J Korean Ophthalmol Soc.  2020 Sep;61(9):1015-1022. 10.3341/jkos.2020.61.9.1015.

Ultraviolet B-induced Senescence Model Using Corneal Fibroblasts and the Anti-aging Effect of Angiogenin

• Sung SM
• Lee SJ
• Lee KW
• Kim JC

• KMID: 2506475
• DOI: http://doi.org/10.3341/jkos.2020.61.9.1015

Abstract

Purpose
To establish a corneal-fibroblast senescence model induced by ultraviolet B (UVB) exposure and to investigate the anti-senescence effect of angiogenin (ANG).
Methods
Fibroblasts were exposed to UVB (1 mJ/cm 2 ) and then cultured with ANG-containing solution for 24 hours. The 24-hour culture procedure was repeated for three days after UVB irradiation. Cell viability was evaluated using the 3-[4, 5–dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) assay. The degree of senescence was assessed using the ratio of senescence-associated (SA)-β-gal-stained cells to total cells. The expression of age-related factors and degree of DNA damage were assessed via Western blot. Samples were divided into a non-UVB group, a UVB group without ANG treatment, and a UVB group with ANG treatment after irradiation (UVB + ANG).
Results
Cell viability in the UVB + ANG group was 11% higher than that in the UVB group (p < 0.05). The UVB + ANG group exhibited a 10% lower degree of SA-β-gal staining compared with the UVB group (p < 0.05). Western blot analysis showed that there was reduced expression of p53, p21, p16, and RB in the UVB + ANG group compared with the UVB group. The expression of phosphorylated histone (_Y-H2AX) and p38 in the UVB + ANG group was less than that in the UVB group.
Conclusions
Senescence in corneal fibroblasts is induced by UVB, and ANG may exert an anti-aging effect by regulating the cell cycle through p53, p21, p16, and RB and reducing DNA damage.

Keyword

Angiogenin; Fibroblast; Senescence