Impact of temperature and humidity on the structural and biocompatibility of 3D-Printed PLA scaffolds for bone regeneration

Fused deposition modeling (FDM) is widely used in medical applications and provides a promising, cost-effective, and user-friendly solution to point-of-care environments. However, this on-site production necessitates extreme process reproducibility in ambient conditions. Although such a requirement is necessary, the effects of environmental temperature and relative humidity during fabrication are poorly understood, especially when complex porous structures are considered. In this paper, we systematically investigate the impact of ambient temperature and relative humidity on the structural, mechanical, and biological performance of porous polylactic acid (PLA) scaffolds fabricated via FDM. Cylindrical porous scaffolds (2.5 cm diameter and heights of 5.3 and 10.3 cm) were printed under controlled conditions in ambient temperature (25-40 °C) and relative humidity (30-70%). Their pore structure (size and density), water-holding capacity, compressive hardness, and in-vitro cytocompatibility were investigated. The geometric fidelity and pore morphology of all scaffolds were similar across fabrication conditions, suggesting that ambient conditions did not influence the qualities in macroscopic visual prints. In comparison, the compressive Young's modulus increased with increasing temperature. A biocompatibility assay showed that variations in relative humidity had minimal effects on the mechanical performance of the scaffolds but affected cell viability and reactive oxygen species (ROS) generation. Conversely, at higher fabrication temperatures, high intracellular ROS activity was observed without affecting the structural integrity of the scaffolds. These findings establish a practical processing window of moderate temperatures (25-35 °C) and low-to-moderate humidity (30-50% RH) that balances mechanical stability with biological compatibility. These insights into environmental dependence can be used to enhance process reliability and repeatability, which are essential for translating scaffolds printed by FDM into low-cost, high-fidelity clinical and point-of-care applications.