Complete α minimum essential medium

Human periodontal ligament tissues were obtained from premolars from 20 donors (aged 13–24 years, 10 men and 10 women, without oral or systematic diseases) during orthodontic procedures under informed consent. Ethics Committee approval was provided by the School of Stomatology, Sun Yat-sen University.
hPDLCs were harvested and cultured as previously reported [29 (link)]. Tissues attached to the middle third of tooth root was collected and cut into small pieces (1 mm³) followed by digestion with 3 mg/mL type 1 collagenase (Life Technologies, Carlsbad, CA, USA) and 4 mg/mL dispase (Life Technologies) at 37°C for 1 h. The tissues were cultured in complete α-minimum essential medium (Gibco, Grand Island, NY, USA) containing 10% (v/v) fetal bovine serum (Gibco), 100 U/mL penicillin, 100 μg/mL streptomycin (HyClone, Logan, UT, USA), and 5 mM L-glutamine (Gibco). The medium was changed every 3 days. When the human periodontal ligament cells reached 80% confluence, they were subcultured.

This research was approved by the Ethics Committee of Affiliated Stomatological Hospital of Sun Yat-sen University (ERC-[2016]-46). Human periodontal ligament tissues were obtained from healthy premolars from 12 donors (12-20 years old, 6 males and 6 females) with orthodontic demands. The middle section tissues of the root surfaces were scraped collected and enzymatically digested with 3 mg/mL collagenase type I and 4 mg/mL dispase for 1 h. Subsequently, the tissues were cultured in a complete α-minimum essential medium (Gibco, Grand Island, NY, USA) containing 10% (v/v) fetal bovine serum (Gibco), 100 U/mL penicillin, 100 μg/mL streptomycin (HyClone, Logan, UT, USA), and 5 mM L-glutamine (Gibco) at 37°C in 5% CO₂. The hPDLSCs used in this study were a mixture of cells collected from 12 donors to decrease individual variation.