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  • Isolation and characterization of human optic nerve head astrocytes and lamina cribrosa cells.

Isolation and characterization of human optic nerve head astrocytes and lamina cribrosa cells.

Experimental eye research (2020-06-12)
Navita N Lopez, Abbot F Clark, Tara Tovar-Vidales
摘要

The lamina cribrosa is the initial site of glaucomatous injury. Pathological changes to the lamina cribrosa include posterior displacement of the lamina cribrosa, loss of trophic support, and remodeling of the extracellular matrix. Optic nerve head (ONH) astrocytes and lamina cribrosa cells synthesize extracellular matrix proteins to support and maintain the lamina cribrosa under physiological conditions. During glaucoma, these cells respond to mechanical strain and other stimuli, which leads to pathological remodeling of the ONH. Although ONH astrocytes and lamina cribrosa cells have been previously cultured, there is no well-accepted, straightforward technique to isolate both cell types from a single dissected human ONH. To better understand the pathophysiology of glaucoma, we obtained and cultured lamina cribrosa explants from human donor eyes. Initially, cells that grew out from the explant were ONH astrocytes and lamina cribrosa cells. Using a specialized medium, we isolated pure populations of lamina cribrosa cells and ONH astrocytes. ONH astrocytes expressed glial fibrillary acidic protein (GFAP). Lamina cribrosa cells expressed alpha-smooth muscle actin (α-SMA), but were negative for GFAP. This method of ONH cell isolation and cell-culture will provide a technique to better understand the molecular and cell-specific changes in glaucomatous damage to the ONH.

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Sigma-Aldrich
杜氏磷酸盐缓冲盐水, Modified, without calcium chloride and magnesium chloride, liquid, sterile-filtered, suitable for cell culture
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青链霉素, Solution stabilized, with 10,000 units penicillin and 10 mg streptomycin/mL, 0.1 μm filtered, BioReagent, suitable for cell culture
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抗层粘连蛋白 兔抗, 0.5 mg/mL, affinity isolated antibody, buffered aqueous solution
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抗-肌动蛋白,α-平滑肌- FITC抗体,小鼠单克隆 小鼠抗, clone 1A4, purified from hybridoma cell culture
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F-10 营养培养基, With sodium bicarbonate, without L-glutamine, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Monoclonal Anti-Neural Cell Adhesion Molecule antibody produced in mouse, clone NCAM-0B11, ascites fluid