Proteomic evaluation of biological nanoparticles isolated from human kidney stones and calcified arteries

Farooq A. Shiekh, Jon E. Charlesworth, Sung Hoon Kim, Larry W. Hunter, Muthuvel Jayachandran, Virginia M. Miller, John C. Lieske

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

Calcifying biological nanoparticles (NPs) develop under cell culture conditions from homogenates of diverse tissue samples displaying extraosseous mineralization, including kidney stones and calcified aneurysms. Probes to definitively identify NPs in biological systems are lacking. Therefore, the aim of this study was to begin to establish a proteomic biosignature of NPs in order to facilitate more definitive investigation of their contribution to disease. Biological NPs derived from human kidney stones and calcified aneurysms were completely decalcified by overnight treatment with ethylenediaminetetraacetic acid or brief incubation in HCl, as evidenced by lack of a calcium shell and of Alizarin Red S staining, by transmission electron microscopy and confocal microscopy, respectively. Decalcified NPs contained numerous proteins, including some from bovine serum and others of prokaryotic origin. Most prominent of the latter group was EF-Tu, which appeared to be identical to EF-Tu from Staphylococcus epidermidis. A monoclonal antibody against human EF-Tu recognized a protein in Western blots of total NP lysate, as well as in intact NPs by immunofluorescence and immunogold EM. Approximately 8% of NPs were quantitatively recognized by the antibody using flow cytometry. Therefore, we have defined methods to reproducibly decalcify biological NPs, and identified key components of their proteome. These elements, including EF-Tu, can be used as biomarkers to further define the processes that mediate propagation of biological NPs and their contribution to disease.

Original languageEnglish (US)
Pages (from-to)4065-4072
Number of pages8
JournalActa Biomaterialia
Volume6
Issue number10
DOIs
StatePublished - Oct 1 2010

Keywords

  • Calcification
  • Elongation factor Tu
  • Nanoparticles
  • Nephrolithiasis
  • Proteomics

ASJC Scopus subject areas

  • Biotechnology
  • Biomaterials
  • Biochemistry
  • Biomedical Engineering
  • Molecular Biology

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