Isolation and Culture of hDPSCs

hDPSCs were isolated as described previously [26 (link)]. All donors provided informed consent. The study was conducted in accordance with the Declaration of Helsinki and applicable local regulatory requirements and laws. All procedures were approved by the Ethics Committee of the University of Valencia (Spain). The dental pulp of human third molars was gently removed under aseptic conditions using cow horn forceps with a small excavator, and immersed in culture tubes filled with culture medium. The specimens were then divided into small pieces using a bistoury blade, immersed in HBSS, and incubated for 2 hours at 37°C in an atmosphere of 5% CO₂ and 95% air. The supernatant was removed, and 0.1% type I collagenase and dispase (Sigma-Aldrich, Madrid, Spain) was added for 15 minutes, followed by centrifugation at 1,500 rpm for 10 minutes. The supernatant was removed, and the cells were plated in 25-cm² flasks in αMEM culture medium (Sigma-Aldrich) containing penicillin/streptomycin, 10% FCS (Sigma-Aldrich), amphotericin B, 2 mM l-glutamine, and 100 μM ascorbic acid (Sigma-Aldrich). The medium was replaced every 3–4 days. Once the cells reached confluence, flow cytometry was performed.