In vitro study of the effect of adipose stem cell-derived exosomes on the biological function of localized scleroderma fibroblasts
10.3760/cma.j.cn114453-20220923-00290
- VernacularTitle:脂肪干细胞来源的外泌体对局限性硬皮病成纤维细胞生物学功能的影响
- Author:
Liquan WANG
1
;
Jiuzuo HUANG
;
Nanze YU
;
Xuda MA
;
Tianhao LI
;
Xiao LONG
Author Information
1. 中国医学科学院北京协和医学院北京协和医院整形外科，北京 100032
- Keywords:
Scleroderma, localized;
Fibrosis;
Adipose-derived stem cells;
Exosome
- From:
Chinese Journal of Plastic Surgery
2023;39(6):655-662
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To explore the regulatory effect of exosomes derived from healthy human adipose stem cells (ADSC) on the fibrosis of localized scleroderma fibroblasts (LSFs) in vitro. Methods:From January 2021 to January 2022, fat from 10 healthy donors in Department of Plastic Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences was collected by liposuction. Adipose stem cells were isolated and cultured in vitro, and exosomes (ADSC-Exo) were collected. Fibroblasts were isolated from skin tissue of 15 patients with localized scleroderma during the same period and cultured in vitro. Induced differentiation and staining, nanoparticle tracking analysis, transmission electron microscopy, PKH26 staining and Western blotting were used to identify ADSC and their exosomes. The effect of ADSC on the expression of fibrosis markers [collagen Ⅰ, collagen Ⅲ, α-smooth muscle actin (α-SMA)] in LSFs through its exosomes was examined by extracellular vesicle secretion inhibition assay. The proliferation and migration abilities of LSFs treated with ADSC-Exo were tested by CCK-8 method and scratch test. Real-time quantitative PCR, immunofluorescence staining and Western blotting were used to detect the expression levels of collagen Ⅰ, collagen Ⅲ, α-SMA, transforming growth factor β (TGF-β) and p-Smad2/3 in LSFs. Independent sample t-test was used to compare between the two groups. One-way ANOVA was used for multi-group comparison, and SNK- q test was used for pairwise comparison. Results:ADSC and LSFs were successfully isolated and cultured in vitro, and ADSC-Exo was extracted. Extracellular vesicle secretion inhibition assay demonstrated that ADSC decreased fibrotic markers of LSFs by secreting extracellular vesicles. Results of CCK-8 and scratch test showed that the proliferation and migration ability of LSFs was decreased by ADSC-Exo treatment. The results of real-time quantitative PCR, immunofluorescence staining and Western blotting showed that compared with the control group, the expressions of collagen Ⅰ, α-SMA, TGF-β and p-Smad 2/3 in the ADSC-Exo treatment group were significantly decreased. Conclusion:In vitro, ADSC-Exo can affect the biological behavior and reduce the expression of fibrosis markers in LSFs by inhibiting the TGF-β/Smad pathway.
