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Updated: Sep 2, 2025
Murine Dermal Fibroblast Isolation by FACS
Published on: January 7, 2016
CD106/VCAM-1 distinguishes a fibroblast subpopulation with high colony-forming capacity and distinct protein
Yizhen Sima1,2,3,4, Junwei Li1, Chengzhen Xiao1
1Department of Gynecology, Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China.
Background:
Pelvic organ prolapse (POP) is a common degenerative disease in women which may diminish quality of life. Investigating the pathological changes of the uterosacral ligament, including the functional changes of fibroblasts, is critical to understanding the pathophysiology of POP. This study was designed to isolate CD106-positive (CD106+) fibroblasts from the human uterosacral ligament and assess the function and expression of this subpopulation.
Methods:
We separated CD106+ fibroblasts and CD106 negative (CD106-) fibroblasts by fluorescence-activated cell sorting (FACS) and cultured them for subsequent experiments. Flow cytometric analysis was used to test the sorting efficiency, CD106 expression, and typical mesenchymal stem cell (MSC) phenotype marker expression. A colony-forming unit (CFU) assay was applied to evaluate the colony-forming ability of the fibroblasts. Trilineage differentiation capacities were assessed after in vitro induction. The protein levels of vimentin, fibroblast specific protein-1 (FSP-1), collagen I (COL 1), matrix metallopeptidase-1 (MMP-1), and α-smooth muscle actin (α-SMA) were detected by western blot analysis. The expression of CD106 was verified by flow cytometric analysis and immunohistochemistry (IHC) in the POP and non-POP groups.
Results:
The CD106+ fibroblasts were isolated with a purity of (93.50±3.91)%. The CD106+ fibroblasts exhibited higher colony-forming capacity than that of CD106- fibroblasts, but neither of them showed adipogenic or osteogenic differentiation similar to that of MSCs. The protein levels of MMP-1 and α-SMA were lower, and the level of COL 1 was higher in the CD106+ fibroblasts than in the CD106- fibroblasts. In addition, we observed a decreased expression of CD106 in the POP group compared with the non-POP group.
Conclusions:
Our results suggest that CD106+ fibroblasts possess a high colony-forming capacity and distinct protein expression, and this subpopulation is reduced in POP.
