Effects of baicalin on ultraviolet A-induced telomere damage in cultured human primary fibroblasts

Abstract

Objective To investigate baicalin effect against ultraviolet A (UVA) induced senescence in cultured human skin fibroblasts (HSF) and influence on telomere pathway. Methods HSF were isolated from the prepuce of neonates and cultured. Subconfluent fibroblasts were classified into blank control group (without treatment), baicalin group (treated with baicalin of 50 μg/ml), UVA group (irradiated with UVA of 10 J/cm2) and UVA + baicalin group (irradiated with UVA of 10 J/cm2 and treated with baicalin of 50 μg/ml before and after the irradiation). After additional culture of various durations, flow cytometry was performed to detect cell cycle, telomere repeat amplification protocol-enzyme linked immunosorbent assay (TRAP-ELISA) to measure telomerase activity, real-time quantitative PCR to determine telomere length, mRNA levels of p53, p16 and c-myc, Western blot to examine the protein expressions of p16 and c-myc. Results UVA irradiation induced cell cycle arrest in G1 phase, and the percentage of HSF at G1 phase increased from 59.94％ in the blank control group to 81.04％ in the UVA group, but was decreased to 65.55％ in the UVA + baicalin group. The length of telomere in HSF in UVA group was shortened to 31.2％ of that in the blank control group, but was restored to 63.9％ in HSF treated with baicalin before and after the irradiation. Compared with the blank control group, the expression level of p53 and p16 mRNA was increased to 2.93 ± 0.21 and 2.14 ± 0.09, respectively, while that of c-myc mRNA decreased to 0.53 ± 0.03 in the UVA group; baicalin could inhibit these changes. Similarly, Western blot showed that after UVA irradiation the protein expression level of p16 increased to 5.84 ± 0.16, while that of c-myc decreased to 0.35 ± 0.04 in HSF compared with that in the blank control group; baicalin treatment before and after the irradiation induced no significant changes in the protein expres sion of c-myc, but a decline in that of p16 (4.09 ± 0.13, P ＜ 0.05). Telomerase activity was undetected in any of these groups. Conclusions Baicalin can delay the photoaging process of HSF, which may be attributed to the regulation of expression of senescence-related genes such as p53, but not to telomerase activity. Key words: Ultraviolet rays; Baicalin; Telomere; Cell aging