Transcriptome Analysis of Reticulated Platelets Reveals a Prothrombotic Profile

Abstract

Reticulated platelets (RPs) are larger, hyperreactive platelets that contain significantly more ribonucleic acid (RNA) compared with mature platelets (MPs). High levels of RPs in peripheral blood are predictors of an insufficient response to dual antiplatelet therapy in cardiovascular patients and of adverse cardiovascular events. However, the mechanisms underlying these correlations remain widely unknown and the biology of RPs has not been investigated yet. Here, we compared for the first time the transcriptomic profiles of RPs and MPs isolated from peripheral blood of healthy donors. Total RNA sequencing revealed 1,744 differentially expressed genes (670 downregulated, 1,074 upregulated) in RPs compared with MPs. In particular, transcripts for the collagen receptor GP6, thromboxane receptor A2 (TBXA2R), thrombin receptor PAR4 (F2RL3), and adenosine triphosphate receptors P2RX1, ORAI2, and STIM1 (both involved in calcium signaling) were significantly upregulated in RPs, whereas several RNA regulators as the ribonuclease PARN, the RISC-component TNRC6A, and the splicing factor LUC7L3 were downregulated in RPs. Gene ontology analysis revealed an enrichment of relevant biological categories in RPs including platelet activation and blood coagulation. Gene Set Enrichment Analysis showed an overrepresentation of several platelet activation pathways like thrombin, thromboxane, and glycoprotein IIb/IIIa signaling in RPs. Small-RNA sequencing reported 9 micro-RNAs significantly downregulated in RPs with targets involved in platelet reactivity. Our data show for the first time an enrichment of several prothrombotic transcripts in RPs providing a first biological explanation for their hyperreactive phenotype.

SARS-CoV-2 infection is associated with a pro-thrombotic platelet phenotype

“…Peripheral venous blood was collected in citrate tubes and immediately processed to produce platelet-rich plasma (PRP) as described before 25,26 . CyTOF staining assay was performed according to the manufacturer's protocols.…”

The multifaceted contribution of platelets in the emergence and aftermath of acute cardiovascular events

“…Another mechanism for increased platelet reactivity is an increased relative number of so-called reticulated platelets, young platelets that contain more αand dense granules and mitochondria and have an increased reactivity compared with older platelets [75,76]. A recent study in mice indicated that high glucose levels led to increased platelet production by the bone marrow megakaryocytes, with the circulating platelets having a high fraction of reticulated platelets [77].…”

Large and small platelets—(When) do they differ?

“…87 In humans, the analysis of reticulated platelets revealed overlapping findings with large platelets. Both subpopulations contain higher amounts of RNA including transcripts for prohemostatic proteins 63,88 and show more P-selectin expression and PS-exposure compared to mature ones after activation. 15,89 Reddy et al also found reticulated human platelets being larger and more prone to expose PS, 90 but a similar size and PS exposure in young and old rabbit platelets distinguished by a biotinylation approach, solidifying differences between species.…”