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Blood, 15 September 2006, Vol. 108, No. 6, pp. 1911-1918.
Prepublished online as a Blood First Edition Paper on May 23, 2006; DOI 10.1182/blood-2006-02-004267.


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HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY

Impaired "outside-in" integrin {alpha}IIbbeta3 signaling and thrombus stability in TSSC6-deficient mice

Matt W. Goschnick, Lai-Man Lau, Janet L. Wee, Yong S. Liu, P. Mark Hogarth, Lorraine M. Robb, Michael J. Hickey, Mark D. Wright, and Denise E. Jackson

From the Kronheimer Building, Burnet Institute incorporating the Austin Research Institute, Heidelberg, Victoria, Australia; The Walter and Eliza Hall Institute, Parkville, Victoria, Australia; and the Centre for Inflammatory Diseases, Department of Medicine, Monash University, Clayton, Victoria, Australia.

We investigated the role of the hematopoietic-specific tetraspanin superfamily member, TSSC6, in platelet function using wild-type mice and TSSC6-deficient mice. TSSC6 is expressed on the surface of murine platelets and is up-regulated by thrombin stimulation, indicating an intracellular pool of TSSC6. Immunoprecipitation/Western blot studies reveal a constitutive physical association of TSSC6 with the integrin {alpha}IIbbeta3 complex under strong detergent conditions. In vivo evaluation of hemostasis by tail bleeding revealed increased bleeding time, volume of blood lost, and evidence of tail rebleeds in TSSC6 null mice, indicating unstable hemostasis. Using ex vivo techniques, we showed that TSSC6-deficient platelets exhibited impaired kinetics of clot retraction, platelet aggregation at lower doses of PAR-4, and collagen and platelet spreading on fibrinogen in the presence of normal integrin {alpha}IIbbeta3 expression. TSSC6-deficient platelets showed normal alpha granule secretion, normal "insideout" integrin {alpha}IIbbeta3 signaling (fluorescein isothiocyanate [FITC]–fibrinogen and JON/A binding), and normal platelet adhesion on fibrinogen. Furthermore, we show that absence of platelet TSSC6 affects the secondary stability of arterial thrombi in vivo upon vascular injury. These data demonstrate that TSSC6 appears to regulate integrin {alpha}IIbbeta3 "outside-in" signaling events in platelets and is necessary for stability of arterial thrombi in vivo.


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