Isolation and Characterization of Human Dental Pulp Stem Cells

The human dental pulp cell (hDPSC) were enzymatically isolated from impacted third molars obtained from 10 adults (18 to 22 years of age), as previously described [28 (link),35 (link)], in compliance with Italian legislation (including informed consent and institutional review board approval of the protocol number 7413). The cells were cultured with α-minimum essential medium (α-MEM) supplemented with 15% fetal bovine serum (FBS), 2 mM L-glutamine, 100 mM L-ascorbic acid-2-phosphate, 100 U/mL penicillin-G, 100 mg/mL streptomycin, and 0.25 mg/mL fungizone (HyClone, Milan, Italy) and maintained in 5% CO₂ at 37 °C. Proliferation, clonogenic potential, and stem cell markers were analyzed. STRO-1+ stem cells were directly sorted from pulp cell at passage 3 with mouse anti-human STRO-1 IgM (Life Technologies, Milan, Italy) with immune magnetic beads according to the manufacturer’s protocol (Dynabeads; Life Technologies, Milan, Italy). After cell sorting, each of the following experiments was performed in triplicate on pooled STRO-1 sorted cells (STRO-1+ cells).