Plasma metabolomic and lipidomic profiles accurately classify mothers of children with congenital heart disease: an observational study.

INTRODUCTION: Congenital heart disease (CHD) is the most common congenital anomaly, representing a significant global disease burden. Limitations exist in our understanding of aetiology, diagnostic methodology and screening, with metabolomics offering promise in addressing these. OBJECTIVE: To evaluate maternal metabolomics and lipidomics in prediction and risk factor identification for childhood CHD. METHODS: We performed an observational study in mothers of children with CHD following pregnancy, using untargeted plasma metabolomics and lipidomics by ultrahigh performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS). 190 cases (157 mothers of children with structural CHD (sCHD); 33 mothers of children with genetic CHD (gCHD)) from the children OMACp cohort and 162 controls from the ALSPAC cohort were analysed. CHD diagnoses were stratified by severity and clinical classifications. Univariate, exploratory and supervised chemometric methods were used to identify metabolites and lipids distinguishing cases and controls, alongside predictive modelling. RESULTS: 499 metabolites and lipids were annotated and used to build PLS-DA and SO-CovSel-LDA predictive models to accurately distinguish sCHD and control groups. The best performing model had an sCHD test set mean accuracy of 94.74% (sCHD test group sensitivity 93.33%; specificity 96.00%) utilising only 11 analytes. Similar test performances were seen for gCHD. Across best performing models, 37 analytes contributed to performance including amino acids, lipids, and nucleotides. CONCLUSIONS: Here, maternal metabolomic and lipidomic analysis has facilitated the development of sensitive risk prediction models classifying mothers of children with CHD. Metabolites and lipids identified offer promise for maternal risk factor profiling, and understanding of CHD pathogenesis in the future.

Sub-5-min RP-UHPLC-TIMS for high-throughput untargeted lipidomics and its application to multiple matrices.

Untargeted lipidomics, with its ability to take a snapshot of the lipidome landscape, is an important tool to highlight lipid changes in pathology or drug treatment models. One of the shortcomings of most untargeted lipidomics based on UHPLC-HRMS is the low throughput, which is not compatible with large-scale screening. In this contribution, we evaluate the application of a sub-5-min high-throughput four-dimensional trapped ion mobility mass spectrometry (HT-4D-TIMS) platform for the fast profiling of multiple complex biological matrices. Human AC-16 cells and mouse brain, liver, sclera, and feces were used as samples. By using a fast 4-min RP gradient, the implementation of TIMS allows us to differentiate coeluting isomeric and isobaric lipids, with correct precursor ion isolation, avoiding co-fragmentation and chimeric MS/MS spectra. Globally, the HT-4D-TIMS allowed us to annotate 1910 different lipid species, 1308 at the molecular level and 602 at the sum composition level, covering 58 lipid subclasses, together with quantitation capability covering more than three orders of magnitude. Notably, TIMS values were highly comparable with respect to longer LC gradients (CV% = 0.39%). These results highlight how HT-4D-TIMS-based untargeted lipidomics possess high coverage and accuracy, halving the analysis time with respect to conventional UHPLC methods, and can be used for fast and accurate untargeted analysis of complex matrices to rapidly evaluate changes of lipid metabolism in disease models or drug discovery campaigns.

The Deep Proteomics Approach Identified Extracellular Vesicular Proteins Correlated to Extracellular Matrix in Type One and Two Endometrial Cancer.

Among gynecological cancers, endometrial cancer is the most common in developed countries. Extracellular vesicles (EVs) are cell-derived membrane-surrounded vesicles that contain proteins involved in immune response and apoptosis. A deep proteomic approach can help to identify dysregulated extracellular matrix (ECM) proteins in EVs correlated to key pathways for tumor development. In this study, we used a proteomics approach correlating the two acquisitions-data-dependent acquisition (DDA) and data-independent acquisition (DIA)-on EVs from the conditioned medium of four cell lines identifying 428 ECM proteins. After protein quantification and statistical analysis, we found significant changes in the abundance (p < 0.05) of 67 proteins. Our bioinformatic analysis identified 26 pathways associated with the ECM. Western blotting analysis on 13 patients with type 1 and type 2 EC and 13 endometrial samples confirmed an altered abundance of MMP2. Our proteomics analysis identified the dysregulated ECM proteins involved in cancer growth. Our data can open the path to other studies for understanding the interaction among cancer cells and the rearrangement of the ECM.

Multi-omics analysis reveals attenuation of cellular stress by empagliflozin in high glucose-treated human cardiomyocytes.

BACKGROUND: Sodium-glucose cotransporter 2 (SGLT2) inhibitors constitute the gold standard treatment for type 2 diabetes mellitus (T2DM). Among them, empagliflozin (EMPA) has shown beneficial effects against heart failure. Because cardiovascular diseases (mainly diabetic cardiomyopathy) are the leading cause of death in diabetic patients, the use of EMPA could be, simultaneously, cardioprotective and antidiabetic, reducing the risk of death from cardiovascular causes and decreasing the risk of hospitalization for heart failure in T2DM patients. Interestingly, recent studies have shown that EMPA has positive benefits for people with and without diabetes. This finding broadens the scope of EMPA function beyond glucose regulation alone to include a more intricate metabolic process that is, in part, still unknown. Similarly, this significantly increases the number of people with heart diseases who may be eligible for EMPA treatment. METHODS: This study aimed to clarify the metabolic effect of EMPA on the human myocardial cell model by using orthogonal metabolomics, lipidomics, and proteomics approaches. The untargeted and multivariate analysis mimicked the fasting blood sugar level of T2DM patients (hyperglycemia: HG) and in the average blood sugar range (normal glucose: NG), with and without the addition of EMPA. RESULTS: Results highlighted that EMPA was able to modulate and partially restore the levels of multiple metabolites associated with cellular stress, which were dysregulated in the HG conditions, such as nicotinamide mononucleotide, glucose-6-phosphate, lactic acid, FA 22:6 as well as nucleotide sugars and purine/pyrimidines. Additionally, EMPA regulated the levels of several lipid sub-classes, in particular dihydroceramide and triacylglycerols, which tend to accumulate in HG conditions resulting in lipotoxicity. Finally, EMPA counteracted the dysregulation of endoplasmic reticulum-derived proteins involved in cellular stress management. CONCLUSIONS: These results could suggest an effect of EMPA on different metabolic routes, tending to rescue cardiomyocyte metabolic status towards a healthy phenotype.

Integrated plasma metabolomics and lipidomics profiling highlights distinctive signature of hepatocellular carcinoma in HCV patients.

BACKGROUND: Early diagnosis of hepatocellular carcinoma (HCC) is essential towards the improvement of prognosis and patient survival. Circulating markers such as α-fetoprotein (AFP) and micro-RNAs represent useful tools but still have limitations. Identifying new markers can be fundamental to improve both diagnosis and prognosis. In this approach, we harness the potential of metabolomics and lipidomics to uncover potential signatures of HCC. METHODS: A combined untargeted metabolomics and lipidomics plasma profiling of 102 HCV-positive patients was performed by HILIC and RP-UHPLC coupled to Mass Spectrometry. Biochemical parameters of liver function (AST, ALT, GGT) and liver cancer biomarkers (AFP, CA19.9 e CEA) were evaluated by standard assays. RESULTS: HCC was characterized by an elevation of short and long-chain acylcarnitines, asymmetric dimethylarginine, methylguanine, isoleucylproline and a global reduction of lysophosphatidylcholines. A supervised PLS-DA model showed that the predictive accuracy for HCC class of metabolomics and lipidomics was superior to AFP for the test set (100.00% and 94.40% vs 55.00%). Additionally, the model was applied to HCC patients with AFP values < 20 ng/mL, and, by using only the top 20 variables selected by VIP scores achieved an Area Under Curve (AUC) performance of 0.94. CONCLUSION: These exploratory findings highlight how metabo-lipidomics enables the distinction of HCC from chronic HCV conditions. The identified biomarkers have high diagnostic potential and could represent a viable tool to support and assist in HCC diagnosis, including AFP-negative patients.

A Multi-Omics Approach Revealed Common Dysregulated Pathways in Type One and Type Two Endometrial Cancers.

Endometrial cancer (EC) is the most frequent gynecologic cancer in postmenopausal women. Pathogenetic mechanisms that are related to the onset and progression of the disease are largely still unknown. A multi-omics strategy can help identify altered pathways that could be targeted for improving therapeutical approaches. In this study we used a multi-omics approach on four EC cell lines for the identification of common dysregulated pathways in type 1 and 2 ECs. We analyzed proteomics and metabolomics of AN3CA, HEC1A, KLE and ISHIKAWA cell lines by mass spectrometry. The bioinformatic analysis identified 22 common pathways that are in common with both types of EC. In addition, we identified five proteins and 13 metabolites common to both types of EC. Western blotting analysis on 10 patients with type 1 and type 2 EC and 10 endometria samples confirmed the altered abundance of NPEPPS. Our multi-omics analysis identified dysregulated proteins and metabolites involved in EC tumor growth. Further studies are needed to understand the role of these molecules in EC. Our data can shed light on common pathways to better understand the mechanisms involved in the development and growth of EC, especially for the development of new therapies.

A Food-Grade Method for Enhancing the Levels of Low Molecular Weight Proanthocyanidins with Potentially High Intestinal Bioavailability.

Proanthocyanidins (PACs) are a group of bioactive molecules found in a variety of plants and foods. Their bioavailability depends on their molecular size, with monomers and dimers being more bioavailable than those that have a higher polymerization degree. This study aimed to develop a method to convert high-molecular-weight PACs to low-molecular-weight ones in a grape seed extract (GSE) from Vitis vinifera L. Therefore, GSE was subjected to alkaline treatment (ATGSE), and its difference in chemical composition, compared to GSE, was evaluated using a molecular networking (MN) approach based on results obtained from HPLC-ESI HRMS/MS characterization analysis. The network analysis mainly noted the PAC cluster with about 142 PAC compounds identified. In particular, the obtained results showed a higher content of monomeric and dimeric PACs in ATGSE compared to GSE, with 58% and 49% monomers and 31% and 24% dimers, respectively. Conversely, trimeric (9%), polymeric (4%), and galloylated PACs (14%) were more abundant in GSE than in ATGSE (6%, 1%, and 4%, respectively). Moreover, in vitro antioxidant and anti-inflammatory activities were investigated, showing the high beneficial potential of both extracts. In conclusion, ATGSE could represent an innovative natural matrix rich in bioavailable and bioaccessible PACs for nutraceutical applications with potential beneficial properties.

Prenatal and Early Postnatal Cerebral d-Aspartate Depletion Influences l-Amino Acid Pathways, Bioenergetic processes, and Developmental Brain Metabolism.

d-Amino acids were believed to occur only in bacteria and invertebrates. Today, it is well known that d-amino acids are also present in mammalian tissues in a considerable amount. In particular, high levels of free d-serine (d-Ser) and d-aspartate (d-Asp) are found in the brain. While the functions of d-Ser are well known, many questions remain unanswered regarding the role of d-Asp in the central nervous system. d-Asp is very abundant at the embryonic stage, while it strongly decreases after birth because of the expression of d-aspartate oxidase (Ddo) enzyme, which catalyzes the oxidation of this d-amino acid into oxaloacetate, ammonium, and hydrogen peroxide. Pharmacologically, d-Asp acts as an endogenous agonist of N-methyl d-aspartate and mGlu5 receptors, which are known to control fundamental brain processes, including brain development, synaptic plasticity, and cognition. In this work, we studied a recently generated knockin mouse model (R26ddo/ddo), which was designed to express DDO beginning at the zygotic stage. This strategy enables d-Asp to be almost eliminated in both prenatal and postnatal lives. To understand which biochemical pathways are affected by depletion of d-Asp, in this study, we carried out a metabolomic and lipidomic study of ddo knockin brains at different stages of embryonic and postnatal development, combining nuclear magnetic resonance (NMR) and high-resolution mass spectrometry (HRMS) techniques. Our study shows that d-Asp deficiency in the brain influences amino acid pathways such as threonine, glycine, alanine, valine, and glutamate. Interestingly, d-Asp is also correlated with metabolites involved in brain development and functions such as choline, creatine, phosphocholine (PCho), glycerophosphocholine (GPCho), sphingolipids, and glycerophospholipids, as well as metabolites involved in brain energy metabolism, such as GPCho, glucose, and lactate.

Indole-3-lactic acid, a metabolite of tryptophan, secreted by Bifidobacterium longum subspecies infantis is anti-inflammatory in the immature intestine.

BACKGROUND: Necrotizing enterocolitis (NEC), a necrotic inflammation of the intestine, represents a major health problem in the very premature infant. Although prevention is difficult, the combination of ingestion of maternal-expressed breastmilk in conjunction with a probiotic provides the best protection. In this study, we establish a mechanism for breastmilk/probiotic protection. METHODS: Ultra-high-performance liquid chromatography-tandem mass spectrometry of Bifidobacterium longum subsp. infantis (B. infantis) secretions was used to identify an anti-inflammatory molecule. Indole-3-lactic acid (ILA) was then tested in an established human immature small intestinal cell line, necrotizing colitis enterocytes, and other immature human enteroids for anti-inflammatory effects and to establish developmental function. ILA was also examined in immature and mature enterocytes. RESULTS: We have identified ILA, a metabolite of breastmilk tryptophan, as the anti-inflammatory molecule. This molecule is developmentally functional in immature but not mature intestinal enterocytes; ILA reduces the interleukin-8 (IL-8) response after IL-1ß stimulus. It interacts with the transcription factor aryl hydrocarbon receptor (AHR) and prevents transcription of the inflammatory cytokine IL-8. CONCLUSIONS: This molecule produced by B. infantis (ATCC No. 15697) interaction with ingested breastmilk functions in a complementary manner and could become useful in the treatment of all at-risk premature infants for NEC if safety and clinical studies are performed.

Chemical profiling of bioactive constituents in hop cones and pellets extracts by online comprehensive two-dimensional liquid chromatography with tandem mass spectrometry and direct infusion Fourier transform ion cyclotron resonance mass spectrometry.

Humulus lupulus L. (hop) is highly interesting from a nutraceutical perspective. The hop phytocomplex contains a wide range of bioactive metabolites, and its characterization is challenging. To tackle such a task, for the first time we applied and compared a combined approach consisting of online comprehensive two-dimensional liquid chromatography with tandem mass spectrometry and direct infusion Fourier transform ion cyclotron mass spectrometry. A reversed phase × reversed phase approach with a shifted gradient in the second dimension ensured selectivity and two-dimensional space coverage. Hyphenation with an ion trap time-of-flight analyzer led to the identification of 83 compounds in 70 min, comprising a novel quercetin derivative and six unknown bitter acids. On the other hand, the direct infusion method was able to identify 40 analytes (except isomers) with high mass accuracy (≤ 0.1 ppm) in less than 1 min analysis time. The developed approach can be used in a complementary way, combining the separation capability and high informative spectra of two-dimensional liquid chromatography tandem mass spectrometry with the ultra-high mass accuracy of direct infusion, for potential compound discovery or the accurate profiling of bioactive compounds in different hop cultivars as well as for monitoring processing and storage of hop-based products.

Fast Profiling of Natural Pigments in Different Spirulina (Arthrospira platensis) Dietary Supplements by DI-FT-ICR and Evaluation of their Antioxidant Potential by Pre-Column DPPH-UHPLC Assay.

Arthrospira platensis, better known as Spirulina, is one of the most important microalgae species. This cyanobacterium possesses a rich metabolite pattern, including high amounts of natural pigments. In this study, we applied a combined strategy based on Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR-MS) and Ultra High-Performance Liquid Chromatography (UHPLC) for the qualitative/quantitative characterization of Spirulina pigments in three different commercial dietary supplements. FT-ICR was employed to elucidate the qualitative profile of Spirulina pigments, in both direct infusion mode (DIMS) and coupled to UHPLC. DIMS showed to be a very fast (4 min) and accurate (mass accuracy ≤ 0.01 ppm) tool. 51 pigments were tentatively identified. The profile revealed different classes, such as carotenes, xanthophylls and chlorophylls. Moreover, the antioxidant evaluation of the major compounds was assessed by pre-column reaction with the DPPH radical followed by fast UHPLC-PDA separation, highlighting the contribution of single analytes to the antioxidant potential of the entire pigment fraction. ß-carotene, diadinoxanthin and diatoxanthin showed the highest scavenging activity. The method took 40 min per sample, comprising reaction. This strategy could represent a valid tool for the fast and comprehensive characterization of Spirulina pigments in dietary supplements, as well as in other microalgae-based products.

α acid fraction from Hop extract exerts an endothelium-derived hyperpolarization vasorelaxant effect through TRPV4 employing the feedforward mechanism of PKCα.

Until now, the beneficial vascular properties of Hop reported in the literature have been mainly attributed to specific compound classes, such as tannins and phenolic acids. However, the potential vascular action of a Hop subfraction containing a high amount of α or ß acids remains completely understood. Therefore, this study aims to investigate the vascular effects of the entire Hop extract and to fraction the Hop extract to identify the main bioactive vascular compounds. A pressure myograph was used to perform vascular reactivity studies on mouse resistance arteries. Phytocomplex fractionation was performed on a semi-prep HPLC system and characterized by UHPLC-PDA-MS/MS coupled to mass spectrometry. Western blot analysis was performed to characterize the phosphorylation site enrolled. The entire Hop extract exerts a direct dose-dependent endothelial vascular action. The B1 subfraction, containing a high concentration of α acids, recapitulates the vascular effect of the crude extract. Its vasorelaxant action is mediated by the opening of Transient Receptor Potential Vanilloid type 4 (TRPV4), potentiated by PKCα, and subsequent involvement of endothelial small-conductance calcium-activated potassium channels (SKCa) and intermediate-conductance calcium-activated potassium channels (IKCa) that drives endothelium-dependent hyperpolarization (EDH) through heterocellular myoendothelial gap junctions (MEGJs). This is the first comprehensive investigation of the vascular function of Hop-derived α acids in resistance arteries. Overall, our data suggest that the B1 subfraction from Hop extracts, containing only α acids, has great potential to be translated into the useful armamentarium of natural bioactive compounds with cardiovascular benefits.

Anticancer Therapies Based on Oxidative Damage: Lycium barbarum Inhibits the Proliferation of MCF-7 Cells by Activating Pyroptosis through Endoplasmic Reticulum Stress.

Lycium barbarum, commonly recognized as goji berry or wolfberry, is highly appreciated not only for its organoleptic and nutritional properties but also as an important source of bioactive compounds such as polysaccharides, carotenoids, phenolics, and various other non-nutritive compounds. These constituents give it a multitude of health benefits, including antioxidant, anti-inflammatory, and anticancer properties. However, the precise biochemical mechanisms responsible for its anticancer effects remain unclear, and the comprehensive composition of goji berry extracts is often insufficiently explored. This study aimed to investigate the biochemical pathways modulated in breast cancer cells by an ethanolic extract of Lycium barbarum fruit (LBE). Following metabolomic profiling using UHPLC-HRMS/MS, we assessed the antitumoral properties of LBE on different breast cancer cell lines. This investigation revealed that LBE exhibited cytotoxic effects, inducing a pro-oxidant effect that triggered pyroptosis activation through endoplasmic reticulum (ER) stress and subsequent activation of the P-IRE1α/XBP1/NLRP3 axis in MCF-7 cells. In addition, LBE did not display cytotoxicity toward healthy human cells but demonstrated antioxidant properties by neutralizing ROS generated by doxorubicin. These findings underscore the potential of LBE as a highly promising natural extract in cancer therapy.

Maternal Exposure to Endocrine-Disrupting Chemicals: Analysis of Their Impact on Infant Gut Microbiota Composition.

Endocrine disruptors (EDCs) are chemicals that interfere with the endocrine system. EDC exposure may contribute to the development of obesity, type 2 diabetes, and cardiovascular diseases by impacting the composition of an infant's gut microbiota during the first 1000 days of life. To explore the relationship between maternal urinary levels of Bisphenol-A and phthalates (UHPLC-MS/MS), and the composition of the infant gut microbiota (16S rDNA) at age 12 months (T3) and, retrospectively, at birth (T0), 1 month (T1), and 6 months (T2), stool samples from 20 infants breastfed at least once a day were analyzed. Metataxonomic bacteria relative abundances were correlated with EDC values. Based on median Bisphenol-A levels, infants were assigned to the over-exposed group (O, n = 8) and the low-exposed group (B, n = 12). The B-group exhibited higher gut colonization of the Ruminococcus torques group genus and the O-group showed higher abundances of Erysipelatoclostridium and Bifidobacterium breve. Additionally, infants were stratified as high-risk (HR, n = 12) or low-risk (LR, n = 8) exposure to phthalates, based on the presence of at least three phthalates with concentrations exceeding the cohort median values; no differences were observed in gut microbiota composition. A retrospective analysis of gut microbiota (T0-T2) revealed a disparity in ß-diversity between the O-group and the B-group. Considering T0-T3, the Linear Discriminant Effect Size indicated differences in certain microbes between the O-group vs. the B-group and the HR-group vs. the LR-group. Our findings support the potential role of microbial communities as biomarkers for high EDC exposure levels. Nevertheless, further investigations are required to deeply investigate this issue.

Novel pyrrole based CB2 agonists: New insights on CB2 receptor role in regulating neurotransmitters' tone.

The cannabinoid system is one of the most investigated neuromodulatory systems because of its involvement in multiple pathologies such as cancer, inflammation, and psychiatric diseases. Recently, the CB2 receptor has gained increased attention considering its crucial role in modulating neuroinflammation in several pathological conditions like neurodegenerative diseases. Here we describe the rational design of pyrrole-based analogues, which led to a potent and pharmacokinetically suitable CB2 full agonist particularly effective in improving cognitive functions in a scopolamine-induced amnesia murine model. Therefore, we extended our study by investigating the interconnection between CB2 activation and neurotransmission in this experimental paradigm. To this purpose, we performed a MALDI imaging analysis on mice brains, observing that the administration of our lead compound was able to revert the effect of scopolamine on different neurotransmitter tones, such as acetylcholine, serotonin, and GABA, shedding light on important networks not fully explored, so far.

Optimization of microwave-assisted extraction of antioxidant compounds from spring onion leaves using Box-Behnken design.

Many studies have explored the extraction of bioactive compounds from different onion solid wastes, such as bulb, skin, and peel. However, onion leaves have received limited attention despite their potential as a valuable source of nutraceutical compounds. This study aimed to valorise, for the first time, the agricultural waste in the form of spring onion leaves (CN, Cipollotto Nocerino) to obtain antioxidant-rich polyphenolic extracts. A Box-Behnken design (BBD) was used to assess the impact of microwave-assisted extraction (MAE) variables (temperature, time, extraction volume, and ethanol concentration) on total polyphenol content (TPC) measured by Folin-Ciocalteu method and the antioxidant power determined by FRAP assay. Response surface methodology (RSM) was applied, and regression equations, analysis of variance, and 3D response curves were developed. Our results highlighted that the TPC values range from 0.76 to 1.43 mg GAE g-1 dw, while the FRAP values range from 8.25 to 14.80 mmol Fe(II)E g-1 dw. The optimal extraction conditions predicted by the model were 60 °C, 22 min, ethanol concentration 51% (v/v), and solvent volume 11 mL. These conditions resulted in TPC and FRAP values of 1.35 mg GAE g-1 dw and 14.02 mmol Fe(II)E g-1 dw, respectively. Furthermore, the extract obtained under optimized conditions was characterized by UHPLC-ESI-Orbitrap-MS analysis. LC/MS-MS platform allowed us to tentatively identify various compounds belonging to the class of flavonoids, saponins, fatty acids, and lipids. Finally, the ability of CN optimal extract to inhibit the intracellular reactive oxygen species (ROS) release in a hepatocarcinoma cell line using an H2O2-induced oxidative stress model, was evaluated. The results highlighted the potential of CN extract as a valuable source of polyphenols with significant antioxidant properties, suitable for various applications in the food and pharmaceutical industries.

Olive oil-derived endocannabinoid-like mediators inhibit palatable food-induced reward and obesity.

N-oleoylglycine (OlGly), a lipid derived from the basic component of olive oil, oleic acid, and N-oleoylalanine (OlAla) are endocannabinoid-like mediators. We report that OlGly and OlAla, by activating the peroxisome proliferator-activated receptor alpha (PPARα), reduce the rewarding properties of a highly palatable food, dopamine neuron firing in the ventral tegmental area, and the obesogenic effect of a high-fat diet rich in lard (HFD-L). An isocaloric olive oil HFD (HFD-O) reduced body weight gain compared to the HFD-L, in a manner reversed by PPARα antagonism, and enhanced brain and intestinal OlGly levels and gut microbial diversity. OlGly or OlAla treatment of HFD-L mice resulted in gut microbiota taxonomic changes partly similar to those induced by HFD-O. We suggest that OlGly and OlAla control body weight by counteracting highly palatable food overconsumption, and possibly rebalancing the gut microbiota, and provide a potential new mechanism of action for the obeso-preventive effects of olive oil-rich diets.

High levels of PD-L1 on platelets of NSCLC patients contributes to the pharmacological activity of Atezolizumab.

Several studies have associated platelets (PLTs) to NSCLC prognosis. To understand the role of PLTs in immunotherapy-treated patients, we used blood samples of NSCLC patients at different TNM stage. We found that PLTs count and the expression of PD-L1 (pPD-L1) were significantly higher in NSCLC patients at Stage IV than Stage I-III and healthy subjects. The presence of high pPD-L1 was associated to upregulated genes for the extracellular matrix organization and tumor immunosuppression. When patients' survival was correlated to the levels of pPD-L1, longer survival rate was observed, but not when progression disease occurred. The in vitro stimulation of pPD-L1 with Atezolizumab induced CXCL4 release, accompanied by higher levels of TGFß at the time of drug resistance when the levels of CD16, CD32 and CD64 significantly increased. Leiden-clustering method defined the phenotype of PLTs which showed that the ezrin-radixin-moesin (ERM) family proteins, underlying the PD-L1 signalosome, were involved in high pPD-L1 and higher survival rate. These data imply that Stage IV NSCLC patients characterized by high pPD-L1 are associated with longer progression-free survival rate because the blockade of pPD-L1 by Atezolizumab avoids the exacerbation of a T cell-mediated immune-suppressive environment. pPD-L1 could be an easy-to-use clinical approach to predict ICI responsiveness.

A Combination of Polymethoxyflavones from Citrus sinensis and Prenylflavonoids from Humulus lupulus Counteracts IL-1ß-Induced Differentiated Caco-2 Cells Dysfunction via a Modulation of NF-κB/Nrf2 Activation.

We here investigated the anti-inflammatory activity of a polymethoxylated flavone-containing fraction (PMFF) from Citrus sinensis and of a prenylflavonoid-containing one (PFF) from Humulus lupulus, either alone or in combination (MIX). To this end, an in vitro model of inflammatory bowel disease (IBD), consisting of differentiated, interleukin (IL)-1ß-stimulated Caco-2 cells, was employed. We demonstrated that non-cytotoxic concentrations of either PMFF or PFF or MIX reduced nitric oxide (NO) production while PFF and MIX, but not PMFF, also inhibited prostaglandin E2 release. Coherently, MIX suppressed both inducible NO synthase and cyclooxygenase-2 over-expression besides NF-κB activation. Moreover, MIX increased nuclear factor erythroid 2-related factor 2 (Nrf2) activation, heme oxygenase-1 expression, restoring GSH and reactive oxygen and nitrogen species (RONs) levels. Remarkably, these effects with MIX were stronger than those produced by PMFF or PFF alone. Noteworthy, nobiletin (NOB) and xanthohumol (XTM), two of the most represented phytochemicals in PMFF and PFF, respectively, synergistically inhibited RONs production. Overall, our results demonstrate that MIX enhances the anti-inflammatory and anti-oxidative effects of the individual fractions in a model of IBD, via a mechanism involving modulation of NF-κB and Nrf2 signalling. Synergistic interactions between NOB and XTM emerge as a relevant aspect underlying this evidence.

Experimental evidence and clinical implications of Warburg effect in the skeletal muscle of Fabry disease.

Skeletal muscle (SM) pain and fatigue are common in Fabry disease (FD). Here, we undertook the investigation of the energetic mechanisms related to FD-SM phenotype. A reduced tolerance to aerobic activity and lactate accumulation occurred in FD-mice and patients. Accordingly, in murine FD-SM we detected an increase in fast/glycolytic fibers, mirrored by glycolysis upregulation. In FD-patients, we confirmed a high glycolytic rate and the underutilization of lipids as fuel. In the quest for a tentative mechanism, we found HIF-1 upregulated in FD-mice and patients. This finding goes with miR-17 upregulation that is responsible for metabolic remodeling and HIF-1 accumulation. Accordingly, miR-17 antagomir inhibited HIF-1 accumulation, reverting the metabolic-remodeling in FD-cells. Our findings unveil a Warburg effect in FD, an anaerobic-glycolytic switch under normoxia induced by miR-17-mediated HIF-1 upregulation. Exercise-intolerance, blood-lactate increase, and the underlying miR-17/HIF-1 pathway may become useful therapeutic targets and diagnostic/monitoring tools in FD.