Protective Effects And Mechanism Of Ginsenoside Re On UVB Irradiation-induced Photo Damage

The ultraviolet,which is divided into three parts according to its wave length,is a kind of electromagnetic wave that the length ranges from 200nm to 400nm.The three parts are UVA（320nm-400nm）,UVB（280nm-320nm）,and UVC（200nm-280nm）.UVA can penetrate the dermis and make the skin premature aging or black;UVB penetrates the skin a few millimeters,the skin will appears sunburn,repeated sunburn will increase the risk of skin cancer.Ginseng is a kind of valuable traditional Chinese medicine,which has been applied to the clinical application for over 2,000 years.The ginseng saponins are the main active ingredients of ginseng,which have strong activity in anti-oxidation,scavenging free radicals,anti-radiation,anti-aging,anti-tumor and enhancing immunity.Metabonomics（metabolomics）is defined as“The living system response to pathophysiological stimuli or perturbation（genetic modification or environmental factors change）with metabolites changed or with metabolites time-varying.LC-MS is particularly well suited for metabolomic analysis since it allowed both high sensitivity and wide dynamic range.RRLC coupled with Q-TOF MS possesses the advantage of high sensitivity,high resolving power and high accuracy.In this study,we screened out ginsenoside Re（G-Re）from protopanaxatriol by vitro cell platform,which has a strong resistance to ultraviolet radiation activity and its UV protection mechanism was studied in-depth as follows:1.Different dose of UVC irradiation were employed to Human RSa cells（RSA）,50 mg·L^-1 panaxatriol saponins（Re,Rf,Rh1,Rg1,Rg2）were treated before and after the irradiation.MTT assay was employed to analyze the cellular viability,apoptosis and cell cycle were assayed by a Flow cytometric meter（FCM）,Superoxide dismutase（SOD）was determined by biochemical methods and colony formation analysis was employed to assay the survival rate of the cells with SOD inhibitors.The results showed that the panaxatriol saponins could suppress the UVC irradiation induced cell damage,G-Re was better than the other panaxatriol saponins on anti-radiation activity,Which could effectively improve the RSA cells survival,reduce UVC irradiation induced cell early apoptosis and cell G2 arrest and increase intracellular SOD activity,the mechanism may be related to its antioxidant,enhancing the vitality of cell proliferation.2 The intervention study that NB-UVB irradiated human skin keratinocytes immortalized（Ha Ca T）cells treated with G-Re showed that G-Re can significantly improve the survival rate of Ha Ca T cells after UVB irradiation.The concentration of G-Re was positively correlated with the survival rate.Panaxatriol is similar to G-Re on the ability of improving cell survival rate after UVB irradiation.Saponin Rf,Rh1,Rg1,Rg2 and F1showed the synergies effects on increasing cell survival rate combined with G-Re.G-Re protected irradiated Ha Ca T cells through enhancing intracellular antioxidant enzyme activity and reducing the content of lipid peroxidation products.3 Pharmacodynamic Study was based on a photo damage model that the rats was irradiated by NB-UVB,Experimental results showed that G-Re was able to inhibit the keratinized layer of the rat skin from UVB irradiation induced thickening and reduce the number of sunburn and inflammatory cells.G-Re could significantly inhibit the protein expression of COX-2 and MMP-9 in the rat skin after UVB radiation and reduce murine skin inflammation and photo damage from UVB radiation.G-Re was significantly increased UVB radiated rats peripheral blood leukocyte counts,murine thymus index and spleen index to enhance immune system,spleen lymphocyte transformation rate stimulated by Con A had significant promoting effects after G-Re treatment.G-Re also effectively reduced the radiated model murine lung index,indicating that the G-Re was possible to reduce the reactive oxygen species and peroxidation products concentration that generated by UVB irradiation in the lungs to relieve inflammation or edema of the lungs.G-Re not only significantly reduced murine serum ALT,AST,TBIL content,but also improved the murine blood,skin and liver SOD activity,which lowered MDA content of the murine blood,skin and liver by serum enzymes and antioxidant assays.G-Re may pass through the protecting cellular membrane,scavenging oxygen free radicals,inhibition of lipid peroxidation,inhibition of immune damage and other pathways to defend murine blood,skin and liver,which further proved that ginsenoside Re could eliminate free radicals and enhance the body’s antioxident level.G-Re was able to lower TNF-alpha and IL-6 levels in the radiated rat skin,liver,serum,which decreased UVB irradiation induced inflammatory reactions in the skin,liver,serum.G-Re could prevent UVB phototoxic from further deepening and block the development of skin cancer.4 In order to effectively evaluate the bioavailability of G-Re,This study employed a RRLC-Q-TOF-MS method to detect the distribution of G-Re in rats tissues,G-RC was used for internal standard,the calibration curve was linear in the range of 10-20000 ng·m L^-1 for tissue homogenates（r>0.99）,The lower limit of detection（LLOD）was 0.03μg·m L^-1,the lower limit of quantification（LLOQ）was 0.1μg·m L^-1,the accuracy,the intra-day and inter-day precision and recovery ratio are in line with the analysis of biological samples.G-Re could quickly widely distributed in organizations with 20 mg·kg^-1G-Re solution by caudal vein injection,which could be detected in many organs（lung,spleen,kidney,heart,brain,liver）and distribute mainly in the lung,spleen,kidney and heart.Lung had a highest affinity to G-Re which was not easy to pass through the blood-brain barrier and hard to accumulate.Tissue distribution showed the highest level was observed in the lungs,the least level was observed in the livers.5 A urinary metabonomics method was developed to investigate the effect of UVB irradiation to the endogenous metabolites and related metabolic pathways,eventually 11potential biomarkers were found and identified.The biological significance analysis showed that the damage caused by UVB radiation may affect sphingolipid metabolism,metabolism of nucleic acids,linoleic acid metabolism,and amino acid metabolism pathway of normal rats.6 In order to get more information about the mechanism of G-Re suppressing the UVB induced photo damage of the murine skin,G-Re was employed to intervene with UVB-induced photo damage,8 related biomarkers were identified by metabolomic approach,G-Re could up-regulate sphingolipid metabolism and linoleic acid metabolism pathway,which were related to UVB induced photo damage.