Inhibitory effect of rapamycin on proliferation,migration and fibrosis of human pterygium fibroblasts in vitro
10.3760/cma.j.issn.2095.0160.2018.12.002
- VernacularTitle:雷帕霉素对体外培养人翼状胬肉成纤维细胞增生、移行和纤维化的抑制作用
- Author:
Di WU
1
;
Xiaonan SUN
;
Lin DU
;
Xiaoyu ZHANG
;
Shanshan LIU
;
Jing SUN
;
Lin XU
;
Shaodan ZHANG
Author Information
1. 110031,沈阳市第四人民医院眼科 沈阳市眼病中心重点实验室
- Keywords:
Pterygium;
Fibroblasts;
Rapamycin;
Anti.fibrosis
- From:
Chinese Journal of Experimental Ophthalmology
2018;36(12):902-907
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the inhibitory effect of rapamycin,an mammalian target of rapamycin (mTOR) pathway inhibitor,on the proliferation,migration and fibrosis of human pterygium fibroblasts (PFBs). Methods Pterygium tissues were collected from patients with primary pterygium who underwent surgical excision in Shenyang Fourth People's Hospital from May to July 2015. The tissues were cultured in vitro and the PFBs were identified by anti.human vimentin immunofluorescence assay. The 3 to 5 generation cells were used for the experiments. The viability of cells treated with different concentrations of rapamycin was detected by methyl thiazolyl tetrazolium ( MTT) . The cells were divided into normal control group and rapamycin group, and the scratch wound healing test was used to evaluate migration of the PFBs. The expressions of MKI67,α.smooth muscle actin (α.SMA), fibronectin,caspase3, mammalian target of rapamycin ( mTOR ) and LC3B mRNA were detected by real.time quantitative PCR. Results The cultured cells showed morphology of long spindle and were vimentin immunopositive. The cell viability in rapamycin treated PFBs demonstrated a dose.dependent decrease. At 24 hours after culture,The cell viability in 30μmol/L rapamycin group was (76. 67±8. 84)％ of that in 0μmol/L rapamycin group ( P<0. 001 ) . The relative residual scratch width in 30μmol/L rapamycin group was ( 35. 40 ± 11. 62 )％ 48 hours after scratch,which was significantly greater than (2. 45±0. 76)％ in the normal control group (P<0. 05). Real.time quantitative PCR showed that the mRNA expressions of MKI67,α.SMA,fibronectin and mTOR in rapamycin group were significantly decreased when compared with those in normal control group (all at P<0. 05). The expression of LC3B mRNA in rapamycin group was significantly higher than that in normal control group (P<0. 05). The mRNA expression of caspase3 was not significantly different between the two groups (P=0. 861). Conclusions Rapamycin can effectively inhibit the proliferation, migration and fibrosis of PFBs without affecting the cell survival. Detailed mechanism remains to be further studied. Rapamycin may serve as an anti.fibrosis agent to prevent the progression and recurrence of pterygium in the future.
