Deliverables from Metabolomics in Kidney Disease: Adenine, New Insights, and Implication for Clinical Decision-Making.

BACKGROUND: Chronic kidney disease (CKD) presents a persistent global health challenge, characterized by complex pathophysiology and diverse progression patterns. Metabolomics has emerged as a valuable tool in unraveling the intricate molecular mechanisms driving CKD progression. SUMMARY: This comprehensive review provides a summary of recent progress in the field of metabolomics in kidney disease with a focus on spatial metabolomics to shed important insights to enhancing our understanding of CKD progression, emphasizing its transformative potential in early disease detection, refined risk assessment, and the development of targeted interventions to improve patient outcomes. KEY MESSAGE: Through an extensive analysis of metabolic pathways and small-molecule fluctuations, bulk and spatial metabolomics offers unique insights spanning the entire spectrum of CKD, from early stages to advanced disease states. Recent advances in metabolomics technology have enabled spatial identification of biomarkers to provide breakthrough discoveries in predicting CKD trajectory and enabling personalized risk assessment. Furthermore, metabolomics can help decipher the complex molecular intricacies associated with kidney diseases for exciting novel therapeutic approaches. A recent example is the identification of adenine as a key marker of kidney fibrosis for diabetic kidney disease using both untargeted and targeted bulk and spatial metabolomics. The metabolomics studies were critical to identify a new biomarker for kidney failure and to guide new therapeutics for diabetic kidney disease. Similar approaches are being pursued for acute kidney injury and other kidney diseases to enhance precision medicine decision-making.

Extracellular Microenvironment Alterations in Ductal Carcinoma In Situ and Invasive Breast Cancer Pathologies by Multiplexed Spatial Proteomics.

Ductal carcinoma in situ (DCIS) is a heterogeneous breast disease that remains challenging to treat due to its unpredictable progression to invasive breast cancer (IBC). Contemporary literature has become increasingly focused on extracellular matrix (ECM) alterations with breast cancer progression. However, the spatial regulation of the ECM proteome in DCIS has yet to be investigated in relation to IBC. We hypothesized that DCIS and IBC present distinct ECM proteomes that could discriminate between these pathologies. Tissue sections of pure DCIS, mixed DCIS-IBC, or pure IBC (n = 22) with detailed pathological annotations were investigated by multiplexed spatial proteomics. Across tissues, 1,005 ECM peptides were detected in pathologically annotated regions and their surrounding extracellular microenvironments. A comparison of DCIS to IBC pathologies demonstrated 43 significantly altered ECM peptides. Notably, eight fibrillar collagen peptides could distinguish with high specificity and sensitivity between DCIS and IBC. Lesion-targeted proteomic imaging revealed heterogeneity of the ECM proteome surrounding individual DCIS lesions. Multiplexed spatial proteomics reported an invasive cancer field effect, in which DCIS lesions in closer proximity to IBC shared a more similar ECM profile to IBC than distal counterparts. Defining the ECM proteomic microenvironment provides novel molecular insights relating to DCIS and IBC.

Complex-Type N-Glycans Are Associated with Cartilage Degeneration within Different Loading Sites of the Tibial Plateau for Knee Osteoarthritis Patients.

Abnormal N-glycosylation has been shown to play an important role in the pathogenesis of multiple diseases. However, little is known about the relationship between N-glycosylation and knee osteoarthritis (KOA) progression at the tissue level. Thus, the aim of this study was to quantify the cartilage histomorphometric changes in formalin-fixed paraffin-embedded (FFPE) tissue collected from the lateral and medial compartments of the tibial plateau KOA patients (n = 8). Subsequently, N-glycans were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) followed by in situ MS/MS fragmentation. Overall, the Osteoarthritis Research Society International (OARSI) histological grade and cartilage surface fibrillation index were significantly higher, and chondrocyte size in the superficial zone was much larger, for the medial high-loaded cartilage compared to the lateral less-loaded cartilage. Among 92 putative N-glycans observed by MALDI-MSI, 3 complex-type N-glycans, (Hex)4(HexNAc)3, (Hex)4(HexNAc)4, and (Hex)5(HexNAc)4, and 1 oligomannose-type N-glycan, (Hex)9(HexNAc)2, were significantly higher in intensity in the medial cartilage compared to the lateral cartilage, whereas 2 tetra-antennary fucosylated-type N-glycans, (Hex)3(HexNAc)6(Fuc)2 and (Hex)3(HexNAc)6(Fuc)3, were significantly higher in intensity in the lateral cartilage than the medial cartilage. Our findings indicate that complex-type N-glycans are associated with higher severity of cartilage degeneration and may influence the cellular processes of KOA.

MALDI mass spectrometry imaging shows a gradual change in the proteome landscape during mouse ovarian folliculogenesis.

Our knowledge regarding the role proteins play in the mutual relationship among oocytes, surrounding follicle cells, stroma, and the vascular network inside the ovary is still poor and obtaining insights into this context would significantly aid our understanding of folliculogenesis. Here, we describe a spatial proteomics approach to characterize the proteome of individual follicles at different growth stages in a whole prepubertal 25-day-old mouse ovary. A total of 401 proteins were identified by nano-scale liquid chromatography-electrospray ionization-tandem mass spectrometry (nLC-ESI-MS/MS), 69 with a known function in ovary biology, as demonstrated by earlier proteomics studies. Enrichment analysis highlighted significant KEGG and Reactome pathways, with apoptosis, developmental biology, PI3K-Akt, epigenetic regulation of gene expression, and extracellular matrix organization being well represented. Then, correlating these data with the spatial information provided by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) on 276 follicles enabled the protein profiles of single follicle types to be mapped within their native context, highlighting 94 proteins that were detected throughout the secondary to the pre-ovulatory transition. Statistical analyses identified a group of 37 proteins that showed a gradual quantitative change during follicle differentiation, comprising 10 with a known role in follicle growth (NUMA1, TPM2), oocyte germinal vesicle-to-metaphase II transition (SFPQ, ACTBL, MARCS, NUCL), ovulation (GELS, CO1A2), and preimplantation development (TIF1B, KHDC3). The proteome landscape identified includes molecules of known function in the ovary, but also those whose specific role is emerging. Altogether, this work demonstrates the utility of performing spatial proteomics in the context of the ovary and offers sound bases for more in-depth investigations that aim to further unravel its spatial proteome.

Structural amyloid plaque polymorphism is associated with distinct lipid accumulations revealed by trapped ion mobility mass spectrometry imaging.

Understanding of Alzheimer's disease (AD) pathophysiology requires molecular assessment of how key pathological factors, specifically amyloid ß (Aß) plaques, influence the surrounding microenvironment. Here, neuronal lipids have been implicated in Aß plaque pathology, though the lipid microenvironment in direct proximity to Aß plaques is still not fully resolved. A further challenge is the microenvironmental molecular heterogeneity, across structurally polymorphic Aß features, such as diffuse, immature, and mature, fibrillary aggregates, whose resolution requires the integration of advanced, multimodal chemical imaging tools. Herein, we used matrix-assisted laser desorption/ionization trapped ion mobility spectrometry time-of-flight based mass spectrometry imaging (MALDI TIMS TOF MSI) in combination with hyperspectral confocal microscopy to probe the lipidomic microenvironment associated with structural polymorphism of Aß plaques in transgenic Alzheimer's disease mice (tgAPPSWE ). Using on tissue and ex situ validation, TIMS MS/MS facilitated unambiguous identification of isobaric lipid species that showed plaque pathology-associated localizations. Integrated multivariate imaging data analysis revealed multiple, Aß plaque-enriched lipid patterns for gangliosides (GM), phosphoinositols (PI), phosphoethanolamines (PE), and phosphatidic acids (PA). Conversely, sulfatides (ST), cardiolipins (CL), and polyunsaturated fatty acid (PUFA)-conjugated phosphoserines (PS), and PE were depleted at plaques. Hyperspectral amyloid imaging further delineated the unique distribution of PA and PE species to mature plaque core regions, while PI, LPI, GM2 and GM3 lipids localized to immature Aß aggregates present within the periphery of Aß plaques. Finally, we followed AD pathology-associated lipid changes over time, identifying plaque- growth and maturation to be characterized by peripheral accumulation of PI (18:0/22:6). Together, these data demonstrate the potential of multimodal imaging approaches to overcome limitations associated with conventional advanced MS imaging applications. This allowed for the differentiation of both distinct lipid components in a complex micro-environment as well as their correlation to disease-relevant amyloid plaque polymorphs. Cover Image for this issue: https://doi.org/10.1111/jnc.15390.

Involvement of α-galactosidase OmAGAL2 in planteose hydrolysis during seed germination of Orobanche minor.

Root parasitic weeds of the Orobanchaceae, such as witchweeds (Striga spp.) and broomrapes (Orobanche and Phelipanche spp.), cause serious losses in agriculture worldwide, and efforts have been made to control these parasitic weeds. Understanding the characteristic physiological processes in the life cycle of root parasitic weeds is particularly important to identify specific targets for growth modulators. In our previous study, planteose metabolism was revealed to be activated soon after the perception of strigolactones in germinating seeds of O. minor. Nojirimycin inhibited planteose metabolism and impeded seed germination of O. minor, indicating a possible target for root parasitic weed control. In the present study, we investigated the distribution of planteose in dry seeds of O. minor by matrix-assisted laser desorption/ionization-mass spectrometry imaging. Planteose was detected in tissues surrounding-but not within-the embryo, supporting its suggested role as a storage carbohydrate. Biochemical assays and molecular characterization of an α-galactosidase family member, OmAGAL2, indicated that the enzyme is involved in planteose hydrolysis in the apoplast around the embryo after the perception of strigolactones, to provide the embryo with essential hexoses for germination. These results indicate that OmAGAL2 is a potential molecular target for root parasitic weed control.

Mass spectrometry imaging spatially identifies complex-type N-glycans as putative cartilage degradation markers in human knee osteoarthritis tissue.

N-Glycan alterations contribute to the pathophysiology and progression of various diseases. However, the involvement of N-glycans in knee osteoarthritis (KOA) progression at the tissue level, especially within articular cartilage, is still poorly understood. Thus, the aim of this study was to spatially map and identify KOA-specific N-glycans from formalin-fixed paraffin-embedded (FFPE) osteochondral tissue of the tibial plateau relative to cadaveric control (CTL) tissues. Human FFPE osteochondral tissues from end-stage KOA patients (n=3) and CTL individuals (n=3), aged >55 years old, were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Overall, it was revealed that 22 N-glycans were found in the cartilage region of KOA and CTL tissue. Of those, 15 N-glycans were more prominent in KOA cartilage than CTL cartilage. We then compared sub-regions of KOA and CTL tissues based on the Osteoarthritis Research Society International (OARSI) histopathological grade (1 to 6), where 1 is an intact cartilage surface and 6 is cartilage surface deformation. Interestingly, three specific complex-type N-glycans, (Hex)4(HexNAc)3, (Hex)4(HexNAc)4, and (Hex)5(HexNAc)4, were found to be localized to the superficial fibrillated zone of degraded cartilage (KOA OARSI 2.5-4), compared to adjacent cartilage with less degradation (KOA OARSI 1-2) or relatively healthy cartilage (CTL OARSI 1-2). Our results demonstrate that N-glycans specific to degraded cartilage in KOA patients have been identified at the tissue level for the first time. The presence of these N-glycans could further be evaluated as potential diagnostic and prognostic biomarkers.

Evaluation of Proteomic and Lipidomic Changes in Aeromonas-Infected Trout Kidney Tissue with the Use of FT-IR Spectroscopy and MALDI Mass Spectrometry Imaging.

Aeromonas species are opportunistic bacteria causing a vast spectrum of human diseases, including skin and soft tissue infections, meningitis, endocarditis, peritonitis, gastroenteritis, and finally hemorrhagic septicemia. The aim of our research was to indicate the molecular alterations in proteins and lipids profiles resulting from Aeromonas sobria and A. salmonicida subsp. salmonicida infection in trout kidney tissue samples. We successfully applied FT-IR (Fourier transform infrared) spectroscopy and MALDI-MSI (matrix-assisted laser desorption/ionization mass spectrometry imaging) to monitor changes in the structure and compositions of lipids, secondary conformation of proteins, and provide useful information concerning disease progression. Our findings indicate that the following spectral bands' absorbance ratios (spectral biomarkers) can be used to discriminate healthy tissue from pathologically altered tissue, for example, lipids (CH2/CH3), amide I/amide II, amide I/CH2 and amide I/CH3. Spectral data obtained from 10 single measurements of each specimen indicate numerous abnormalities concerning proteins, lipids, and phospholipids induced by Aeromonas infection, suggesting significant disruption of the cell membranes. Moreover, the increase in the content of lysolipids such as lysophosphosphatidylcholine was observed. The results of this study suggest the application of both methods MALDI-MSI and FT-IR as accurate methods for profiling biomolecules and identifying biochemical changes in kidney tissue during the progression of Aeromonas infection.

[Spatial distribution characteristics of metabolities in rhizome of Paris polyphylla var. yunnanensis: based on MALDI-MSI].

In this study, a method was established for in-situ visualization of metabolite distribution in the rhizome of Paris polyphylla var. yunnanensis. To be specific, through matrix-assisted laser desorption/ionization-mass spectrometry imaging(MALDI-MSI), the spatial locations of steroidal saponins, amino acids, organic acids, phytosterols, phytoecdysones, nucleosides, and esters in rhizome of the medicinal plant were directly analyzed, and six unknown compounds with differential distribution in rhizome tissues were identified. The specific procedure is as follows: preparation of rhizome tissue section, matrix screening and optimization, and MALDI-MSI analysis. The results showed that the steroidal saponins were mainly distributed in the central, amino acids in epidermis and cortex, low-molecular-weight organic acids in central epidermis, phytosterols in the epidermis and lateral cortex, the phytoecdysones in epidermis and cortex, nucleosides(uneven distribution) in epidermis and cortex, growth hormones around the epidermis and cortex, particularly outside the cortex, and esters in cortex with unobvious difference among different tissues. In this study, the spatial distribution of meta-bolites in the rhizome of P. polyphylla var. yunnanensis was characterized for the first time. The result can serve as a reference for identifying and extracting endogenous metabolites of P. polyphylla var. yunnanensis, exploring the synthesis and metabolism mechanisms of the metabolites, and evaluating the quality of medicinal materials.

Tissue-specific analysis of lipid species in Drosophila during overnutrition by UHPLC-MS/MS and MALDI-MSI.

Diets high in calories can be used to model metabolic diseases, including obesity and its associated comorbidities, in animals. Drosophila melanogaster fed high-sugar diets (HSDs) exhibit complications of human obesity including hyperglycemia, hyperlipidemia, insulin resistance, cardiomyopathy, increased susceptibility to infection, and reduced longevity. We hypothesize that lipid storage in the high-sugar-fed fly's fat body (FB) reaches a maximum capacity, resulting in the accumulation of toxic lipids in other tissues or lipotoxicity. We took two approaches to characterize tissue-specific lipotoxicity. Ultra-HPLC-MS/MS and MALDI-MS imaging enabled spatial and temporal localization of lipid species in the FB, heart, and hemolymph. Substituent chain length was diet dependent, with fewer odd chain esterified FAs on HSDs in all sample types. By contrast, dietary effects on double bond content differed among organs, consistent with a model where some substituent pools are shared and others are spatially restricted. Both di- and triglycerides increased on HSDs in all sample types, similar to observations in obese humans. Interestingly, there were dramatic effects of sugar feeding on lipid ethers, which have not been previously associated with lipotoxicity. Taken together, we have identified candidate endocrine mechanisms and molecular targets that may be involved in metabolic disease and lipotoxicity.

Detecting Proteomic Indicators to Distinguish Diabetic Nephropathy from Hypertensive Nephrosclerosis by Integrating Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging with High-Mass Accuracy Mass Spectrometry.

INTRODUCTION: Diabetic nephropathy (DN) and hypertensive nephrosclerosis (HN) represent the most common causes of chronic kidney disease (CKD) and many patients progress to -end-stage renal disease. Patients are treated primarily through the management of cardiovas-cular risk factors and hypertension; however patients with HN have a more favorable outcome. A noninvasive clinical approach to separate these two entities, especially in hypertensive patients who also have diabetes, would allow for targeted treatment and more appropriate resource allocation to those patients at the highest risk of CKD progression. Meth-ods: In this preliminary study, high-spatial-resolution matrix-assisted laser desorption/ion-ization (MALDI) mass spectrometry imaging (MSI) was integrated with high-mass accuracy MALDI-FTICR-MS and nLC-ESI-MS/MS analysis in order to detect tissue proteins within kidney biopsies to discriminate cases of DN (n = 9) from cases of HN (n = 9). RESULTS: Differences in the tryptic peptide profiles of the 2 groups could clearly be detected, with these becoming even more evident in the more severe histological classes, even if this was not evident with routine histology. In particular, 4 putative proteins were detected and had a higher signal intensity within regions of DN tissue with extensive sclerosis or fibrosis. Among these, 2 proteins (PGRMC1 and CO3) had a signal intensity that increased at the latter stages of the disease and may be associated with progression. DISCUSSION/CONCLUSION: This preliminary study represents a valuable starting point for a future study employing a larger cohort of patients to develop sensitive and specific protein biomarkers that could reliably differentiate between diabetic and hypertensive causes of CKD to allow for improved diagnosis, fewer biopsy procedures, and refined treatment approaches for clinicians.

Comparison of Dietary Oils with Different Polyunsaturated Fatty Acid n-3 and n-6 Content in the Rat Model of Cutaneous Wound Healing.

Dietary supplementation with polyunsaturated fatty acids (PUFA) n-3 can affect cutaneous wound healing; however, recent findings demonstrate the variable extent of their influence on the quality of healing. Here, we compare the effect of several dietary oils, containing different levels of PUFA n-3 and PUFA n-6, on wound healing in the rat model. Rats were fed the feed mixture with 8% palm oil (P), safflower oil (S), fish oil (F) or Schizochytrium microalga extract (Sch) and compared to the animals fed by control feed mixture (C). Dorsal full-thickness cutaneous excisions were performed after 52 days of feeding and skin was left to heal for an additional 12 days. Histopathological analysis of skin wounds was performed, including immune cells immunolabeling and the determination of hydroxyproline amount as well as gene expression analyses of molecules contributing to different steps of the healing. Matrix-assisted-laser-desorption-ionization mass-spectrometry-imaging (MALDI-MSI) was used to determine the amount of collagen α-1(III) chain fragment in healing samples. Treatment by Schizochytrium extract resulted in decrease in the total wound area, in contrast to the safflower oil group where the size of the wound was larger when comparing to control animals. Diet with Schizochytrium extract and safflower oils displayed a tendency to increase the number of new vessels. The number of MPO-positive cells was diminished following any of oil treatment in comparison to the control, but their highest amount was found in animals with a fish oil diet. On the other hand, the number of CD68-positive macrophages was increased, with the most significant enhancement in the fish oil and safflower oil group. Hydroxyproline concentration was the highest in the safflower oil group but it was also enhanced in all other analyzed treatments in comparison to the control. MALDI-MSI signal intensity of a collagen III fragment decreased in the sequence C > S > Sch > P > F treatment. In conclusion, we observed differences in tissue response during healing between dietary oils, with the activation of inflammation observed following the treatment with oil containing high eicosapentaenoic acid (EPA) level (fish oil) and enhanced healing features were induced by the diet with high content of docosahexaenoic acid (DHA, Schizochytrium extract).

Tissue Distribution and Penetration of Isavuconazole at the Site of Infection in Experimental Invasive Aspergillosis in Mice with Underlying Chronic Granulomatous Disease.

Isavuconazole, the active moiety of the prodrug isavuconazonium sulfate, has potent activity against a wide spectrum of fungal pathogens and is approved for the treatment of invasive aspergillosis, yet little is known about the tissue penetration of isavuconazole at the target sites of infection. Here, we explored the spatial and quantitative distribution of isavuconazole in tissue lesions in experimental pulmonary aspergillosis established in mice with chronic granulomatous disease (CGD) (gp91phox-). Matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) and laser capture microdissection (LCM)-directed high-pressure liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) were used to analyze infected lungs and brain tissues collected 1, 3, 6, and 24 h after a single oral administration of the prodrug at a dose of 256 mg/kg of body weight (corresponding to 122.9 mg/kg of isavuconazole). Drug enrichment within granulomatous lesions was observed in lung tissue at 1 h postdose, although drug levels quickly equilibrated afterwards between lesion and nonlesion areas. A prominent antifungal effect in the infected lung tissue was revealed by histopathological analysis. Isavuconazole also penetrated into the brain with high efficiency. These data further support the value of isavuconazole to treat patients with invasive aspergillosis.

Complementarity of molecular and elemental mass spectrometric imaging of Gadovist™ in mouse tissues.

Drug biodistribution analyses can be considered a key issue in pharmaceutical discovery and development. Here, mass spectrometric imaging can be employed as a powerful tool to investigate distributions of drug compounds in biologically and medically relevant tissue sections. Both matrix-assisted laser desorption ionization-mass spectrometric imaging as molecular method and laser ablation inductively coupled plasma-mass spectrometric imaging as elemental detection method were applied to determine drug distributions in tissue thin sections. Several mouse organs including the heart, kidney, liver, and brain were analyzed with regard to distribution of Gadovist™, a gadolinium-based contrast agent already approved for clinical investigation. This work demonstrated the successful detection and localization of Gadovist™ in several organs. Furthermore, the results gave evidence that gadolinium-based contrast agents in general can be well analyzed by mass spectrometric imaging methods. In conclusion, the combined application of molecular and elemental mass spectrometry could complement each other and thus confirm analytical results or provide additional information.

Time-dependent regional brain distribution of methadone and naltrexone in the treatment of opioid addiction.

Opioid addiction is a serious public health concern with severe health and social implications; therefore, extensive therapeutic efforts are required to keep users drug free. The two main pharmacological interventions, in the treatment of addiction, involve management with methadone an mu (µ)-opioid agonist and treatment with naltrexone, µ-opioid, kappa (κ)-opioid and delta (δ)-opioid antagonist. MET and NAL are believed to help individuals to derive maximum benefit from treatment and undergo a full recovery. The aim of this study was to determine the localization and distribution of MET and NAL, over a 24-hour period in rodent brain, in order to investigate the differences in their respective regional brain distributions. This would provide a better understanding of the role of each individual drug in the treatment of addiction, especially NAL, whose efficacy is controversial. Tissue distribution was determined by using mass spectrometric imaging (MSI), in combination with quantification via liquid chromatography tandem mass spectrometry. MSI image analysis showed that MET was highly localized in the striatal and hippocampal regions, including the nucleus caudate, putamen and the upper cortex. NAL was distributed with high intensities in the mesocorticolimbic system including areas of the cortex, caudate putamen and ventral pallidum regions. Our results demonstrate that MET and NAL are highly localized in the brain regions with a high density of µ-receptors, the primary sites of heroin binding. These areas are strongly implicated in the development of addiction and are the major pathways that mediate brain stimulation during reward.

Anti-Myocardial Infarction Effects of Radix Aconiti Lateralis Preparata Extracts and Their Influence on Small Molecules in the Heart Using Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry Imaging.

Radix Aconiti Lateralis Preparata (fuzi) is the processed product of Aconitum carmichaelii Debeaux tuber, and has great potential anti-myocardial infarction effects, including improving myocardial damage and energy metabolism in rats. However, the effects of Radix Aconiti Lateralis Preparata extracts in a rat model of myocardial infarction have not yet been fully illustrated. Herein, Radix Aconiti Lateral Preparata was used to prepare Radix Aconiti Lateralis Preparata extract (RAE), fuzi polysaccharides (FPS), and fuzi total alkaloid (FTA). Then, we aimed to compare the effects of RAE, FPS, and FTA in MI rats and further explore their influence on small molecules in the heart. We reported that Radix Aconiti Lateralis Preparata extract (RAE) and fuzi total alkaloid (FTA) significantly improved left ventricular function and structure, and reduced myocardial damage and infarct size in rats with myocardial infarction by the left anterior descending artery ligation. In contrast, fuzi polysaccharides (FPS) was less effective than RAE and FTA, indicating that alkaloids might play a major role in the treatment of myocardial infarction. Moreover, via matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI), we further showed that RAE and FTA containing alkaloids as the main common components regulated myocardial energy metabolism-related molecules and phospholipids levels and distribution patterns against myocardial infarction. In particular, it was FTA, not RAE, that could also regulate potassium ions and glutamine to play a cardioprotective role in myocardial infarction, which revealed that an appropriate dose of alkaloids generated more obvious cardiotonic effects. These findings together suggested that Radix Aconiti Lateralis Preparata extracts containing an appropriate dose of alkaloids as its main pharmacological active components exerted protective effects against myocardial infarction by improving myocardial energy metabolism abnormalities and changing phospholipids levels and distribution patterns to stabilize the cardiomyocyte membrane structure. Thus, RAE and FTA extracted from Radix Aconiti Lateralis Preparata are potential candidates for the treatment of myocardial infarction.

Metabolomic insights into the effects of thyroid hormone on Rana [Lithobates] catesbeiana metamorphosis using whole-body Matrix Assisted Laser Desorption/Ionization-Mass Spectrometry Imaging (MALDI-MSI).

Anuran metamorphosis involves the transformation of an aquatic tadpole into a juvenile frog. This process is completely dependent upon thyroid hormones (THs). Although much research has been focused on changes in gene expression programs during this postembryonic developmental period, transitions in the metabolic profiles are relatively poorly understood. Matrix Assisted Laser Desorption/Ionization-Mass Spectrometry Imaging (MALDI-MSI) is a technique that generates highly multiplexed mass spectra while retaining spatial location information on a thin tissue section. Reconstructed ion heat maps are correlated with morphology of the tissue section for biological interpretation. The present study is the first to use whole-body MALDI-MSI on tadpoles to gain insights into anuran metamorphosis. Approximately 1000 features were detected in each of five tissues examined (brain, eye, liver, notochord, and tail muscle) from premetamorphic North American bullfrog (Rana [Lithobates] catesbeiana) tadpoles. Of these detected metabolites, 1700 were unique and 136 were significantly affected by exposure to 50 nM thyroxine for 48 h. Of the significantly-affected metabolites, 64 features were tentatively identified using the MassTRIX annotation tool. All tissues revealed changes in lipophilic compounds including phosphatidylcholines, phosphatidylinositols, phosphatidylglycerols, phosphatidylethanolamines, and phosphatidylserines. These lipophilic compounds made up the largest portion of significantly-affected metabolites indicating that lipid signaling is a major target of TH action in frog tadpoles.

Evaluation of non-supervised MALDI mass spectrometry imaging combined with microproteomics for glioma grade III classification.

An integrated diagnosis using molecular features is recommended in the 2016 World Health Organization (WHO) classification. Our aim was to explore non-targeted molecular classification using MALDI mass spectrometry imaging (MALDI MSI) associated to microproteomics in order to classify anaplastic glioma by integration of clinical data. We used fresh-frozen tissue sections to perform MALDI MSI of proteins based on their digestion peptides after in-situ trypsin digestion of the tissue sections and matrix deposition by micro-spraying. The generated 70µm spatial resolution image datasets were further processed by individual or global segmentation in order to cluster the tissues according to their molecular protein signature. The clustering gives 3 main distinct groups. Within the tissues the ROIs (regions of interest) defined by these groups were used for microproteomics by micro-extraction of the tryptic peptides after on-tissue enzymatic digestion. More than 2500 proteins including 22 alternative proteins (AltProt) are identified by the Shotgun microproteomics. Statistical analysis on the basis of the label free quantification of the proteins shows a similar classification to the MALDI MSI segmentation into 3 groups. Functional analysis performed on each group reveals sub-networks related to neoplasia for group 1, glioma with inflammation for group 2 and neurogenesis for group 3. This demonstrates the interest on these new non-targeted large molecular data combining both MALDI MSI and microproteomics data, for tumor classification. This analysis provides new insights into grade III glioma organization. This specific information could allow a more accurate classification of the biopsies according to the prognosis and the identification of potential new targeted therapeutic options. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann.

Matrix-assisted laser desorption/ionization mass spectrometry imaging for quorum sensing.

Quorum sensing (QS) is a complex communication system in bacteria, directing their response to the environment. QS is also one of the main regulators of bacterial biofilms' formation, maturation and dispersion. Matrix-assisted laser desorption ionization (MALDI) mass spectrometry imaging (MSI) is a molecular imaging technique that allows the mapping of QS molecules in bacterial biofilms. Here, we highlight the latest advances in MALDI-MSI in recent years and how this technology can improve QS understanding at the molecular level.

Integrated Mechanism of Immune Response Modulation by Arctium Lappa L. Fructans Based on Microbiome and Metabolomics Technologies.

Arctium lappa L. is widely consumed for its various biological effects, and polysaccharides are its main functional components. The present study aimed to evaluate the immunoregulatory effects of the main polysaccharides from burdock (ALP-1) and reveal the underlying mechanisms. ALP-1 consisted of fructose and glucose (14.57:1) and had a molecular weight of 2757 Da, with typical characteristics of (1 → 2)-linked linear fructans. Oral intake of ALP-1 significantly increased the number of colonic goblet cells, serum immunoglobulin A and immunoglobulin G levels, and fecal secretory immunoglobulin A content as well as up-regulated antioxidant enzymes and increased short chain fatty acid production. In addition, ALP-1 administration regulated pro/anti-inflammatory cytokines (i.e., interleukin (IL)-1ß, IL-6, tumor necrosis factor-α, interferon-γ, and IL-10), intestinal microbiota structure, and the spatial information on key metabolites. Some gut-microbiota-mediated metabolic processes were also significantly altered. These results indicated that ALP-1 could exert beneficial effects on immune responses and intestinal health in healthy mice.