紫外线A诱导的人真皮成纤维细胞光老化及小鼠皮肤光老化模型的构建与评价OA

Objective To establish photoaging models of human dermal fibroblasts(HDFs)and mouse skin induced by ul-traviolet A(UVA),so as to provide the optimized models for exploring mechanisms of skin photoaging and preventing and treating skin photoaging.Methods Enzyme digestion was used to isolate HDFs and surface markers were examined.UVA at a dose of 5 J/cm2 was utilized to irradiate HDFs for 15 minutes each day for three consecutive days,and then in vitro HDF photoaging models were estab-lished.Twelve female Kunming mice were divided into a control group and a model group(UVA irradiation group)with six mice in each group.A dose of 20 J/cm2 of UVA was applied for 1 hour daily,5 times a week,which was continued for eight weeks to establish a mouse skin photoaging model.Furthermore,the model efficacies were assessed by observing the cellular morphology,detecting the cell proliferation ability and the activity of senescence-related β-galactosidase,observing the gross appearance of the dorsal skin of mice,performing hematoxylin-eosin staining and Masson staining of the skin tissues,detecting the expression of P16INK4a and P21 mR-NA via RT-qPCR,and examining the expression of type Ⅰ collagen(Collagen-Ⅰ)and matrix metalloproteinase 1(MMP1)by West-ern blot and immunohistochemical staining.Results HDFs were long fusiform-shaped,which expressed vimentin but did not express E-cadherin.After UVA irradiation,the size of HDFs became larger with spreading morphology,along with attenuated proliferation abil-ity.And the positive rate of senescent cells and the expression levels of P16INK4a and P21 mRNA were up-regulated.Meanwhile,Colla-gen-Ⅰ expression was declined,while MMP1 expression was elevated.Compared with the control group,the dorsal skin of mice was rough with pigmentation after UVA irradiation.UVA exposure gave rise to thickening of the epidermis and thinning of the dermis with decreased and disorganized collagen.Moreover,the expression levels of P16INK4a and P21 mRNA in UVA irradiation group were dra-matically higher than those in the control group.And the expression of Collagen-Ⅰ diminished,while the expression of MMP1 en-hanced.Conclusion The UVA-induced HDF photoaging model and mouse skin photoaging model are successfully established,which provide an experimental basis for exploring the mechanism of skin photoaging and developing therapeutic strategies.