Zein/Fucoidan microcarriers promote myogenic differentiation via topographical cues and hydrodynamic modulation

Microcarrier surface topography and fluid shear stress (FSS) critically regulate cellular behavior. An edible zein/fucoidan microcarrier with 200 μm grooves was designed for this study. Computational fluid dynamics (CFD) simulations and experiments were combined to analyze surface microfluidic characteristics during dynamic culture and their cellular effects. The research demonstrates controlled myogenic differentiation through groove topography and FSS modulation for scalable cultured meat production. The results demonstrated that the grooved microcarriers exhibited excellent cell attachment and proliferation capacity in spinner flask dynamic culture, achieving a maximum cell density of 1.16 × 10⁶ cells/mL, comparable to commercial Cultispher-S microcarriers. The groove structure promoted cell alignment through contact guidance, significantly enhancing the gene expression of myogenic differentiation markers (myogenic differentiation 1, MyoD1; α-actinin; myosin heavy chain, MHC) and cell fusion (myomaker, MYMK). CFD simulations revealed that the grooves created a low-shear microenvironment (minimum average FSS: 3.93 × 10^-2 Pa, maximum: 1.18 × 10^-1 Pa), which effectively avoided high FSS-induced damage while maintaining mechanical stimulation. This optimal mechanical microenvironment further activated the expression of key genes involved in early-stage (MyoD1; myogenin, MyoG; myocyte enhancer factor 2C, MEF2C) and late-stage (α-actinin; myosin heavy chain 2, Myh2) myogenic differentiation. Flat and spherical microcarriers showed lower myogenic differentiation efficiency. This study elucidates the synergistic mechanism between groove structures and the low FSS microenvironment within grooves, providing a novel scaffold design rationale that combines biomimetic topology with fluid dynamics compatibility for large-scale cultured meat production.