Nanofibril guided spheroid formation for enhanced pluripotency and differentiation of human induced pluripotent stem cells

The spheroid formation derived from human-induced pluripotent stem cells (hiPSCs) generates a threedimensional (3D) microenvironment that fosters cell-cell interactions, modulates epigenetic regulation, and imitates early embryonic development, all of which contribute to biomedical applications including drug discovery, disease modeling, and cell replacement therapy. In this study, we developed 3D hiPSC spheroids by incorporating surface-engineered nanofibrils to enhance their pluripotency and differentiation capability. Polymeric nanofibrils were chemically immobilized with cell adhesion peptides that mimic the properties of vitronectin and laminin. The incorporation of nanofibrils into hiPSC spheroids significantly improved their viability and undifferentiated state, which may be due to the facilitated mass transport across the spheroids, enabled by the loosely held structure of cells and nanofibrils. By harnessing the enhanced pluripotency of hiPSC spheroids with nanofibrils, we induced differentiation into the hepatic lineage and found a significant upregulation of mature hepatocyte-specific markers in spheroids with nanofibrils compared with those without nanofibrils. Therefore, the combination of 3D hiPSC spheroids with surface-engineered nanofibrils offers a promising approach to enhancing pluripotency and hepatic differentiation, with potential implications for regenerative medicine.