| Project/Area Number |
23K19731
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| Research Category |
Grant-in-Aid for Research Activity Start-up
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| Allocation Type | Multi-year Fund |
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| Review Section |
0907:Oral science and related fields
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| Research Institution | Matsumoto Dental University |
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Principal Investigator |
何 治鋒 松本歯科大学, 総合歯科医学研究所, 助教 (10980047)
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| Project Period (FY) |
2023-08-31 – 2025-03-31
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| Project Status |
Granted (Fiscal Year 2023)
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| Budget Amount *help |
¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000)
Fiscal Year 2024: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2023: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
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| Keywords | macrophage / bone repair / Wnt signal / Macrophage / Bone repair |
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| Outline of Research at the Start |
Wnt signaling has been reported to be involved in bone mass regulation. The applicant's laboratory has revealed that the Wnt signaling network is involved bone metabolism. This time I came up with an idea about connects the skeletal stem cells, Wnt signaling network and macrophages (Mφ) (which have been considered as the dominant immune cell taking part in bone healing) together to establish the new strategy for investigating the complex and fascinating molecule interactions in the bone regeneration process.
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| Outline of Annual Research Achievements |
We identified the surface macrophage marker "A". Utilizing flow cytometry and staining with the A antibody, we were able to segregate F4/80+ Csf1r- cells into 3 distinct subpopulations: A high, A intermediate, and A low. Through comparison of cell frequencies and their responsiveness to clodronate liposome, we determined that A high F4/80+ Csf1r- cells are essential in bone repair.
We also identified the secreted protein factor X. In our RNA-seq data, where we compared F4/80+ Csf1r+ and F4/80+ Csf1r- cells, factor X emerged as one of the top 20 genes. Previous studies have associated its expression with granulocytes and macrophages. We observed its upregulation following bone injury and subsequent downregulation upon macrophage depletion, which was concurrent with changes in Wnt signaling.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
Most of the objectives outlined in the grant application have been met successfully. Our research concepts are well-defined, and the experimental methodologies employed are strongly backed by resources available at our research institute.
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| Strategy for Future Research Activity |
After identifying factor "X", we will proceed with in vitro experiments. Our strategy involves introducing recombinant "X" into the culture medium of bone marrow stromal cells, with and without specific osteogenic agents. We will assess the impact of "X" on osteoblast differentiation through assays such as Alp or alizarin red staining. Additionally, we will evaluate the mRNA expression and protein levels of Wnts.
Although we initially planned to generate "A"-Cre "X" flox/flox conditional knockout (cKO) mice and observe their bone regeneration phenotype, we intend to start with Vav1-Cre instead. Finally, we will assess the effect of recombinant "X", with macrophage ablation through injection or Matrigel embedding at the bone injury site, to investigate its role in accelerating bone healing.
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