Activation induced morphological changes and integrin alpha IIb beta 3 activity of living platelets

作者：Posch Sandra; Neundlinger Isabel; Leitner Michael; Siostrzonek Peter; Panzer Simon; Hinterdorfer Peter; Ebner Andreas^*

来源：Methods, 2013, 60(2): 179-185.

DOI：10.1016/j.ymeth.2013.03.034

• Platelets
• Single molecule force spectroscopy
• Atomic force microscopy
• Integrin alpha IIb beta 3

Platelets are essential in hemostasis. Upon activation they undergo a shape-change accompanied with receptor presentation. Atomic force microscopy (AFM) imaging and single molecule force spectroscopy (SMFS) were used as powerful tools for exploring morphological changes as well as receptor activities of platelets. Imaging time series was accomplished with and without fixation steps at the single platelet level. Hereby the response of mechanical stimulation of the platelet by the AFM cantilever tip was directly observed. We demonstrate that living and fixed platelets develop filopodia after a short activation time followed by their disappearance including cellular bleb formation. Thereafter a second filopodia formation (filopodia extrusion) was observed; those filopodia subsequently disappeared again, and finally platelets detached from the support due to cell death. We determined the influence of mechanical stress on the chronology of morphological changes of platelets and demonstrated shear force induced filopodia formation. Through recordings over several hours, topographical AFM images over the full platelet lifetime - from early activation up to apoptosis - are presented. SMFS measurements on living platelets allowed determining the activation state of the most prominent membrane receptor integrin alpha Ilb beta 3 at all different phases of activation. alpha IIb beta 3 was fully activated, independent of the morphological state.

• 出版日期2013-4-1