Center for Biomedical Engineering, Brown University, Providence, RI, 02912, USA.
Department of Pathology and Laboratory Medicine, Brown University, Providence, RI, 02912, USA.
Cell Mol Life Sci. 2022 May 27;79(6):320. doi: 10.1007/s00018-022-04351-w.
Cellular mechanophenotype is often a defining characteristic of conditions like cancer malignancy/metastasis, cardiovascular disease, lung and liver fibrosis, and stem cell differentiation. However, acquiring living cells based on mechanophenotype is challenging for conventional cell sorters due to a lack of biomarkers. In this study, we demonstrate a workflow for surface protein discovery associated with cellular mechanophenotype. We sorted heterogeneous adipose-derived stem/stromal cells (ASCs) into groups with low vs. high lamin A/C, an intracellular protein linked to whole-cell mechanophenotype. Proteomic data of enriched groups identified surface protein candidates as potential biochemical proxies for ASC mechanophenotype. Select surface biomarkers were used for live-cell enrichment, with subsequent single-cell mechanical testing and lineage-specific differentiation. Ultimately, we identified CD44 to have a strong inverse correlation with whole-cell elastic modulus, with CD44 cells exhibiting moduli three times greater than that of CD44 cells. Functionally, these stiff and soft ASCs showed enhanced osteogenic and adipogenic differentiation potential, respectively. The described workflow can be replicated for any phenotype with a known correlated intracellular protein, allowing for the acquisition of live cells for further characterization, diagnostics, or therapeutics.
细胞力学表型通常是癌症恶性/转移、心血管疾病、肺和肝纤维化以及干细胞分化等疾病的特征。然而，由于缺乏生物标志物，传统的细胞分选器很难根据力学表型获取活细胞。在这项研究中，我们展示了一种与细胞力学表型相关的表面蛋白发现的工作流程。我们将异质脂肪来源的干细胞/基质细胞（ASCs）分为低和高核纤层蛋白 A/C 的组，核纤层蛋白 A/C 是一种与整个细胞力学表型相关的细胞内蛋白。富集组的蛋白质组学数据鉴定出表面蛋白候选物作为 ASC 力学表型的潜在生化替代物。选择表面生物标志物用于活细胞富集，随后进行单细胞力学测试和谱系特异性分化。最终，我们发现 CD44 与整个细胞弹性模量呈强烈负相关，CD44 细胞的模量比 CD44 细胞高三倍。功能上，这些硬和软 ASC 显示出增强的成骨和成脂分化潜力。该描述的工作流程可复制到任何具有已知相关细胞内蛋白的表型，允许获取活细胞以进行进一步的表征、诊断或治疗。
