Metabolism of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) to the neurotoxin MPP⁺ in the brain causes permanent Parkinson’s disease-like symptoms by destroying dopaminergic neurons in the pars compacta of the substantia nigra in humans and non-human primates. However, the complete molecular pathology underlying MPTP-induced parkinsonism remains poorly understood. We used dual polarity matrix-assisted laser desorption/ionization mass spectrometry imaging to thoroughly image numerous glycerophospholipids and sphingolipids in coronal brain tissue sections of MPTP-lesioned and control non-human primate brains (Macaca mulatta). The results revealed specific distributions of several sulfatide lipid molecules based on chain-length, number of double bonds, and importantly, hydroxylation stage. More specifically, certain long-chain hydroxylated sulfatides with polyunsaturated chains in the molecular structure were depleted within motor-related brain regions in the MPTP-lesioned animals, e.g., external and internal segments of globus pallidus and substantia nigra pars reticulata. In contrast, certain long-chain non-hydroxylated sulfatides were found to be elevated within the same brain regions. These findings demonstrate region-specific dysregulation of sulfatide metabolism within the MPTP-lesioned macaque brain. The depletion of long-chain hydroxylated sulfatides in the MPTP-induced pathology indicates oxidative stress and oligodendrocyte/myelin damage within the pathologically relevant brain regions. Hence, the presented findings improve our current understanding of the molecular pathology of MPTP-induced parkinsonism within primate brains, and provide a basis for further research regarding the role of dysregulated sulfatide metabolism in PD.

Neonatal hypoxic-ischaemic (HI) encephalopathy is among the most serious complications in neonatology. In the present study, we studied the immediate (0 hour), subacute (36 hours) and late (144 hours) responses of the neonatal brain to experimental HI insult in laboratory rats. At the striatal level, the mass spectrometry imaging revealed an aberrant plasma membrane distribution of Na⁺/K⁺ ions in the oedema-affected areas. The failure of the Na⁺/K⁺ gradients was also apparent in the magnetic resonance imaging measurements, demonstrating intracellular water accumulation during the acute phase of the HI insult. During the subacute phase, compared with the control brains, an incipient accumulation of an array of N-acylphosphatidylethanolamine (NAPE) molecules was detected in the HI-affected brains, and both the cytotoxic and vasogenic types of oedema were detected. In the severely affected brain areas, abnormal distributions of the monosialogangliosides GM2 and GM3 were observed in two-thirds of the animals exposed to the insult. During the late stage, a partial restoration of the brain tissue was observed in most rats in both the in vivo and ex vivo studies. These specific molecular changes may be further utilized in neonatology practice in proposing and testing novel therapeutic strategies for the treatment of neonatal HI encephalopathy.

Infection metallomics is a mass spectrometry (MS) platform we established based on the central concept that microbial metallophores are specific, sensitive, noninvasive, and promising biomarkers of invasive infectious diseases. Here we review the in vitro, in vivo, and clinical applications of metallophores from historical and functional perspectives, and identify under‐studied and emerging application areas with high diagnostic potential for the post‐COVID era. MS with isotope data filtering is fundamental to infection metallomics; it has been used to study the interplay between “frenemies” in hosts and to monitor the dynamic response of the microbiome to antibiotic and antimycotic therapies. During infection in critically ill patients, the hostile environment of the host's body activates secondary bacterial, mycobacterial, and fungal metabolism, leading to the production of metallophores that increase the pathogen's chance of survival in the host. MS can reveal the structures, stability, and threshold concentrations of these metal‐containing microbial biomarkers of infection in humans and model organisms, and can discriminate invasive disease from benign colonization based on well‐defined thresholds distinguishing proliferation from the colonization steady state.

Pseudomonas aeruginosa is recognized as a significant cause of morbidity and mortality among nosocomial pathogens. In respiratory infections, P. aeruginosa acts not only as a single player but also collaborates with the opportunistic fungal pathogen Aspergillus fumigatus. This study introduced a QS molecule portfolio as a potential new biomarker that affects the secretion of virulence factors and biofilm formation. The quantitative levels of QS molecules, including 3-o-C12-HSL, 3-o-C8-HSL, C4-HSL, C6-HSL, HHQ, PQS, and PYO, measured using mass spectrometry in a monoculture, indicated metabolic changes during the transition from planktonic to sessile cells. In the co-cultures with A. fumigatus, the profile of abundant QS molecules was reduced to 3-o-C12-HSL, C4-HSL, PQS, and PYO. A decrease in C4-HSL by 50% to 170.6 ± 11.8 ng/mL and an increase 3-o-C12-HSL by 30% up to 784.4 ± 0.6 ng/mL were detected at the stage of the coverage of the hyphae with bacteria. Using scanning electron microscopy, we showed the morphological stages of the P. aeruginosa biofilm, such as cell aggregates, maturated biofilm, and cell dispersion. qPCR quantification of the genome equivalents of both microorganisms suggested that they exhibited an interplay strategy rather than antagonism. This is the first study demonstrating the quantitative growth-dependent appearance of QS molecule secretion in a monoculture of P. aeruginosa and a co-culture with A. fumigatus.

A procedure for processing frozen rat lung tissue sections for scanning electron microscopy (SEM) from deeply frozen samples initially collected and stored for matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was developed. The procedure employed slow thawing of the frozen sections while floating on the surface and melting in a fixative solution. After the float-washing step, the sections were dehydrated in a graded ethanol series and dried in a critical point dryer. The SEM generated images with well-preserved structures, allowing for monitoring of bacterial cells and fungal hyphae in the infected tissue. Importantly, the consecutive nonfixed frozen sections were fully compatible with MALDI-MSI, providing molecular biomarker maps of Pseudomonas aeruginosa. The protocol enables bimodal image fusion in the in-house software CycloBranch, as demonstrated by SEM and MALDI-MSI.

Liver is endowed with high regenerative activity, so that the tissue regrows in mouse after partial hepatectomy within days. We reason that this requires de novo pyrimidine synthesis to support rapid progression via the cell cycle. We find that suppression of de novo pyrimidine synthesis prevents proliferation in regenerating liver, suppressing liver regrowth. Tracing studies and spatial metabolomics reveal a metabolic shift such that ammonia, normally detoxified to urea in the periportal region under homeostasis, is redirected for generating aspartate and carbamoyl phosphate periportally, and glutamine pericentrally, and these products are utilized as precursors by the de novo pyrimidine synthesis pathway. Our research uncovers a metabolic reprogramming leading to utilization of a toxic byproduct for anabolic pathways that are essential for liver regeneration.