Abstract

Background/Aim: Sensitive bioactive coatings, such as polycaprolactone with cholecalciferol, present handling challenges in animal models due to their susceptibility to mechanical damage. This study aimed to develop and validate a refined surgical protocol for inserting such polycaprolactone-cholecalciferol-coated titanium microscrews into the proximal tibia of Sprague-Dawley rats, ensuring primary stability and preserving the integrity of the nanofibrous coating. Materials and Methods: Fourteen male Sprague-Dawley rats (300-350 g) were used: one pilot and 13 animals in the main trial. A 3D anatomical tibia model was created for in-vitro validation of the surgical technique. The refined protocol incorporated enlarged cortical drilling (1.8 mm) before inserting microscrews (1.5x7 mm) to minimize friction on the outer cortical bone. Multimodal anesthesia, postoperative analgesia, and systematic clinical monitoring were implemented. Coating integrity, primary stability, wound healing, and animal welfare were evaluated through clinical observation, micro-computed tomography, and scanning electron microscopy (SEM). Results: The refined protocol achieved 100% procedural success. All implants reached immediate primary stability with uneventful first-intention healing and no critical adverse events. SEM confirmed preservation of morphology of the nanofibrous coating exclusively in the refined protocol group. Micro-computed tomography demonstrated consistent bone-to-implant contact in both cortical and trabecular compartments. Clinical recovery was rapid and with all animals welfare indicators Conclusion: The refined protocol safeguards the structural integrity of functionalized surfaces, ensures primary stability, and promotes early osseointegration, while complying with the principles of replacement, reduction, and refinement (3Rs). This model provides a valid experimental platform for investigations into osseointegration dynamics and the controlled local release of therapeutic agents.