Desorption Electrospray Ionization Mass Spectrometry Research Articles

Cover Feature: Are Phosphatidic Acids Ubiquitous in Mammalian Tissues or Overemphasized in Mass Spectrometry Imaging Applications? (Anal. Sens. 5/2023)

The cover feature image shows a cartoon representation of matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), which can produce in-source fragmentation of lipids and cause artificial annotations from biological samples, particularly for phosphatidic acids. Whether these annotations are endogenous or artificial may be assessed by comparison with nanospray desorption electrospray ionization mass spectrometry imaging (nano-DESI-MSI). More information can be found in the Research Article by Christopher Anderton and co-workers.

Rapid Molecular Evaluation of Human Kidney Tissue Sections by In Situ Mass Spectrometry and Machine Learning to Classify the Nephrotic Syndrome.

Nephrotic syndrome (NS) is classified based on morphological changes of glomeruli in biopsied kidney tissues evaluated by time-consuming microscopy methods. In contrast, we employed desorption electrospray ionization mass spectrometry (DESI-MS) directly on renal biopsy specimens obtained from 37 NS patients to rapidly differentiate lipid profiles of three prevalent forms of NS: IgA nephropathy (n = 9), membranous glomerulonephritis (n = 7), and lupus nephritis (n = 8), along with other types of glomerular diseases (n = 13). As we noted molecular heterogeneity in regularly spaced renal tissue regions, multiple sections from each biopsy specimen were collected, providing a total of 973 samples for investigation. Using multivariate analysis, we report differential expressions of glycerophospholipids, sphingolipids, and glycerolipids among the above four classes of NS kidneys, which were otherwise overlooked in several past studies correlating lipid abnormalities with glomerular diseases. We developed machine learning (ML) models with the top 100 features using the support vector machine, which enabled us to discriminate the concerned glomerular diseases with 100% overall accuracy in the training, validation, and holdout test set. This DESI-MS/ML-based tissue analysis can be completed in a few minutes, in sharp contrast to a daylong procedure followed in the conventional histopathology of NS.

Immuno‐Desorption Electrospray Ionization Mass Spectrometry Imaging Identifies Functional Macromolecules by Using Microdroplet‐Cleavable Mass Tags

AbstractWe present immunoassay‐based desorption electrospray ionization mass spectrometry imaging (immuno‐DESI‐MSI) to visualize functional macromolecules such as drug targets and cascade signaling factors. A set of boronic acid mass tags (BMTs) were synthesized to label antibodies as MSI probes. The boronic ester bond is employed to cross‐link the BMT with the galactosamine‐modified antibody. The BMT can be released from its tethered antibody by ultrafast cleavage of the boronic ester bond caused by the acidic condition of sprayed DESI microdroplets containing water. The fluorescent moiety enables the BMT to work in both optical and MS imaging modes. The positively charged quaternary ammonium group enhances the ionization efficiency. The introduction of the boron element also makes mass tags readily identified because of its unique isotope pattern. Immuno‐DESI‐MSI provides an appealing strategy to spatially map macromolecules beyond what can be observed by conventional DESI‐MSI, provided antibodies are available to the targeted molecules of interest.

Immuno-Desorption Electrospray Ionization Mass Spectrometry Imaging Identifies Functional Macromolecules by Using Microdroplet-Cleavable Mass Tags.

We present immunoassay-based desorption electrospray ionization mass spectrometry imaging (immuno-DESI-MSI) to visualize functional macromolecules such as drug targets and cascade signaling factors. A set of boronic acid mass tags (BMTs) were synthesized to label antibodies as MSI probes. The boronic ester bond is employed to cross-link the BMT with the galactosamine-modified antibody. The BMT can be released from its tethered antibody by ultrafast cleavage of the boronic ester bond caused by the acidic condition of sprayed DESI microdroplets containing water. The fluorescent moiety enables the BMT to work in both optical and MS imaging modes. The positively charged quaternary ammonium group enhances the ionization efficiency. The introduction of the boron element also makes mass tags readily identified because of its unique isotope pattern. Immuno-DESI-MSI provides an appealing strategy to spatially map macromolecules beyond what can be observed by conventional DESI-MSI, provided antibodies are available to the targeted molecules of interest.

Endocannabinoid 2-Arachidonoylglycerol Levels in the Anterior Cingulate Cortex, Caudate Putamen, Nucleus Accumbens, and Piriform Cortex Were Upregulated by Chronic Restraint Stress.

Endocannabinoid 2-arachidonoylglycerol (2-AG) has been implicated in habituation to stress, and its augmentation reduces stress-induced anxiety-like behavior. Chronic restraint stress (CRS) changes the 2-AG levels in some gross brain areas, such as the forebrain. However, the detailed spatial distribution of 2-AG and its changes by CRS in stress processing-related anatomical structures such as the anterior cingulate cortex (ACC), caudate putamen (CP), nucleus accumbens (NAc), and piriform cortex (PIR) are still unclear. In this study, mice were restrained for 30 min in a 50 mL-centrifuge tube for eight consecutive days, followed by imaging of the coronal brain sections of control and stressed mice using desorption electrospray ionization mass spectrometry imaging (DESI-MSI). The results showed that from the forebrain to the cerebellum, 2-AG levels were highest in the hypothalamus and lowest in the hippocampal region. 2-AG levels were significantly (p < 0.05) upregulated and 2-AG precursors levels were significantly (p < 0.05) downregulated in the ACC, CP, NAc, and PIR of stressed mice compared with control mice. This study provided direct evidence of 2-AG expression and changes, suggesting that 2-AG levels are increased in the ACC CP, NAc, and PIR when individuals are under chronic stress.

Leaves of Cannabis sativa and their trichomes studied by DESI and MALDI mass spectrometry imaging for their contents of cannabinoids and flavonoids.

In recent years, industrial production of Cannabis sativa has increased due to increased demand of medicinal products based on the plant. In these medicinal products, it is mainly the contents of cannabinoids like THCA and CBDA which are of interest, but also the flavonoids of C. sativa have pharmaceutical interest. The primary aim is to study the distribution of the different cannabinoids in leaves of C. sativa and specifically to which extent they are located on the trichomes found on the surface of C. sativa leaves. Desorption electrospray ionization (DESI) and matrix assisted laser desorption ionization (MALDI) mass spectrometry imaging (MSI) provide non-targeted imaging of numerous compounds in the same experiment. Therefore, the distribution of flavonoids is also mapped in the same experiments. Fan leaves from C. sativa were imaged in the lateral dimension using direct DESI-MSI as well as indirect DESI-MSI via a porous PTFE surface using pixel sizes of 150-200 μm. For cross sections of sugar leaves, MALDI-MSI was performed at 20 μm pixel size. From indirect DESI-MSI experiments, a connection was made between the cannabinoid CBGA and capitate-stalked trichomes. Other cannabinoids like THCA/CBDA (isomers, which are not resolved in an MSI experiment) were also detected in the capitate-stalked trichomes, but in addition to this also in the small glandular trichomes. MALDI-MSI experiments on cross sections of sugar leaves confirmed that the cannabinoids were not an integral part of the leaf tissue itself, but originated from the trichomes on the surface of the leaf. The study provides visual evidence that the cannabinoids are produced and accumulated in the trichomes of C. sativa leaves.

Long-chain polyunsaturated lipids associated with responsiveness to anti-PD-1 therapy are colocalized with immune infiltrates in the tumor microenvironment

The programmed cell death protein-1 (PD-1) is highly expressed on the surface of antigen-specific exhausted T cells and, upon interaction with its ligand PD-L1, can result in inhibition of the immune response. Anti-PD-1 treatment has been shown to extend survival and result in durable responses in several cancers, yet only a subset of patients benefit from this therapy. Despite the implication of metabolic alteration following cancer immunotherapy, mechanistic associations between antitumor responses and metabolic changes remain unclear. Here, we used desorption electrospray ionization mass spectrometry imaging to examine the lipid profiles of tumor tissue from three syngeneic murine models with varying treatment sensitivity at the baseline and at three time points post-anti-PD-1 therapy. These imaging experiments revealed specific alterations in the lipid profiles associated with the degree of response to treatment and allowed us to identify a significant increase of long-chain polyunsaturated lipids within responsive tumors following anti-PD-1 therapy. Immunofluorescence imaging of tumor tissues also demonstrated that the altered lipid profile associated with treatment response is localized to dense regions of tumor immune infiltrates. Overall, these results indicate that effective anti-PD-1 therapy modulates lipid metabolism in tumor immune infiltrates, and we thereby propose that further investigation of the related immune-metabolic pathways may be useful for better understanding success and failure of anti-PD-1 therapy.

DESI-MSI-based technique to unravel spatial distribution of COMT inhibitor Tolcapone

Ascertaining compound exposure and its spatial distribution are essential steps in the drug development process. Desorption electrospray ionization mass spectrometry (DESI-MSI) is a label-free imaging technique capable of simultaneously identify and visualize the distribution of a diverse range of biomolecules.In this study, DESI-MSI was employed to investigate spatial distribution of tolcapone in rat liver and brain coronal – frontal and striatal –sections after a single oral administration of 100 mg/Kg of tolcapone, brain-penetrant compound.Tolcapone was evenly distributed in liver tissue sections whereas in the brain it showed differential distribution across brain regions analyzed, being mainly located in the olfactory bulb, basal forebrain region, striatum, and pre-frontal cortex (PFC; cingulate, prelimbic and infralimbic area). Tolcapone concentration in tissues was compared using DESI-MSI and liquid-chromatography mass spectrometry (LC-MS/MS). DESI-MSI technique showed a higher specificity on detecting tolcapone in liver sections while in the brain samples DESI-MSI did not allow a feasible quantification. Indeed, DESI-MSI is a qualitative technique that allows to observe heterogeneity on distribution but more challenging regarding accurate measurements.Overall, tolcapone was successfully localized in liver and brain tissue sections using DESI-MSI, highlighting the added value that this technique could provide in assisting tissue-specific drug distribution studies.

Identification of end‐stage renal disease metabolic signatures from human perspiration

AbstractEnd‐stage renal disease (ESRD), characterized by cessation in kidney function, has been linked to severe metabolic disturbances, caused by the buildup of toxic solutes in blood. To remove these solutes, ESRD patients undergo dialysis. As a proof of concept, we tested whether ESRD‐related metabolic signatures can be detected in perspiration samples using a combined methodology. Our rapid methodology involves swabbing a glass slide across the patient's forehead, detecting the metabolites in the imprint using desorption electrospray ionization mass spectrometry, and identifying the key differences using machine learning methods. Based on collecting 42 healthy and 27 ESRD samples, we find saturated fatty acids are consistently suppressed in ESRD patients, with little change after dialysis. Moreover, our method enables the detection of uremic solutes, where we find elevated levels of uric acid (6.7 fold higher on average) that sharply decrease after dialysis. Beyond the study of individual metabolites, we find that a lasso model, which selects for 8 m/z fragments from 24,602 detected analytes, achieves area under the curve performance of 0.85 and 0.87 on training (n = 52) and validation sets (n = 17), respectively. Together, these results suggest that this methodology is promising for detecting signatures relevant for precision health.Key points Combines physical measurements with statistical analysis based on machine learning. Uses results to make medical predictions. Learns about biological pathways.

Low-cost, automated reaction screening for energetic precursor cage compounds by a benchtop liquid handling robot and desorption electrospray ionization mass spectrometry

This study reports a low-cost, open source, and commercial-off-the-shelf automated pipetting robot coupled with a homebuilt desorption electrospray ionization mass spectrometer for the screening of energetic precursor cage compounds.

Desorption electrospray ionization mass spectrometry imaging (DESI-MSI) in disease diagnosis: an overview.

Tissue analysis, which is essential to histology and is considered the benchmark for the diagnosis and prognosis of many illnesses, including cancer, is significant. During surgery, the surgical margin of the tumor is assessed using the labor-intensive, challenging, and commonly subjective technique known as frozen section histopathology. In the biopsy section, large numbers of molecules can now be visualized at once (ion images) following recent developments in [MSI] mass spectrometry imaging under atmospheric conditions. This is vastly superior to and different from the single optical tissue image processing used in traditional histopathology. This review article will focus on the advancement of desorption electrospray ionization mass spectrometry imaging [DESI-MSI] technique, which is label-free and requires little to no sample preparation. Since the proportion of molecular species in normal and abnormal tissues is different, DESI-MSI can capture ion images of the distributions of lipids and metabolites on biopsy sections, which can provide rich diagnostic information. This is not a systematic review but a summary of well-known, cutting-edge and recent DESI-MSI applications in cancer research between 2018 and 2023.

Multimodal high-resolution nano-DESI MSI and immunofluorescence imaging reveal molecular signatures of skeletal muscle fiber types.

The skeletal muscle is a highly heterogeneous tissue comprised of different fiber types with varying contractile and metabolic properties. The complexity in the analysis of skeletal muscle fibers associated with their small size (30-50 μm) and mosaic-like distribution across the tissue tnecessitates the use of high-resolution imaging to differentiate between fiber types. Herein, we use a multimodal approach to characterize the chemical composition of skeletal fibers in a limb muscle, the gastrocnemius. Specifically, we combine high-resolution nanospray desorption electrospray ionization (nano-DESI) mass spectrometry imaging (MSI) with immunofluorescence (IF)-based fiber type identification. Computational image registration and segmentation approaches are used to integrate the information obtained with both techniques. Our results indicate that the transition between oxidative and glycolytic fibers is associated with shallow chemical gradients (<2.5 fold change in signals). Interestingly, we did not find any fiber type-specific molecule. We hypothesize that these findings might be linked to muscle plasticity thereby facilitating a switch in the metabolic properties of fibers in response to different conditions such as exercise and diet, among others. Despite the shallow chemical gradients, cardiolipins (CLs), acylcarnitines (CAR), monoglycerides (MGs), fatty acids, highly polyunsaturated phospholipids, and oxidized phospholipids, were identified as molecular signatures of oxidative metabolism. In contrast, histidine-related compounds were found as molecular signatures of glycolytic fibers. Additionally, the presence of highly polyunsaturated acyl chains in phospholipids was found in oxidative fibers whereas more saturated acyl chains in phospholipids were found in glycolytic fibers which suggests an effect of the membrane fluidity on the metabolic properties of skeletal myofibers.

Mapping of Fatty Aldehydes in the Diabetic Rat Brain Using On-Tissue Chemical Derivatization and Air-Flow-Assisted Desorption Electrospray Ionization-Mass Spectrometry Imaging.

Fatty aldehydes (FALs) are involved in various biological processes, and their abnormal metabolism is related to the occurrence and development of neurological diseases. Because of their low ionization efficiency, methods for in situ detection and mass spectrometry imaging (MSI) analysis of FALs remain underreported. On-tissue chemical tagging of hardly ionizable target analytes with easily ionized moieties can improve ionization efficiency and detection sensitivity in MSI experiments. In this study, an on-tissue chemical derivatization-air-flow-assisted desorption electrospray ionization-MSI method was developed to visualize FALs in the rat brain. The method showed high sensitivity and specificity, allowing the use of in situ high-resolution MS3 to identify FALs. The methodology was applied to investigate the region-specific distribution of FALs in the brains of control and diabetic encephalopathy (DE) rats. In DE rats, FALs were found to be significantly enriched in various brain regions, especially in the cerebral cortex, hippocampus, and amygdala. Thus, increased FAL levels and oxidative stress occurred in a region-dependent manner, which may contribute to cognitive function deficits in DE. In summary, we provide a novel method for the in situ detection of FALs in biological tissues as well as new insights into the potential pathogenesis of DE.

Expression and Kinetics of Endogenous Cannabinoids in the Brain and Spinal Cord of a Spare Nerve Injury (SNI) Model of Neuropathic Pain

The role of endogenous cannabinoids in neuropathic pain has been actively studied, among which 2-arachidonoyl glycerol (2-AG) has received the most attention. However, owing to its chemical properties, direct detection of 2-AG distribution in tissues is difficult. Moreover, although desorption electrospray ionization mass spectrometry imaging (DESI-MSI) has enabled the detection of 2-AG, its distribution in the brain and spinal cord of neuropathic pain models has not been reported. In this study, the expression and distribution of 2-AG in the brain and spinal cord of a spare nerve injury (SNI) mice model of neuropathic pain was examined using DESI-MSI. The brain and lumbar spinal cord were collected and analyzed on days 3, 7, and 21 after treatment. On days 3 and 7 after treatment, 2-AG expression in the SNI model was decreased in the hypothalamus, midbrain, and especially in the periaqueductal gray (PAG) region but increased in the lumbar spinal cord. On day 21, the SNI model showed decreased 2-AG expression in the hypothalamus, but the difference from the control was not significant. Furthermore, there were no differences in 2-AG expression between the lumbar spinal cord, midbrain, or PAG. These data suggest that 2-AG might be involved in pain control.

Visualization of Metabolites Identified in the Spatial Metabolome of Traditional Chinese Medicine Using DESI-MSI.

The medicinal use of traditional Chinese medicine is mainly due to its secondary metabolites. Visualization of the distribution of these metabolites has become a crucial topic in plant science. Mass spectrometry imaging can extract huge volumes of data and provide spatial distribution information about these by analyzing tissue slices. With the advantage of high throughput and higher accuracy, desorption electrospray ionization mass spectrometry imaging (DESI-MSI) is often used in biological research and in the study of traditional Chinese medicine. However, the procedures used in this research are complicated and not affordable. In this study, we optimized sectioning and DESI imaging procedures and developed a more cost-effective method to identify the distribution of metabolites and categorize these compounds in plant tissues, with a special focus on traditional Chinese medicines. The study will promote the utilization of DESI in metabolite analysis and standardization of traditional Chinese medicine/ethnic medicine for research-related technologies.

Investigation on property differences of ginseng and American ginseng by spatial metabolomics of neurochemicals with desorption electrospray ionization mass spectrometry imaging

Ethnopharmacological relevanceThe properties are the body's response to traditional Chinese medicine (TCM). The essence of traditional Chinese medicine properties are cold, hot, warm, and cool. In the theory of traditional Chinese medicine, ginseng is warm and American ginseng is cool, they present two opposite properties. The material basis of property differences and effect mechanism of property degree need further investigation. Aim of the studyThe aim of this work was to screen out the neurochemicals related to warm and cool properties of ginseng and American ginseng, and investigate the distributions of identified neurochemicals in rat brain and the metabolic mechanism. Materials and methodsSpatial metabolomics was applied to study the effects of ginseng and American ginseng on the distributions of neurochemicals in rat brain by desorption electrospray ionization mass spectrometry imaging (DESI-MSI). Based on discriminant coefficients in partial least square discriminant analysis (PLS-DA) processing, neurochemicals related to warm and cool properties were classified. In addition, the score contributions of the neurochemicals markers could be used to evaluate the warm and cool property degrees. ResultsA total of 25 neurochemicals were imaged and identified in brain section. The distribution regions of main neurochemicals were consistent with in sagittal and coronal sections of brain reported in literature. 17 neurochemicals were classified as warm markers. Meanwhile, 8 neurochemicals were identified as cool markers, correlated with the cool properties of American ginseng. It demonstrated that the score contributions of the 25 neurochemicals markers could be used to evaluate the warm and cool property degrees. Based on the regulatory effects of neurochemicals, the warm markers could promote the body's energy metabolism, improve the function of endocrine system, and enhance the excitability of central nervous system. The cool property markers have reduced excitability of central nervous system, weakened metabolism and stress response ability, thus presented the biological activity of cool and cold. ConclusionsOur findings provided a rapid and effective visualization method for the spatial distribution and metabolism of small molecular neurochemicals in rat brain. DESI-MSI was a reference methodology for evaluating the properties of TCM.

High accumulation of microplastic fibers in fish hindgut induces an enhancement of triphenyl phosphate hydroxylation

Fiber shedding from artificial textiles is among the primary sources of pervasive microplastics in various aquatic habitats. To avoid molten drop burning, triphenyl phosphate (TPhP), a typical flame retardant additive, is commonly incorporated into textile fibers. However, the role of microplastic fibers (MFs) as a vehicle for TPhP remains largely unknown. In this study, we investigated the effects of MFs on the bioaccumulation and metabolism of TPhP in zebrafish. We applied the compound spinning technique for a non-disruptive in situ measurement of fluorescent MFs in fish, and the desorption electrospray ionization mass spectrometry (DESI-MS) to display the tissue distribution of TPhP and its metabolites vividly. Laboratory results showed that ingested MFs did not change the TPhP distribution in fish; however, they statistically increased the metabolite p-OH-TPhP concentration in the fish hindgut, which was probably because the high accumulation of MFs there enhanced the TPhP hydroxylation. Field investigation further supported the lab-based analyses. Higher concentrations of MFs did cause a higher ratio of [p-OH-TPhP]/[TPhP] in the wild fish gut, particularly in the hindgut. Collectively, our results demonstrated that MFs can change the distribution and bioavailability of TPhP metabolites, which was confirmed by both laboratory and fieldwork. Therefore, the ingestion of MFs can indirectly but substantially influence the bioaccumulation and biotransformation of co-existing pollutants.

2022 Canadian Surgery Forum Sept. 15–17, 202201. Operative classification of ventral abdominal hernias: new and practical classification02. Watchful waiting for large primary splenic cysts03. Transversus abdominis plane (TAP) blocks with and without dexamethasone in colorectal surgery04. What factors determine publication of resident research day projects?05. Characterization of near-infrared imaging and indocyanine green use amongst general surgeons06. Variation in opioid prescribing

# 01. Operative classification of ventral abdominal hernias: new and practical classification {#article-title-2} Ventral hernias of the abdomen are defined as a noninguinal, nonhiatal defect in the fascia of the abdominal wall. Unfortunately, there is not currently a universal classification system

Identification and localization of morphological feature-specific metabolites in Reynoutria multiflora roots

Reynoutria multiflora roots are a classical herbal medicine with unique nourishing therapeutic effects. Anomalous vascular bundle (AVB) forming “cloudy brocade patterns” is a typical morphological feature of R. multiflora roots and has been empirically linked to its quality classification. However, scientific evidence, especially for AVB-specific specialised metabolites, has not been comprehensively revealed thus far. Herein, desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) analysis was applied to carry out an in situ analysis of specialised metabolites distributed specifically at the AVB and cork of R. multiflora roots. To enlarge the scope of compounds by DESI detection, various solvent systems including acetone, acetonitrile, methanol, and water were used to assist in the discoveries of 40 specialised metabolites with determined localization. A series of bioactive constituents including stilbenes, flavonoids, anthraquinones, alkaloids, and naphthalenes were found specifically around the brocade patterns. Notably, phospholipids were detected from R. multiflora roots by in situ analysis for the first time and were found mainly in the phloem of AVB (PAB). This is the first study to use gradient solvent systems in DESI-MSI analysis to locate the specialised metabolites distribution. The discovery of feature-specific compounds will bridge the empirical identification to precision quality control of R. multiflora roots.

Spatial metabolomics identifies lipid profiles of human carotid atherosclerosis

Background and aimsCarotid atherosclerosis is an important cause of ischemic stroke. Lipids play a key role in the progression of atherosclerosis. To date, the spatial lipid profile of carotid atherosclerotic plaques related to histology has not been systematically investigated. MethodsCarotid atherosclerosis samples from 12 patients were obtained and classified into four classical pathological stages (preatheroma, atheroma, fibroatheroma and complicated lesion) by histological staining. Desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) was used to investigate the lipid profile of carotid atherosclerosis, and correlated it with histological information. Bioinformatics technology was used to process MSI data among different pathological stages of atherosclerosis lesions. ResultsA total of 55 lipids (26 throughout cross-section regions [TCSRs], 13 in lipid-rich regions [LRRs], and 16 in collagen-rich regions [CRRs]) were initially identified in carotid plaque from one patient. Subsequently, 32 of 55 lipids (12 in TCSRs, eight in LRRs, and 12 in CRRs) were further screened in 11 patients. Pathway enrichment analysis showed that multiple metabolic pathways, such as fat digestion and absorption, cholesterol metabolism, lipid and atherosclerosis, were enriched in TCSRs; sphingolipid signaling pathway, necroptosis pathway were enriched in LRRs; and glycerophospholipid metabolism, ether lipid metabolism pathway were mainly enriched in CRRs. ConclusionsThis study comprehensively showed the spatial lipid metabolism footprint in human carotid atherosclerotic plaques. The lipid profiles and related metabolism pathways in three regions of plaque with disease progression were different markedly, suggesting that the different metabolic mechanisms in these regions of carotid plaque may be critical in atherosclerosis progression.