Retrovirus-mediated conditional immortalization and analysis of established cell lines of osteoclast precursor cells.
Osteoclast precursor cells (OPCs) have previously been established from bone marrow cells of SV40 temperature-sensitive T antigen-expressing transgenic mice. Here, we use retrovirus-mediated gene transfer to conditionally immortalize OPCs by expressing temperature-sensitive large T antigen (tsLT) from wild type bone marrow cells. The immortalized OPCs proliferated at the permissive temperature of 33.5 degrees C, but stopped growing at the non-permissive temperature of 39 degrees C. In the presence of receptor activator of NFkappaB ligand (RANKL), the OPCs differentiated into tartrate-resistant acid phosphatase (TRAP)-positive cells and formed multinucleate osteoclasts at 33.5 degrees C. From these OPCs, we cloned two types of cell lines. Both differentiated into TRAP-positive cells, but one formed multinucleate osteoclasts while the other remained unfused in the presence of RANKL. These results indicate that the established cell lines are useful for analyzing mechanisms of differentiation, particularly multinucleate osteoclast formation. Retrovirus-mediated conditional immortalization should be a useful method to immortalize OPCs from primary bone marrow cells.
Kawata
Shigehisa
S
Laboratory of Molecular Oncology, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0101, Japan.
Suzuki
Jun
J
Maruoka
Masahiro
M
Mizutamari
Megumi
M
Ishida-Kitagawa
Norihiro
N
Yogo
Keiichiro
K
Jat
Parmjit S
PS
Shishido
Tomoyuki
T
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002470:Cell Survival; D051379:Mice; D010010:Osteoclasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D053245:RANK Ligand / Q000494:pharmacology; D012190:Retroviridae / Q000235:genetics*; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects; D013696:Temperature
D053245:RANK Ligand