miRNA expression profile in platelets after ex vivo activation. miRNA expression profile in platelets after ex vivo activation

We performed microRNA sequencing (miRNA-seq) according to Illumina sequencing protocols. Platelet-rich plasma (PRP) has been obtained by centrifuging whole blood at 160 x g for 15 min at room temperature. Platelets have been activated with 3 different physiologic agonists, namely ADP (20uM), collagen (60ug/mL), or Thrombin Receptor Activating Peptide (TRAP 25uM) under continuous stirring. An aliquot of platelets has been obtained at 120 minutes following ADP, collagen and TRAP and then processed to determine miRNA expression profiles. Time 0, indicating samples before any agonist treatment, was used as baseline. Total RNA was extracted by using mirVana Paris kit (life technologies) and then was subjected to size selection library preparation and retrotranscribed. Obtained cDNA was sequenced with the Illumina HiSeq 1000 sequencer .