Mediating human stem cell behaviour via defined fibrous architectures by melt electrospinning writing.

The architecture within which cells reside is key to mediating their specific functions within the body. In this study, we use melt electrospinning writing (MEW) to fabricate cell micro-environments with various fibrous architectures to study their effect on human stem cell behaviour. We designed, built and optimised a MEW apparatus and used it to fabricate four different platform designs of 10.4 ± 2 μm fibre diameter, with angles between fibres on adjacent layers of 90°, 45°, 10° and R (random). Mechanical characterisation was conducted via tensile testing, and human skeletal stem cells (hSSCs) were seeded to scaffolds to study the effect of architecture on cell morphology and mechanosensing (nuclear YAP). Cell morphology was significantly altered between groups, with cells on 90° scaffolds having a lower aspect ratio, greater spreading, greater cytoskeletal tension and nuclear YAP expression. Long term cell culture studies were then conducted to determine the differentiation potential of scaffolds in terms of alkaline phosphatase activity, collagen and mineral production. Across these studies, an increased cell spreading in 3-dimensions is seen with decreasing alignment of architecture correlated with enhanced osteogenesis. This study therefore highlights the critical role of fibrous architecture in regulating stem cell behaviour with implications for tissue engineering and disease progression. STATEMENT OF SIGNIFICANCE: This is the first study which has investigated the effect of controlled fibrous architectures fabricated via melt electrospinning writing on stem cell behaviour and differentiation. After optimising the fabrication process and characterising scaffolds via SEM and mechanical testing, skeletal stem cells were seeded onto fibrous scaffolds with various micro-architectures. These architectures drove cell shape changes resulting in architecture dependent nuclear YAP localisation, suggesting altered mechanosensing at early time points. In agreement with these early markers, long term cell culture studies revealed for the first time that a 90° fibrous architecture is optimal for the osteogenic differentiation of skeletal stem cells.