Synthesis of proteoglycans by suspension and monolayer cultures of adult chondrocytes and de novo cartilage nodules-the effect of hyaluronic acid

Abstract

Chondrocytes were isolated from adult laryngeal cartilage by an enzymic procedure that included 6 h digestion with collagenase. The level of 35SO4(2-) incorporation into cetylpyridinium chloride-precipitable material by these cells depended upon the subsequent culturing conditions. Suspension cultures incorporated more 35SO4(2-)/cell than monolayer cultures. Hyaluronic acid in the medium inhibited 35SO4(2-) incorporation only when the cells were in primary suspension cultures. It had no effect on monolayer cultures, or monolayers organized into nodules, or suspension cultures derived from monolayers. Mild pretreatment with EDTA, however, rendered these susceptible to hyaluronic acid inhibition. In contrast EDTA abolished the inhibitory effect of hyaluronic acid on primary suspension cultures. Oligosaccharides, derived from hyaluronidase digestion of hyaluronic acid that were larger than decassaccharide, had some inhibitor effect on 35SO4(2-) incorporation by monolayer cultures. The total 35SO4(2-) incorporation was less in primary suspension cultures of chondrocytes isolated after 12 h than after 6 h digestion of cartilage and the inhibition by hyaluronic acid was also less. These differences persisted during 12 days of culture. It is suggested that the method of isolating chondrocytes and subsequent culture conditions may modify the cell surface and mask or abolish specific binding sites for hyaluronic acid.