Effect of Sterilization and Disinfection Methods on the Physicochemical, Mechanical, and Biological Properties of Highly Porous Thermoset Polycaprolactone-Based Scaffolds

Tissue engineering scaffolds intended for clinical translation must fulfill strict sterilization requirements to ensure patient safety and regulatory compliance. However, alcohol-based disinfection remains widely used in in vitro studies, despite failing to reflect clinically relevant terminal sterilization conditions and potentially misrepresenting scaffold performance. Here, we systematically evaluated the effects of terminal sterilization and commonly employed disinfection methods on highly porous thermoset polycaprolactone methacrylate (4PCLMA) polymerized high internal phase emulsion (PolyHIPE) scaffolds from a translational perspective. Steam autoclaving under dry, humid, and wet conditions and dry heat treatment were investigated as terminal sterilization approaches, while ultraviolet (UVC) exposure and 70% alcohol treatment were assessed as disinfection methods. Scaffold physicochemical, mechanical, and biological properties were comprehensively characterized, including thermal stability, chemical structure, pore morphology, compressive mechanical performance, sterility, and in vitro cell response using L929 fibroblasts. Terminal sterilization by autoclaving and dry heat preserved the chemical integrity, highly porous architecture, and compressive mechanical properties of the thermoset 4PCLMA PolyHIPE scaffolds. Crucially, only terminal sterilization methods achieved complete sterility throughout the scaffold volume, whereas UVC exposure and alcohol treatment failed to eradicate microbial contamination in these highly porous constructs. Sterilization or disinfection treatments didn't adversely affect cell viability, attachment, or proliferation. Collectively, these findings demonstrate that alcohol-based disinfection is insufficient for highly porous tissue engineering scaffolds and that 4PCLMA PolyHIPE scaffolds are compatible with clinically relevant terminal sterilization methods. This work highlights the importance of integrating appropriate sterilization strategies early in scaffold design to enhance translational relevance and clinical readiness.