Dulbecco s modified eagle s medium dmem ham s f 12

Unless otherwise stated, all chemicals, including MTT and mouse monoclonal anti-β-actin, were obtained from the Sigma Chemical Company, St. Louis, MO, USA. The mouse anti-rat TfR1 antibody was purchased from Invitrogen, Carlsbad, CA, USA, and the rabbit polyclonal anti-mouse Fpn1 antibody from Chemicon International, Temecula, CA, USA. The rabbit anti-rat DMT1 + IRE and DMT1 - IRE polyclonal antibodies were bought from the Alpha Diagnostic International Company, San Antonio, TX, USA, and goat anti-rabbit or anti-mouse IRDye 800 CW secondary antibodies from LI-COR Biosciences, Lincoln, NE, USA. Dulbecco’s Modified Eagle’s Medium (DMEM)/Ham’s F-12 was obtained from GIBCO, Grand Island, NY, USA and BSA from the Calbiochem-Novabiochem Corporation, San Diego, CA, USA. The scintillation cocktail and tubes were obtained from the Beckman Coulter Company, Fullerton, CA, USA. All experimental protocols were performed according to the Animal Management Rules of the Ministry of Health of China, and approved by the Animal Ethics Committees of Fudan University and The Chinese University of Hong Kong.

The hIDPSCs used in this study were isolated from the dental pulp of deciduous teeth of a 6-year-old individual (male), using a novel and disruptive technology developed by Kerkis et al. [18 (link)] and protected by the US patent number US20160184366A1. The hIDPSCs were cultivated until the fifth passage in basal medium (Dulbecco’s modified Eagle’s medium (DMEM)/Ham’s F12, supplemented with 15% fetal bovine serum, 100 U/mL penicillin, 100 μg/mL streptomycin, 2 mM L-glutamine and 2 nM nonessential amino acids, all from Gibco, Carlsbad, CA, USA) [23 (link)]. The MSC phenotype of these cells was confirmed using the criteria defined by the International Society for Cellular Therapy (ISCT) [24 (link),25 (link)]. The hIDPSCs employed in this study are CD105-, CD73- and CD90-positive, and CD45-, CD34-, CD11b- and HLA-DR-negative, as previously described by Kerkis et al. [18 (link)]. These cells also express high levels of BDNF, as previously demonstrated [26 (link)]. The hIDPSCs employed in this study were produced by the Brazilian facility Cellavita Pesquisas Científicas Ltda. (Valinhos-SP, Brazil) according to the good manufacturing practices (GMP) required by the Brazilian Health Regulatory Agency (ANVISA, RDC 508/21) for advanced therapy products [23 (link)]. These cells comprise the active component of the NestaCell® product [23 (link)].