Effect of SRSF2 on the biological characteristics of keloid fibroblasts
10.3760/cma.j.issn.1009-4598.2019.04.005
- VernacularTitle: 剪接因子SRSF2对瘢痕疙瘩成纤维细胞生物学特性的作用研究
- Author:
Jing PAN
1
;
Hu JIAO
2
;
Ran XIAO
1
Author Information
1. Research Center of Plastic Surgery Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Beijing 100144, China
2. The 9th Department of Plastic and Reconstructive Surgery, Plastic Surgery Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Beijing 100144, China
- Publication Type:Journal Article
- Keywords:
SRSF2;
Keloid;
Fibroblast;
Biological characteristics
- From:
Chinese Journal of Plastic Surgery
2019;35(4):347-354
- CountryChina
- Language:Chinese
-
Abstract:
Objective:This study aims to explore the influence of the serine and arginine rich splicing factor 2 (SRSF2) on the biological characteristics of keloid fibroblast, by comparing the expression levels of SRSF2 in normal skin and keloid, with the purpose to provide a new method to study the pathogenesis of keloid.
Methods:Samples of normal skin were derived from excess tissue of skin grafts, collected from 8 patients, aged 8-53 years old, while the specimens of keloid were from 12 keloid patients, aged 18-24 years old. All the patients were admitted in the Plastic Surgery Hospital, Chinese Academy of Medical Sciences. The expression of SRSF2 was assessed by immunohistochemistry and immunofluorescent staining in both normal and keloid tissue. Keloid fibroblasts were cultured in vitro to detect the relationship between TGF-β1 stimulation and SRSF2 expression. After constructing the lentiviral sh-RNA-expression vector, targeting SRSF2 and infecting keloid fibroblast, the apoptosis and proliferation of cells were analyzed by the MuseTM Cell Analyzer. The expression of CyclinE1 was analyzed by real-time PCR and western blot, and the secretion and expression of extracellular matrix and TGF-β1 were detected by real-time PCR and ELISA.The software SPSS 21.0 was used to do the statistical analysis.
Results:The expression of SRSF2 was significantly higher in the keloid samples than in the normal skin samples. The percentage of SRSF2 positive cells in keloid was (77.04±4.37)% , while the percentage of SRSF2 positive cells in normal skin was (25.10±1.24)%. TGF-β1 promotes the expression of SRSF2 to (159.73±17.03)% times in keloid fibroblasts. After the SRSF2 knock-out, the apoptosis rate of the keloid fibroblasts increased, its cell cycle arrest was observed at the G0/G1 phase, and the expression of Cyclin E1 decreased. Apoptotic cells in control group was about (18.83±1.24)% , while (25.81±7.09)% and (26.71±6.14)% were in the two knock-out groups respectively. Cells in G0/G1 phase was about (58.97±1.73)% in control group, while (63.95±2.07)% and (64.65±3.23)% were in the two knock-out groups respectively. Compared to the control group, the expression of Cyclin E1 mRNA in the two knock-out groups were (31.60±6.81)% and (33.01±11.39)% respectively. Additionally, the expression of COL3A1 FN1 mRNA decreased, and the expression and secretion of TGF-β1 was declined. Compared to the control group, the expression of COL3A1 mRNA in the two knock-out groups were (64.90±23.71)% and (67.97±13.50)%, the expression of FN1 mRNA in the two knock-out groups were (59.10±8.11)% and (70.70±18.26)%, the expression of TGFB1 mRNA in the two knock-out groups were (53.37±15.51)% and (67.53±19.33)% respectively. The concentration of TGF-β1 in the medium of control group was (115.60±18.17)pg/ml, while (75.35±12.25) pg/ml and (72.06±14.66) pg/ml were in the two knock-out groups respectively.
Conclusions:The expression of SRSF2 increased in keloid. The inhibition of SRSF2 in keloid fibroblast resulted in the growth restriction and apoptosis of cells, and decreased the secretion of extracellular matrix and TGF-β1, which providing a new insight into keloid pathogenesis, and suggesting that SRSF2 could be a new therapeutic target for keloid conditions.
