Effects Of Cigarette Smoking Extract Combined With UVB Irradiation On The Telomere Length And Telomerase Activity In Human Skin Fibroblasts

Objective:To observe the effects of cigarette smoke extract (CSE) combined with UVB irradiation on telomere length and telomerase activity in human skin fibroblasts, and further to explore the smoke fumes and UVB irradiation could promote the occurrence of skin aging.Methods:Using the ratio of cell-cultured old liquid with cell-cultured new liquid was1:2to culture human skin fibroblasts and drew the curve of cell growth. MTT assay was used to detect the effect of cigarette smoke extract on the proliferation of human skin fibroblast. After the treatment with three different concentrations of CSE and UVB irradiation (30mJ/cm2) on human skin fibroblasts, the real-time quantitative polymerase chain reaction(RT-PCR) was used to detect the changes of telomere length, while using the method of polymerase chain reaction-enzyme linked immunosorbent assay (PCR-ELISA) to detect the changes of telomerase activity.Results:Compared to common cell-cultured method, using the ratio of cell-cultured old liquid with cell-cultured new liquid was1:2to culture human skin fibroblasts that had relatively short doubling time, The incubation period was0-ld, growing period was2-4d, the cell proliferation became slow after4d, and entered into a relatively stable plateau, the common cell-cultured method: The incubation period was0-2d, growing period was2-5d, and after5d was a platform period. Cell proliferation with the treatment of cigarette smoke extract (CSE) presented promoting effects in light concentration (<37.5μL/mL) and inhibitory action in high concentration (>75μL/mL), but compared to no cigarette smoke extract group the difference was no statistical significance (p>0.05). Compared to undisposed group, treated with different concentrations of CSE combined with UVB irradiation group and single UVB irradiation group on human skin fibroblasts, the changes of telomere length and telomerase activity had statistically significant (p<0.05). and also presented with the increasing of the concentration of CSE, the effect was significant.Conclusions:Using the ratio of cell-cultured old liquid with cell-cultured new liquid was1:2to culture human skin fibroblasts had relatively short doubling time, we could obtain enough fibroblasts with high reproductive activity and stable growth for subsequent experiments in a relatively short period of time. Compared to no cigarette smoke extract group, the proliferation of human skin fibroblast was no statistical significance in the group with cigarette smoke extract. CSE and UVB irradiation could shorten telomere length, and reduce telomerase activity, and also presented dose-dependent effects. We concluded that cigarette smoking extract combined with UVB irradiation had a role in promoting skin aging through the mechanism of telomere-telomerase.