Viscoelasticity-tuned hyaluronic acid–gelatin microcarriers with thermoresponsive polymer grafts for scalable cell expansion and gentle harvesting

Abstract

Three-dimensional cell culture using microcarriers is an effective strategy for scalable cell expansion. However, conventional enzymatic detachment can compromise cell viability, surface proteins, and native signaling. We report viscoelasticity-tunable hyaluronic acid (HA)-gelatin microspheres as microcarriers, engineered with a thermoresponsive polymer coating to enhance cell attachment and enable gentle harvesting. Gelatin-only (GLA), gelatin-HA (G-HA), and gelatin-HA-L-lysine (G-HA-L) microspheres were fabricated. HA incorporation and lysine functionalization were used to tune microsphere mechanics and interfacial stability. Frequency-sweep rheology revealed that HA-containing formulations exhibited higher elastic dominance (G′ > G″) and a broader, more stable viscoelastic response than gelatin-only and a commercial gelatin microcarrier benchmark, with G-HA-L showing the most favorable balance of stiffness and damping (highest G′/ G″ across the tested window). The microspheres were subsequently coated with poly (N-isopropylacrylamide-co-acrylic acid) (P(NIPAM-AAc)), producing a temperature-responsive interface. Importantly, the thermoresponsive coating enhanced early cell attachment, particularly on G-HA-L (reaching ∼70% within 4 h and approaching ∼90% by 24 h), outperforming both coated G-HA and commercial microcarriers. For harvesting, low-temperature conditioning markedly improved cell release and recovery compared to trypsin-only controls, consistent with temperature-triggered polymer swelling, which facilitated detachment. Collectively, these results demonstrate that coupling viscoelastic microcarrier design with thermoresponsive surface engineering provides a promising platform for efficient cell growth and gentle, process-friendly harvesting, with potential applications in tissue engineering and regenerative medicine.