Scaffold Fiber Architecture and Uniaxial Stretch Differentially Regulate Cell and Nuclear Morphology in Human Dermal...

Cellular and nuclear morphology, together with directional alignment, are fundamental determinants of cell function and play critical roles in the formation, organization, and maintenance of functional fibrous connective tissues. In tissue-engineered systems, these characteristics emerge from the combined influence of scaffold microarchitecture and mechanical loading, which regulate cellular mechanosensing, cytoskeletal organization, and extracellular matrix remodeling, thereby directly affecting tissue regeneration outcomes. In this study, the coupled effects of scaffold fiber architecture and uniaxial stretching on scaffold organization and cell morphology were investigated using bilayer electrospun scaffolds composed of an aligned-fiber layer (AFL) that gradually transformed into a random-fiber layer (RFL). Scaffold morphology and strain-induced fiber alignment were characterized by scanning electron microscopy and two-dimensional fast Fourier transform analysis at elongation levels of 0%, 10%, and 20%. Adult human dermal fibroblasts were cultured on both AFL and RFL scaffold surfaces, and cellular and nuclear morphologies were quantified using shape descriptors under both unstretched conditions and sustained uniaxial stretching of 10%. Uniaxial stretching promoted alignment of initially random fibers while preserving the orientation of pre-aligned fiber architectures. At the cellular level, nuclear morphology exhibited little dependence on scaffold architecture under unstretched conditions but showed pronounced sensitivity to applied strain. In contrast, cell morphology was governed primarily by fiber alignment and displayed only limited sensitivity to sustained mechanical loading. These findings indicate that scaffold architecture and mechanical stretching regulate structural organization (from scaffold microstructure to cellular and nuclear morphology) through distinct yet complementary mechanisms. The ability to independently and complementarily control structural and mechanical cues offers an effective design strategy for bilayer or graded scaffolds intended for the regeneration of complex tissue interfaces, where both isotropic and anisotropic cellular organization are required.