2002 Fiscal Year Final Research Report Summary
Function analysis of Na^+ dependent ion transporters in bone metabolism
| Project/Area Number |
12671815
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional basic dentistry
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| Research Institution | Showa university |
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Principal Investigator |
AMANO Hitoshi Showa Univ. School of Dentistry assistant Prof., 歯学部, 講師 (90212571)
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| Co-Investigator(Kenkyū-buntansha) |
SAKAI Nobuhiro Showa Univ. School of Dentistry assistant, 歯学部, 助手 (90286849)
SUZUKI Keiko Showa Univ. School of Dentistry assistant Prof., 歯学部, 講師 (50119187)
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| Project Period (FY) |
2000 – 2002
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| Keywords | mechanical stress / osteoblast / RT-PCR / osteoclast / resorption pit / co-culture / Na^+ / Ca^<2+> exchanger / NPT2 / Pi輸送体 |
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| Research Abstract |
Na^+/Ca^<2+> exchanger (NCX) catalyzes the electrogenic exchange of 3 Na^+ for 1 Ca2^+ across the plasma membrane in many mammalian cells. We have previously reported that the Na^+/Ca^<2+> exchange activity had an important role for CSF-1-induced osteoclast differentiation. In the present study, we sought the effect of mechanical stress to confirm the role of NCX1, in osteoblastic bone formation and osteoclastic bone resorption formation.
Osteoblast were obtained from calvalia of new bone rat. Mechanical stress induced NCX1 mRNA and F-actin polyme ralization in osteoblast. The application of Gadlinium ion, a blocker of SAchannel, inhibited stretch-activated cell responces.
Bone marrow cells were obtained from tibiae and femuor of 5- to 8-week-old male ddy mice. We used Sephadex G-10 column (G10) in order to remove macrophages and stroma cells. Non adherent G10-passed cells were cultured in a-MEM containing the final concentration of 15 % FBS, 100 ng/ml RANKL, 20 ng/ml CSF-1. KB-R7943 inhibited the osteoclast differentiation and bone resorption in vitro dose-dependently. In addition, NCX1 antisense oligo deoxy nucleotide (ODN) reduced the number of tartrate-resistant acid phosphatase-positive multinucleated cells and decreased the amount of NCX1 in protein level. Eight week-old female ddY mice received sham-operation or ovar iectomy (OVX) and were fed a laboratory chow for 4 weeks after the operation. Animals were given subcutaneously 337.5 mg/kg KB-R 7943 or DMSO every other day for 4 weeks. After 4 weeks treatment with KB-R7943, the BMD in the femur of OVX group was significantly decreased compared with sham group. However the BMD in KB-R7943 treated OVX group was quantitatively similar to the sham group. These results suggest that bone resorption was also blocked by treatment with KB-R7943 in vivo.
In conclusion, the activation of NCX1 is essential for osteoclast differentiation and activation.
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