Abstract
Hyaluronan (HA), an essential component of the extracellular matrix of the skin, is synthesized by HA synthases (HAS1-3). To date, epidermal HA has been considered a major player in regulating cell proliferation and differentiation. However, a previous study reported that depletion of epidermal HA by Streptomyces hyaluronidase (St-HAase) has no influence on epidermal structure and function. In the present study, to further explore roles of epidermal HA, we examined effects of siRNA-mediated knockdown of HAS3, as well as conventional HA-depletion methods using St-HAase and 4-methylumbelliferone (4MU), on epidermal turnover and architecture in reconstructed skin or epidermal equivalents. Consistent with previous findings, HA depletion by St-HAase did not have a substantial influence on the epidermal architecture and turnover in skin equivalents. 4MU treatment resulted in reduced keratinocyte proliferation and epidermal thinning but did not seem to substantially decrease the abundance of extracellular HA. In contrast, siRNA-mediated knockdown of HAS3 in epidermal equivalents resulted in a significant reduction in epidermal HA content and thickness, accompanied by decreased keratinocyte proliferation and differentiation. These results suggest that HAS3-mediated HA production, rather than extracellularly deposited HA, may play a role in keratinocyte proliferation and differentiation, at least in the developing epidermis in reconstructed epidermal equivalents.
Highlights
Received: 3 February 2022Hyaluronan (HA) is a linear and extra-large glycosaminoglycan that is composed of disaccharide repeats of glucuronic acid and N-acetylglucosamine (GlcNAc), and is abundant in the extracellular matrix of connective tissues
Previous studies have indicated a strong interaction between epidermal HA and keratinocyte proliferation or differentiation [15,16,17,18,19,20,23], St-HAase treatment was reported to have no effect on keratinocyte proliferation and differentiation in reconstructed human epidermal equivalents [24]
We investigated the role of HA in regulating epidermal function and structure using three different experimental methods: HA digestion by St-HAase, inhibition of HA synthesis by 4MU, and HA depletion by siRNA-mediated HAS3 knockdown
Summary
Hyaluronan (HA) is a linear and extra-large glycosaminoglycan that is composed of disaccharide repeats of glucuronic acid and N-acetylglucosamine (GlcNAc), and is abundant in the extracellular matrix of connective tissues. Expression patterns of HAS1-3 mRNAs depend on cell type, and we and other research groups have previously reported that HAS3 plays a crucial role in HA production in normal human epidermal keratinocytes [9,10,11,12,13]. Epidermal function and structure depend on a tightly regulated balance between keratinocyte proliferation and differentiation [14], and multiple lines of evidence have suggested that HA plays an important role in regulating epidermal integrity by controlling cell proliferation and differentiation. Enhanced epidermal HA stimulated by keratinocyte growth factor, epidermal growth factor, or retinoic acid is Accepted: 21 February 2022
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