A steady supply of platelets maintains their levels in the blood, and this is achieved by the generation of progeny from platelet intermediates. Recent emerging evidence demonstrates that platelets are equipped with elaborate intracellular machinery, and their organization is far more complex than previously regarded. However, their ultrastructural organization in platelets in the different stages of maturation process remains unclear due to the difficulty in determining and quantifying the ultrastructure of platelets by diffraction-limited conventional light microscopy. To overcome this limit, we used stochastic optical reconstruction microscopy to resolve the native ultrastructure and their interaction of different organelles in platelets. The recent development of super-resolution fluorescence microscopy allows the location of molecules to be determined with nanometer-scale spatial resolution. Using systematic and integrative super resolution microscopy, we examined the ultrastructural organization of various organelles in different platelet intermediates to understand the mechanism of organelle redistribution and sorting in the platelet intermediate maturation as the early step of platelet progeny production. We observed the interconversion between the intermediates via various pathways and found that microtubules were responsible for controlling the shape of platelet intermediates. Platelet intermediates showed increase of their protein contents using their protein synthesis machinery to prepare their maturation and gradually distribute them equally into two-sided swellings of proplatelet with the help of cytoskeletal elements. Furthermore, super-resolution images showed that most of the organelles were located near the cell periphery in oval preplatelets and confined to the bulbous tips in proplatelets. We also found that the distribution of the dense tubular system and α-granules was regulated by actin, whereas that of mitochondria and dense granules was governed by microtubules. Altogether, our results call for a reassessment of organelle redistribution in platelet intermediates.

Super-resolution imaging of cytoskeleton-dependent organelle rearrangement in platelet maturation and its quantitative analysis

ACS Fall 2021

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