MALDI mass spectrometry based molecular phenotyping of CNS glial cells for prediction in mammalian brain tissue
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The development of powerful analytical techniques for specific molecular characterization of neural cell types is of central relevance in neuroscience research for elucidating cellular functions in the central nervous system (CNS). This study examines the use of differential protein expression profiling of mammalian neural cells using direct analysis by means of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). MALDI-MS analysis is rapid, sensitive, robust, and specific for large biomolecules in complex matrices. Here, we describe a newly developed and straightforward methodology for direct characterization of rodent CNS glial cells using MALDI-MS-based intact cell mass spectrometry (ICMS). This molecular phenotyping approach enables monitoring of cell growth stages, (stem) cell differentiation, as well as probing cellular responses towards different stimulations. Glial cells were separated into pure astroglial, microglial, and oligodendroglial cell cultures. The intact cell suspensions were then analyzed directly by MALDI-TOF-MS, resulting in characteristic mass spectra profiles that discriminated glial cell types using principal component analysis. Complementary proteomic experiments revealed the identity of these signature proteins that were predominantly expressed in the different glial cell types, including histone H4 for oligodendrocytes and S100-A10 for astrocytes. MALDI imaging MS was performed, and signature masses were employed as molecular tracers for prediction of oligodendroglial and astroglial localization in brain tissue. The different cell type specific protein distributions in tissue were validated using immunohistochemistry. ICMS of intact neuroglia is a simple and straightforward approach for characterization and discrimination of different cell types with molecular specificity.
KeywordsIntact cell mass spectrometry (ICMS) MALDI-TOF-MS Imaging mass spectrometry (IMS) Glial cells
The Swedish Research Council Grants 342-2004-3944 (JB), 621-2008-3562 (JB), 522-2006-6416 (MA), 521-2007-5407 (MA), and 2006-4268 (ÅFS); The Royal Swedish Academy of Sciences (MA, JH); Gyllenstiernska Krapperupstiftelsen (ÅFS); Åhlenstiftelsen, Hjärnfonden (GW, postdoctoral); and the Swedish Chemical Society (JH) are gratefully acknowledged for financial support.
- 10.Mittelbronn M, Dietz K, Schluesener HJ, Meyermann R (2001) Acta Neuropathol 101(3):249–255Google Scholar
- 20.Chung YC, Ko HW, Bok E, Park ES, Huh SH, Nam JH, Jin BK (2010) BMB Rep 43(4):225–232Google Scholar
- 32.Paxinos G, Watson C (2007) The rat brain in stereotaxic coordinates, 6th edn. Elsevier Academic, San DiegoGoogle Scholar