Dihydrocortisone

Standards of prednisolone, prednisone, cortisone, cortisol, dihydrocortisone, aldosterone, allotetrahydrocortisol, urocortisol, tetrahydrocortisone, corticosterone, deoxycorticosterone, α-cortolone and 6β-hydroxycortisol were purchased from Sigma-Aldrich (St. Louis, MO, USA). Internal standards were prednisolone-d₈ (TRC, Canada), cortisol-d₄ and prednisolonde-d₄ (Sigma-Aldrich). Reagents were of analytical grade (Merck, Darmstadt, Germany) when used for extraction purposes and of liquid chromatography-mass spectrometry (LC-MS) Optima grade (Fisher Scientific, Loughborough, UK) for ultra-high performance LC (UHPLC) tandem MS (MS/MS) and high-resolution MS (HRMS) applications. Ultrapure water was obtained by usage of a purified-water system (Sartorius AG, Goettingen, Germany). Stock solutions were prepared in ethanol at a concentration of 200 μg mL⁻¹ and stored in dark glass bottles at −20 °C.

For detailed description, please refer to https://web.expasy.org/cellosaurus/. Briefly: K562: erythroleukemia immortalized myelogenous cell line. MEG-01: human megakaryoblastic leukemic cell line. SUP-B15: Human B cell precursor leukemia. HEL: Human Erythroleukemic cell line. SET-2: human essential thrombocythemic. UKE1: human essential thrombocythemic cells. B-ALL BCR-ABL1⁺ GFP⁺: cell line established from Cdkn2a^−/− murine bone marrow cells infected with a pMIG retrovirus expressing a p190^BCR-ABL1-GFP cassette (a gift from Prof. Ghysdael, Paris Diderot University). K562 and HEL cell lines were grown in RPMI medium (Lonza) supplemented with 10% of FBS (Fetal Bovine Serum) (Sigma); MEG-01 and SET-2 were grown in RPMI medium (Lonza) supplemented with 20% of FBS (Fetal Bovine Serum); UKE1 were grown in IMDM medium (EuroClone) supplemented with 10% FBS (Fetal Bovine Serum) (Sigma) and 10% Horse Serum (Invitrogen), 1uM di HydroCortisone (Sigma); SUP-B15 were grown in RPMI medium (Lonza) supplemented with 10% of FBS (Usa Origin Sigma); B-ALL BCR-ABL1⁺ GFP⁺ were grown in RPMI (Lonza) with 15% FBS (Sigma) and 7,5% NaHCO₃. The medium of all the cell lines was supplemented with 100 μg/ml penicillin/streptomycin (EuroClone), 4mM L-glutamine (EuroClone). Cells were grown at an optimal concentration of 0,5 × 10⁶ cells/mL, in a humidified 5% CO₂ atmosphere at 37 °C.