Hyaluronate Accumulation in Human Epidermis Treated with Retinoic Acid in Skin Organ Culture

Abstract

Retinoic acid (RA) has been shown to retard the differentiation of epidermal keratinocytes by several morphologic and biochemical criteria. In this study, the epidermal content and localization of hyaluronate (HA), as well as its synthesis and disappearance in human skin organ culture, were characterized to test the idea that some of the RA influences on epidermal differentiation are associated with keratinocyte HA metabolism. RA stimulated the incorporation of 3H-glucosamine into HA by up to 60% at concentrations between 50 nM and 5 microM, while pulse-chase experiments revealed little change in its disappearance rate from epidermis. After 5 d in culture, the chemically quantified HA was more than doubled in the treated epidermis. The accumulation of HA was substantiated by light and electron microscopy with a specific probe prepared from the HA binding region of cartilage proteoglycan. The staining was particularly enhanced between the upper spinous cell layers, where the terminal differentiation into corneocytes normally takes place. A patchy, discontinuous staining was also seen in stratum granulosum and corneum layers, which are not stained at all in control cultures. The present study demonstrates that RA leads to an accumulation of HA in the superficial layers of epidermis by stimulating its synthesis in keratinocytes. This may account for the delay in terminal differentiation, and the weakened cohesion of the keratinocytes previously observed both in vivo and vitro.

Keratinocyte Growth Factor Stimulates Migration and Hyaluronan Synthesis in the Epidermis by Activation of Keratinocyte Hyaluronan Synthases 2 and 3

“…An inverse correlation between the content of hyaluronan in the spinous cell layer and the indicators of epidermal differentiation has also been noted with other effectors like vitamin A (38) and EGF (39), both of which stimulate hyaluronan synthesis and inhibit differentiation. Conversely, we have found that pharmacological concentrations of hydrocortisone enhance differentiation but inhibit hyaluronan synthesis (40) and Has2 expression.…”

Hyaluronan-dependent pericellular matrix☆

“…In human skin organ cultures, factors like all-trans-retinoic acid which increase hyaluronan synthesis lead to delayed differentiation [132], like in human skin in vivo. Likewise, organotypic epidermal cultures show that keratinocyte differentiation is stimulated and inhibited by factors that decrease and increase hyaluronan synthesis, respectively [133].…”

The Human Hyaluronan Synthase 2 Gene Is a Primary Retinoic Acid and Epidermal Growth Factor Responding Gene

“…Hyaluronan is synthesized by the Has enzymes Has1, Has2, and Has3 at the plasma membrane (2). In skin epidermis, the narrow extracellular space surrounding keratinocytes contains a high concentration of hyaluronan, but it is found mainly between the basal and spinous cell layers of normal human epidermis and much less in terminally differentiated layers (3). Both in normal and diseased epidermis, keratinocyte growth and differentiation are regulated by paracrine and endocrine signaling molecules, such as EGF 1 and the nuclear hormone all-trans-RA.…”

“…Both in normal and diseased epidermis, keratinocyte growth and differentiation are regulated by paracrine and endocrine signaling molecules, such as EGF 1 and the nuclear hormone all-trans-RA. Interestingly, hyaluronan synthesis rate is stimulated by EGF in epidermal keratinocytes in monolayer (4) and organotypic cultures (5) and by all-trans-RA in human skin organ cultures (3). Direct evidence for the biological role of hyaluronan in epidermal keratinocytes emerged by the finding that Has2-mediated hyaluronan synthesis controls the migration rate of keratinocytes in scratch-wounded monolayer cultures (6).…”

“…In this way, activated EGFRs induce via the tyrosine kinase Src the phosphorylation of STAT proteins 1, 3, and 5 (10), which then translocate to the nucleus. STAT homodimers recognize REs within promoter regions of EGF-responding genes of the sequence TTCN [2][3][4] GAA with a preference for three spacing nucleotides (11).…”