Influence of Scaffold Topography and Culture Duration on Fibroblast Morphology in Tissue Engineering

Cell morphology is not only integral to its function within the body but also plays a critical role in cellular behavior and fate. In tissue engineering, cell-scaffold interactions play a critical role because scaffold physical and biochemical characteristics, such as pore size, fiber alignment, and surface architecture, directly influence cellular morphology and behavior. These interactions impact key biological processes, including adhesion, proliferation, migration, and differentiation of the cells, ultimately influencing tissue formation and regeneration. This study investigated how scaffold topography and culture time influence fibroblast morphology and behavior in a bilayer scaffold consisting of randomly oriented fiber layer and aligned fiber layer. Fibroblasts were seeded onto the scaffolds and cultured for 1, 3, 6, or 9 days, and nuclear and cytoskeletal morphologies were quantified using shape descriptors, including nuclear and cellular roundness, eccentricity, aspect ratio, and area ratio. The results demonstrate that scaffold fiber alignment significantly modulates cellular morphology, with aligned fibers promoting elongated, aligned morphologies and randomly oriented fibers favoring branched, multidirectional spreading. Culture time emerged as a key factor, as cells on both surfaces exhibited more rounded, stabilized morphologies by day 6, suggesting time-dependent remodeling and interaction with the scaffold microarchitecture. Specifically, aligned fiber-like scaffold surfaces may benefit regeneration of uniaxially aligned tissues, such as tendon, ligament, or nerve, whereas random fiber-like scaffold surfaces may support stromal or bone environments requiring isotropic spreading. Furthermore, the bilayer scaffold architecture holds promise for complex tissue interfaces, such as the periodontium or osteochondral units, where region-specific topographical cues are essential for functional tissue integration.