Proteomic profiling of the thrombin-activated canine platelet secretome (CAPS)

PLoS One. 2019 Nov 13;14(11):e0224891. doi: 10.1371/journal.pone.0224891. eCollection 2019.

Abstract

Domestic dogs share the same environment as humans, and they represent a valuable animal model to study naturally-occurring human disease. Platelet proteomics holds promise for the discovery of biomarkers that capture the contribution of platelets to the pathophysiology of many disease states, however, canine platelet proteomic studies are lacking. Our study objectives were to establish a protocol for proteomic identification and quantification of the thrombin-activated canine platelet secretome (CAPS), and to compare the CAPS proteins to human and murine platelet proteomic data. Washed platelets were isolated from healthy dogs, and stimulated with saline (control) or gamma-thrombin (releasate). Proteins were separated by SDS-page, trypsin-digested and analyzed by liquid chromatography and tandem mass spectrometry (MS). CAPS proteins were defined as those with a MS1-abundance ratio of two or more for releasate vs. unstimulated saline control. A total of 1,918 proteins were identified, with 908 proteins common to all dogs and 693 characterized as CAPS proteins. CAPS proteins were similar to human and murine platelet secretomes and were highly represented in hemostatic pathways. Differences unique to CAPS included replacement of platelet factor 4 with other cleavage products of platelet basic protein (e.g. interleukin-8), novel proteins (e.g. C-C motif chemokine 14), and proteins in relatively high (e.g. protease nexin-1) or low (e.g. von Willebrand factor) abundance. This study establishes the first in-depth platelet releasate proteome from healthy dogs with a reference database of 693 CAPS proteins. Similarities between CAPS and the human secretome confirm the utility of dogs as translational models of human disease, but we also identify differences unique to canine platelets. Our findings provide a resource for further investigations into disease-related CAPS profiles, and for comparative pathway analyses of platelet activation among species.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomarkers
  • Blood Platelets / metabolism*
  • Chromatography, Liquid
  • Computational Biology / methods
  • Dogs
  • Female
  • Humans
  • Mice
  • Molecular Sequence Annotation
  • Platelet Activation*
  • Proteome*
  • Proteomics* / methods
  • Tandem Mass Spectrometry
  • Thrombin / metabolism*

Substances

  • Biomarkers
  • Proteome
  • Thrombin

Grant support

This work was supported by the Independent Research Fund Denmark through an individual postdoc grant to SEC (ID: DFF-6111-00123) and an individual Sapare Aude grant to SEC (ID: DFF-6111-00123B). https://dff.dk/en. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.