Primary cell culture of mengiothelial cells ‐ a new model for future studies of optic nerve disease

Purpose Our previous study revealed that the occurrence and amount of meningothelial cell nests in the subarachnoid space were significantly increased in glaucoma histologic examination. For further investigation of the possible role of meningothelial cells in the pathogenesis of optic nerve diseases in vitro, pure meningothelial cells are needed. Accordingly, we developed a culture method of meningothelial cells harvested from the arachnoid layer covering the optic nerve. Methods Menigiothelial cells were prepared from porcine optic nerve sheaths by scraping cells from the arachnoid layer into cell culture media. After outgrowth of cells, contaminating fibroblasts were eliminated using MACS. The remaining cells were characterized using immunofluorescence staining of fibroblast and meningothelial cell markers. Results Primary cells grew within 1‐3 weeks and formed a monolayer composed of two different cell types, flattened cells with round nuclei and spindle shaped cells. Cells were separated by automatic fibroblast microbeads. Positive cells labelled with anti‐fibroblast microbeads showed a characteristic elongated fibroblast‐like cytoplasm whereas negative cells against fibroblast bead presented a flattened cell type. We used GFAP as well as keratan sulphate antibodies as markers for identifying meningothelial cells. Cells not Conclusion Our method for primary meningothelial cell culture combines harvesting of superficial arachnoid cells from the optic nerve sheath with a purification process based on magnetic beads and is effective in providing pure and rich meningothelial cells.