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Thromb Haemost 2017; 117(03): 529-542
DOI: 10.1160/TH16-04-0322
Cellular Haemostasis and Platelets
Schattauer GmbH

Hyperglycaemia suppresses microRNA expression in platelets to increase P2RY12 and SELP levels in type 2 diabetes mellitus

Zsolt Fejes
1   Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
Szilárd Póliska
2   Department of Biochemistry and Molecular Biology, Genomic Medicine and Bioinformatics Core Facility, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
Zsolt Czimmerer
2   Department of Biochemistry and Molecular Biology, Genomic Medicine and Bioinformatics Core Facility, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
Miklós Káplár
3   Institute of Internal Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
András Penyige
4   Department of Human Genetics, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
5   Faculty of Pharmacy, University of Debrecen, Debrecen, Hungary
,
Gabriella Gál Szabó
1   Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
Ildikó Beke Debreceni
1   Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
Satya P. Kunapuli
6   Department of Physiology and Sol Sherry Thrombosis Center, Temple University School of Medicine, Philadelphia, Pennsylvania, USA
,
János Kappelmayer
1   Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
,
Béla Nagy Jr
1   Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
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Publikationsverlauf

Received: 23. April 2016

Accepted after major revision: 26. November 2016

Publikationsdatum:
22. November 2017 (online)

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Summary

Megakaryocyte (MK)-derived miRNAs have been detected in platelets. Here, we analysed the expression of platelet and circulating miR-223, miR-26b, miR-126 and miR-140 that might be altered with their target mRNAs in type 2 diabetes mellitus (DM2). MiRNAs were isolated from leukocyte-depleted platelets and plasma samples obtained from 28 obese DM2, 19 non-DM obese and 23 healthy individuals. The effect of hyperglycaemia on miRNAs was also evaluated in MKs using MEG-01 and K562 cells under hyperglycaemic conditions after 8 hours up to four weeks. Quantitation of mature miRNA, pre-miRNAs and target mRNA levels (P2RY12 and SELP) were measured by RT-qPCR. To prove the association of miR-26b and miR-140 with SELP (P-selectin) mRNA level, overexpression or inhibition of these miRNAs in MEG-01 MKs was performed using mimics or anti-miRNAs, respectively. The contribution of calpain substrate Dicer to modulation of miRNAs was studied by calpain inhibition. Platelet activation was evaluated via surface P-selectin by flow cytometry. Mature and pre-forms of investigated miRNAs were significantly reduced in DM2, and platelet P2RY12 and SELP mRNA levels were elevated by two-fold at increased platelet activation compared to controls. Significantly blunted miRNA expressions were observed by hyperglycaemia in MEG-01 and K562-MK cells versus baseline values, while the manipulation of miR-26b and miR-140 expression affected SELP mRNA level. Calpeptin pretreatment restored miRNA levels in hyperglycaemic MKs. Overall, miR-223, miR-26b, miR-126 and miR-140 are expressed at a lower level in platelets and MKs in DM2 causing upregulation of P2RY12 and SELP mRNAs that may contribute to adverse platelet function.

Supplementary Material to this article is available online at www.thrombosis-online.com.

Keywords

Platelet activation - microRNA - megakaryocyte - diabetes mellitus - hyperglycaemia

 

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