Effect of the hydroxyl radical on fibroblast-mediated collagen remodelling in vitro

Abstract

1. It has been reported that free radicals prevent wound healing. However, the mechanism of this effect is not yet clear. We attempted to clarify the influence of hydroxyl radicals on wound healing in vitro. 2. We used an ascorbate-copper ion system (ACS) to produce hydroxyl radicals in accordance with variables of time elapsed and concentration of copper ion. The effects of hydroxyl radical on fibroblast-mediated collagen remodelling, cell viability, the functions of fibroblasts and collagen fibrils were studied. 3. With a copper ion concentration of 100 mumol/L ACS significantly reduced contraction, while 10 mumol/L stimulated contraction. Hydrogen peroxide (H2O2) was employed in observing these findings. ACS did not influence cell viability, the expression of alpha 2 beta 1 integrin and cellular fibronectin, or the cytoskeletal organization of fibroblasts involving actin until 3 h. A concentration of ACS at 10 mumol/L of copper ion induced the polymerization of collagen after 30 min, while ACS at 100 mumol/L induced collagen degradation; this finding was also established by using H2O2. Collagen reduced the amount of formaldehyde produced by trapping hydroxyl radical with dimethyl sulfoxide. 4. Our findings suggest that collagen is denatured by scavenging the hydroxyl radical before fibroblasts are damaged, so that the radical may influence the remodelling of collagen.