Abstract
Scaling up cell therapy requires efficient expansion of high-quality cells. Microcarrier(MC)-based systems offer high surface-to-volume ratios and reduce culture media usage. In this study, we developed BrushGel, a temperature-responsive MC composed of gelatin methacryloyl (GelMA) hydrogel particles coated with poly(N-isopropyl acrylamide) (PNIPAM) polymer brushes via covalent grafting. BrushGel was fabricated using a flow-focusing droplet microfluidic device and functionalized using carbodiimide chemistry ( 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide-N-Hydroxysuccinimide, EDC-NHS). The degree of PNIPAM coating was tuned by varying the degree of methacrylation (DOM) of GelMA and the concentration of PNIPAM. Human dermal fibroblast (HNDF) cells cultured on the BrushGel under dynamic conditions showed a 4.9 fold increase in cell density, 12-fold upregulation in COL1A1 gene expression and elevated procollagen protein secretion compared to static culture. Low temperature detachment (4 °C) yeilded up to 65% detachment efficiency with >95% post-detachment viability. Clinical grade human bone marrow-derived mesenchymal stromal/stem cells (MSCs) expnaded 5.3 fold over five days on BrushGel with 69% detachment efficiency and 80% post-harvest viability using 10-fold less enzyme. BrushGel supported over 10 days of culture in spinner flasks, enabling enzyme-minimized, scalable cell expansion. These findings position BrushGel as a promising platfrom for dyanmic cell culture systems in regenerative medicine.