Abstract
The release of nucleotides into extracellular space is triggered by insults like wounding and ultraviolet radiation, resulting in stimulatory or inhibitory signals via plasma membrane nucleotide receptors. As similar insults are known to activate hyaluronan synthesis we explored the possibility that extracellular UTP or its breakdown products UDP and UMP act as mediators for hyaluronan synthase (HAS) activation in human epidermal keratinocytes. UTP increased hyaluronan both in the pericellular matrix and in the culture medium of HaCaT cells. 10-100 μm UTP strongly up-regulated HAS2 expression, although the other hyaluronan synthases (HAS1, HAS3) and hyaluronidases (HYAL1, HYAL2) were not affected. The HAS2 response was rapid and transient, with the maximum stimulation at 1.5 h. UDP exerted a similar effect, but higher concentrations were required for the response, and UMP showed no stimulation at all. Specific siRNAs against the UTP receptor P2Y2, and inhibitors of UDP receptors P2Y6 and P2Y14, indicated that the response to UTP was mediated mainly through P2Y2 and to a lesser extent via UDP receptors. UTP increased the phosphorylation of p38, ERK, CREB, and Ser-727 of STAT3 and induced nuclear translocation of pCaMKII. Inhibitors of PKC, p38, ERK, CaMKII, STAT3, and CREB partially blocked the activation of HAS2 expression, confirming the involvement of these pathways in the UTP-induced HAS2 response. The present data reveal a selective up-regulation of HAS2 expression by extracellular UTP, which is likely to contribute to the previously reported rapid activation of hyaluronan metabolism in response to tissue trauma or ultraviolet radiation.
Highlights
The release of nucleotides into extracellular space is triggered by insults like wounding and ultraviolet radiation, resulting in stimulatory or inhibitory signals via plasma membrane nucleotide receptors
Extracellular UTP Enhances Hyaluronan Production—The influence of UTP on hyaluronan metabolism of human keratinocytes was studied by treating HaCaT cells with 100 M UTP and analyzing hyaluronan staining of the cultures and the amount of hyaluronan secreted in the growth medium
The present study shows for the first time that the extracellular uridine nucleotides UTP and UDP strongly up-regulate HAS2 expression, leading to the accumulation of hyaluronan in the pericellular matrix and later in the culture medium
Summary
The release of nucleotides into extracellular space is triggered by insults like wounding and ultraviolet radiation, resulting in stimulatory or inhibitory signals via plasma membrane nucleotide receptors. The present data reveal a selective up-regulation of HAS2 expression by extracellular UTP, which is likely to contribute to the previously reported rapid activation of hyaluronan metabolism in response to tissue trauma or ultraviolet radiation. Cells subjected to mechanical pressure, osmotic shock, bacterial infection, chemical irritation, UV radiation, and tissue wounding rapidly release nucleotides like UTP, UDP, ATP, and ADP to the extracellular space. Extracellular nucleotides signal via specific ligand-gated or G protein-coupled receptor families (P2X and P2Y, respectively) These receptors are expressed in keratinocytes in a differentiation-dependent manner, members of the P2Y2 family are mainly found in the basal cell compartment, representing the proliferative cell population (reviewed in Ref. 13). The newly synthesized hyaluronan chain remains associated with the pericellular matrix, either bound to the synthase, or plasma membrane receptors like CD44, but is later released into the extracellular matrix, where it is often associ-
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