| Co-Investigator(Kenkyū-buntansha) |
HONDA Shigenori Osaka University Medical School, Assistant Professor, 医学部, 助手 (00303959)
KANAKURA Yuzuru Osaka University Medical School, Professor, 医学部, 教授 (20177489)
KURATA Yoshiyuki Osaka University Hospital, Instructor, 医学部・附属病院, 講師 (80127224)
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| Research Abstract |
In this research project, we have investigated both inside-out and outside-in signaling through the integrin alphaIIbbeta3.
Concerning alphaIIbbeta3 activation via inside-out signaling, we have further investigated our hypothesis that Na^+/Ca^<2+> exchanger (NCX) may be involved as a common inside-out signaling.We cloned platelet NCX cDNA and showed that platelet NCX is BD type, while cardiac NCX is BCDEF.Next, employing two unrelated NCX inhibitors, DCB and bepridil, we investigated the role of NCX in platelet activation in detail.Both inhibitors abolished platelet alphaIIbbeta3 activation induced by all agonists examined.However, these inhibitors did not inhibit protein phosphorylation or increase in intracellular calcium concentration.These findings further support our hypothesis that NCX is involves in the common steps of inside-out signaling through alphaIIbbeta3.
We characterized a Japanese variant of Glanzmann thrombasthenia whose platelets express normal amounts of alphaIIbbeta3 and newly showed that a two-amino acid insertion in the Cysl46-Cysl67 loop of the alphaIIb subunit is responsible for this phenotype.In addition, Asp163 is a critical residue for ligand binding function.Concerning outside-in signaling, we showed alphaIIbbeta3 antagonists can be divided into two groups.In group I, antagonists can induce LIBS (ligand-induced binding site) on both alphaIIb and beta3, while in group II, antagonists can induce LIBS on alphaIIb, but not on beta3.Interestingly, only group I antagonists can initiate alphaIIbbeta3-mediated intracellular Ca^<2+> signaling.Our findings strongly sugget the association between beta3 LIBS expression and alphaIIbbeta3-mediated intracellular Ca^<2+> signaling.
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