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Background and Objectives Metabolomics studies have revealed transition points in metabolic signatures of red cells during storage in SAGM, whose clinical significance is unclear. We set out to investigate whether these transition points occur independent of storage media and define differences in the metabolism of red cells in additive solutions. Materials and Methods Red cell concentrates were stored in SAGM, AS-1, AS-3 or PAGGSM, and sampled fourteen times spanning Day 1-46. Following quality control, the samples were split into extracellular and intracellular aliquots. These were analysed with ultra-high-performance liquid chromatography coupled to mass spectrometry analysis affording quantitative metabolic profiles of both intra- and extracellular red cell metabolites. Results Differences were observed in glycolysis, purine salvage, glutathione synthesis and citrate metabolism on account of the storage solutions. Donor variability however hindered the accurate characterization of metabolic transition time-points. Intracellular citrate concentrations were increased in red cells stored in AS-3 and PAGGSM media. The metabolism of citrate in red cells in SAGM was subsequently confirmed using 13C citrate isotope labelling and shown to originate from citrate anticoagulant. Conclusion Metabolic signatures that discriminate between 'fresh' and 'old' stored red cells are dependent upon additive solutions. Specifically, the incorporation and metabolism of citrate in additive solutions with lower chloride ion concentration is altered and impacts glycolysis. [ABSTRACT FROM AUTHOR]

Background. This study aims to evaluate the effects of HYADD 4-G, a not chemically cross-linked HA (Hyaluronic Acid) derivative, and hylan G-F 20, a cross-linked HA product, on synovial matrix metalloproteinases and cytokines in patients suffering from knee osteoarthritis. Methods. 31 patients were randomized to receive HYADD 4-G or hylan G-F 20. Synovial fluid was collected before and one week after the first injection. Activity of MMP-3, MMP-2, MMP-13, IL-6 and other cytokines were measured. Changes in synovial fluid neutrophils and lymphocytes were analyzed. The VAS, the WOMAC questionnaire, and a Physician Global Assessment (PhGA VAS) were recorded. Results. A trend towards a greater decrease in MMP-3 activity was observed in the Hymovis® group. Active MMP-2 and MMP-13 decreased in both groups. IL-6 levels also decreased significantly in both groups. Median change in neutrophils from baseline was significantly different between the treatment groups. No differences between the treatment groups were observed for the WOMAC, VAS and PhGA VAS scores. No serious adverse events were reported. Conclusions. These findings demonstrate that intra-articular HA injections in patients with knee osteoarthritis tend to protect cartilage structural integrity reducing the activity of key proteolytic enzymes implicated in cartilage degradation as well as inflammatory mediators. [ABSTRACT FROM AUTHOR]

Supplemental oxygen (hyperoxia) improves physical performance during hypoxic exercise. Based on the analysis of metabolome and iron homeostasis from human athlete blood samples, we show that hyperoxia during recovery periods interferes with metabolic alterations following hypoxic exercise. This may impair beneficial adaptations to exercise and/or hypoxia and highlights risks of oxygen supplementation in hypoxia., Competing Interests: Declaration of competing interest The authors declare no competing interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: PI4KA-related disorder is a highly clinically variable condition characterized by neurological (limb spasticity, developmental delay, intellectual disability, seizures, ataxia, nystagmus) and gastrointestinal (inflammatory bowel disease and multiple intestinal atresia) manifestations. Although features consistent with immunodeficiency (autoimmunity/autoinflammation and recurrent infections) have been reported in a subset of patients, the burden of B-cell deficiency and hypogammaglobulinemia has not been extensively investigated. We sought to describe the clinical presentation and manifestations of patients with PI4KA-related disorder and to investigate the metabolic consequences of biallelic PI4KA variants in B cells., Methods: Clinical data from patients with PI4KA variants were obtained. Multi-omics analyses combining transcriptome, proteome, lipidome and metabolome analyses in conjunction with functional assays were performed in EBV-transformed B cells., Results: Clinical and laboratory data of 13 patients were collected. Recurrent infections (7/13), autoimmune/autoinflammatory manifestations (5/13), B-cell deficiency (8/13) and hypogammaglobulinemia (8/13) were frequently observed. Patients' B cells frequently showed increased transitional and decreased switched memory B-cell subsets. Pathway analyses based on differentially expressed transcripts and proteins confirmed the central role of PI4KA in B cell differentiation with altered B-cell receptor (BCR) complex and signalling. By altering lipids production and tricarboxylic acid cycle regulation, and causing increased endoplasmic reticulum stress, biallelic PI4KA mutations disrupt B cell metabolism inducing mitochondrial dysfunction. As a result, B cells show hyperactive PI3K/mTOR pathway, increased autophagy and deranged cytoskeleton organization., Conclusion: By altering lipid metabolism and TCA cycle, impairing mitochondrial activity, hyperactivating mTOR pathway and increasing autophagy, PI4KA-related disorder causes a syndromic inborn error of immunity presenting with B-cell deficiency and hypogammaglobulinemia., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: Autoimmune encephalitis includes a heterogeneous group of rare and complex diseases, usually presenting with severe and disabling symptoms, such as behavioral changes, cognitive deficits, and seizures., Method: This report presents the case of a 26-year-old man who was diagnosed with autoimmune encephalitis following SARS-CoV-2 vaccination (<40 days). Symptoms first appeared in February 2022 with a temporal seizure, associated with confusion and memory loss. Psychiatric manifestations such as disorientation and altered thought contents emerged soon after., Results: Neuroimaging testing showed signs of hypometabolism in occipital, prefrontal, and temporal regions, whereas an extensive neuropsychological assessment revealed the presence of multiple alterations in memory, executive, and visuoconstructive processes., Conclusions: In this case, a combination of neuroimaging testing, psychiatric evaluation, and neuropsychological assessment provided evidence for a diagnosis of autoimmune encephalitis post-vaccination. Early recognition is essential in order to prevent clinical progression; avoid intractable epilepsy, brain atrophy, and cognitive impairment; and improve prognosis., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objective: Treatment-resistant depression (TRD) affects around 20-30% of people with major depressive disorder. In 2019, esketamine nasal spray was approved for TRD by both the US Food and Drug Administration and the European Medicines Agency. While its clinical efficacy and safety are proven, the mechanisms underlying its antidepressant effect remain unclear. The use of metabolomics may allow understanding the metabolic effects of esketamine and predicting biological features associated with clinical response in TRD. Nonetheless, there is a lack of studies exploring the predictive value of metabolomics. The Resistant Depression Response to Esketamine Assessing Metabolomics (ReDREAM) project aims at identifying metabolic biosignatures that may represent novel correlates of response to esketamine treatment., Study Design: This is the protocol of an observational, prospective study., Methods: We plan to select 60 people with TRD from 3 clinical sites in Italy. The participants will be administered with esketamine nasal spray, following standard clinical practice, twice a week for 4 weeks ("induction phase"), then once a week for 4 additional weeks ("maintenance phase"). We will test the correlations between baseline metabolic profile and depressive symptom improvement at study endpoints (weeks 4 and 8) and we will explore the likelihood of different metabolic phenotypes between responders and non-responders., Expected Results: An involvement of energy metabolism, amino acid metabolism, urea cycle, and nitric oxide synthesis in response to treatment with esketamine nasal spray is hypothesized., Conclusion: Unbiased data from untargeted metabolomics associated with clinical changes after esketamine treatment may contribute to define new paradigms for precision psychiatry-oriented, personalized care of TRD., Competing Interests: Declaration of Interests: Francesco Bartoli is Editor-in-Chief of Alpha Psychiatry and was not involved in the processing of this article. In the last three years, he has received consultant fees by IQVIA Solutions Italy and Edra S.p.A., and carried out paid editorial activities for Elsevier and AVES. In the last three years Renato de Filippis has received speaker fee from Janssen, Angelini, Lundbeck, and Rovi, and travel support from Rovi, Otsuka, Lundbeck, and Janssen. In the last three years Umberto Albert received speaker fee from Angelini Pharma, Janssen Pharmaceutica, OM Pharma Suisse, Lundbeck, Fidia, and Boehringer. The other authors declare no conflicts of interest., (2024 authors.)

The antidepressant effects of ketamine and esketamine are well-documented. Nonetheless, most of the underlying molecular mechanisms have to be uncovered yet. In the last decade, metabolomics has emerged as a useful means to investigate the metabolic phenotype associated with depression as well as changes induced by antidepressant treatments. This mini-review aims at summarizing the main findings from preclinical and clinical studies that used metabolomics to investigate the metabolic effects of subanesthetic, antidepressant doses of ketamine and esketamine and their relationship with clinical response. Both animal and human studies report alterations in several metabolic pathways - including the tricarboxylic acid cycle, glycolysis, the pentose phosphate pathway, lipid metabolism, amino acid metabolism, the kynurenine pathway, and the urea cycle - following the administration of ketamine or its enantiomers. Although more research is needed to clarify commonalities and differences in molecular mechanisms of action between the racemic compound and its enantiomers, these findings comprehensively support an influence of ketamine and esketamine on mitochondrial and cellular energy production, membrane homeostasis, neurotransmission, and signaling. Metabolomics may thus represent a promising strategy to clarify molecular mechanisms underlying treatment-resistant depression and related markers of clinical response to ketamine and esketamine. This body of preclinical and clinical evidence, if further substantiated, has the potential to guide clinicians towards personalized approaches, contributing to new paradigms in the clinical management of depression., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Blood serves as the primary global biological matrix for health surveillance, disease diagnosis, and response to drug treatment, holding significant promise for personalized medicine. The diverse array of lipids and metabolites in the blood provides a snapshot of both physiological and pathological processes, with many routinely monitored during conventional wellness checks. The conventional method involves intravenous blood collection, extracting a few milliliters via venipuncture, a technique limited to clinical settings due to its dependence on trained personnel. Microsampling methods have evolved to be less invasive (collecting ≤150 µL of capillary blood), user-friendly (enabling self-collection), and suitable for remote collection in longitudinal studies. Dried blood spot (DBS), a pioneering microsampling technique, dominates clinical and research domains. Recent advancements in device technology address critical limitations of classical DBS, specifically variations in hematocrit and volume. This review presents a comprehensive overview of state-of-the-art microsampling devices, emphasizing their applications and potential for monitoring metabolites and lipids in blood. The scope extends to diverse areas, encompassing population studies, nutritional investigations, drug discovery, sports medicine, and multi-omics research.

The protein disulfide isomerase A3 (PDIA3) is directly or indirectly involved in various physiopathological processes and participates in cancer initiation, progression and chemosensitivity. However, little is known about its involvement in glioblastoma. To obtain specific information, we performed cellular experiments in the T98G and U-87 MG glioblastoma cell lines to evaluate the role of PDIA3. The loss of PDIA3 functions, either through inhibition or silencing, reduced glioblastoma cells spreading by triggering cytotoxic phenomena. PDIA3 inhibition led to a redistribution of PDIA3, resulting in the formation of protein aggregates visualized through immunofluorescence staining. Concurrently, cell cycle progression underwent arrest at the G 1 /S checkpoint. After PDIA3 inhibition, ROS-independent DNA damage and the activation of the repair system occurred, as evidenced by the phosphorylation of H2A.X and the overexpression of the Ku70 protein. We also demonstrated through a clonogenic assay that PDIA3 inhibition could increase the chemosensitivity of T98G and U-87 MG cells to the approved glioblastoma drug temozolomide (TMZ). Overall, PDIA3 inhibition induced cytotoxic effects in the analyzed glioblastoma cell lines. Although further in vivo studies are needed, the results suggested PDIA3 as a novel therapeutic target that could also be included in already approved therapies.

Triple-negative breast cancer (TNBC) is a very aggressive disease even in its early stages and is characterized by a severe prognosis. Neoadjuvant chemotherapy is one of the milestones of treatment, and paclitaxel (PTX) is among the most active drugs used in this setting. However, despite its efficacy, peripheral neuropathy occurs in approximately 20-25% of cases and represents the dose-limiting toxicity of this drug. New deliverable strategies to ameliorate drug delivery and reduce side effects are keenly awaited to improve patients' outcomes. Mesenchymal stromal cells (MSCs) have recently been demonstrated as promising drug delivery vectors for cancer treatment. The aim of the present preclinical study is to explore the possibility of a cell therapy approach based on the use of MSCs loaded with PTX to treat TNBC-affected patients. For this purpose, we in vitro evaluated the viability, migration and colony formation of two TNBC cell lines, namely, MDA-MB-231 and BT549, treated with MSC-PTX conditioned medium (MSC-CM PTX) in comparison with both CM of MSCs not loaded with PTX (CTRL) and free PTX. We observed stronger inhibitory effects on survival, migration and tumorigenicity for MSC-CM PTX than for CTRL and free PTX in TNBC cell lines. Further studies will provide more information about activity and potentially open the possibility of using this new drug delivery vector in the context of a clinical study.

Lipid-lowering therapies are widely used to prevent the development of atherosclerotic cardiovascular disease (ASCVD) and related mortality worldwide. "Omics" technologies have been successfully applied in recent decades to investigate the mechanisms of action of these drugs, their pleiotropic effects, and their side effects, aiming to identify novel targets for future personalized medicine with an improvement of the efficacy and safety associated with the treatment. Pharmacometabolomics is a branch of metabolomics that is focused on the study of drug effects on metabolic pathways that are implicated in the variation of response to the treatment considering also the influences from a specific disease, environment, and concomitant pharmacological therapies. In this review, we summarized the most significant metabolomic studies on the effects of lipid-lowering therapies, including the most commonly used statins and fibrates to novel drugs or nutraceutical approaches. The integration of pharmacometabolomics data with the information obtained from the other "omics" approaches could help in the comprehension of the biological mechanisms underlying the use of lipid-lowering drugs in view of defining a precision medicine to improve the efficacy and reduce the side effects associated with the treatment.

In the present study, we used a mouse model of Alzheimer's disease (AD) (3×Tg-AD mice) to longitudinally analyse the expression level of PDIA3, a protein disulfide isomerase and endoplasmic reticulum (ER) chaperone, in selected brain limbic areas strongly affected by AD-pathology (amygdala, entorhinal cortex, dorsal and ventral hippocampus). Our results suggest that, while in Non-Tg mice PDIA3 levels gradually reduce with aging in all brain regions analyzed, 3×Tg-AD mice showed an age-dependent increase in PDIA3 levels in the amygdala, entorhinal cortex, and ventral hippocampus. A significant reduction of PDIA3 was observed in 3×Tg-AD mice already at 6 months of age, as compared to age-matched Non-Tg mice. A comparative immunohistochemistry analysis performed on 3×Tg-AD mice at 6 (mild AD-like pathology) and 18 (severe AD-like pathology) months of age showed a direct correlation between the cellular level of Aβ and PDIA3 proteins in all the brain regions analysed, even if with different magnitudes. Additionally, an immunohistochemistry analysis showed the presence of PDIA3 in all post-mitotic neurons and astrocytes. Overall, altered PDIA3 levels appear to be age- and/or pathology-dependent, corroborating the ER chaperone's involvement in AD pathology, and supporting the PDIA3 protein as a potential novel therapeutic target for the treatment of AD.

Volumetric absorptive microsampling (VAMS) is a recently developed sample collection method that enables single-drop blood collection in a minimally invasive manner. Blood biomolecules can then be extracted and processed for analysis using several analytical platforms. The integration of VAMS with conventional mass spectrometry (MS)-based metabolomics approaches is an attractive solution for human studies representing a less-invasive procedure compared to phlebotomy with the additional potential for remote sample collection. However, as we recently demonstrated, VAMS samples require long-term storage at -80 °C. This study investigated the stability of VAMS samples during short-term storage and compared the metabolome obtained from capillary blood collected from the fingertip to those of plasma and venous blood from 22 healthy volunteers. Our results suggest that the blood metabolome collected by VAMS samples is stable at room temperature only for up to 6 h requiring subsequent storage at -80 °C to avoid significant changes in the metabolome. We also demonstrated that capillary blood provides better coverage of the metabolome compared to plasma enabling the analysis of several intracellular metabolites presented in red blood cells. Finally, this work demonstrates that with the appropriate pre-analytical protocol capillary blood can be successfully used for untargeted metabolomics studies.

The European Food Safety Authority asked for novel approaches for identifying mechanically separated meat (MSM) in meat products, due to food safety concern. In this study, a novel approach based on multivariate analysis of 43 trace elements in meat products is described. Overall, 27 trace elements and 16 rare earth elements were determined by using ICP-MS after sample mineralization of 100 meat samples, composed of different percentages of MSM, obtained at low and high pressure, and without MSM. After development and optimization, the multivariate approach was validated by analyzing and then classifying 10 "blind" meat samples, obtaining method accuracy equal to 90%. Thus, the applicability of this new analytical approach was demonstrated. The method represents a significant improvement for this type of determination, especially when MSM is obtained at low pressure, since this product is characterized by chemical characteristics very similar to fresh meat., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Dual functionalized liposomes were developed to cross the blood−brain barrier (BBB) and to release their cargo in a pathological matrix metalloproteinase (MMP)-rich microenvironment. Liposomes were surface-functionalized with a modified peptide deriving from the receptor-binding domain of apolipoprotein E (mApoE), known to promote cargo delivery to the brain across the BBB in vitro and in vivo; and with an MMP-sensitive moiety for an MMP-triggered drug release. Different MMP-sensitive peptides were functionalized at both ends with hydrophobic stearate tails to yield MMP-sensitive lipopeptides (MSLPs), which were assembled into mApoE liposomes. The resulting bi-functional liposomes (i) displayed a < 180 nm diameter with a negative ζ-potential; (ii) were able to cross an in vitro BBB model with an endothelial permeability of 3 ± 1 × 10−5 cm/min; (iii) when exposed to functional MMP2 or 9, efficiently released an encapsulated fluorescein dye; (iv) showed high biocompatibility when tested in neuronal cultures; and (v) when loaded with glibenclamide, a drug candidate with poor aqueous solubility, reduced the release of proinflammatory cytokines from activated microglial cells.

Mass spectrometry imaging (MSI) is an emerging technology that is capable of mapping various biomolecules within their native spatial context, and performing spatial multiomics on formalin-fixed paraffin-embedded (FFPE) tissues may further increase the molecular characterization of pathological states. Here we present a novel workflow which enables the sequential MSI of lipids, N-glycans, and tryptic peptides on a single FFPE tissue section and highlight the enhanced molecular characterization that is offered by combining the multiple spatial omics data sets. In murine brain and clear cell renal cell carcinoma (ccRCC) tissue, the three molecular levels provided complementary information and characterized different histological regions. Moreover, when the spatial omics data was integrated, the different histopathological regions of the ccRCC tissue could be better discriminated with respect to the imaging data set of any single omics class. Taken together, these promising findings demonstrate the capability to more comprehensively map the molecular complexity within pathological tissue.

Researchers worldwide are taking advantage of novel, commercially available, technologies, such as ion mobility mass spectrometry (IM-MS), for metabolomics and lipidomics applications in a variety of fields including life, biomedical, and food sciences. IM-MS provides three main technical advantages over traditional LC-MS workflows. Firstly, in addition to mass, IM-MS allows collision cross-section values to be measured for metabolites and lipids, a physicochemical identifier related to the chemical shape of an analyte that increases the confidence of identification. Second, IM-MS increases peak capacity and the signal-to-noise, improving fingerprinting as well as quantification, and better defining the spatial localization of metabolites and lipids in biological and food samples. Third, IM-MS can be coupled with various fragmentation modes, adding new tools to improve structural characterization and molecular annotation. Here, we review the state-of-the-art in IM-MS technologies and approaches utilized to support metabolomics and lipidomics applications and we assess the challenges and opportunities in this growing field., (© 2021 John Wiley & Sons Ltd.)

Arrhythmogenic cardiomyopathy (ACM) is a genetic disease associated with sudden cardiac death and cardiac fibro-fatty replacement. Over the last years, several works have demonstrated that different epigenetic enzymes can affect not only gene expression changes in cardiac diseases but also cellular metabolism. Specifically, the histone acetyltransferase GCN5 is known to facilitate adipogenesis and modulate cardiac metabolism in heart failure. Our group previously demonstrated that human primary cardiac stromal cells (CStCs) contribute to adipogenesis in the ACM pathology. Thus, this study aims to evaluate the role of GCN5 in ACM intracellular lipid accumulation. To do so, CStCs were obtained from right ventricle biopsies of ACM patients and from samples of healthy cadaveric donors (CTR). GCN5 expression was increased both in ex vivo and in vitro ACM samples compared to CTR. When GCN5 expression was silenced or pharmacologically inhibited by the administration of MB-3, we observed a reduction in lipid accumulation and a mitigation of reactive oxygen species (ROS) production in ACM CStCs. In agreement, transcriptome analysis revealed that the presence of MB-3 modified the expression of pathways related to cellular redox balance. Altogether, our findings suggest that GCN5 inhibition reduces fat accumulation in ACM CStCs, partially by modulating intracellular redox balance pathways., (© 2022 EURAC Research. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Metabolites are intermediates or end products of biochemical processes involved in both health and disease. Here, we take advantage of the well-characterized Cooperative Health Research in South Tyrol (CHRIS) study to perform an exome-wide association study (ExWAS) on absolute concentrations of 175 metabolites in 3294 individuals. To increase power, we imputed the identified variants into an additional 2211 genotyped individuals of CHRIS. In the resulting dataset of 5505 individuals, we identified 85 single-variant genetic associations, of which 39 have not been reported previously. Fifteen associations emerged at ten variants with >5-fold enrichment in CHRIS compared to non-Finnish Europeans reported in the gnomAD database. For example, the CHRIS-enriched ETFDH stop gain variant p.Trp286Ter (rs1235904433-hexanoylcarnitine) and the MCCC2 stop lost variant p.Ter564GlnextTer3 (rs751970792-carnitine) have been found in patients with glutaric acidemia type II and 3-methylcrotonylglycinuria, respectively, but the loci have not been associated with the respective metabolites in a genome-wide association study (GWAS) previously. We further identified three gene-trait associations, where multiple rare variants contribute to the signal. These results not only provide further evidence for previously described associations, but also describe novel genes and mechanisms for diseases and disease-related traits.

The aim of this study was to determine alterations of the metabolome in blood plasma in response to concentric-eccentric leg exercise performed at a simulated altitude of 3,500 m. To do so, we recruited 11 well-trained subjects and performed an untargeted metabolomics analysis of plasma samples obtained before, 20 min after as well as on day 8 after five sets of maximal, concentric-eccentric leg exercises that lasted 90 s each. We identified and annotated 115 metabolites through untargeted liquid chromatography-mass spectrometry metabolomics and used them to further calculate 20 sum/ratio of metabolites. A principal component analysis (PCA) revealed differences in-between the overall metabolome at rest and immediately after exercise. Interestingly, some systematic changes of relative metabolite concentrations still persisted on day 8 after exercise. The first two components of the PCA explained 34% of the relative concentrations of all identified metabolites analyzed together. A volcano plot indicates that 35 metabolites and two metabolite ratios were significantly changed directly after exercise, such as metabolites related to carbohydrate and TCA metabolism. Moreover, we observed alterations in the relative concentrations of amino acids (e.g., decreases of valine, leucine and increases in alanine) and purines (e.g., increases in hypoxanthine, xanthine and uric acid). In summary, high intensity concentric-eccentric exercise performed at simulated altitude systematically changed the blood metabolome in trained athletes directly after exercise and some relative metabolite concentrations were still changed on day 8. The importance of that persisting metabolic alterations on exercise performance should be studied further., Competing Interests: WS was employed by Tirol-Kliniken GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Dünnwald, Paglia, Weiss, Denti, Faulhaber, Schobersberger and Wackerhage.)

Neural tissue has high metabolic requirements. Following spinal cord injury (SCI), the damaged tissue suffers from a severe metabolic impairment, which aggravates axonal degeneration and neuronal loss. Impaired cellular energetic, tricarboxylic acid (TCA) cycle and oxidative phosphorylation metabolism in neuronal cells has been demonstrated to be a major cause of neural tissue death and regeneration failure following SCI. Therefore, rewiring the spinal cord cell metabolism may be an innovative therapeutic strategy for the treatment of SCI. In this study, we evaluated the therapeutic effect of the recovery of oxidative metabolism in a mouse model of severe contusive SCI. Oral administration of TCA cycle intermediates, co-factors, essential amino acids, and branched-chain amino acids was started 3 days post-injury and continued until the end of the experimental procedures. Metabolomic, immunohistological, and biochemical analyses were performed on the injured spinal cord sections. Administration of metabolic precursors enhanced spinal cord oxidative metabolism. In line with this metabolic shift, we observed the activation of the mTORC1 anabolic pathway, the increase in mitochondrial mass, and ROS defense which effectively prevented the injury-induced neural cell apoptosis in treated animals. Consistently, we found more choline acetyltransferase (ChAT)-expressing motor neurons and increased neurofilament-positive corticospinal axons in the spinal cord parenchyma of the treated mice. Interestingly, oral administration of the metabolic precursors increased the number of activated microglia expressing the CD206 marker suggestive of a pro-resolutive, M2-like phenotype. These molecular and histological modifications observed in treated animals ultimately led to a significant, although partial, improvement of the motor functions. Our data demonstrate that rewiring the cellular metabolism can represent an effective strategy to treat SCI., (Copyright © 2022. Published by Elsevier Ltd.)

The ERp57/PDIA3 protein is a pleiotropic member of the PDIs family and, although predominantly located in the endoplasmic reticulum (ER), has indeed been found in other cellular compartments, such as the nucleus or the cell membrane. ERp57/PDIA3 is an important research target considering it can be found in various subcellular locations. This protein is involved in many different physiological and pathological processes, and our review describes new data on its functions and summarizes some ligands identified as PDIA3-specific inhibitors., (© 2022. The Author(s).)

Objectives: This study aimed to determine the prevalence of ultrasound-detected tendon abnormalities in healthy subjects (HS) across the age range., Methods: Adult HS (age 18-80 years) were recruited in 23 international Outcome Measures in Rheumatology ultrasound centres and were clinically assessed to exclude inflammatory diseases or overt osteoarthritis before undergoing a bilateral ultrasound examination of digit flexors (DFs) 1-5 and extensor carpi ulnaris (ECU) tendons to detect the presence of tenosynovial hypertrophy (TSH), tenosynovial power Doppler (TPD) and tenosynovial effusion (TEF), usually considered ultrasound signs of inflammatory diseases. A comparison cohort of patients with rheumatoid arthritis (RA) was taken from the Birmingham Early Arthritis early arthritis inception cohort., Results: 939 HS and 144 patients with RA were included. The majority of HS (85%) had grade 0 for TSH, TPD and TEF in all DF and ECU tendons examined. There was a statistically significant difference in the proportion of TSH and TPD involvement between HS and subjects with RA (HS vs RA p<0.001). In HS, there was no difference in the presence of ultrasound abnormalities between age groups., Conclusions: Ultrasound-detected TSH and TPD abnormalities are rare in HS and can be regarded as markers of active inflammatory disease, especially in newly presenting RA., Competing Interests: Competing interests: There are no declared competing interests from authors except the following: CC declared grants from NIHR Versus Arthritis, Lilly sponsored EULAR conference travel, Modern Biosciences payment as DSM committee member, Roche consultancy fee and Novartis sponsored writing of one medical paper. KI declared a Mitsubishi-Tanabe research grant for RA; Abbvie, Eli Lilly, Mitsubishi-Tanabe, Bristol-Myers-Squib and Novartis speaker’s fees; and participation on a DSM board for Abbvie, Eli Lilly and Mitsubishi-Tanabe. RK declared support from Abbvie, Roche, Novartis and UCB with payments for travel to meetings/lectures, presentations, speakers’ bureaus, manuscript writing/educational events., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY. Published by BMJ.)

Mass spectrometry (MS)-based metabolomics approaches have been used for characterizing the metabolite content and composition of biological samples in drug discovery and development, as well as in metabolic engineering, and food and plant sciences applications. Here, we describe a protocol routinely used in our laboratory to conduct a metabolic profiling of small polar metabolites from biological samples. Metabolites can be extracted from each sample using a methanol-based single-phase extraction procedure. The combination of LC-based hydrophilic interaction liquid chromatography (HILIC) and a hybrid quadrupole-time of flight (Q-ToF) mass spectrometer allows the comprehensive analysis of small polar metabolites including sugars, phosphorylated compounds, purines and pyrimidines, nucleotides, nucleosides, acylcarnitines, carboxylic acids, hydrophilic vitamins and amino acids. Retention times and accurate masses of metabolites involved in key metabolic pathways are annotated for routine high-throughput screening in both untargeted and targeted metabolomics analyses., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

The β-isomer of hexachlorocyclohexane (β-HCH) is a globally widespread pollutant that embodies all the physicochemical characteristics of organochlorine pesticides, constituting an environmental risk factor for a wide range of noncommunicable diseases. Previous in vitro studies from our group disclosed the carcinogenic potential of β-HCH, which contributes to neoplastic transformation by means of multifaceted intracellular mechanisms. Considering the positive evidence regarding the protective role of natural bioactive compounds against pollution-induced toxicity, micronutrients from olive and tomato endowed with the capability of modulating β-HCH cellular targets were tested. For this purpose, the solution obtained from a patented food supplement (No. EP2851080A1), referred to as Tomato and Olive Bioactive Compounds (TOBC), was administered to the androgen-sensitive prostate cancer cells LNCaP and different biochemical and cellular assays were performed to evaluate its efficiency. TOBC shows a dose-dependent significant chemoprotection by contrasting β-HCH-induced intracellular responses such as STAT3 and AhR activation, disruption of AR signaling, antiapoptotic and proliferative activity, and increase in ROS production and DNA damage. These experimental outcomes identified TOBC as a suitable functional food to be included in a diet regimen aimed at defending cells from β-HCH negative effects, recommending the development of tailored enriched formulations for exposed individuals.

In a previous work, it was shown that punicalagin, an active ingredient of pomegranate, is able to bind to PDIA3 and inhibit its disulfide reductase activity. Here we provide evidence that punicalagin can also bind to PDIA1, the main expressed form of protein disulfide isomerase (PDI). In this comparative study, the affinity and the effect of punicalagin binding on each protein were evaluated, and a computational approach was used to identify putative binding sites. Punicalagin binds to either PDIA1 or PDIA3 with a similar affinity, but the inhibition efficacy on protein reductase activity is higher for PDIA3. Additionally, punicalagin differently affects the thermal denaturation profile of both proteins. Molecular docking and molecular dynamics simulations led to propose a punicalagin binding mode on PDIA1 and PDIA3, identifying the binding sites at the redox domains a' in two different pockets, suggesting different effects of punicalagin on proteins' structure. This study provides insights to develop punicalagin-based ligands, to set up a rational design for PDIA3 selective inhibitors, and to dissect the molecular determinant to modulate the protein activity.

Predicting the prognosis of colorectal cancer (CRC) patients remains challenging and a characterisation of the tumour immune environment represents one of the most crucial avenues when attempting to do so. For this reason, molecular approaches which are capable of classifying the immune environments associated with tumour infiltrating lymphocytes (TILs) are being readily investigated. In this proof of concept study, we aim to explore the feasibility of using spatial lipidomics by MALDI-MSI to distinguish CRC tissue based upon their TIL content. Formalin-fixed paraffin-embedded tissue from human thymus and tonsil was first analysed by MALDI-MSI to obtain a curated mass list from a pool of single positive T lymphocytes, whose putative identities were annotated using an LC-MS-based lipidomic approach. A CRC tissue microarray (TMA, n = 30) was then investigated to determine whether these cases could be distinguished based upon their TIL content in the tumour and its microenvironment. MALDI-MSI from the pool of mature T lymphocytes resulted in the generation of a curated mass list containing 18 annotated m / z features. Initially, subsets of T lymphocytes were then distinguished based on their state of maturation and differentiation in the human thymus and tonsil tissue. Then, when applied to a CRC TMA containing differing amounts of T lymphocyte infiltration, those cases with a high TIL content were distinguishable from those with a lower TIL content, especially within the tumour microenvironment, with three lipid signals being shown to have the greatest impact on this separation ( p < 0.05). On the whole, this preliminary study represents a promising starting point and suggests that a lipidomics MALDI-MSI approach could be a promising tool for subtyping the diverse immune environments in CRC.

Recently, 1-DNJ has been widely studied by scientists for its capacity to inhibit α-glucosidase and reduce postprandial blood glucose and fat accumulation. To the best of our knowledge, this is the first analytical determination of 1-DNJ in Morus sp.pl. leaves carried out on Italian crops, and it could be used as a reference to assess the quality of the plant material in comparison to Far Eastern Asia cultivations. The effects of two thermal treatments were compared to test the incidence of the drying process on the 1-DNJ extractability. In addition, two harvesting seasons in the same year (2017) and two subsequent harvesting years (2017-2018) were considered. The amount of 1-DNJ herein found was comparable to that reported in the scientific literature for Asian cultivations. The increase in 1-DNJ along the summer and the higher level of this compound in the apical leaves also complies with previous findings. However, a strong implication for the climatic conditions in the different years and a significant interaction between climate and genotypes suggest exploring very carefully the agronomic practices and selecting cultivars according to different environmental conditions with a view to standardize the 1-DNJ amount in leaves.

Organochlorine pesticides constitute the majority of the total environmental pollutants, and a wide range of compounds have been found to be carcinogenic to humans. Among all, growing interest has been focused on β-hexachlorocyclohexane (β-HCH), virtually the most hazardous and, at the same time, the most poorly investigated member of the hexachlorocyclohexane family. Considering the multifaceted biochemical activities of β-HCH, already established in our previous studies, the aim of this work is to assess whether β-HCH could also trigger cellular malignant transformation toward cancer development. For this purpose, experiments were performed on the human normal bronchial epithelium cell line BEAS-2B exposed to 10 µM β-HCH. The obtained results strongly support the carcinogenic potential of β-HCH, which is achieved through both non-genotoxic (activation of oncogenic signaling pathways and proliferative activity) and indirect genotoxic (ROS production and DNA damage) mechanisms that significantly affect cellular macroscopic characteristics and functions such as cell morphology, cell cycle profile, and apoptosis. Taking all these elements into account, the presented study provides important elements to further characterize β-HCH, which appears to be a full-fledged carcinogenic agent.

Arrhythmogenic cardiomyopathy (ACM) is a genetic-based cardiac disease accompanied by severe ventricular arrhythmias and a progressive substitution of the myocardium with fibro-fatty tissue. ACM is often associated with sudden cardiac death. Due to the reduced penetrance and variable expressivity, the presence of a genetic defect is not conclusive, thus complicating the diagnosis of ACM. Recent studies on human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs) obtained from ACM individuals showed a dysregulated metabolic status, leading to the hypothesis that ACM pathology is characterized by an impairment in the energy metabolism. However, despite efforts having been made for the identification of ACM specific biomarkers, there is still a substantial lack of information regarding the whole metabolomic profile of ACM patients. The aim of the present study was to investigate the metabolic profiles of ACM patients compared to healthy controls (CTRLs). The targeted Biocrates AbsoluteIDQ ® p180 assay was used on plasma samples. Our analysis showed that ACM patients have a different metabolome compared to CTRLs, and that the pathways mainly affected include tryptophan metabolism, arginine and proline metabolism and beta oxidation of fatty acids. Altogether, our data indicated that the plasma metabolomes of arrhythmogenic cardiomyopathy patients show signs of endothelium damage and impaired nitric oxide (NO), fat, and energy metabolism.

Organochlorine pesticides (OCPs) belong to a heterogeneous class of organic compounds blacklisted by the Stockholm Convention in 2009 due to their harmful impact on human health. Among OCPs, β-hexachlorocyclohexane (β-HCH) is one of the most widespread and, at the same time, poorly studied environmental contaminant. Due to its physicochemical properties, β-HCH is the most hazardous of all HCH isomers; therefore, clarifying the mechanisms underlying its molecular action could provide further elements to draw the biochemical profile of this OCP. For this purpose, LNCaP and HepG2 cell lines were used as models and were subjected to immunoblot, immunofluorescence, and RT-qPCR analysis to follow the expression and mRNA levels, together with the distribution, of key biomolecules involved in the intracellular responses to β-HCH. In parallel, variations in redox homeostasis and cellular bioenergetic profile were monitored to have a complete overview of β-HCH effects. Obtained results strongly support the hypothesis that β-HCH could be an endocrine disrupting chemical as well as an activator of AhR signaling, promoting the establishment of an oxidative stress condition and a cellular metabolic shift toward aerobic glycolysis. In this altered context, β-HCH can also induce DNA damage through H2AX phosphorylation, demonstrating its multifaceted mechanisms of action.

Formalin-fixed paraffin-embedded (FFPE) tissue represents the primary source of clinical tissue and is routinely used in MALDI-MSI studies. However, it is not particularly suitable for lipidomics imaging given that many species are depleted during tissue processing. Irrespective, a number of solvent-resistant lipids remain, but their extraction may be hindered by the cross-link between proteins. Therefore, an antigen retrieval step could enable the extraction of a greater number of lipids and may provide information that is complementary to that which can be obtained from other biomolecules, such as proteins. In this short communication, we aim to address the effect of performing antigen retrieval prior to MALDI-MSI of lipids in FFPE tissue. As a result, an increased number of lipid signals could be detected and may have derived from lipid species that are known to be implicated in the lipid-protein cross-linking that is formed as a result of formalin fixation. Human renal cancer tissue was used as a proof of concept to determine whether using these detected lipid signals were also able to highlight the histopathological regions that were present. These preliminary findings may highlight the potential to enhance the clinical relevance of the lipidomic information obtained from FFPE tissue.