| Project/Area Number |
17591597
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Orthopaedic surgery
|
|---|
| Research Institution | KEIO UNIVERSITY |
|---|
Principal Investigator |
MATSUZAKI Kenichiro KEIO UNIVERSITY, School of Medicine, assistant professor (30317169)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
TOYAMA Yoshiaki Keio University, School of Medicine, Professor (40129549)
|
|---|
| Project Period (FY) |
2005 – 2006
|
| Project Status |
Completed (Fiscal Year 2006)
|
| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2006: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2005: ¥2,500,000 (Direct Cost: ¥2,500,000)
|
|---|
| Keywords | Osteoclast / Bone metabolism / Cell biology |
|---|
| Research Abstract |
We hypothesized that osteoclast precursors can be immortalized by inactivating P53 and RB, which are known as tumor suppressor genes. We chose CFU-GM (colony- forming units of granulocytes/macrophages) derived from human bone marrow as a target of immortalization. They were infected with human papiloma virus E6/E7 using lenti-viral vector. E6 inactivate P53 and E7 inactivate RB (retinoblastoma). We got some cell lines that continued to proliferate, but they failed to differentiate into osteoclast.
Some of them turned out to be stromal cells by immuno-staining with Stro-1. Therefore we thought that human CFU-GM was not suitable for immortalization because they were contaminated with stromal cells. We next chose human CD14 positive peripheral blood monocytes as a target of immortalization.
It might be difficult for the osteoclast precursors to be immortalized, because the termination of cell cycle cannot happen due to continuing expression of immortalizing genes. So we developed the inducible lenti-viral vector with Tet on-off system. Cells continue to proliferate at maintaining stages by keeping the immortalizing genes on, and once the immortalizing gene are switched off by Tet on-off system, cells are ready to differentiate into osteoclast. Immortalized osteoclast precursors have not yet to be established so far, but we are continuing to try to get them by optimizing Tet on/off system with a young investigator.
|
