Effect of two-dimensional spatial confinement on platelet mechanics.

Platelets are small, anucleate cells critical for hemostasis and thrombosis. Within platelet plugs and narrow capillaries, they often encounter spatially confining microenvironments. To examine how confinement influences platelet properties, we employed microcontact printing to generate fibrinogen micropatterns of varying sizes and shapes. Platelets cultured on these micropatterns adapted both their morphology and mechanical characteristics. Scanning ion conductance microscopy revealed changes in area, aspect ratio, and height, while fluorescence microscopy showed F-actin redistribution toward the periphery. Confinement reduced platelet stiffness in a size-dependent but shape-independent manner. To explore the role of intracellular signaling, we examined cyclic guanosine monophosphate (cGMP), a key inhibitor of platelet activation, adhesion, and aggregation. Treatment with a cGMP analog preserved F-actin redistribution but prevented stiffness changes in response to confinement, indicating that cGMP inhibits stiffness modulation without affecting cytoskeletal reorganization.