| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2005: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2004: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Research Abstract |
Class IA phosphoinositide 3-kinase (PI3K) is heavily involved in platelet function by regulating cytosolic protein kinases. PI3K inhibitors like wortmannin have been shown to exhibit anti-platelet function in vitro, but have inherent drawback in specificity. In order to elucidate the significance of the PI3K pathway as a target for anti-platelet therapy, platelet function from gene knockout mice lacking components of the PI3K pathway (PI3K p85α subunit and Bruton's tyrosine kinase : BTK) was studied. Collagen (or collagen related peptide : CRP)-induced platelet function (triggered by collagen receptor GPVI crosslinking) was readily impaired in either p85α or BTK-deficient (-/-) mice. Although BTK activation downstream of B cell receptor engagement is controlled predominantly by PI3K, this is not the case with platelets ; Functional phenotype was much severer in BTK-/- than p85α-/- platelets. Moreover, unlike B cells, as compared with Btk-/- platelets, only subtle functional defect was observed in Xid platelets in which PI3K-dependent Btk activation is selectively lacking due to a point mutation of the gene encoding pleckstrin homology domain of the kinase. These results suggested the existence of PI3K-independent pathway for BTK activation. To further study the notion above, double deficient (p85α-/- BTK-/-) mice were examined. Indeed, CRP-induced cellular responses (aggregation, P-selectin expression, fibrinogen binding, adhesion and phospholipase Cγ2 phosphorylation) were most severely defective in double knockout platelets. Furthermore, BTk was brought down with the LAT/Gads/SLP76 adaptor complex in a PI3K-independent manner. These, results clearly demonstrated there were two pathways toward BTK activation ; one is PI3K-dependent and another independent in collagen-induced platelet function. Thus, BTK is a promising target for anti-platelet therapy.
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