Topography‐Regulated Screening Platform for Myoblast Contact Guidance

ABSTRACT Identification and development of physiologically relevant in vitro models are critical for the advancement of creating platforms suitable for tissue engineering, drug testing, and mechanistic research on pathologies. This study focuses on skeletal muscle pathologies and establishes a topography‐integrated screening platform combining 96‐well plates with an aligned surface topography with pattern dimensions ranging from 0 to 23.3 µm in wavelength and 0 to 3.15 µm in height. The system enables simultaneous analysis of myoblast responses on 12 different patterns while preventing cross‐talk between cells when they are grown on gradient surfaces for screening. Mechanistic analysis demonstrates topography‐dependent modulation of focal adhesion distribution and cytoskeletal reorganization. Surface topography directs myotube alignment through contact guidance, achieving orientation control in optimal configurations and wrinkle‐induced polarization mimics native tissue architecture more effectively than planar substrates. Micropattern dimensions differentially regulate cellular morphogenesis, with micro‐size topography features producing minimal myotubes (reduced area, quantity, and length) versus enhanced differentiation at larger topography dimensions. This model enables rapid generation of morphologically distinct myotube phenotypes through dimensional tuning, providing a physio‐mimetic solution for objectively evaluating muscle therapeutics and is expandable toward other cell culture approaches to identify optimum cell stimulation for specialized culture.