Culturing Mammalian Cell Lines for Experimental Studies

The MS-5 cells [25 (link)], OP-9 cells, distributed by RIKEN BRC through the National Bio-Resource of the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT), and MC3T3-E1 cells, were maintained as described [10 (link)]. Mouse BM-derived primary mesenchymal stem/stromal cells (MSCs; Gibco), and Med1^+/+ and Med1^−/− mouse embryonic fibroblasts (MEFs) that harbored a p53 inactivation [10 (link)], were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS) [10 (link)]. The MB-1 niche-dependent myeloblastoma cells [26 (link), 27 (link)] were maintained by coculturing with mitomycin C (MMC)-treated OP-9 cells and used by coculturing with MMC-treated either MS-5 or OP-9 cells [12 (link)]. In some experiments, 0.5 μM LDN-193189 (Selleck) or 10 μM warfarin (Wako) was added to the coculture [24 (link)].

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Kuronuma K., Yokoi A., Fukuoka T., Miyata M., Maekawa A., Tanaka S., Matsubara L., Goto C., Matsuo M., Han H.W., Tsuruta M., Murata H., Okamoto H., Hasegawa N., Asano S, & Ito M. (2020). Matrix Gla protein maintains normal and malignant hematopoietic progenitor cells by interacting with bone morphogenetic protein-4. Heliyon, 6(4), e03743.

Publication 2020

LDN-193189 warfarin

Cells Coculture Cultured medium Embryonic Fetal bovine serum Fibroblasts Ldn 193189 Med1 Mesenchymal stem cells Mitomycin c Mouse Warfarin

Corresponding Organization : Waseda University

Other organizations : Japan Science and Technology Agency

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Variable analysis

independent variables

Presence of LDN-193189 (0.5 μM)
Presence of warfarin (10 μM)

dependent variables

Cell culture and maintenance of MS-5 cells, OP-9 cells, MC3T3-E1 cells, mouse BM-derived primary mesenchymal stem/stromal cells (MSCs), Med1+/+ and Med1−/− mouse embryonic fibroblasts (MEFs), and MB-1 niche-dependent myeloblastoma cells

control variables

Culture medium (Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS))
Mitomycin C (MMC)-treated OP-9 cells for co-culture with MB-1 cells
Mitomycin C (MMC)-treated MS-5 or OP-9 cells for co-culture with MB-1 cells

controls

Positive control: Co-culture of MB-1 cells with MMC-treated OP-9 cells
Negative control: Co-culture of MB-1 cells with MMC-treated MS-5 or OP-9 cells

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