Sabine Werner

The Ca2+-binding proteins S100A8 and S100A9 are encoded by novel injury-regulated genes

To gain insight into the molecular mechanisms underlying cutaneous wound repair, we performed a large scale screen to identify novel injury-regulated genes. Here we show a strong up-regulation of the RNA and protein levels of the two Ca2+-binding proteins S100A8 and S100A9 in the hyperthickened epidermis of acute murine and human wounds and of human ulcers. Furthermore, both genes were expressed by inflammatory cells in the wound. The increased expression of S100A8 and S100A9 in wound keratinocytes is most likely related to the activated state of the keratinocytes and not secondary to the inflammation of the skin, since we also found up-regulation of S100A8 and S100A9 in the epidermis of activin-overexpressing mice, which develop a hyperproliferative and abnormally differentiated epidermis in the absence of inflammation. Furthermore, S100A8 and S100A9 expression was found to be associated with partially differentiated keratinocytes in vitro. Using confocal microscopy, both proteins were shown to be at least partially associated with the keratin cytoskeleton. In addition, cultured keratinocytes efficiently secreted the S100A8/A9 dimer. These results together with previously published data suggest that S100A8 and S100A9 are novel players in wound repair, where they might be involved in the reorganization of the keratin cytoskeleton in the wounded epidermis, in the chemoattraction of inflammatory cells, and/or in the defense against microorganisms.

Unexpected Down-regulation of the hIK1 Ca2+-activated K+ Channel by Its Opener 1-Ethyl-2-benzimidazolinone in HaCaT Keratinocytes INVERSE EFFECTS ON CELL GROWTH AND PROLIFERATION

We used a combination of electrophysiological and cell and molecular biological techniques to study the regulation and functional role of the intermediate conductance Ca2+-activated K+ channel, hIK1, in HaCaT keratinocytes. When we incubated cells with the hIK1 opener, 1-ethyl-2-benzimidazolinone (1-EBIO), to investigate the cellular consequences of prolonged channel activity, an unexpected down-regulation of channels occurred within a few hours. The same effect was produced by the hIK1 openers chlorzoxazone and zoxazolamine and was also observed in a different cell line (C6 glioma cells). After 3 days of treatment with 1-EBIO, mRNA levels of hIK1 were substantially diminished and no channel activity was detected. Down-regulation of hIK1 was accompanied by a loss of mitogenic activity and a strong increase in cell size. After withdrawal of 1-EBIO, hIK1 mRNA and channel activity fully recovered and the cells resumed mitogenic activity. Our data present evidence for a novel feedback mechanism of hIK1 expression that appears to result from the paradoxical action of its pharmacological activator during prolonged application. Because the down-regulation of hIK1 bears immediate significance on the biological fate of keratinocytes, 1-EBIO and related compounds might emerge as potent tools to influence the proliferation of various non-excitable cells endowed with IK channels.