Establishing an Improved Cellular Photoaging Model by Repeated UVA Exposure of Human Skin Fibroblasts

Repeated ultraviolet A (UVA) exposure-induced cellular photoaging models had been widely applicated to study skin photoaging. However, there is no standard protocol to prepare these models and cell passaging is inevitably needed when fibroblasts are cultured in conventional media for 5-7 days.

The adhesion ability of fibroblasts was tested after UVA irradiation. And both the cell culture medium and the UVA dose were optimized to help fibroblasts surviving a 7-day culture period without cell subculture and to establishing an improved cellular photoaging model. Finally, senescence-associated β-galactosidase staining, intracellular reactive oxygen species (ROS) detection with the fluorescent redox probe dichlorodihydrofluorescein diacetate (DCFH-DA), and Western blot analysis were performed to compare the improved model with the classical model.

UVA exposure induced a significant decrease in the adhesion ability of fibroblasts, and thus implied that cell passaging might be a screen pressure against the photodamaged cells. And fibroblasts incubated in the medium supplemented with 1% fetal calf serum could survive the 7-day culture period without cell subculture and tolerate UVA irradiation up to a dose of 5.8 J/cm² daily for 7 days. No significant differences were found between the improved model and the classical model in intracellular ROS production; however, our model demonstrated a significantly higher percentage of senescence-associated β-galactosidase positive cells. Moreover, both p53 and p21 were up-regulated in our model, while in the classical model only p21 was up-regulated.

An improved cellular photoaging model was established, which seems to be more suitable than the classical model for elucidating the underlying molecular mechanisms of skin photoaging.