Establishment and characterization of a novel human immature megakaryoblastic leukemia cell line, M-MOK, dependent on fibroblasts for its viability.
A novel fibroblast-dependent human immature megakaryoblastic leukemia cell line (M-MOK) was established from the bone marrow of a girl with acute megakaryoblastic leukemia, and its growth was determined to be completely dependent on the presence of human embryonic lung-derived fibroblasts, HEL-O. Adhesive interaction between M-MOK and HEL-O was crucial for viability; once HEL-O was removed from the culture, mortality was total within a few days. On HEL-O cells, M-MOK could be passaged for more than 2 years. With regard to surface marker profile, the established cells were positive for CD11a, CD13, CD18, CD33, CD34, CD41b, CD42b, CD54, and c-kit antigens, but negative for HLA class II antigen and glycophorin. Histochemically, the cells were negative for myeloperoxidase, nonspecific esterase, and naphthol ASD chloroacetate esterase staining. Electron-microscope examination revealed the cells to be negative for platelet peroxidase (PPO). After induction of differentiation by a phorbol ester, however, the cells were demonstrated to be positive for PPO with a morphological change to megakaryocytes. From these results, M-MOK was considered to represent an immature cell line of megakaryocyte lineage. Studies of the mechanisms sustaining the HEL-O-dependent continuous in vitro growth of M-MOK cells revealed the following results: (1) M-MOK could grow even when separated from HEL-O by a nucleopore membrane; (2) conditioned medium (CM) from HEL-O supported the growth of M-MOK for more than 1 month without feeder cells; (3) the growth of M-MOK on HEL-O or CM supplement was nearly entirely inhibited by anti-GM-CSF (1 microgram/mL); (4) GM-CSF mRNA was detected in HEL-O cells; and (5) HEL-O was found to secrete GM-CSF into the culture medium. Taken together, the growth of M-MOK might therefore be driven by a soluble factor, that is, GM-CSF secreted from HEL-O cells. The presence of HEL-O, however, inhibited anti-GM-CSF-induced M-MOK death. Co-culture of M-MOK and HEL-O cells thus offers a useful experimental model for analysis of interactions between hematopoietic stem cells and stromal cells.
Itano
M
M
Department of Pediatric Oncology, Tohoku University, Sendai, Japan.
Tsuchiya
S
S
Minegishi
N
N
Fujie
H
H
Minegishi
M
M
Morita
S
S
Yambe
T
T
Ohashi
Y
Y
Masuda
T
T
Koike
T
T
D001483:Base Sequence; D001792:Blood Platelets / Q000201:enzymology; D001853:Bone Marrow / Q000473:pathology; D002448:Cell Adhesion; D002470:Cell Survival; D017077:Culture Media, Conditioned; D004622:Embryo, Mammalian; D005260:Female; D005347:Fibroblasts / Q000502:physiology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology; D006651:Histocytochemistry; D006801:Humans; D016130:Immunophenotyping; D007223:Infant; D007947:Leukemia, Megakaryoblastic, Acute / Q000473:pathology*; D008168:Lung; D008969:Molecular Sequence Data; D009195:Peroxidase / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D014407:Tumor Cells, Cultured
D017077:Culture Media, Conditioned; D012333:RNA, Messenger; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D009195:Peroxidase