Protective Effect Of EGCG Against Oxidative Damages Induced By UVA In Human Skin Fibroblast

Human skin, like all other organs, undergoes chronological aging with age. However, skin is in direct contact with the environment and therefore undergoes aging as a consequence of environmental damage. Photo-aging, which refers to the skin aging superimposed by long-term UV exposure and sun damage. Many of the functions of skin, that are declined with age, show an accelerated decline in photo-aged skin. Besides, the activity of antioxidant enzymes and the levels of nonenzymatic antioxidants decline with age, leading to oxidative damage.UVA can cause direct biological damage or indirect damage by induction of reactive oxygen species (ROS), including superoxide anion, peroxide, and singlet oxygen. ROS damages cellular DNA as well as lipids and proteins such as collagen. Furthermore, mutagenesis by UVA may involve the production of trans-urocanic acid and result in changes of gene expression.Recent advances in skin biological studies have elucidated mechanisms by which photo-aging occurs. There are great demands to search for natural bioactives for preventing or reversing this process. Green tea catechins and its major component epigallocatechin gal late (EGCG), are considered to be potent antioxidants which can be used in various skin care products.In this study, human skin fibroblast (HSF) was incubated in medium containing EGCG and then irradiated with UVA so as to investigate the functions of EGCG in prevention of skin photo-aging by testing oxidative damages, and expressions of genes of VEGF, collagen I and collagen III.In this study, a cell photo-aging model was established by irradiating HSF using UVA. HSF cells were plated in100mm dishs or96well plates with an equal number of cells and cultured in DMEM medium (Dulbecco’s Modified Eagle’s Medium) supplemented with10%fetal bovin serum (FBS) under5%CO2at37℃. The cultured cells were randomly divided into three groups, i.e., control group, UVA-treated group, and EGCG+UVA treated group. EGCG was added into the medium before and after UVA irradiation. Cell proliferation and cellular activity was tested by methyl thiazolyl tetrazolium (MTT) method. Antioxidant activities of the cell were investigated by meassuring activities of glutathione peroxidase (GSH-Px), anion radicas (O2-·) and maleic dialdehyde (MDA). Expressions of genes VEGF, collagen I and collagen III were detected by real-time fluorescent quantitative PCR. Statistical analysis one-way ANOVA was carried out on softwere SPSS17.0,.The major results of the study were as follows:1. EGCG alleviating cell proliferation suppression induced by UVA irradiationUVA irradiation suppressed the cell proliferation in a dose-dependent manner. However, EGCG at a concentration40u.g/mL alleviated the suppression induced by UVA irradiation.2. EGCG increasing anti-oxidant activities of HSF cells irradiated by UVAUVA irradiation dereased the activities of GSH-Px and increased the level of malonyldialdehyde (MDA) in HSF cells. EGCG treatment upregulated the activity of GSH-Px, leading to a lower level in MDA.3. EGCG accelerating the expression of type Ⅰ and type Ⅲ collagen genesCompared with control group, expressions of type Ⅰ and type III collagen genes in the UVA-induced HSF cells were suppressed. When the cells after UVA irradiation, were incubated in medium containing a certain concentration of EGCG, the expression level of ollagen genes type Ⅰ and Ⅲ increased, indicating that EGCG can alleviate this inhibition partially, and protecte skin from photo-aging.4. EGCG suppressing over expression of VEGF gene in HSF cellsAfter UVA irradiation24h, the expression of VEGF increased significantly in HSF cells, indicating that UVA radiation was an important factor to stimulate the secretion of vascular endothelial growth factor. EGCG had the effect inhibiting the overexpression of VEGF gene induced by UVA irradiation.It is concluded that EGCG inhibited the HSF cell proliferation suppression induced by UVA irradiation. The mechanism of EGCG protecting HSF cells from UVA-induced damages might be related to its increasing anti-oxidative enzymes and regulating the expressions of genes encoding collagens type I and III. and VEGF.