Ham s f12

Early passages of human brain tumor initiating cells (BTICs), were used and previously validated by Johns Hopkins Genetic Resources Core Facility11 (link). GBM612 cells isolated from intraoperative tissue are multipotent and are able to form diffuse tumors when implanted into animal models11 (link)12 13 . BTICs were grown in culture flasks coated with poly-L-ornithine and laminin and cultured with stem cell media composed of DMEM/F12, B27 supplements, EGF, and FGF14 (link). The molecular subtypes of the GBM cell lines were characterized previously using a metagene score based approach15 (link). Three different GBM subtypes including proneural (GBM 612 and 276), mesenchymal (GBM 626 and 253), and classical (GBM 965) were selected for this study. U87 cells were acquired from ATCC and cultured as recommended in EMEM supplemented with 10% fetal bovine serum. Primary fetal neural progenitor cells F54 were obtained as described previously16 (link) and were maintained in 2:1 high-glucose DMEM (Invitrogen)/Ham’s F-12 (Cellgro), 1X B-27, 1% anti–anti, 20 ng/mL bFGF, 20 ng/mL EGF, 20 ng/mL leukemia inhibitory factor (LIF, Millipore, Billerica, MA), and 5 μg/mL heparin (Sigma).

IPSCs-derived human alveolar type 2 cells (iAT2s) (differentiated from the SPC2-ST-B2 iPSC line) were expanded as alveolospheres in growth factor reduced Matrigel (Corning, 354230) as described elsewhere (Jacob et al., 2019 (link)). The iAT2s were maintained in IMDM medium containing 25% Ham’s F12 (Cellgro, 10–080-CV), 1% B27 supplement, 0.5% N2 supplement, 0.05% BSA (Invitrogen, 15260037), 200 ng/ml Primocin (Invivogen, NC9141851), 1X GlutaMAX, 50 μg/ml ascorbic acid (Sigma, A4544), 0.45 mM monothioglycerol (Sigma, M6145), 3 μM CHIR99021, 10 ng/ml rhKGF (R&D, 251-KG-010), 50 nM Dexamethasone (Sigma, D4902), 10 μM Y27632 (for first 3 days of culture), 0.1 mM 8-bromoadenosine 3′,5′-cyclic monophosphate sodium salt (Sigma, B7880), and 0.1 mM 3-isobutyl-1-methylxanthine (Sigma, I5879).

MDA-MB-231, Hs578t, MCF-7, and MDA-MB-468 cells were purchased from American Type Culture Collection (ATCC). MDA-MB-231 and MCF-7 were cultured in Dulbecco’s Modified Eagle Medium (DMEM) (Corning CellGro., Manassas, VA, USA) supplemented with 10% fetal bovine serum (Sigma-Aldrich, St. Louis, MO, USA), 1% pen–strep, and 1% L-glutamine (Corning, CellGro). MDA-MB-468 was cultured in DMEM (Corning CellGro) supplemented with 10% fetal bovine serum, 1% pen–strep, 1% L-glutamine, 1% sodium pyruvate, and 1% Minimum Essential Medium (MEM) amino acids (Corning CellGro). Hs578t was cultured in MEM supplemented with 10% fetal bovine serum, 1% pen–strep, and 1% L-glutamine (Corning CellGro). SUM159PT cells were obtained from Asterand and cultured in Ham’s/F-12 (Corning CellGro) supplemented with 10% fetal bovine serum, 1% pen–strep, 1% L-glutamine, 10 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), 1 μg/mL hydrocortisone, and 5 μg/mL insulin. All cell lines were maintained in a humidified incubator at 5% CO₂ and 37 °C.

HeLa, COS7, HEK293, and PAC-1 (rat vascular smooth muscle cells) cells were cultured in DMEM medium (Invitrogen) supplemented with 10% FBS (Sigma) and penicillin/streptomycin (Invitrogen). WT, ldlA, pgsa-745, Lec1, Lec2, and Lec8 Chinese hamster ovary (CHO) cells were cultured in Ham’s F-12 (Cellgro) with 10% FBS and penicillin/streptomycin. For secreted recombinant protein expression in media, transfected cells were grown in OPTI-MEM medium (Invitrogen) with 2% IgG-free FBS (Hyclone). Total cell extracts were prepared by solubilizing cells in a cell lysis buffer [0.5% Triton X-100, 0.5% sodium deoxycholate, 50 mM Tris-HCl (pH 7.5), 250 mM NaCl, 5 mM EDTA, and 50 mM sodium fluoride] in the presence of complete protease inhibitor cocktail (Roche).