Quantitative analysis of chondroitin sulfate disaccharides from human and rodent fixed brain tissue by electrospray ionization-tandem mass spectrometry

Kimberly M Alonge; Aric F Logsdon; Taylor A Murphree; William A Banks; C Dirk Keene; J Scott Edgar; Dale Whittington; Michael W Schwartz; Miklos Guttman

doi:10.1093/glycob/cwz060

Quantitative analysis of chondroitin sulfate disaccharides from human and rodent fixed brain tissue by electrospray ionization-tandem mass spectrometry

Glycobiology. 2019 Nov 20;29(12):847-860. doi: 10.1093/glycob/cwz060.

Authors

Kimberly M Alonge¹, Aric F Logsdon^{2

3}, Taylor A Murphree⁴, William A Banks^{2

3}, C Dirk Keene⁵, J Scott Edgar⁴, Dale Whittington⁴, Michael W Schwartz¹, Miklos Guttman⁴

Affiliations

¹ University of Washington Medicine Diabetes Institute, Department of Medicine, Seattle, WA, USA.
² Department of Geriatric Research Education and Clinical Center (GRECC), Veterans Affairs Puget Sound Health Care System, Seattle, WA, USA.
³ Division of Gerontology and Geriatric Medicine, Department of Medicine, University of Washington, Seattle, WA, USA.
⁴ Department of Medicinal Chemistry, University of Washington, Seattle, WA, USA.
⁵ Division of Neuropathology, Department of Pathology, University of Washington, Seattle, WA, USA.

Abstract

Chondroitin sulfates (CS) are long, negatively charged, unbranched glycosaminoglycan (GAG) chains attached to CS-proteoglycan (CSPG) core proteins that comprise the glycan component in both loose interstitial extracellular matrices (ECMs) and in rigid, structured perineuronal net (PNN) scaffolds within the brain. As aberrant CS-PNN formations have been linked to a range of pathological states, including Alzheimer's disease (AD) and schizophrenia, the analysis of CS-GAGs in brain tissue at the disaccharide level has great potential to enhance disease diagnosis and prognosis. Two mass-spectrometry (MS)-based approaches were adapted to detect CS disaccharides from minute fixed tissue samples with low picomolar sensitivity and high reproducibility. The first approach employed a straightforward, quantitative direct infusion (DI)-tandem mass spectrometry (MS/MS) technique to determine the percentages of Δ4S- and Δ6S-CS disaccharides within the 4S/6S-CS ratio, while the second used a comprehensive liquid chromatography (LC)-MS/MS technique to determine the relative percentages of Δ0S-, Δ4S-, Δ6S-, Δ4S6S-CS and Δ2S6S-CS disaccharides, with internal validation by full chondroitin lyase activity. The quantitative accuracy of the five primary biologically relevant CS disaccharides was validated using a developmental time course series in fixed rodent brain tissue. We then analyzed the CS disaccharide composition in formalin-fixed human brain tissue, thus providing the first quantitative report of CS sulfation patterns in the human brain. The ability to comprehensively analyze the CS disaccharide composition from fixed brain tissue provides a means with which to identify alterations in the CS-GAG composition in relation to the onset and/or progression of neurological diseases.

Keywords: chondroitin sulfate; extracellular matrix; glycosaminoglycan; mass spectrometry; perineuronal nets.

Publication types

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

MeSH terms

Animals
Brain Chemistry*
Chondroitin Sulfates / analysis*
Female
Humans
Male
Mice
Mice, Inbred C57BL
Spectrometry, Mass, Electrospray Ionization
Tandem Mass Spectrometry

Substances

chondroitin sulfate glycosaminoglycan
Chondroitin Sulfates

Abstract

Publication types

MeSH terms

Substances

Grants and funding