Quantitative proteomic analysis of hepatocyte-secreted extracellular vesicles reveals candidate markers for liver toxicity

J Proteomics. 2014 May 30:103:227-40. doi: 10.1016/j.jprot.2014.04.008. Epub 2014 Apr 18.


Extracellular vesicles have created great interest as possible source of biomarkers for different biological processes and diseases. Although the biological function of these vesicles is not fully understood, it is clear that they participate in the removal of unnecessary cellular material and act as carriers of various macromolecules and signals between the cells. In this report, we analyzed the proteome of extracellular vesicles secreted by primary hepatocytes. We used one- and two-dimensional liquid chromatography combined with data-independent mass spectrometry. Employing label-free quantitative proteomics, we detected significant changes in vesicle protein expression levels in this in vitro model after exposure to well-known liver toxins (galactosamine and Escherichia coli-derived lipopolysaccharide). The results allowed us to identify candidate markers for liver injury. We validated a number of these markers in vivo, providing the basis for the development of novel methods to evaluate drug toxicity. This report strongly supports the application of proteomics in the study of extracellular vesicles released by well-controlled in vitro cellular systems. Analysis of such systems should help to identify specific markers for various biological processes and pathological conditions.

Biological significance: Identification of low invasive candidate marker for hepatotoxicity. Support to apply proteomics in the study of extracellular vesicles released by well-controlled in vitro cellular systems to identify low invasive markers for diseases.

Keywords: Exosomes; Hepatocytes; LC–MS; Label-free quantitation; Liver; Microvesicles.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomarkers / analysis*
  • Chemical and Drug Induced Liver Injury / metabolism*
  • Exosomes / chemistry
  • Galactosamine / toxicity
  • Hepatocytes / metabolism*
  • Lipopolysaccharides / toxicity
  • Liver / chemistry
  • Liver / drug effects
  • Male
  • Mass Spectrometry
  • Proteomics / methods
  • Rats, Sprague-Dawley
  • Secretory Vesicles / chemistry*


  • Biomarkers
  • Lipopolysaccharides
  • Galactosamine