Trioxsalen in the presence of UVA is able to induce nuclear factor kappa B binding activity in HaCaT keratinocytes

It has been described that treatment of cells with high dose psoralen and UVA induce the production of Reactive Oxygen Species (ROS) leading to DNA damage. Transcription factor nuclear factor kappa B (NFkappaB) plays a crucial role in regulating not only cell growth but also cell differentiation, and ROS seem to be partly involved in these mechanisms. The aim of this research was to find out the effect of a combined treatment with trioxsalen (TMP)/UVA on NFkappaB binding activity in HaCaT keratinocytes. HaCaT keratinocytes were treated with 27 microg/l TMP. This concentration did not affect the proliferation rates, nor was it toxic, as shown by cytotoxicity assays. After treatment with TMP with or without UVA (1 J/cm(2)), NFkappaB binding activity in nuclear protein extracts was measured by electrophoretic mobility shift assays. The effect on cytokines and cytokine receptor genes was investigated using cDNA expression arrays. An inhibitory effect on NFkappaB binding activity was found between 30 and 60 min after TMP supplementation of the culture media. UVA irradiation induced a 2-fold increase in NFkappaB binding activity in TMP supplemented HaCaT keratinocytes compared with the non-irradiated control. In addition, NFkappaB binding activity was higher after UVA irradiation with TMP than in UVA irradiated cells in the absence of TMP. TGF-alpha, IL-1R, IL-2Ralpha, IL-12beta and PDGF expression was induced by UVA. However, all of them except PDGF were inhibited by combined TMP/UVA treatment. Using an inhibitor of NFkappaB activation, we found out that under these conditions, these cytokines or cytokine receptor genes are apparently not regulated by NFkappaB. Our results indicate that a combined TMP/UVA treatment of HaCaT keratinocytes induces NFkappaB binding activity, and that this is a synergistic effect. The investigated cytokines, and cytokine receptor genes do not seem to be NFkappaB regulated; however, TMP shows anti-inflammatory capacities in vitro.