Ning Li1, Ran Zhang2. 1. Department of Ophthalmology, The First Affiliated Hospital of Anhui Medical University, Hefei, 230022, China. 2. Department of Dermatology and Institute of Dermatology, The First Affiliated Hospital of Anhui Medical University, Hefei, 230022, China. liningnature@126.com.
Abstract
PURPOSE: Growth factors are considered as key molecules that participating in fibrosis formation. This research aimed to clarify potential effects of p53 on regulation of transforming growth factor β (TGF-β) and fibrosis formation and investigate the associated mechanisms. METHODS: Vimentin was examined to identify human Tenon's fibroblasts (HTFs). p53-targeting small interfere RNA (siRNA) was synthesis and transfected into HTFs. Real-time PCR assay was utilized to evaluate p53 and microRNA-29b (miR-29b) expression. Immunocytochemical assay was used to observe TGF-β expression. The wound healing assay was conducted to evaluate migration of HTFs. Dual-luciferase assay was employed to identify interaction between p53 and miR-29b in HTFs. RESULTS: Vimentin was extensively distributed in HTFs cells. HTFs at density of 5 × 104 cells/ml and 6 days exhibited the best growth. The p53 level in TGF-β treatment group was significantly higher compared to that in blank group (p < 0.01). miR-29b level in siRNA targeting p53 group was significantly increased compared to that in blank group (p < 0.01). siRNA targeting p53 could significantly inhibit HTFs migration compared to that in single TGF-β treating HTFs group (p < 0.01). Relative luciferase activity was significantly increased in p53 overexpressed HTFs compared to that in cells transfected with empty pcDNA3.0 plasmid (p < 0.01). CONCLUSIONS: p53 inhibited expression of TGF-β, suppressed HTFs migration and inhibited HTFs growth, by reducing miR-29b expression and interacting with miR29b gene in HTFs.
PURPOSE: Growth factors are considered as key molecules that participating in fibrosis formation. This research aimed to clarify potential effects of p53 on regulation of transforming growth factor β (TGF-β) and fibrosis formation and investigate the associated mechanisms. METHODS:Vimentin was examined to identify human Tenon's fibroblasts (HTFs). p53-targeting small interfere RNA (siRNA) was synthesis and transfected into HTFs. Real-time PCR assay was utilized to evaluate p53 and microRNA-29b (miR-29b) expression. Immunocytochemical assay was used to observe TGF-β expression. The wound healing assay was conducted to evaluate migration of HTFs. Dual-luciferase assay was employed to identify interaction between p53 and miR-29b in HTFs. RESULTS:Vimentin was extensively distributed in HTFs cells. HTFs at density of 5 × 104 cells/ml and 6 days exhibited the best growth. The p53 level in TGF-β treatment group was significantly higher compared to that in blank group (p < 0.01). miR-29b level in siRNA targeting p53 group was significantly increased compared to that in blank group (p < 0.01). siRNA targeting p53 could significantly inhibit HTFs migration compared to that in single TGF-β treating HTFs group (p < 0.01). Relative luciferase activity was significantly increased in p53 overexpressed HTFs compared to that in cells transfected with empty pcDNA3.0 plasmid (p < 0.01). CONCLUSIONS:p53 inhibited expression of TGF-β, suppressed HTFs migration and inhibited HTFs growth, by reducing miR-29b expression and interacting with miR29b gene in HTFs.
Entities:
Keywords:
Human Tenon's fibroblasts; Transforming growth factor β; miR-29b; p53