Ma Zhiyao, Chawla Shikha, Lan Xiaoyi, Zhou Eva, Mulet-Sierra Aillette, Kunze Melanie, Sommerfeldt Mark, Adesida Adetola B
Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada.
Department of Biomedical Engineering, Faculty of Engineering, University of Alberta, Edmonton, Alberta, Canada.
Cell Prolif. 2025 Jan;58(1):e13735. doi: 10.1111/cpr.13735. Epub 2024 Oct 8.
Collagenase digestion (d) and cellular outgrowth (og) are the current modalities of meniscus fibrochondrocytes (MFC) isolation for bioengineering and mechanobiology-related studies. However, the impact of these modalities on study outcomes is unknown. Here, we show that og- and d-isolated MFC have distinct proliferative capacities, transcriptomic profiles via RNA sequencing (RNAseq), extracellular matrix (ECM)-forming, and migratory capacities. Our data indicate that microtissue pellet models developed from og-isolated MFC display a contractile phenotype with higher expressions of alpha-smooth muscle actin (ACTA2) and transgelin (TAGLN) and are mechanically stiffer than their counterparts from d-MFC. Moreover, we introduce a novel method of MFC isolation designated digestion-after-outgrowth (dog). The transcriptomic profile of dog-MFC is distinct from d- and og-MFC, including a higher expression of mechanosensing caveolae-associated caveolin-1 (CAV1). Additionally, dog-MFC were superior chondrogenically and generated larger-size microtissue pellet models containing a higher frequency of smaller collagen fibre diameters. Thus, we demonstrate that the modalities of MFC isolation influence the downstream outcomes of bioengineering and mechanobiology-related studies.
胶原酶消化法（d）和细胞生长法（og）是目前用于生物工程和力学生物学相关研究的半月板纤维软骨细胞（MFC）分离方法。然而，这些方法对研究结果的影响尚不清楚。在此，我们表明，通过og法和d法分离的MFC具有不同的增殖能力、通过RNA测序（RNAseq）得到的转录组图谱、细胞外基质（ECM）形成能力和迁移能力。我们的数据表明，由og法分离的MFC构建的微组织颗粒模型表现出收缩表型，α-平滑肌肌动蛋白（ACTA2）和转胶蛋白（TAGLN）表达较高，并且在机械性能上比d法分离的MFC构建的模型更硬。此外，我们介绍了一种新的MFC分离方法，即生长后消化法（dog）。dog法分离的MFC的转录组图谱与d法和og法分离的MFC不同，包括机械传感小窝相关的小窝蛋白-1（CAV1）表达较高。此外，dog法分离的MFC在软骨形成方面更具优势，并且能生成更大尺寸的微组织颗粒模型，其中较小直径胶原纤维的频率更高。因此，我们证明了MFC分离方法会影响生物工程和力学生物学相关研究的下游结果。
