Ultrastructural, transcriptional, and functional differences between human reticulated and non-reticulated platelets

Abstract

Background: Reticulated platelets (RP) are the youngest circulating platelets in blood. An increased amount of this subpopulation is associated with higher cardiovascular risk and mortality. Objectives: It is unknown to what extent intrinsic properties of RP contribute to their hyperreactive features. This study is the first providing a multifactorial approach based on ultrastructural, transcriptional, and functional analysis of RP compared to non-RP sorted by flow cytometry. Methods: Reticulated platelets and non-RP were sorted after platelet staining with SYTO 13. Employing transmission electron microscopy, 1089 micrographs were analyzed for platelet size, amounts of intracellular structures, and anatomical surrogates indicating activation. Long and small RNA-sequencing (RNA-seq) were performed for analyzing differential gene expression. Functional analysis of P-selectin—an upregulated mRNA in RP—was performed in healthy subjects and patients on P2Y12-inhibitors. Results: Electron micrographs uncovered distinct ultrastructural differences in RP versus non-RP. Cross sections were 1.9-fold larger in RP (P