Effects of UV Rays and Thymol/Thymus vulgaris L. Extract in an ex vivo Human Skin Model: Morphological and Genotoxicological Assessment.

Ultraviolet (UV) radiation is the major environmental factor affecting functions of the skin. Compounds rich in polyphenols, such as Thymus vulgaris leaf extract and thymol, have been proposed for the prevention of UV-induced skin damage. We compared the acute effects induced by UVA and UVB rays on epidermal morphology and proliferation, cytotoxicity, and genotoxicity. Normal human skin explants were obtained from young healthy women (n = 7) after informed consent and cultured at the air-liquid interface overnight. After 24 h, the samples were divided in 2 groups: the former exposed to UVA (16 or 24 J/cm2) and the latter irradiated with UVB (0.24 or 0.72 J/cm2). One hour after the end of irradiation, supernatants were collected for evaluation of the lactate dehydrogenase activity. Twenty-four hours after UVB exposure, biopsies were processed for light and transmission electron microscopy analysis, proliferation, cytotoxicity, and genotoxicity. UVB and UVA rays induced early inhibition of cell proliferation and DNA damage compared to controls. In particular, UVB rays were always more cytotoxic and genotoxic than UVA ones. For this reason, we evaluated the effect of either T. vulgaris L. extract (1.82 µg/ml) or thymol (1 µg/ml) on all samples treated for 1 h before UVB irradiation. While Thymus had a protective action for all of the endpoints evaluated, the action of the extract was less pronounced on epidermal proliferation and morphological features. The results presented in this study could be the basis for investigating the mechanism of thymol and T. vulgaris L. extract against the damage induced by UV radiation.

Thymol and Thymus Vulgaris L. activity against UVA- and UVB-induced damage in NCTC 2544 cell line.

Many authors focused on the research of natural compounds in order to protect skin from indirect (UVA) and direct (UVB) ultraviolet radiation side effects. The aim of this study to evaluate the protective effect of a dry extract from T. vulgaris L. and of its major synthetic compound thymol (about 60%), against oxidative and genotoxic UVA- and UVB damage. Experiments were reproduced in a low differentiated keratinocytes cell line (NCTC 2544) Cells were pretreated for 1h, in serum-free medium, with thymol (1µg/mL) or T. vulgaris L. (1.82µg/mL) then exposed to different UVA (8-24J/cm(2)) or UVB doses (0.016-0.72J/cm(2)). Immediately after the UV exposure the intracellular redox status was evaluated by ROS quantification and by LPO. Genotoxic aspects were evaluated 24h after the end of irradiations using the alkaline comet assay, the micronucleus formation assay and the immunostaining of phosphorylated H2AX histone protein (detected 1h after the end of UV exposure). Thymol and T. vulgaris L. extract inhibited ROS generation in UVA and UVB-irradiated cells. On the contrary, MDA formation was reduced only in UVA treated cells. Both agents decreased the DNA damage evaluated by the alkaline comet assay, but not in the micronucleus and H2AX tests probably because of the severity of damage (double strands) detected.

Protective effect of Vaccinium myrtillus extract against UVA- and UVB-induced damage in a human keratinocyte cell line (HaCaT cells).

Recently, the field of skin protection have shown a considerable interest in the use of botanicals. Vaccinium myrtillus contains several polyphenols and anthocyanins with multiple pharmacological properties. The purpose of our study was to examine whether a water-soluble V. myrtillus extract (dry matter 12.4%; total polyphenols 339.3mg/100 g fw; total anthocyanins 297.4 mg/100 g fw) was able to reduce UVA- and UVB-induced damage using a human keratinocyte cell line (HaCaT). HaCaT cells were pretreated for 1h with extract in a serum-free medium and then irradiated with UVA (8-40 J/cm(2)) and UVB (0.008-0.72 J/cm(2)) rays. All experiments were performed 24h after the end of irradiation, except for oxidative stress tests. The extract was able to reduce the UVB-induced cytotoxicity and genotoxicity (studied by comet and micronucleous assays) at lower doses. V. myrtillus extract reduced lipid peroxidation UVB-induced, but had no effect against the ROS UVB-produced. With UVA-induced damage V. myrtillus reduced genotoxicity as well as the unbalance of redox intracellular status. Moreover our extract reduced the UVA-induced apoptosis, but had no effect against the UVB one. V. myrtillus extract showed its free radical scavenging properties reducing oxidative stress and apoptotic markers, especially in UVA-irradiated cells.

Genotoxic effects of polychlorinated biphenyls (PCB 153, 138, 101, 118) in a fish cell line (RTG-2).

Polychlorinated biphenyls (PCBs) are persistent pollutants in aquatic environments, often causing the decline or disappearance of wild populations. The primary aim of this study was to investigate the genotoxic effects of some PCBs (PCB153 (2,2',4,4',5,5'-hexachlorobiphenyl) and 138 (2,2',3,4,4',5'-hexachloro-biphenyl), both non-dioxin-like compounds, and the pentachlorobiphenyls PCB118 (2,3',4,4',5-) and 101 (2,2',4',5,5'-), the former an ortho-substituted, low-affinity dioxin-like compound and the latter a non-coplanar congener classified as non-dioxin-like) in fish cells (RTG-2). These congeners are mostly present in surface waters and in edible aquatic organisms and the loss of DNA integrity in vitro serves as a sensitive biomarker of cytogenetic alterations and is considered as an initial step for the identification of genotoxic effects. The alkaline comet assay and the micronucleus test show clear genotoxic damage after short and longer exposure (2 and 24h) to maximum soluble, non-cytotoxic doses, evident sooner with PCBs 101 and 118. Oxidative stress situations involving ROS release, reduction in total GSH, lipid peroxidation and alteration to superoxide dismutase, seen after exposure with all the congeners, though with different kinetics, seem the most likely explanation for the genotoxic damage. This appears to be confirmed by the modified comet assay (pH 10) for detection of oxidized bases using endonuclease III. The increased generation of intracellular ROS might explain the apoptosis seen after treatment with the single PCBs and evaluated on the basis of the rise in 3-7 caspase activity. Therefore both the non-coplanar, non-dioxin-like PCBs (153, 138, 101) and the low-affinity dioxin-like compound PCB118 cause evident genotoxic damage, probably as a consequence of oxidative stress.

Protective effect of erdosteine metabolite I against hydrogen peroxide-induced oxidative DNA-damage in lung epithelial cells.

It has been shown that the mucolytic agent erdosteine (N-carboxymethylthio-acetyl-homocysteine thiolactone, CAS 84611-23-4) has anti-inflammatory and anti-oxidant properties, and an active metabolite I (MET I) containing pharmacologically active sulphydryl group has been found to have a free radical scavenging activity. The aim of this study was to assess the ability of erdosteine metabolite I to protect A549 human lung adenocarcinoma cell against hydrogen peroxide (H2O2)-mediated oxidative stress and oxidative DNA damage. When A549 cells were pre-treated with the active metabolite I (2.5-5-10 microg/ml) for 10-30 min and then exposed to H2O2 (1-4 mM) for two additional hours at 37 degrees C, 5% at CO2, the intracellular peroxide production, reflected by dichlorofluorescein (DCF) fluorescence, decreased in a concentration-dependent manner. Furthermore, using a comet assay as an indicator for oxidative DNA damage, it was found that the metabolite I prevented damage to cells exposed to shortterm H2O2 treatment. The data suggest that this compound is effective in preventing H2O2-induced oxidative stress and DNA damage in A549 cells. The underlying mechanisms involve the scavenging of intracellular reactive oxygen species (ROS).