Abstract

Background: Upon wound formation, platelets adhere to the neighboring extracellular matrix and spread on it, a process which is critical for physiological wound healing. Multiple external factors, such as the molecular composition of the environment and its mechanical properties, play a key role in this process and direct its speed and outcome. Methods: We combined live cell imaging, quantitative interference reflection microscopy and cryo-electron tomography to characterize, at a single platelet level, the differential spatiotemporal dynamics of the adhesion process to fibrinogen- and collagen IV-functionalized surfaces. Results: Initially, platelets sense both substrates by transient rapid extensions of filopodia. On collagen IV, a short-term phase of filopodial extension is followed by lamellipodia-based spreading. This transition is preceded by the extension of a single or couple of microtubules into the platelet's periphery and their apparent insertion into the core of the filopodia. On fibrinogen surfaces, the filopodia-to-lamellipodia transition was partial and microtubule extension was not observed leading to limited spreading, which could be restored by manganese or thrombin. Conclusions: Based on these results, we propose that interaction with collagen IV stimulate platelets to extend microtubules to peripheral filopodia, which in turn, enhances filopodial-to-lamellipodial transition and overall lamellipodia-based spreading. Fibrinogen, on the other hand, fails to induce these early microtubule extensions, leading to full lamellipodia spreading in only a fraction of the seeded platelets. We further suggest that activation of integrin αIIbβ3is essential for filopodial-to-lamellipodial transition, based on the capacity of integrin activators to enhance lamellipodia spreading on fibrinogen.

Highlights

  • Upon wound formation, platelets adhere to the neighboring extracellular matrix and spread on it, a process which is critical for physiological wound healing

  • The filopodia-to-lamellipodia transition was partial and microtubule extension was not observed leading to limited spreading, which could be restored by manganese or thrombin

  • We further suggest that activation of integrin αIIbβ3 is essential for filopodial-to-lamellipodial transition, based on the capacity of integrin activators to enhance lamellipodia spreading on fibrinogen

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Summary

Introduction

Upon wound formation, platelets adhere to the neighboring extracellular matrix and spread on it, a process which is critical for physiological wound healing. Methods: We combined live cell imaging, quantitative interference reflection microscopy and cryo-electron tomography to characterize, at a single platelet level, the differential spatiotemporal dynamics of the adhesion process to fibrinogen- and collagen IVfunctionalized surfaces. On collagen IV, a short-term phase of filopodial extension is followed by lamellipodia-based spreading This transition is preceded by the extension of a single or couple of microtubules into the platelet’s periphery and their apparent insertion into the core of the filopodia. We further suggest that activation of integrin αIIbβ is essential for filopodial-to-lamellipodial transition, based on the capacity of integrin activators to enhance lamellipodia spreading on fibrinogen

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