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Oxygen Reduces Accumulation of Type IV Collagen in Endothelial Cell Subcellular Matrix via Oxidative Stress

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Anchorage-dependent cells in culture attach initially to proteins adsorbed to the culture substrate from the medium, and produce and deposit a subcellular matrix during the course of the cultivation. The aim of this study was to determine whether the concentration of O2 in the culture atmosphere affects the accumulation of type IV collagen and laminin under human endothelial-cell monolayers. Enzyme-linked immunoassays on decellularized polystyrene substrates showed less type IV collagen, but not less laminin, under cells incubated in the standard atmosphere (5% CO2 in air, i.e., ∼20% O2) compared to an atmosphere of 5% O2 and 5% CO2 in N2. Type IV collagen accumulation was inhibited via oxidative stress, because the inhibitory effect of 20% O2 was antagonized by antioxidant ascorbic acid, and mimicked by prooxidant pyrogallol and exogenous H2O2. Measurements of endogenous H2O2 accumulation demonstrated that endothelial cells partially adapt to the high O2 concentration. These results may have implications in endothelium modeling in vitro and in engineering of endothelial cell sheets and endothelialized vascular grafts.
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Keywords: Endothelium; Extracellular matrix; Hydrogen peroxide; Laminin; Oxygen; Tissue engineering; Type IV collagen

Document Type: Research Article

Affiliations: 1: Research & Development, Nunc A/S, Roskilde; and 2: Mammalian Cell Biology, Bioneer A/S, Hørsholm, Denmark

Publication date: December 1, 2006

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