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PURPOSE: To validate a new culture model of primary human trabecular meshwork cells (p-hTMCs) using Matrigel®, in order to mimic in vitro 3D-TM organization, and to investigate the proinflammatory effect of benzalkonium chloride (BAK) in 3D p-hTMC cultures. METHODS: p-hTMCs, seeded onto Matrigel®-coated inserts were stimulated with BAK (10(-4)%), dexamethasone (DEX) (10(-6)M) or transforming growth factor-beta 2 (TGF-β2) (5ng/ml) for 48h and observed with confocal microscopy. The BAK effect at 10(-4)% or 5.10(-3)% on the gene expressions of interleukin-6 (IL-6), interleukin-8 (IL-8) and matrix metalloproteinase (MMP-9) was investigated using qRT-PCR in 2D and 3D p-hTMC cultures. RESULTS: p-hTMCs seeded in Matrigel® were able to organize themselves in a 3D-spatial conformation in the different conditions tested with cross-linked actin network (CLAN) formation in presence of DEX or TGF-β2 and intercellular space contraction with TGF-β2. IL-6 and IL-8 gene expressions increased in presence of BAK in 2D and in 3D p-hTMC cultures. BAK 10(-4)% only showed a tendency to stimulate MMP-9 expression in p-hTMCs after 24h-recovery. CONCLUSIONS: We investigated this new 3D-TM in vitro model in Matrigel(®) matrix for pathophysiological and toxicological purposes. It appears as a new promising tool for a better understanding of TM behavior in physiological and stress conditions, as well as toxicological evaluations of antiglaucoma eyedrops and preservatives.
Research Laboratory LR99ES11, Department of Biochemistry, Rabta Hospital, Tunis, Tunisia. Electronic address: bouchemimeryem@yahoo.fr.
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