Stem cells are widely explored in regenerative medicine as a source to produce diverse cell types. Despite the wide usage of stem cells like mesenchymal stem cells (MSCs) and induced pluripotent stem cells (iPSCs), there is a lack of robust methods to rapidly discern the phenotypic and functional heterogeneity of stem cells. The organization of actin cytoskeleton has been previously used to discern divergent stem cell differentiation pathways. In this paper, we highlight the versatility of a cell profiling method for actin turnover dynamics. Actin filaments in live stem cells are labeled using SiR-actin, a cell permeable fluorogenic probe, to determine the endogenous actin turnover. Live MSC imaging after days of induction successfully demonstrated lineage specific change in actin turnover. Next, we highlighted the differences in the cellular heterogeneity of actin dynamics during adipogenic or osteogenic MSC differentiation. Next, we applied the method to differentiating iPSCs in culture, and detected a progressive slowdown in actin turnover during differentiation upon stimulation with neural or cardiac media. Finally, as a proof of concept, the actin dynamic profiling was used to isolate MSCs via flow cytometry prior to sorting into three distinct sub-populations with low, intermediate or high actin dynamics. A greater fraction of MSCs with more rapid actin dynamics demonstrated increased inclination for adipogenesis, whereas, slower actin dynamics correlated with increased osteogenesis. Together, these results show that actin turnover can serve as a versatile biomarker to not only track cellular phenotypic heterogeneity but also harvest live cells with potential for differential phenotypic fates.

中文翻译：

---

基于荧光的干细胞肌动蛋白转换动力学作为干细胞功能进化、异质性和表型谱系解析的分析方法

干细胞在再生医学中被广泛探索作为产生多种细胞类型的来源。尽管间充质干细胞 (MSCs) 和诱导多能干细胞 (iPSCs) 等干细胞得到广泛使用，但缺乏可靠的方法来快速识别干细胞的表型和功能异质性。肌动蛋白细胞骨架的组织以前已被用于辨别不同的干细胞分化途径。在本文中，我们强调了用于肌动蛋白周转动力学的细胞分析方法的多功能性。使用可渗透细胞的荧光探针 SiR-actin 标记活干细胞中的肌动蛋白丝，以确定内源性肌动蛋白转换。诱导数天后的活 MSC 成像成功地证明了肌动蛋白转换的谱系特异性变化。下一个，我们强调了在脂肪形成或成骨 MSC 分化过程中肌动蛋白动力学的细胞异质性差异。接下来，我们将该方法应用于区分培养中的 iPSC，并检测到在神经或心脏介质刺激后分化过程中肌动蛋白转换的逐渐减慢。最后，作为概念验证，肌动蛋白动态分析用于通过流式细胞术分离 MSC，然后分类为具有低、中或高肌动蛋白动力学的三个不同亚群。更大比例的具有更快肌动蛋白动力学的 MSCs 表明脂肪生成的倾向增加，而较慢的肌动蛋白动力学与成骨增加相关。一起，