized by a higher mitogenic activity and had a different morphology when compared to cells derived from the central cornea. Cultured cells seeded on allogeneic stromal buttons adhered and proliferated to form a nice and flat corneal endothelial monolayer similar to that observed in vivo. Cells seeded on Descemet’s membrane adhered more strongly than those seeded directly on the stroma. The presence or absence of living keratocytes seemed to have no effect on the adhesion and proliferation of the seeded endothelial cells.

Conclusions: : This study provides evidence for the feasibility of an autologous endothelial graft by the seeding of endothelial cells on stromal buttons. Moreover, these experiments lead to the optimization of the conditions for the reconstruction of a partial thickness posterior cornea. They also provide informations on the role of Descemet’s membrane and living keratocytes for the generation of highly cellular and healthy endothelium on the donor corneal button.

Keywords: cornea: endothelium • cornea: basic science • transplantation 
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