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Quantitative Proteomics Using Formalin-fixed, Paraffin-embedded Biopsy Tissues in Inflammatory Disease.

Authors
  • Amarnani, Abhimanyu1, 2
  • Capri, Joseph R3
  • Souda, Puneet3
  • Elashoff, David A4
  • Lopez, Ivan A1
  • Whitelegge, Julian P3
  • Singh, Ram R2, 5, 6, 7
  • 1 Department of Head and Neck Surgery, UCLA, Los Angeles, CA 90095, USA.
  • 2 Department of Medicine/Rheumatology, UCLA, Los Angeles, CA 90095, USA.
  • 3 The Pasarow Mass Spectrometry Laboratory, Semel Institute for Neuroscience and Human Behavior, UCLA, Los Angeles, CA 90024, USA.
  • 4 Department of Medicine/Statistics Core, UCLA, Los Angeles, CA 90095, USA.
  • 5 Department of Pathology and Laboratory Medicine, UCLA, Los Angeles, CA 90095, USA.
  • 6 Molecular Toxicology Interdepartmental Program, UCLA, Los Angeles, CA 90095, USA.
  • 7 Jonsson Comprehensive Cancer Center, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095, USA.
Type
Published Article
Journal
Journal of Proteomics & Bioinformatics
Publisher
OMICS Publishing Group
Publication Date
Jan 01, 2019
Volume
12
Issue
7
Pages
104–112
Identifiers
DOI: 10.35248/0974-276X.12.19.503
PMID: 32431480
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

Investigations in human disease pathogenesis have been hampered due to paucity of access to fresh-frozen tissues (FFT) for use in global, data-driven methodologies. As an alternative, formalin-fixed, paraffin-embedded (FFPE) tissues are readily available in pathology banks. However, the use of formalin for fixation can lead to the loss of proteins that appear during inflammation, thus introducing an inherent sample bias. To address this, we compared FF and FFPE tissue proteomics to determine whether FFPE-tissue can be used effectively in inflammatory diseases. Adjacent kidney slices from lupus nephritic mice were processed as FFPE or FFTs. Their tissue lysates were run together using proteomics workflow involving filter-aided sample preparation, in-solution dimethyl isotope labeling, StageTip fractionation, and nano-LC MS/MS through an Orbitrap XL MS. We report a >97% concordance in protein identification between adjacent FFPE and FFTs in murine lupus nephritic kidneys. Specifically, proteins representing pathways, namely, 'systemic lupus erythematosus', 'interferon-α', 'TGF-β', and 'extracellular matrix', were reproducibly quantified between FFPE and FFTs. However, 12%-29% proteins were quantified differently in FFPE compared to FFTs, but the differences were consistent across experiments. In particular, certain proteins represented in pathways, including 'inflammatory response' and 'innate immune system' were quantified less in FFPE than in FFTs. In a pilot study of human FFPE tissues, we identified proteins relevant to pathogenesis in lupus nephritic kidney biopsies compared to control kidneys. This is the first report of lupus nephritis kidney proteomics using FFPE tissue. We concluded that archived FFPE tissues can be reliably used for proteomic analyses in inflammatory diseases, with a caveat that certain proteins related to immunity and inflammation may be quantified less in FFPE than in FFTs.

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