Hmscs

Bone marrow-derived hMSCs (Lonza, Walkersville, MD) were purchased at passage 2 and expanded for use at passage 4. Cells were cultured and passaged according to manufacturer’s protocol. Cell suspensions containing 1 million cells/ml were frozen in cryovials in LN₂ for further use. Cryovials were then thawed and hMSCs were seeded onto 182 cm² tissue culture-treated plastic at 5,000 cells/cm². Maintenance media (MSCGM, Lonza) was added and hMSCs were propagated at 37 °C, 5% CO₂, and 85% humidity for 7 days or until ~90% confluence, with media changes every 3 days.

hMSCs (Lonza) were expanded at low density on T175 tissue culture polystyrene (TCPS) flasks to maintain their multipotency and used for experiments by passage 6. hMSCs were grown in growth medium comprising of α-Eagle Essential Media (α-MEM) supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 100 U/mL penicillin/streptomycin at 37°C and 5% CO₂. Media was exchanged every other day and hMSCs were passaged or harvested at 75% confluency.

HUVECs and hMSCs used in the study were obtained from Lonza, and were each used until passage 10. HUVECs were cultured in EGM-2 (Lonza), and hMSCs were cultured in mesenchymal stem cell growth medium (MSCGM) (Lonza). MDA-MB-231 cells were obtained from ATCC and were cultured in DMEM supplemented with 10% FBS and 2 mM L-Glutamine. HEK293T cells were obtained from ATCC and were cultured in DMEM supplemented with 10% FBS. The Med411-FH medulloblastoma patient-derived xenograft line [77 (link),81 (link)] was maintained in the Wechsler-Reya lab. Tumors were harvested and dissociated into single cells the day of culture, and were subsequently encapsulated in their respective growth environments. For Med411-FH, media was composed of NeuroCult basal medium supplemented with Proliferation Kit (StemCell Technologies), 2 μg/mL heparin (Sigma), 20 ng/mL epidermal growth factor (Sigma), and 20 ng/mL fibroblast growth factor (Lonza).