Modulation of the UV-B-induced Oxidative Stress and Apoptosis in HaCaT Cell Line with Calluna vulgaris Extract

Abstract

The reactive oxygen species (ROS) production due to ultraviolet B (UV-B) exposure is extremely harmful to the skin. It causes lesions of DNA, proteins and lipids and leads to cellular death. In the present study the UV-B-induced ROS and subsequent apoptosis in the human keratinocyte cell line (HaCaT) were counterbalanced by a plant extract with antioxidant capacity. Some molecules modulated by common heather (Calluna vulgaris) (CV) extract through which this may exert its photoprotective effects were also identified. The ROS were evaluated with CM-H2DCFDA assay, while apoptosis and Bax-α/Bcl-xL molecules with ELISA. The extract was standardized according to its polyphenolic content and the most important biologically active compounds, such as hyperozid, quercetin, isoquercetin, kampferol were evidenced by high-performance liquid chromatography. The UV-B induced ROS production occurred at its highest level at 2 h after the exposure of the HaCaT cells, while apoptosis later, at 4 h. The most significant changes in Bax-α and Bcl-XL proteins induced by UV-B, as well as the highest effect of the extract on apoptosis, were both registered at 4 h. The CV extract decreased concentration- and time-dependently the UV-B-induced ROS production and prevented apoptosis. These effects of CV occurred, at least to a certain extent, due to the modulation of Bax-α/Bcl-XL proteins. These findings suggest that skin cells could be protected from some of the UV-B-induced harmful effects by the administration of the CV extract, which may be further exploited as a potential photoprotective agent.

Vitis vinifera seeds extract for the modulation of cytosolic factors Bax-α and NF-kB involved in UVB-induced oxidative stress and apoptosis of human skin cells

“…The ROS production after the exposure of keratinocytes to 100 mJ/cm 2 UVB radiation was evaluated earlier at several time points following the exposure (1, 2 and 4 h). Results showed significant ROS productions at 1 and 2 h and slighter differences as compared to control at 4 h [ 17 ]. None of the selected concentrations of the grape seed extract showed ROS reducing effects at 1 or 2 h after UVB radiation (data not shown).…”

“…Because the highest level of the ROS was registered 2 h after UVB exposure [ 17 ], apoptosis, quantified as intracellular level of ccK18 was evaluated at this time point and later. Thus, at 2 h this molecule increased from 253±16.43 U/L to 411.8±4.43 U/L, corresponding to 1.62 fold increase (p<0.001) as compared to control.…”

“…UVB exposure resulted in statistically significant changes in the Bax-α pro-apoptotic protein level at 4 h after UVB, when this molecule increased from 0.39±0.0007 to 0.84±0.06 pg/ml, representing 2.13 fold increase vs control (p<0.05) [ 17 ]. Similarly to apoptosis results, the higher concentration of the plant extract (75 μgEqGA/ml) determined statistically significant changes in Bax- α protein level.…”

“…It was previously established that the exposure of HaCaT cells to UVB radiation within the physiological range of exposure and equivalent to the minimal erythema dose (100 mJ/cm 2 ) results in significant ROS production and consequently apoptosis, one of the contributing molecules appearing to be the Bax-α pro-apoptotic protein [ 17 ]. As the present study showed, another element which undergoes changes and most likely contributes to the complexity of the events experienced by human skin exposed to UVB radiation is the NF-kB p65 protein.…”

“…The apoptosis regulatory protein (Bax-α) and a transcription factor (NF-kB) were also assessed in order to decipher some of the molecules and pathways through which this extract might exert its beneficial effects. The study represents the continuation of a previous work of the authors in which the effects of the UVB radiation on ROS production, apoptosis induction and apoptosis regulatory proteins were assessed in the human keratinocytes cell line (HaCaT) [ 17 ]. Here we demonstrated that an important amount of the UVB-induced deleterious effects could be prevented by the administration of the GSE, which exerted ROS reducing and apoptosis preventive effects, due to the modulation of Bax-α anti-apoptotic protein and re-establishment of the UV-B affected NF-kB level.…”