| Project/Area Number |
16590229
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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 |
General medical chemistry
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| Research Institution | Kobe University |
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Principal Investigator |
TOHYAMA Yumi Kobe University, Graduate school of Medicine, Associate Professor, 大学院・医学系研究科, 助教授 (70362770)
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| Co-Investigator(Kenkyū-buntansha) |
YAMAMURA Hirohei Kobe University, Graduate school of Medicine, Professor, 大学院・医学系研究科, 教授 (90030882)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2005: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2004: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Keywords | phagocytosis / tyrosine kinase / Syk / complement / cytoskeleton / RhoA / infection / innate immunity / 食作用 / C3bi |
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| Research Abstract |
Phagocytosis is a central event in the innate immune responses that are triggered by the association between ligands on the surface of pathogens and receptors on the membrane of phagocytes. A protein-tyrosine kinase, Syk plays a central role in Fcγ-mediated phagocytosis in the adaptive immune system. Here we show that Syk is also essential for the process of phagocytosis via complement activation. Phagocytosis assay was performed using macrophage-like differentiated HL60 cells as phagocytes and C3bi-opsonized zymosan as a pathogen. Syk was promptly activated and accumulation of Syk around the nascent phagosomes was detected. Dominant-negative Syk(DN-Syk) or Syk-siRNA was transferred to HL60 cells. Quantitative analysis using flow cytometry showed that transfer of either DN-Syk or Syk-siRNA resulted in the inhibition of phagocytosis. Quenching assay of fluorescence-labeled-zymosan revealed that Syk is required for engulfment of C3bi-bound zymosan but not for attachment of C3bi-zymosan to complement receptor 3 (CR3).
To clarify the molecular mechanism how Syk affects the process of phagosome formation and its engulfment, the change of actin dynamics and the activation of RhoA were investigated. At the early stage of phagocytosis, there was a marked accumulation of actin around phagosomes, which was followed by rapid dissociation. On the other hand, transfer of DN-Syk or Syk-siRNA led to attenuated accumulation of actin around the nascent phagosomes. In addition, RhoA was clearly activated in HL60 cells but the activation was decreased in both mutant cells. These results indicate that Syk plays an essential role in complement-mediated phagocytosis in the process of phagosome engulfment in related to actin dynamics.
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