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Francoise Brignole-Baudouin, Meryem Bouchemi, Luisa RIANCHO, Christophe Roubeix, Karima Kessal, Anne-Laure RAVEU, Hayet SOUALMIA, Christophe BAUDOUIN; Benzalkonium chloride cytotoxic and proinflammatory effects in a new 3D-trabecular-meshwork model. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3289.
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© ARVO (1962-2015); The Authors (2016-present)
Several trabecular meshwork (TM) cells are available for research purposes in the field of glaucoma and are classically cultured in 2D. As TM function is to filter aqueous humor, the 3D organization is an important parameter to consider when designing in vitro models. Our aim was to validate a new culture model of TM cells using Matrigel extracellular compound in order to mimic in vitro, the in vivo 3D-TM organization, for a better understanding of TM behavior in physiological and stress conditions.
We cultured primary trabecular cells (ScienceCell) at 105cells/mL in Matrigel® medium diluted in DMEM (1/3). Cells were treated with benzalkonium chloride (BAK) from 10-4 %-5.10-3% for 30mn to 48h. IL-6, CXCL8, CXCL12, CXCR4, CXCR3, alpha-SMA were investigated for protein expression in fluorescence immunostainings and confocal microscopy and for gene expressions using qRT-PCR.
BAK disorganized the trabecular cell meshwork, increasing spaces between cells by cell loss and cell shrinkage. It also induced pro-inflammatory responses of IL-6, CXCL8, CXCL12, CXCR4, and CXCR3 and stimulated alpha-SMA, in a concentration dependent manner with higher responses for IL-6 vs CXCL8 and CXCR3 vs CXCR4.
In this study we could confirm the interest of 3D-TM cultures in Matrigel matrix that could offer a new tool to investigate pharmacological compounds, chemical or physical stress conditions to better understand the pathophysiology of this important structure in the normal intraocular pressure regulation avoiding glaucoma disease.
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