Kaylan Kerim B, Ermilova Viktoriya, Yada Ravi Chandra, Underhill Gregory H
Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA.
Sci Rep. 2016 Mar 30;6:23490. doi: 10.1038/srep23490.
The bipotential differentiation of liver progenitor cells underlies liver development and bile duct formation as well as liver regeneration and disease. TGFβ and Notch signaling are known to play important roles in the liver progenitor specification process and tissue morphogenesis. However, the complexity of these signaling pathways and their currently undefined interactions with other microenvironmental factors, including extracellular matrix (ECM), remain barriers to complete mechanistic understanding. Utilizing a series of strategies, including co-cultures and cellular microarrays, we identified distinct contributions of different Notch ligands and ECM proteins in the fate decisions of bipotential mouse embryonic liver (BMEL) progenitor cells. In particular, we demonstrated a cooperative influence of Jagged-1 and TGFβ1 on cholangiocytic differentiation. We established ECM-specific effects using cellular microarrays consisting of 32 distinct combinations of collagen I, collagen III, collagen IV, fibronectin, and laminin. In addition, we demonstrated that exogenous Jagged-1, Delta-like 1, and Delta-like 4 within the cellular microarray format was sufficient for enhancing cholangiocytic differentiation. Further, by combining Notch ligand microarrays with shRNA-based knockdown of Notch ligands, we systematically examined the effects of both cell-extrinsic and cell-intrinsic ligand. Our results highlight the importance of divergent Notch ligand function and combinatorial microenvironmental regulation in liver progenitor fate specification.
肝祖细胞的双潜能分化是肝脏发育、胆管形成以及肝脏再生和疾病的基础。已知转化生长因子β（TGFβ）和Notch信号通路在肝祖细胞的特化过程和组织形态发生中发挥重要作用。然而，这些信号通路的复杂性以及它们目前与包括细胞外基质（ECM）在内的其他微环境因子尚未明确的相互作用，仍然是全面理解其机制的障碍。我们利用包括共培养和细胞微阵列在内的一系列策略，确定了不同的Notch配体和ECM蛋白在双潜能小鼠胚胎肝（BMEL）祖细胞命运决定中的不同作用。特别是，我们证明了Jagged-1和TGFβ1对胆管细胞分化具有协同影响。我们使用由I型胶原、III型胶原、IV型胶原、纤连蛋白和层粘连蛋白的32种不同组合组成的细胞微阵列，确定了ECM的特异性作用。此外，我们证明在细胞微阵列形式下，外源性Jagged-1、Delta样1和Delta样4足以增强胆管细胞分化。此外，通过将Notch配体微阵列与基于短发夹RNA（shRNA）的Notch配体敲低相结合，我们系统地研究了细胞外和细胞内配体的作用。我们的结果突出了不同的Notch配体功能和组合微环境调节在肝祖细胞命运特化中的重要性。
