Whole-cell MALDI-TOF Mass Spectrometry is an Accurate and Rapid Method to Analyze Different Modes of Macrophage Activation

Ouedraogo, Richard; Daumas, Aurélie; Capo, Christian; Mège, Jean‐Louis; Textoris, Julien

doi:10.3791/50926

Cited by 8 publications

(7 citation statements)

References 26 publications

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“…Mixed volume and dried droplet method have been shown to improve the MS signal intensity and the peak reproducibility. As an example, the mixed volume was more effective in the analysis of intact Fusarium spores while the dried droplet was shown to be more effective in the analysis of macrophages . During the mixed volume method, mixing T. cruzi parasite cells with a matrix solution in an Eppendorff tube generated a precipitate which was analyzed separately from the supernatant (Supplementary Figure S2a and S2b).…”

Section: Resultsmentioning

confidence: 99%

Direct identification of trypanosomatids by matrix-assisted laser desorption ionization-time of flight mass spectrometry (DIT MALDI-TOF MS)

2016

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Accurate and rapid determination of trypanosomatids is essential in epidemiological surveillance and therapeutic studies. Matrix-assisted laser desorption ionization/time of flight mass spectrometry (MALDI-TOF MS) has been shown to be a useful and powerful technique to identify bacteria, fungi, metazoa and human intact cells with applications in clinical settings. Here, we developed and optimized a MALDI-TOF MS method to profile trypanosomatids. trypanosomatid cells were deposited on a MALDI target plate followed by addition of matrix solution. The plate was then subjected to MALDI-TOF MS measurement to create reference mass spectra library and unknown samples were identified by pattern matching using the BioTyper software tool. Several m/z peaks reproducibly and uniquely identified trypanosomatids species showing the potentials of direct identification of trypanosomatids by MALDI-TOF MS. Moreover, this method discriminated different life stages of Trypanosoma cruzi, epimastigote and bloodstream trypomastigote and Trypanosoma brucei, procyclic and bloodstream. T. cruzi Discrete Typing Units (DTUs) were also discriminated in three clades. However, it was not possible to achieve enough resolution and software-assisted identification at the strain level. Overall, this study shows the importance of MALDI-TOF MS for the direct identification of trypanosomatids and opens new avenues for mass spectrometry-based detection of parasites in biofluids. Copyright © 2016 John Wiley & Sons, Ltd.

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Section: Resultsmentioning

confidence: 99%

Direct identification of trypanosomatids by matrix-assisted laser desorption ionization-time of flight mass spectrometry (DIT MALDI-TOF MS)

2016

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“…The monocyte isolation procedure was the same as in a previous method. 39 In brief, blood samples were diluted in saline at a 1:10 ratio, and then the mixture was further incubated with an OptiPrep gradient medium (07820; StemCell Technologies, Cambridge, MA, USA) containing Ficoll. After centrifuging at 700 × g for 20 min, the peripheral blood mononuclear cells (PBMCs) were collected.…”

Section: Methodsmentioning

confidence: 99%

The Monocyte-Derived Exosomal CLMAT3 Activates the CtBP2-p300-NF-κB Transcriptional Complex to Induce Proinflammatory Cytokines in ALI

Chen

Dong

Qiu

et al. 2020

Molecular Therapy - Nucleic Acids

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Monocytes and macrophages are the two major cell types involved in innate immunity. Exosomes act as signaling molecules to regulate cell-to-cell communication by releasing proteins, mRNAs, microRNAs (miRNAs), and long noncoding RNAs (lncRNAs). However, it is still unclear whether monocyte-derived exosomes are involved in the communication between monocytes and macrophages. In this study, we analyzed the differentially expressed lncRNA profiles in monocytes isolated from blood samples of healthy controls and acute lung injury (ALI) patients. We focused our study on investigating the signaling downstream of CLMAT3 (colorectal liver metastasis-associated transcript 3), a lncRNA that regulated proinflammatory cytokine genes. We revealed that CLMAT3 specifically targeted CtBP2 (C-terminal binding protein 2) and repressed its expression. Elevated CtBP2 acted as a coactivator to assemble a transcriptional complex with histone acetyltransferase p300 and NF-κB (nuclear factor κB) subunits. In vitro coculture and in vivo injection of ALI monocyte-derived exosomes increased the production of proinflammatory cytokines. Importantly, the administration of two CtBP2 inhibitors, NSC95397 and MTOB, could significantly reverse CtBP2-mediated transactivation. Collectively, our results support a model in which monocyte-derived exosomal CLMAT3 activates the CtBP2-p300-NF-κB complex to induce proinflammatory cytokines, thus contributing to the pathogenesis of ALI.

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“…After thawing of the PBMCs, 1 μL of cell suspension was added to 1 μL of matrix solution (saturated solution of α-cyano-4-hydroxy-cynnamic acid in a mixture of 50% acetonitrile, 25% trifluoroacetic acid and water) as previously described [ 17 , 20 , 23 ]. The mixture was deposited on the MALDI target.…”

Section: Methodsmentioning

confidence: 99%

MALDI-TOF MS monitoring of PBMC activation status in sepsis

et al. 2018

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BackgroundMALDI-TOF mass spectrometry (MS) on whole cells enables the detection of different cell types and cell activation. Here, we wondered whether this approach would be useful to investigate the host response in sepsis.MethodsPeripheral blood mononuclear cells (PBMCs) from patients with severe sepsis and healthy donors were analyzed with MALDI-TOF MS. PBMCs from healthy donors were also stimulated with lipopolysaccharide, peptidoglycan, CpG oligonucleotides, polyinosinic polycytidylic acid, and with heat-inactivated bacteria. Averaged spectra of PBMCs stimulated in vitro by different agonists were generated from the database using the Biotyper software and matching scores between each spectrum from patients and averaged spectra from the database were calculated.ResultsWe show that the MALDI-TOF MS signature of PBMCs from septic patients was specific, compared with healthy controls. As the fingerprints observed in patients may be related to PBMC activation, PBMCs from healthy controls were stimulated with cytokines, soluble Pathogen-Associated Molecular Patterns (PAMPs) and heat-killed bacteria, and we created a database of reference spectra. The MALDI-TOF MS profiles of PBMCs from septic patients were then compared with the database. No differences were found between patients with documented infection (n = 6) and those without bacteriological documentation (n = 6). The spectra of PBMCs from septic patients matched with those of interferon-γ- and interleukin-10-stimulated PBMCs, confirming that sepsis is characterized by both inflammatory and immunoregulatory features. Interestingly, the spectra of PBMCs from septic patients without documented infection matched with the reference spectrum of PBMCs stimulated by CpG oligonucleotides, suggesting a bacterial etiology in these patients.ConclusionsDespite the limits of this preliminary study, these results indicate that the monitoring of functional status of PBMCs in peripheral blood by whole cell MALDI-TOF MS could provide unique opportunities to assess disease progression or resolution in clinical settings.Electronic supplementary materialThe online version of this article (10.1186/s12879-018-3266-7) contains supplementary material, which is available to authorized users.

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Whole-cell MALDI-TOF Mass Spectrometry is an Accurate and Rapid Method to Analyze Different Modes of Macrophage Activation

Cited by 8 publications

References 26 publications

Direct identification of trypanosomatids by matrix-assisted laser desorption ionization-time of flight mass spectrometry (DIT MALDI-TOF MS)

Direct identification of trypanosomatids by matrix-assisted laser desorption ionization-time of flight mass spectrometry (DIT MALDI-TOF MS)

The Monocyte-Derived Exosomal CLMAT3 Activates the CtBP2-p300-NF-κB Transcriptional Complex to Induce Proinflammatory Cytokines in ALI

MALDI-TOF MS monitoring of PBMC activation status in sepsis

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