July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Biomimetic modelling of the lamina cribrosa region using tissue engineered scaffolds - a novel 3D model for glaucoma research
Author Affiliations & Notes
  • Deirdre Brennan
    Anatomy, School of Medicine, University College Dublin, Belfield, Dublin 4, Ireland
  • Deirdre Clissmann
    Anatomy, School of Medicine, University College Dublin, Belfield, Dublin 4, Ireland
  • Rory Murphy
    Clinical Research Centre, School of Medicine, University College Dublin, Nelson Street, Dublin 7, Ireland
  • Deborah Wallace
    Clinical Research Centre, School of Medicine, University College Dublin, Nelson Street, Dublin 7, Ireland
  • Irina Pascu
    Tissue Engineering Research Group (TERG), Royal College of Surgeons in Ireland, 123 St. Stephen's Green, Dublin 2, Ireland
  • Alan Hibbitts
    Tissue Engineering Research Group (TERG), Royal College of Surgeons in Ireland, 123 St. Stephen's Green, Dublin 2, Ireland
  • Fergal J O'Brien
    Tissue Engineering Research Group (TERG), Royal College of Surgeons in Ireland, 123 St. Stephen's Green, Dublin 2, Ireland
  • Colm J O'Brien
    Clinical Research Centre, School of Medicine, University College Dublin, Nelson Street, Dublin 7, Ireland
    Institute of Ophthalmology, Mater Misericordiae University Hospital, 60 Eccles Street, Dublin 7, Ireland
  • Footnotes
    Commercial Relationships   Deirdre Brennan, None; Deirdre Clissmann, None; Rory Murphy, None; Deborah Wallace, None; Irina Pascu, None; Alan Hibbitts, None; Fergal O'Brien, None; Colm O'Brien, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 6168. doi:
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      Deirdre Brennan, Deirdre Clissmann, Rory Murphy, Deborah Wallace, Irina Pascu, Alan Hibbitts, Fergal J O'Brien, Colm J O'Brien; Biomimetic modelling of the lamina cribrosa region using tissue engineered scaffolds - a novel 3D model for glaucoma research. Invest. Ophthalmol. Vis. Sci. 2019;60(9):6168.

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

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Abstract

Purpose : Fibrotic remodeling and loss of compliance (increased stiffness) are characteristic hallmarks of glaucomatous tissue change in the lamina cribrosa (LC) region of the optic nerve head. Mediating these glaucoma-related changes at the cellular level, are resident fibroblast-like LC cells. To accurately study the pro-fibrotic activity of LC cells, requires an appropriate cell culture model system to replicate the complex glaucoma tissue microenvironment. Thus, the purpose of this study was to create a novel, 3D in-vitro model of the stiffened glaucomatous LC utilizing bioengineered scaffolds.

Methods : A suspension of 84% collagen type I, 7% chondroitin-6-sulphate and 9% elastin in 0.05M acetic acid was freeze-dried to fabricate LC scaffolds (porosity 80-100µm). Scaffolds were EDAC chemically crosslinked to produce ‘normal’ and glaucomatous ‘stiff’ scaffold compliance reference values. Normal primary human LC cells were seeded onto scaffolds at a density of 3500cells/mm2 and cultured for 5 days (n=2 biological replicates, n=11 scaffolds/group) and processed for histological evaluation.

Results : The microporous 3D appearance of fabricated LC scaffolds was comparable in both ‘normal’ and ‘stiff’ groups. (Fig.1) Micro-CT scanning provided a representative overview of the mesh-like structure of the bioengineered scaffolds (1A). The collagen type I composition (gold), confirmed using picrosirius red staining with birefringence, was abundantly found throughout the scaffold structure (1B). The elastin content (green, arrows) was validated with immunofluorescence (1C), and was localized to the scaffold beams using Verhoff-van Gieson stain (black, arrows, 1D). Histological analysis using eosin staining (1E) and scanning electron microscopy (1F) confirmed fabricated scaffolds had a porous ultrastructure. Analysis of cell-seeded scaffolds showed similar successful infiltration and adherence of primary LC cells within both ‘normal’ and ‘stiff’ scaffolds groups (arrows, Fig.2).

Conclusions : Novel 3D biomimetic scaffolds were developed to have similar structure and composition to the native LC. Primary LC cells have successfully been grown for the first time within these glaucomatous ‘stiff’ and ‘normal’ 3D scaffolds. This new in-vitro model of glaucoma has significant potential for deciphering the underlying fibrotic response of LC cells to stiffened glaucomatous conditions.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

 

 

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