Sima Yizhen, Li Junwei, Xiao Chengzhen, Xu Leimei, Wang Ling, Chen Yisong
Department of Gynecology, Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China.
Laboratory for Reproductive Immunology, Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China.
Ann Transl Med. 2022 May;10(9):511. doi: 10.21037/atm-21-5136.
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.
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 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.
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.
Our results suggest that CD106 fibroblasts possess a high colony-forming capacity and distinct protein expression, and this subpopulation is reduced in POP.
盆腔器官脱垂（POP）是女性常见的退行性疾病，可能会降低生活质量。研究子宫骶韧带的病理变化，包括成纤维细胞的功能变化，对于理解POP的病理生理学至关重要。本研究旨在从人子宫骶韧带中分离出CD106阳性（CD106⁺）成纤维细胞，并评估该亚群的功能和表达。
我们通过荧光激活细胞分选（FACS）分离出CD106⁺成纤维细胞和CD106阴性（CD106⁻）成纤维细胞，并将它们培养用于后续实验。采用流式细胞术分析来测试分选效率、CD106表达以及典型间充质干细胞（MSC）表型标志物表达。应用集落形成单位（CFU）试验来评估成纤维细胞的集落形成能力。诱导后评估三系分化能力。通过蛋白质印迹分析检测波形蛋白、成纤维细胞特异性蛋白-1（FSP-1）、胶原蛋白I（COL 1）、基质金属肽酶-1（MMP-1）和α-平滑肌肌动蛋白（α-SMA）的蛋白水平。通过流式细胞术分析和免疫组织化学（IHC）在POP组和非POP组中验证CD106的表达。
分离出的CD106⁺成纤维细胞纯度为（93.50±3.91）%。CD106⁺成纤维细胞表现出比CD106⁻成纤维细胞更高的集落形成能力，但它们均未表现出与MSC相似的成脂或成骨分化。CD106⁺成纤维细胞中MMP-1和α-SMA的蛋白水平较低，而COL 1的水平高于CD106⁻成纤维细胞。此外，我们观察到与非POP组相比，POP组中CD106的表达降低。
我们的结果表明，CD106⁺成纤维细胞具有高集落形成能力和独特的蛋白表达，并且该亚群在POP中减少。
