Biomedical application of human pluripotent stem cell-derived hepatocyte-like cells (hPSC-HLCs) relies on efficient large-scale differentiation, which is commonly performed by a suspension culture of three-dimensional (3D) multicellular spheroids in bioreactors. However, this approach requires large amounts of growth factors (GFs) and the need to overcome limited diffusional transport posed by the inherent 3D structure of hPSC spheroids. Here, we have hypothesized that localized delivery of GFs by incorporation of GF-laden degradable polymeric microparticles (MPs) within the hPSC spheroids would circumvent such limitations. In this study, GFs for hepatocytic differentiation were encapsulated in gelatin-coated poly (l-lactic acid)/poly (DL-lactic-co-glycolic acid) (PLLA/PLGA) MPs which were subsequently incorporated into the hPSC spheroids. Gene expression analyses demonstrated that MP delivery of the GFs resulted in similar expression levels of hepatocytic markers despite the use of 10-fold less total GFs. The differentiated HLCs in the MP group exhibited ultrastructure and functional characteristics comparable with the conventional soluble GF group. The generated HLCs in the MP group were successfully engrafted in an acute liver injury mouse model and maintained hepatocytic function after implantation. These results suggested that sustained and localized delivery of GFs using MPs might offer a novel approach towards scalable technologies for hepatocytic differentiation and engineer a better 3D microenvironment for cells.

人多能干细胞衍生的肝细胞样细胞（hPSC-HLCs）的生物医学应用依赖于有效的大规模分化，这通常是通过在生物反应器中进行三维（3D）多细胞球体的悬浮培养来完成的。但是，这种方法需要大量的生长因子（GFs），并且需要克服hPSC椭球的固有3D结构所造成的有限的扩散运输。在这里，我们假设通过在hPSC椭球体中掺入载有GF的可降解聚合物微粒（MPs）来实现GF的局部递送将避免这种局限性。在这项研究中，用于肝细胞分化的GFs被封装在明胶包被的聚乳酸（l-乳酸）/聚（DL-乳酸-乙醇酸共聚物）（PLLA / PLGA）MP，随后将其掺入hPSC球体中。基因表达分析表明，尽管使用的总GF少了10倍，但GF的MP传递导致了相似的肝细胞标志物表达水平。MP组中分化的HLC表现出与常规可溶性GF组相当的超微结构和功能特性。MP组中生成的HLC已成功植入急性肝损伤小鼠模型中，并在植入后保持了肝细胞功能。这些结果表明，使用MP持续和局部递送GFs可能为实现可扩展的肝细胞分化技术提供新方法，并为细胞设计更好的3D微环境。