Platelet microRNA expression and association with platelet maturity and function in patients with essential thrombocythemia

Abstract

Essential thrombocythemia (ET) is characterized by persistently elevated platelet counts and an increased risk of thromboembolic events. Dysregulated expression of small noncoding microRNAs (miRNAs) have been shown in ET and may influence platelet maturity and function in ET patients. In this study, we included 22 ET patients and 19 healthy controls to investigate the expression of 12 platelet miRNAs previously reported to be dysregulated in ET. Further, we investigated the correlation between the expression of selected miRNAs and platelet maturity and platelet function. Total RNA was isolated from platelets, and expression analyses were performed using TaqMan quantitative PCR (qPCR). Mean platelet volume (MPV) and immature platelet count and -fraction (IPC and IPF) were measured using the Sysmex XE-5000 automated haematology system. Platelet function was investigated by multiple electrode aggregometry (agonists: arachidonic acid (AA), thrombin-receptor-activating-peptide (TRAP) and adenosine diphosphate (ADP)), while platelet activation was determined by multi-colour flow cytometry (antibodies: bound-fibrinogen, CD63 and P-selectin (CD62p), agonists: AA, TRAP and ADP). We showed that miR-9 and miR-490 were significantly upregulated in ET patients compared with healthy controls (p-values < 0.01), while miR-10a, miR-28, miR-126, miR-155, miR-221, miR-222, miR-223 and miR-431 were significantly downregulated in ET patients (all p-values < 0.001). A significant positive correlation was observed between miR-431 and MPV, IPC and IPF (all p-values < 0.05). The expression of miR-126 was negatively correlated with platelet aggregation induced by AA and TRAP (p < 0.05). In addition, we found the expression of miR-9 and miR-490 to be negatively correlated with the percentage of fibrinogen-, CD63- and P-selectin- positive platelets using TRAP as agonist (p < 0.05). In conclusion, our data indicate that platelet microRNAs may play a role in ET and that specific microRNAs are correlated with platelet maturity and platelet function.

Keywords:

• Blood platelets
• essential thrombocythemia
• microRNA
• platelet aggregation
• platelet maturity

Acknowledgements

Mai Stenulm Veirup and Vivi Bo Mogensen are acknowledged for expert technical assistance.

We thank Søren Bønløkke, Peter Buur van Kooten Niekerk and Jesper Stentoft from the Department of Haematology, Aarhus University Hospital, for help with identification of the ET patients.

Declaration of interest

None related to the present manuscript.

Statement of Contribution

PHN, AMH, OHP and JQDT conceived and designed the study. OHP and MLL performed clinical work-up of ET patients. JQDT, OHP and MLL performed data collection and data analysis. JQDT, OHP and PHN wrote the manuscript. PHN, MLL, ELG, SDK and AMH revised the manuscript. All authors accept responsibility for the entire content of this submitted manuscript and approved submission.

Additional information

Funding

We received financial support from Aarhus University, The Danish Medical Association Research Foundation, Aase and Ejnar Danielsens Foundation, Oda and Hans Svenningsens Foundation and The Danish Heart Association. The funding sources had no influence on the study.