Quantitative whole-cell MALDI-TOF MS fingerprints distinguishes human monocyte sub-populations activated by distinct microbial ligands

Abstract

BackgroundConventionally, human monocyte sub-populations are classified according to surface marker expression into classical (CD14++CD16−), intermediate (CD14++CD16+) and non-classical (CD14+CD16++) lineages. The involvement of non-classical monocytes, also referred to as proinflammatory monocytes, in the pathophysiology of diseases including diabetes mellitus, atherosclerosis or Alzheimer’s disease is well recognized. The development of novel high-throughput methods to capture functional states within the different monocyte lineages at the whole cell proteomic level will enable real time monitoring of disease states.ResultsWe isolated and characterized (pan-) monocytes, mostly composed of classical CD16− monocytes, versus autologous CD16+ subpopulations from the blood of healthy human donors (n = 8) and compared their inflammatory properties in response to lipopolysaccharides and M.tuberculosis antigens by multiplex cytokine profiling. Following resting and in vitro antigenic stimulation, cells were recovered and subjected to whole-cell mass spectrometry analysis. This approach identified the specific presence/absence of m/z peaks and therefore potential biomarkers that can discriminate pan-monocytes from their CD16 counterparts. Furthermore, we found that semi-quantitative data analysis could capture the subtle proteome changes occurring upon microbial stimulation that differentiate resting, from lipopolysaccharides or M. tuberculosis stimulated monocytic samples.ConclusionsWhole-cell mass spectrometry fingerprinting could efficiently distinguish monocytic sub-populations that arose from a same hematopoietic lineage. We also demonstrate for the first time that mass spectrometry signatures can monitor semi-quantitatively specific activation status in response to exogenous stimulation. As such, this approach stands as a fast and efficient method for the applied immunology field to assess the reactivity of potentially any immune cell types that may sustain health or promote related inflammatory diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s12896-015-0140-1) contains supplementary material, which is available to authorized users.

Monocyte Subsets: Phenotypes and Function in Tuberculosis Infection

“…Gren et al using single-cell PCR gene expression identified 22 genes that are expressed by the classical population, 8 that characterize the intermediate population, and 6 that distinguish the non-classical population ( 32 ). Resting and activated monocyte subsets can be effectively distinguished using whole cell mass spectrometry fingerprinting ( 33 ). All these studies provide a platform to study further key details about monocyte phenotype and function, their behavior during disease and their interactions with pathogens.…”

Whole Cell MALDI Fingerprinting Is a Robust Tool for Differential Profiling of Two-Component Mammalian Cell Mixtures

“…More advanced applications were aimed at distinguishing different cell types originating from the same cell lineage: identification of two different pancreatic hormone-secreting cell lines [36], the comparison of primary human blood cells and blood cell lines [37,38], molecular phenotyping of central nervous system (CNS) glial cells (astroglial, microglial and oligodendroglial) [39], and MALDI-MS fingerprinting of different melanoma cell lines [40]. Further applications of mammalian fingerprinting has focused on physiological changes of a single cell, reflecting its specific cell states or cell transformations such as differentiation of human colon carcinoma [41] or leukemia [38] cell lines, multifaceted activation of human macrophages [42], identification of resting and activated human monocyte subsets [43], rapid detection of apoptosis/necrosis signature [44], and monitoring of histone deacetylase drug target engagement [45]. Regardless of the scope of the aforementioned studies, no consistency in method parameters were observed (such as matrix, cell density, cell media, sample application technique, laser frequency/number of shots, etc) for either cell authentication [35][36][37][38][39][40] or close monitoring of a single cell changes applications [41][42][43][44][45].…”

Profiling embryonic stem cell differentiation by MALDI TOF mass spectrometry: development of a reproducible and robust sample preparation workflow

“…Characterising the protein signatures of mammalian cells by MALDI-MS is less common when compared with lipid analysis; however it has been used for phenotypic screening of human cancer cell lines, 31,32 identification of cells within a co-culture 33,34 or tissues 35 and detection of transient changes within a specific cell type, such as immune cells. [36][37][38][39] However, many of these studies list dramatically different experimental procedures with several being adapted from existing biotyping protocols. The huge range of experimental parameters could therefore be problematic for translation of published assays to the pharmaceutical industry.…”