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Lilangi S Ediriwickrema, Rebecca Fawcett, Neil R Miller, Steven L Bernstein; Decellularization of Porcine and Primate Optic Nerve Lamina towards Cell Culture with Neural Progenitor Cells. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3324.
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© ARVO (1962-2015); The Authors (2016-present)
The optic nerve lamina cribrosa is a mesh-like collagenous structure consisting of an extracellular matrix with varying porosity and glial cell distribution through which axons of retinal ganglion cells travel en route to the lateral geniculate nucleus. There has been significant interest in its structure with respect to its possible role in the pathophysiology of glaucomatous optic neuropathy. Our goal was to evaluate decellularized porcine and primate lamina as three-dimensional matrices for stem cell growth, regeneration, and delivery.
Optic nerve laminae were dissected from porcine and primate cadaveric enucleations. The tissue specimens were embedded into a sugar moiety and subsequently sectioned into variable thicknesses using a cryotome. They were then decellularized using a chemical detergent buffer (Tris-SDS-EDTA) over several pre-determined time-points. Electron microscopy and histological analysis using Gömöri trichrome staining confirmed complete decellularization. Select decellularized scaffolds were then sterilized, seeded with either squamous cell carcinoma cells (SCC) or human neural progenitor cells, and maintained in stem cell culture.
Gömöri trichrome staining confirmed that porcine or primate optic nerve lamina sectioned at 50 µm, 100 µm, or 180 µm thicknesses were completely decellularized at 45 minutes, 1 hour, or 2 hours, respectively. Sterilized porcine laminar sections tolerated SCC and neural progenitor cell adhesion, survival, and three-dimensional growth in culture, confirming biocompatibility.
Chemical detergent buffer immersion results in complete decellularization of porcine and primate optic nerve lamina. These porous extracellular matrices may have a potential role as a scaffold for cellular growth and neural regeneration in treating optic neuropathy.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
Figure 1. Decellularization of the Porcine Optic Nerve Lamina
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