Hyaluronan is an abundant and rapidly turned over matrix molecule between the vital cell layers of the epidermis. In this study, epidermal growth factor (EGF) induced a coat of hyaluronan and a 3-5-fold increase in its rate of synthesis in a rat epidermal keratinocyte cell line that has retained its ability for differentiation. EGF also increased hyaluronan in perinuclear vesicles, suggesting concurrent enhancement in its endocytosis. Cell-associated hyaluronan was most abundant in elongated cells that were stimulated to migrate by EGF, as determined in vitro in a wound healing assay. Large fluctuations in the pool size of UDP-N-acetylglucosamine, the metabolic precursor of hyaluronan, correlated with medium glucose concentrations but not with EGF. Reverse transcriptase-polymerase chain reaction (RT-PCR) showed no increase in hyaluronan synthases 1 and 3 (Has1 and Has3), whereas Has2 mRNA increased 2-3-fold in less than 2 h following the introduction of EGF, as estimated by quantitative RT-PCR with a truncated Has2 mRNA internal standard. The average level of Has2 mRNA increased from ϳ6 copies/ cell in cultures before change of fresh medium, up to ϳ54 copies/cell after 6 h in EGF-containing medium. A control medium with 10% serum caused a maximum level of ϳ21 copies/cell at 6 h. The change in the Has2 mRNA levels and the stimulation of hyaluronan synthesis followed a similar temporal pattern, reaching a maximum level at 6 h and declining toward 24 h, a finding in line with a predominantly Has2-dependent hyaluronan synthesis and its transcriptional regulation.Hyaluronan is a large glycosaminoglycan found in the extracellular space of most animal tissues. It forms a loose, highly hydrated, gel-like matrix that contributes to the maintenance of the extracellular space and facilitates nutrient diffusion. Furthermore, hyaluronan is involved in cell proliferation and differentiation, produces an environment favorable for migration (1), and stimulates cell locomotion (2, 3). Elevated tissue levels of hyaluronan occur during embryonic growth of tissues and organs (1), wound healing (4, 5), inflammation (6), and invasion of certain cancers (7-10).In skin epidermis, the narrow extracellular space surrounding keratinocytes contains a high concentration of hyaluronan (11, 12), as do other stratifying squamous epithelia (13, 14). The half-life of labeled epidermal hyaluronan in human skin organ culture is ϳ1 day (15), indicating fast local turnover by keratinocytes. The importance of the strikingly high concentration and turnover of hyaluronan in the multilayered squamous epithelia is not completely understood, but we have hypothesized that the former is necessary to maintain an extracellular space for the nutritional needs of the more superficial cell layers, whereas the latter allows the dramatic modulation of cell shape that occurs during differentiation and for the high migratory potential of keratinocytes that is activated, e.g. in wound healing (16).Unlike other glycosaminoglycans, hyaluronan is synthesized at the inne...

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During maturation, significant modulation of tissue structure, composition and mechanical properties takes place. Importantly, the present study provides insight into the mechanical, chemical and structural interactions that lead to functional properties of mature articular cartilage.