Switching from healthy to unhealthy oxidative stress - does the radical type can be used as an indicator?

Abstract

Ultraviolet (UV) radiation leads to the formation of free radicals, which may cause immunological modulations, skin aging or skin cancer. Sunlight exposure in the UVA region according to CIE 85 promotes almost 46% of radical formation in skin. A critical radical concentration characterized by the inversion of the domination of primary ROS (reactive oxygen species) to an excess of secondary LOS (lipid oxygen species) is proven for the spectral regions UV and or VIS light and is intended to be a marker for an imbalance in the redox system, which can no longer compensate harmful effects. To investigate whether this transition point is also universally valid for one spectral region, the radical formation during and after targeted UVA in situ-irradiation at 365 ± 5 nm and three different irradiances (31, 94 and 244 mW/cm²) was investigated in ex vivo porcine skin using x-band electron paramagnetic resonance (EPR) spectroscopy. The quantification was performed with the spin probe 3-(carboxy)-2,2,5,5-tetramethylpyrrolidin-1-oxyl (PCA), the spin trap 5,5-Dimethyl-1-Pyrroline-N-Oxide (DMPO) was used to characterize the radical species. Furthermore, the viability of the skin cells after irradiation was controlled by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, skin integrity was examined by histological analysis. A significant dose dependence in the radical formation is given at higher irradiance. The transition point was detected in the range of 0.5 MED after irradiation with the highest irradiance. From this point on the proportion of LOS increases with increasing dose and the proportion of ROS decreases. After switching off the UVA irradiation no further quantitative changes were detected, but rapid changes in the radical pattern were observed demonstrating the importance of in situ irradiation during the use of spin traps. Heat-pre-stressed skin showed more LOS than ROS already at the beginning of the irradiation, leading to the assumption that the transition point to the distress-level has already been reached. In summary, a postulated transition point could be verified for the UVA spectral region using only one spin trap combined with in-situ irradiation. A certain degree of stress is necessary to detect an inversion of the ratio of ROS to LOS. This reversal indicates an imbalance in the redox status. However, at low intensities no changes at all in radical pattern appeared over time (dose), probably it can be compensated by adaptation processes of the skin.

Keywords: Electron paramagnetic resonance (EPR) spectroscopy; Lipid oxygen species (LOS); Oxidative stress; Reactive oxygen species (ROS); UVA irradiation.