Human keloid fibroblast-derived exosomes promote the phenotypic transformation of fibroblasts to myofibroblasts and their significance
10.3760/cma.j.cn114453-20240318-00070
- VernacularTitle:人瘢痕疙瘩成纤维细胞来源外泌体促进成纤维细胞向肌成纤维细胞表型转化及其意义
- Author:
Jinming LI
1
;
Zhibo XIAO
1
Author Information
1. 哈尔滨医科大学附属第二医院整形外科，哈尔滨 150086
- Publication Type:Journal Article
- Keywords:
Fibroblasts;
Keloid fibroblasts;
Myofibroblasts;
Exosomes;
Transforming growth factor-β1
- From:
Chinese Journal of Plastic Surgery
2025;41(4):356-365
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the role of keloid fibroblast-derived exosomes (KF-Exos) in inducing the phenotypic transformation of normal fibroblasts (NFs) into myofibroblasts and to assess their significance in keloid formation.Methods:Normal skin tissues and keloid tissues were collected from female patients aged 20-49 years who underwent surgery at the Department of Plastic Surgery, the Second Hospital of Harbin Medical University from October 2022 to March 2023. Normal fibroblasts and keloid fibroblasts (KFs) were extracted, and the supernatant of KFs was collected to isolate their exosomes. The exosomes were characterized using nanoparticle size analysis (NTA) and transmission electron microscopy (TEM). The samples were divided into the control, KEFS, and KF-Exos groups, then cell proliferation and migration abilities of groups were assessed by scratch assay and Edu assay. The samples were divided into the control, KEFS, KF-Exos, and KFs groups, real-time fluorescence quantitative PCR was used to measure the gene expression levels of Col1a1, Col1a2, Col3a1, α-SMA, TGF-β1, TGF-β2, and TGF-β3. The samples were divided into the control, and KF-Exos groups, Western blot analysis was conducted to evaluate the protein expression levels of Col-1, α-SMA, TGF-β1, Smad2/3, and Smad4. Statistical analysis and plotting were conducted using GraphPad Prism 9.0 software and Image J. Comparisons among multiple groups were analyzed using one-way ANOVA, then comparison of subgroups with the control group was performed using the Dunnett- t test. Independent samples t-test was used to compare the control group with the KF-Exos group in Western blot. P<0.05 was considered statistically significant. Results:The extracted KF-Exos was identified by TEM as a bilayer lipid membrane vesicles. NTA revealed that the average diameter of KF-Exos was 105.4 nm. Scratch and Edu assays demonstrated statistically significant differences in proliferation and migration abilities among the control, KEFS, and KF-Exos groups (all P<0.05). Compared to the control group, the KF-Exos group exhibited statistically significant differences in both proliferation and migration abilities (both P<0.05), while the KEFS group did not show statistically significant differences in these abilities (both P>0.05). Quantitative PCR results showed that, among the control, KEFS, KF-Exos, and KFs groups, there were significant differences in the mRNA expression levels of Col1a1, Col1a2, Col3a1, α-SMA, TGF-β1, TGF-β2, and TGF-β3 (all P<0.05). Compared with the control group, the Col3a1 mRNA level in the KEFS group was significantly different ( P<0.05), whereas the differences in Col1a1, Col1a2, and α-SMA levels were not statistically significant (all P>0.05). The expression levels of Col1a1, Col1a2, Col3a1, α-SMA, TGF-β1, TGF-β2, and TGF-β3 mRNA of the KFs group, and the expression levels of Col1a1, Col1a2, Col3a1, α-SMA, TGF-β1, and TGF-β3 mRNA of the KFs-Exos group had significant differences compared with the control group (all P<0.05), while the difference of TGF-β2 between the KF-Exos group and the control group was not statistically significant ( P>0.05). Western blotting results showed that the protein expression levels of TGF-β1, Smad2/3 and Smad4 were significantly up-regulated in the KF-Exos group compared to the control group (all P<0.05). Conclusion:KF-Exos successfully induced the differentiation of NFs into myofibroblasts and enhanced the expression of extracellular matrix-related factors. This phenomenon may be mediated by the upregulation of the TGF-β1-Smad2/3 and Smad4 signaling pathways.
