Table2_Chemical Pretreatment Activated a Plastic State Amenable to Direct Lineage Reprogramming.xlsx

Somatic cells can be chemically reprogrammed into a pluripotent stem cell (CiPSC) state, mediated by an extraembryonic endoderm- (XEN-) like state. We found that the chemical cocktail applied in CiPSC generation initially activated a plastic state in mouse fibroblasts before transitioning into XEN-like cells. The plastic state was characterized by broadly activated expression of development-associated transcription factors (TFs), such as Sox17, Ascl1, Tbx3, and Nkx6-1, with a more accessible chromatin state indicating an enhanced capability of cell fate conversion. Intriguingly, introducing such a plastic state remarkably improved the efficiency of chemical reprogramming from fibroblasts to functional neuron-like cells with electrophysiological activity or beating skeletal muscles. Furthermore, the generation of chemically induced neuron-like cells or skeletal muscles from mouse fibroblasts was independent of the intermediate XEN-like state or the pluripotency state. In summary, our findings revealed a plastic chemically activated multi-lineage priming (CaMP) state at the onset of chemical reprogramming. This state enhanced the cells’ potential to adapt to other cell fates. It provides a general approach to empowering chemical reprogramming methods to obtain functional cell types bypassing inducing pluripotent stem cells.

体细胞可经化学重编程进入化学诱导多能干细胞（CiPSC, chemical-induced pluripotent stem cell）状态，该过程由胚外内胚层（XEN）样细胞状态介导。我们发现，在制备CiPSC过程中使用的化学诱导剂组合，最初会在小鼠成纤维细胞中激活一种可塑性状态，随后该细胞才会转化为XEN样细胞。该可塑性状态的特征为：发育相关转录因子（TFs, transcription factors）的表达被广泛激活，涵盖Sox17、Ascl1、Tbx3及Nkx6-1等靶点，同时伴随染色质可及性显著提升，这提示细胞命运转换的能力得到增强。值得注意的是，引入此种可塑性状态可显著提升将成纤维细胞经化学重编程为具有电生理活性的功能性神经元样细胞，或是可自主收缩的骨骼肌的效率。此外，从小鼠成纤维细胞制备化学诱导的神经元样细胞或骨骼肌，无需经历中间的XEN样细胞状态或多能性状态。综上，我们的研究揭示了化学重编程启动阶段存在一种化学激活的多谱系致敏可塑性（CaMP, chemically activated multi-lineage priming）状态。该状态可增强细胞适配其他细胞命运的潜能，为优化化学重编程技术提供了通用策略，使其能够绕过多能干细胞诱导阶段，直接获取功能性细胞类型。