LPS-induced lipid alterations in microglia revealed by MALDI mass spectrometry-based cell fingerprinting in neuroinflammation studies

Abstract

Pathological microglia activation can promote neuroinflammation in many neurodegenerative diseases, and it has therefore emerged as a potential therapeutic target. Increasing evidence suggests alterations in lipid metabolism as modulators and indicators in microglia activation and its effector functions. Yet, how lipid dynamics in activated microglia is affected by inflammatory stimuli demands additional investigation to allow development of more effective therapies. Here, we report an extensive matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) whole cell fingerprinting workflow to investigate inflammation-associated lipid patterns in SIM-A9 microglial cells. By combining a platform of three synergistic MALDI MS technologies we could detect substantial differences in lipid profiles of lipopolysaccharide (LPS)- stimulated and unstimulated microglia-like cells leading to the identification of 21 potential inflammation-associated lipid markers. LPS-induced lipids in SIM-A9 microglial cells include phosphatidylcholines, lysophosphatidylcholines (LysoPC), sphingolipids, diacylglycerols and triacylglycerols. Moreover, MALDI MS-based cell lipid fingerprinting of LPS-stimulated SIM-A9 microglial cells pre-treated with the non-selective histone deacetylase inhibitor suberoylanilide hydroxamic acid revealed specific modulation of LPS-induced-glycerolipids and LysoPC(18:0) with a significant reduction of microglial inflammation response. Our study introduces MALDI MS as a complementary technology for fast and label-free investigation of stimulus-dependent changes in lipid patterns and their modulation by pharmaceutical agents.

Downregulation of SF3B2 protects CNS neurons in models of multiple sclerosis

“…[16][17][18] SIM-A9 microglia cell line was stimulated with LPS to mimic activated microglia. 19,20 First, we confirmed the neurotoxicity of MCM on Neuro-2A cells (N2A) as demonstrated by increase in cleaved caspase-3 and reduction in ATP levels (Fig. S4).…”

“…When microglia are exposed to LPS, they release pro‐inflammatory cytokines, such as interferon‐ γ (IFN‐γ), tumor necrosis factor‐α (TNF‐α), and interleukin‐6 (IL‐6), which are important mediators of MS pathogenesis 16–18 . SIM‐A9 microglia cell line was stimulated with LPS to mimic activated microglia 19,20 . First, we confirmed the neurotoxicity of MCM on Neuro‐2A cells (N2A) as demonstrated by increase in cleaved caspase‐3 and reduction in ATP levels (Fig.…”

Lipid signatures and inter-cellular heterogeneity of naïve and lipopolysaccharide-stimulated human microglia-like cells

“…[19][20][21] Mass spectrometry imaging (MSI) further enables the spatial discrimination of phenotypes both in tissue and in vitro, based on lipidomic signatures. 19,22,23 Although lipidomic analyses of microglia cell lines have already been performed, 24,25 the approach or spatial resolution in previous studies did not allow for interrogation of single-cell heterogeneity.…”

Lipid Signatures and Inter-Cellular Heterogeneity of Naı̈ve and Lipopolysaccharide-Stimulated Human Microglia-like Cells

“…29 These MSI methods enable the spatial discrimination of cellular phenotypes and heterogeneity on a sub-cellular level, both in tissue and in vitro, based on lipid signatures. 17,30,31 Although lipidomic analyses of microglia cell lines have already been performed, 32,33 the approach or lateral resolution in previous studies did not allow for interrogation of single-cell heterogeneity.…”

“…In another approach, lateral resolution and sensitivity were improved to achieve sub-cellular lateral resolution on the low micrometer scale using MALDI MSI, applying specialized laser optics or boosting ion signal using post-ionization techniques. − For visualizing these cellular features, optimized sample preparation is key to preserve cellular morphology and bioanalytical composition as demonstrated by Bien and co-workers . These MSI methods enable the spatial discrimination of cellular phenotypes and heterogeneity on a sub-cellular level, both in tissue and in vitro, based on lipid signatures. ,, Although lipidomic analyses of microglia cell lines have already been performed, , the approach or lateral resolution in previous studies did not allow for interrogation of single-cell heterogeneity.…”