Laser-patterned microgroove polystyrene culture dishes for engineering 3D raised texture cell sheets

Abstract Techniques for fabricating cell sheets with two-dimensional (2D) patterns have advanced significantly over the years. However, creating cell sheets featuring three-dimensional (3D) raised textures that mimic the complex architecture of natural tissues continues to present a considerable challenge. This study introduces a versatile methodology for fabricating cell sheets with both 2D patterns and 3D microstructures, utilizing picosecond laser-induced microgroove-patterned polystyrene (PS) culture dishes. This technology leverages the sequential interaction of the laser with the photosensitive coating and the PS substrate to create microgrooves with exceptional precision in both width and depth. Both direct ablation and coating-assisted ablation result in patterned culture dishes that demonstrate excellent cell compatibility, an absence of cytotoxicity, and the ability to regulate cell proliferation. The patterned PS dishes create distinct 3D microenvironments that guide cell contact and adhesion arrangements, thereby modulating gene expression and protein secretion in normal human dermal fibroblasts. Notably, key proteins such as type I alpha1 collagen ( Col-1 ), type VI alpha1 collagen ( Col-6 ), Elastin, fibronectin, and matrix metalloproteinase 2 ( MMP-2 ) are significantly influenced by the structure of pattern. Furthermore, cell sheets with raised textures (CSRTs) can be detached from the patterned PS culture dishes while preserving their 3D structure for over 72 h, with structural longevity dependent on feature size. To our knowledge, this study represents the first successful fabrication of CSRT using a laser-induced micro-patterning technique. This approach provides foundational insights into the development of biomimetic tissues for regenerative medicine and advanced in vitro models, offering a promising platform for future applications in tissue engineering and biomedical research.