Biofunctional thermoresponsive gelatin-PNIPAm microcarriers with embedded zero-valent iron nanoparticles for enhanced human adipose-derived stem cell expansion and differentiation.

Human adipose-derived stem cells (ADSCs) hold substantial promise for regenerative medicine due to their multipotency and immunomodulatory capacity; however, efficient and scalable expansion remains a critical bottleneck. Conventional microcarrier culture systems typically rely on enzymatic dissociation for cell harvesting, which can compromise stemness, reduce viability, and limit process robustness. Here we report a thermoresponsive, magnetically responsive microcarrier platform (ZVI-GMC) comprising a gelatin core functionalized with poly(N-isopropylacrylamide)-allylamine (PNIPAm-ALA) and embedded zero-valent iron nanoparticles (ZVI NPs). The gelatin-PNIPAm-ALA hybrid interface enables temperature-triggered, enzyme-free cell detachment, supporting repeated passaging of ADSCs while preserving their viability, phenotype, and differentiation potential. Incorporation of ZVI NPs enhances structural stability, imparts magnetic responsiveness for facile handling, and promotes ADSCs proliferation under dynamic culture conditions. ADSCs cultured on ZVI-GMC microcarriers retained robust trilineage differentiation capacity toward osteogenic, adipogenic, and chondrogenic lineages, as confirmed by Alizarin Red S, Oil Red O, and Alcian Blue staining, respectively. Collectively, this macromolecule-nanoparticle hybrid microcarrier system provides a scalable and bioactive strategy for ADSCs expansion and lineage maintenance. By integrating thermoresponsive polymers, biofunctional macromolecular matrices, and magnetic nanostructures within a single platform, ZVI-GMC offers strong potential for incorporation into automated, closed, and high-throughput stem cell biomanufacturing workflows, positioning it as a promising next-generation microcarriers for regenerative medicine applications.