Great Ormond Street Institute of Child Health, University College London, WC1N1EH London, UK.
Shanghai Institute for Advanced Immunochemical Studies (SIAIS), ShanghaiTech University, 201210 Shanghai, China.
Cell Rep. 2020 Dec 1;33(9):108453. doi: 10.1016/j.celrep.2020.108453.
The specification of the hepatic identity during human liver development is strictly controlled by extrinsic signals, yet it is still not clear how cells respond to these exogenous signals by activating secretory cascades, which are extremely relevant, especially in 3D self-organizing systems. Here, we investigate how the proteins secreted by human pluripotent stem cells (hPSCs) in response to developmental exogenous signals affect the progression from endoderm to the hepatic lineage, including their competence to generate nascent hepatic organoids. By using microfluidic confined environment and stable isotope labeling with amino acids in cell culture-coupled mass spectrometry (SILAC-MS) quantitative proteomic analysis, we find high abundancy of extracellular matrix (ECM)-associated proteins. Hepatic progenitor cells either derived in microfluidics or exposed to exogenous ECM stimuli show a significantly higher potential of forming hepatic organoids that can be rapidly expanded for several passages and further differentiated into functional hepatocytes. These results prove an additional control over the efficiency of hepatic organoid formation and differentiation for downstream applications.
人类肝脏发育过程中肝本质的特化受到外在信号的严格调控，但细胞如何通过激活分泌级联反应来响应这些外源性信号仍不清楚，而分泌级联反应在 3D 自我组织系统中尤为重要。在这里，我们研究了人多能干细胞 (hPSC) 分泌的蛋白质如何响应发育性外源性信号，从而影响从内胚层到肝谱系的进展，包括它们生成新生肝类器官的能力。通过使用微流控限制环境和稳定同位素标记与细胞培养结合的质谱 (SILAC-MS) 定量蛋白质组学分析，我们发现细胞外基质 (ECM) 相关蛋白的丰度很高。在微流控中衍生的或暴露于外源性 ECM 刺激的肝祖细胞显示出形成肝类器官的潜力显著提高，这些肝类器官可以快速扩增多个传代，并进一步分化为功能性肝细胞。这些结果证明了对下游应用中肝类器官形成和分化效率的额外控制。
