Anti-photosensitive Agent On UVB-induced Changes Affect HaCaT Cell Proliferation, Apoptosis And Cell Cycle

photosensitive dermatosis,is a group of skin disease caused by sunlight on the skin contain photosensitive sex material,belongs to the common disease, frequently-occurring disease of dermatology, Chinese medicine called "sun boils".Its pathogenesis is closely related to the ultraviolet radiation damage skin cutin cells.Anti—Photosensitivity mixture made by beijing hospital of traditional Chinese medicine is composed of Artemisinin, Herba Artemisiae Scopariae(HAS), Radix Paeoniae Rubra, Campsis grandiflora spray etc,and has a good curative effect to photosensitive dermatosis in clinical application。 The research of protective effect of Anti—Photosensitivity mixture which can protect the skin from ultraviolet damage have not been reported yet, both in China and abroad.The subject of the proposed establishment of ultraviolet B irradiation HaCaT damage model is to simulate Ultraviolet radiation damage on the skin keratinocyte,and test the protective effect of Anti—Photosensitivity mixture on UVB radiation damage to HaCaT cells, and investigate the protection mechanism, if any protection.ObjectiveTo establish ultraviolet B irradiation HaCaT damage model,and explore the protective mechanism of Anti—Photosensitivity mixture to HaCaT cells.Methods1.Cell cultureWith containing10%fetal bovine serum and1%of the double resistance to cultivate HaCaT cell MEM broth, quantitative inoculation in petri dishes or96well plates.2.The preparation of medicine serum containing Anti—Photosensitivity mixtureAccording to experimental design preparation of the same amount of drug-containing serum and biank serum.3.Making Ultraviolet radiation damage model on the skin keratinocyteCultivate HaCaT cell growth to a certain extent, according to the experimental design timing quantitative cells with UVB radiation, and joined the relevant drug intervention treatment in before and after irradiation.4.Hydroxychloroquine concentration detectionCCK-8method to detect Optimal concentration.5.effect of Anti—Photosensitivity mixture on HaCaT cell proliferation activity CCK-8method to detect cell proiiferation activity.6.Effect of Anti—Photosensitivity mixture to HaCaT cell proliferation activity after UVB radiationCCK-8method to detect cell proliferation activity.7.Apoptosis index and cell cycle detectionUsing flow cytometry instrument detecting apoptosis index under different processing conditions and the change of the cell cycle.Result1.Influence of Hydroxychloroquine to HaCaT eel! proliferation activity after UVB radiationComparing the HaCaT cell survival rate of treatment group which radiated by different doses of hydroxychloroquine with the exposure control group, the difference was statistically significant (P<0.05),Join after irradiation doses of hydroxychloroquine compared with, suggesting that after dealing with the hydroxychloroquine,the HaCaT cell activity can be recovered to a certain degree, among which the survival rate of20ug/ml hydroxychloroquine group is the highest.2.the effect of Anti—Photosensitivity mixture on HaCaT cell proliferation activityComparing the HaCaT cell survival rate of dosing group with the blank group, there were significant differences (P<0.01), suggesting that the drug-containing serum of Anti—Photosensitivity mixture can promote the HaCaT cell growth; cell survival rate in the Positive medicine control group compared with the blank group, was no significant difference (P>0.05), showed that20ug/ml hydroxychloroquine group was no effect on HaCaT cell activity.3.Effect of Anti—Photosensitivity mixture to HaCaT cell proliferation activity after UVB radiationComparing the experimental HaCaT cell survival rate with model group, there were significant differences (P<0.01), suggesting that the drug-containing serum of Anti—Photosensitivity mixture can make the HaCaT cell activity after UVB radiation has a certain degree of recovery; cell survival rate in the Positive medicine control group compared with the experimental group,there were significant differences(P<0.01), suggesting that the effect of drug-containing serum of Anti—Photosensitivity mixture on HaCaT cell in protection was not as well as20ug/ml hydroxychloroquine group.4. Effect of Anti—Photosensitivity mixture to HacaT cell apoptosis after UVB irradiation The apoptosis rate of experimental HaCaT cell compared with the model group has a certain decline, but no obvious difference (P>0.05), showed that the apoptosis inhibition of drug-containing serum of Anti—Photosensitivity mixture to HaCaT call is not obvious.5. Effect of Anti—Photosensitivity mixture to HacaT cell cycle after UVB irradiationAfter30mJ/cm2UVB radiation, the HacaT cell cycle has been a marked S phase retardation, and after join the the drug-containing serum of Anti Photosensitivity mixture, the S phase cell number can have a certain decline, but no significant difference (P>0.05).ConclusionThis study results show that a certain dose of Anti—Photosensitivity mixture can restrain the cell cycle change mildly,that caused by UVB radiation, and then reduce the radiation of HaCaT cell caused by ultraviolet (uv) B, and has a mild inhibition to apoptosis caused by UVB radiation, which means protection effectiveness.