Search

To date, little is known concerning the circulating levels of biochemically relevant metabolites (antioxidants, oxidative/nitrosative stress biomarkers, purines, and pyrimidines) in patients with primary myelofibrosis (PMF), a rare form of myeloproliferative tumor causing a dramatic decrease in erythropoiesis and angiogenesis. In this study, using a targeted metabolomic approach, serum samples of 22 PMF patients and of 22 control healthy donors were analyzed to quantify the circulating concentrations of hypoxanthine, xanthine, uric acid (as representative purines), uracil, β-pseudouridine, uridine (as representative pyrimidines), reduced glutathione (GSH), ascorbic acid (as two of the main water-soluble antioxidants), malondialdehyde, nitrite, nitrate (as oxidative/nitrosative stress biomarkers) and creatinine, using well-established HPLC method for their determination. Results showed that PMF patients have dramatic depletions of both ascorbic acid and GSH (37.3- and 3.81-times lower circulating concentrations, respectively, than those recorded in healthy controls, p < 0.0001), accompanied by significant increases in malondialdehyde (MDA) and nitrite + nitrate (4.73- and 1.66-times higher circulating concentrations, respectively, than those recorded in healthy controls, p < 0.0001). Additionally, PMF patients have remarkable alterations of circulating purines, pyrimidines, and creatinine, suggesting potential mitochondrial dysfunctions causing energy metabolism imbalance and consequent increases in these cell energy-related compounds. Overall, these results, besides evidencing previously unknown serum metabolic alterations in PMF patients, suggest that the determination of serum levels of the aforementioned compounds may be useful to evaluate PMF patients on hospital admission for adjunctive therapies aimed at recovering their correct antioxidant status, as well as to monitor patients' status and potential pharmacological treatments. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD) is characterized by the accumulation of lipids within hepatocytes, which compromises liver functionality following mitochondrial dysfunction and increased production of reactive oxygen species (ROS). Lipoic acid is one of the prosthetic groups of the pyruvate dehydrogenase complex also known for its ability to confer protection from oxidative damage because of its antioxidant properties. In this study, we aimed to investigate the effects of lipoic acid on lipotoxicity and mitochondrial dynamics in an in vitro model of liver steatosis. HepG2 cells were treated with palmitic acid and oleic acid (1:2) to induce steatosis, without and with 1 and 5 µM lipoic acid. Following treatments, cell proliferation and lipid droplets accumulation were evaluated. Mitochondrial functions were assessed through the evaluation of membrane potential, MitoTracker Red staining, expression of genes of the mitochondrial quality control, and analysis of energy metabolism by HPLC and Seahorse. We showed that lipoic acid treatment restored membrane potential to values comparable to control cells, as well as protected cells from mitochondrial fragmentation following PA:OA treatment. Furthermore, our data showed that lipoic acid was able to determine an increase in the expression of mitochondrial fusion genes and a decrease in mitochondrial fission genes, as well as to restore the bioenergetics of cells after treatment with palmitic acid and oleic acid. In conclusion, our data suggest that lipoic acid reduces lipotoxicity and improves mitochondrial functions in an in vitro model of steatosis, thus providing a potentially valuable pharmacological tool for NAFLD treatment. [ABSTRACT FROM AUTHOR]

In a previous study, we showed that various low-molecular-weight compounds in follicular fluid (FF) samples of control fertile females (CFF) have different concentrations compared to those found in FF of infertile females (IF), before and after their categorization into different subgroups, according to their clinical diagnosis of infertility. Using the same FF samples of this previous study, we here analyzed the FF concentrations of free and bound bilirubin and compared the results obtained in CFF, IF and the different subgroups of IF (endometriosis, EM, polycystic ovary syndrome, PCOS, age-related reduced ovarian reserve, AR-ROR, reduced ovarian reserve, ROR, genetic infertility, GI and unexplained infertility, UI). The results clearly indicated that CFF had lower values of free, bound and total bilirubin compared to the respective values measured in pooled IF. These differences were observed even when IF were categorized into EM, PCOS, AR-ROR, ROR, GI and UI, with EM and PCOS showing the highest values of free, bound and total bilirubin among the six subgroups. Using previous results of ascorbic acid, GSH and nitrite + nitrate measured in the same FF samples of the same FF donors, we found that total bilirubin in FF increased as a function of decreased values of ascorbic acid and GSH, and increased concentrations of nitrite + nitrate. The values of total bilirubin negatively correlated with the clinical parameters of fertilization procedures (number of retrieved oocytes, mature oocytes, fertilized oocytes, blastocysts, high-quality blastocysts) and with clinical pregnancies and birth rates. Bilirubin concentrations in FF were not linked to those found in serum samples of FF donors, thereby strongly suggesting that its over production was due to higher activity of heme oxygenase-1 (HO-1), the key enzyme responsible for bilirubin formation, in granulosa cells, or cumulus cells or oocytes of IF and ultimately leading to bilirubin accumulation in FF. Since increased activity of HO-1 is one of the main enzymatic intracellular mechanisms of defense towards external insults (oxidative/nitrosative stress, inflammation), and since we found correlations among bilirubin and oxidative/nitrosative stress in these FF samples, it may reasonably be supposed that bilirubin increase in FF of IF is the result of protracted exposures to the aforementioned insults evidently playing relevant roles in female infertility. [ABSTRACT FROM AUTHOR]

To date, there is no information on the effect of TBI on the changes in brain CoQ levels and possible variations in its redox state. In this study, we induced graded TBIs (mild TBI, mTBI and severe TBI, sTBI) in male rats, using the weight-drop closed-head impact acceleration model of trauma. At 7 days post-injury, CoQ9, CoQ10 and α-tocopherol were measured by HPLC in brain extracts of the injured rats, as well as in those of a group of control sham-operated rats. In the controls, about the 69% of total CoQ was in the form of CoQ9 and the oxidized/reduced ratios of CoQ9 and CoQ10 were, respectively, 1.05 ± 0.07 and 1.42 ± 0.17. No significant changes in these values were observed in rats experiencing mTBI. Conversely, in the brains of sTBI-injured animals, an increase in reduced and a decrease in oxidized CoQ9 produced an oxidized/reduced ratio of 0.81 ± 0.1 (p < 0.001 compared with both controls and mTBI). A concomitant decrease in both reduced and oxidized CoQ10 generated a corresponding oxidized/reduced ratio of 1.38 ± 0.23 (p < 0.001 compared with both controls and mTBI). An overall decrease in the concentration of the total CoQ pool was also found in sTBI-injured rats (p < 0.001 compared with both controls and mTBI). Concerning α-tocopherol, whilst no differences compared with the controls were found in mTBI animals, a significant decrease was observed in rats experiencing sTBI (p < 0.01 compared with both controls and mTBI). Besides suggesting potentially different functions and intracellular distributions of CoQ9 and CoQ10 in rat brain mitochondria, these results demonstrate, for the first time to the best of knowledge, that sTBI alters the levels and redox states of CoQ9 and CoQ10, thus adding a new explanation to the mitochondrial impairment affecting ETC, OXPHOS, energy supply and antioxidant defenses following sTBI. [ABSTRACT FROM AUTHOR]

Metabolic changes of malignant plasma cells (PCs) and adaptation to tumour microenvironment represent one of the hallmarks of multiple myeloma (MM). We previously showed that MM mesenchymal stromal cells are more glycolytic and produce more lactate than healthy counterpart. Hence, we aimed to explore the impact of high lactate concentration on metabolism of tumour PCs and its impact on the efficacy of proteasome inhibitors (PIs). Lactate concentration was performed by colorimetric assay on MM patient's sera. The metabolism of MM cell treated with lactate was assessed by seahorse and real time Polymerase Chain Reaction (PCR). Cytometry was used to evaluate mitochondrial reactive oxygen species (mROS), apoptosis and mitochondrial depolarization. Lactate concentration resulted increased in MM patient's sera. Therefore, PCs were treated with lactate and we observed an increase of oxidative phosphorylation‐related genes, mROS and oxygen consumption rate. Lactate supplementation exhibited a significant reduction in cell proliferation and less responsive to PIs. These data were confirmed by pharmacological inhibition of monocarboxylate transporter 1 (MCT1) by AZD3965 which was able to overcame metabolic protective effect of lactate against PIs. Consistently, high levels of circulating lactate caused expansion of Treg and monocytic myeloid derived suppressor cells and such effect was significantly reduced by AZD3965. Overall, these findings showed that targeting lactate trafficking in TME inhibits metabolic rewiring of tumour PCs, lactate‐dependent immune evasion and thus improving therapy efficacy. [ABSTRACT FROM AUTHOR]

Notwithstanding the great improvement of ART, the overall rate of successful pregnancies from implanted human embryos is definitely low. The current routine embryo quality assessment is performed only through morphological criteria, which has poor predictive capacity since only a minor percentage of those in the highest class give rise to successful pregnancy. Previous studies highlighted the potentiality of the analysis of metabolites in human embryo culture media, useful for the selection of embryos for implantation. In the present study, we analyzed in blind 66 human embryo culture media at 5 days after in vitro fertilization with the aim of quantifying compounds released by cell metabolism that were not present as normal constituents of the human embryo growth media, including purines, pyrimidines, nitrite, and nitrate. Only some purines were detectable (hypoxanthine and uric acid) in the majority of samples, while nitrite and nitrate were always detectable. When matching biochemical results with morphological evaluation, it was found that low grade embryos (n = 12) had significantly higher levels of all the compounds of interest. Moreover, when matching biochemical results according to successful (n = 17) or unsuccessful (n = 25) pregnancy, it was found that human embryos from the latter group released higher concentrations of hypoxanthine, uric acid, nitrite, and nitrate in the culture media. Additionally, those embryos that developed into successful pregnancies were all associated with the birth of healthy newborns. These results, although carried out on a relatively low number of samples, indicate that the analysis of the aforementioned compounds in the culture media of human embryos is a potentially useful tool for the selection of embryos for implantation, possibly leading to an increase in the overall rate of ART. [ABSTRACT FROM AUTHOR]

Background: Acute and chronic neurodegenerative diseases represent an immense socioeconomic burden that drives the need for new disease modifying drugs. Common pathogenic mechanisms in these diseases are evident, suggesting that a platform neuroprotective therapy may offer effective treatments. Here we present evidence for the mode of pharmacological action of a novel neuroprotective low molecular weight dextran sulphate drug called ILB®. The working hypothesis was that ILB® acts via the activation of heparin-binding growth factors (HBGF). Methods: Pre-clinical and clinical (healthy people and patients with ALS) in vitro and in vivo studies evaluated the mode of action of ILB®. In vitro binding studies, functional assays and gene expression analyses were followed by the assessment of the drug effects in an animal model of severe traumatic brain injury (sTBI) using gene expression studies followed by functional analysis. Clinical data, to assess the hypothesized mode of action, are also presented from early phase clinical trials. Results: ILB® lengthened APTT time, acted as a competitive inhibitor for HGFGlypican-3 binding, effected pulse release of heparin-binding growth factors (HBGF) into the circulation and modulated growth factor signaling pathways. Gene expression analysis demonstrated substantial similarities in the functional dysregulation induced by sTBI and various human neurodegenerative conditions and supported a cascading effect of ILB® on growth factor activation, followed by gene expression changes with profound beneficial effect on molecular and cellular functions affected by these diseases. The transcriptional signature of ILB® relevant to cell survival, inflammation, glutamate signaling, metabolism and synaptogenesis, are consistent with the activation of neuroprotective growth factors as was the ability of ILB® to elevate circulating levels of HGF in animal models and humans. Conclusion: ILB® releases, redistributes and modulates the bioactivity of HBGF that target disease compromised nervous tissues to initiate a cascade of transcriptional, metabolic and immunological effects that control glutamate toxicity, normalize tissue bioenergetics, and resolve inflammation to improve tissue function. This unique mechanism of action mobilizes and modulates naturally occurring tissue repair mechanisms to restore cellular homeostasis and function. The identified pharmacological impact of ILB® supports the potential to treat various acute and chronic neurodegenerative disease, including sTBI and ALS. [ABSTRACT FROM AUTHOR]

In a previous study, we found that administration of ILB®, a new low molecular weight dextran sulphate, significantly improved mitochondrial functions and energy metabolism, as well as decreased oxidative/nitrosative stress, of brain tissue of rats exposed to severe traumatic brain injury (sTBI), induced by the closed-head weight-drop model of diffused TBI. Using aliquots of deproteinized brain tissue of the same animals of this former study, we here determined the concentrations of 24 amino acids of control rats, untreated sTBI rats (sacrificed at 2 and 7 days post-injury) and sTBI rats receiving a subcutaneous ILB® administration (at the dose levels of 1, 5 and 15 mg/kg b.w.) 30 min post-impact (sacrificed at 2 and 7 days post-injury). Additionally, in a different set of experiments, new groups of control rats, untreated sTBI rats and ILB®-treated rats (administered 30 min after sTBI at the dose levels of 1 or 5 mg/kg b.w.) were studied for their neurocognitive functions (anxiety, locomotor capacities, short- and long-term memory) at 7 days after the induction of sTBI. Compared to untreated sTBI animals, ILB® significantly decreased whole brain glutamate (normalizing the glutamate/glutamine ratio), glycine, serine and γ-aminobutyric acid. Furthermore, ILB® administration restored arginine metabolism (preventing nitrosative stress), levels of amino acids involved in methylation reactions (methionine, L-cystathionine, S-adenosylhomocysteine), and N-acetylaspartate homeostasis. The macroscopic evidences of the beneficial effects on brain metabolism induced by ILB® were the relevant improvement in neurocognitive functions of the group of animals treated with ILB® 5 mg/kg b.w., compared to the marked cognitive decline measured in untreated sTBI animals. These results demonstrate that ILB® administration 30 min after sTBI prevents glutamate excitotoxicity and normalizes levels of amino acids involved in crucial brain metabolic functions. The ameliorations of amino acid metabolism, mitochondrial functions and energy metabolism in ILB®-treated rats exposed to sTBI produced significant improvement in neurocognitive functions, reinforcing the concept that ILB® is a new effective therapeutic tool for the treatment of sTBI, worth being tested in the clinical setting. [ABSTRACT FROM AUTHOR]

Aims: Several studies reported that astrocytes support neuronal communication by the release of gliotransmitters, including ATP and glutamate. Astrocytes also play a fundamental role in buffering extracellular glutamate in the synaptic cleft, thus limiting the risk of excitotoxicity in neurons. We previously demonstrated that extracellular tau oligomers (ex‐oTau), by specifically targeting astrocytes, affect glutamate‐dependent synaptic transmission via a reduction in gliotransmitter release. The aim of this work was to determine if ex‐oTau also impair the ability of astrocytes to uptake extracellular glutamate, thus further contributing to ex‐oTau‐dependent neuronal dysfunction. Methods: Primary cultures of astrocytes and organotypic brain slices were exposed to ex‐oTau (200 nM) for 1 h. Extracellular glutamate buffering by astrocytes was studied by: Na+ imaging; electrophysiological recordings; high‐performance liquid chromatography; Western blot and immunofluorescence. Experimental paradigms avoiding ex‐oTau internalisation (i.e. heparin pre‐treatment and amyloid precursor protein knockout astrocytes) were used to dissect intracellular vs extracellular effects of oTau. Results: Ex‐oTau uploading in astrocytes significantly affected glutamate‐transporter‐1 expression and function, thus impinging on glutamate buffering activity. Ex‐oTau also reduced Na‐K‐ATPase activity because of pump mislocalisation on the plasma membrane, with no significant changes in expression. This effect was independent of oTau internalisation and it caused Na+ overload and membrane depolarisation in ex‐oTau‐targeted astrocytes. Conclusions: Ex‐oTau exerted a complex action on astrocytes, at both intracellular and extracellular levels. The net effect was dysregulated glutamate signalling in terms of both release and uptake that relied on reduced expression of glutamate‐transporter‐1, altered function and localisation of NKA1A1, and NKA1A2. Consequently, Na+ gradients and all Na+‐dependent transports were affected. [ABSTRACT FROM AUTHOR]

Down Syndrome (DS) is a neurodevelopmental disorder that is characterized by an accelerated aging process, frequently associated with the development of Alzheimer's disease (AD). Previous studies evidenced that DS patients have various metabolic anomalies, easily measurable in their serum samples, although values that were found in DS patients were compared with those of age-matched non-DS patients, thus hampering to discriminate the physiologic age-related changes of serum metabolites from those that are truly caused by the pathologic processes associated with DS. In the present study we performed a targeted metabolomic evaluation of serum samples from DS patients without dementia of two age classes (Younger DS Patients, YDSP, aging 20–40 years; Aged DS Patients, ADSP, aging 41–60 years), comparing the results with those that were obtained in two age classes of non-DS patients (Younger non-DS Patients, YnonDSP, aging 30–60 years; Aged-nonDS Patients, AnonDSP, aging 75–90 years). Of the 36 compounds assayed, 30 had significantly different concentrations in Pooled non-DS Patients (PnonDSP), compared to Pooled DS Patients (PDSP). Age categorization revealed that 11/30 compounds were significantly different in AnonDSP, compared to YnonDSP, indicating physiologic, age-related changes of their circulating concentrations. A comparison between YDSP and ADSP showed that 19/30 metabolites had significantly different values from those found in the corresponding classes of non-DS patients, strongly suggesting pathologic, DS-associated alterations of their serum levels. Twelve compounds selectively and specifically discriminated PnonDSP from PDSP, whilst only three discriminated YDSP from ADSP. The results allowed to determine, for the first time and to the best of our knowledge, the true, age-independent alterations of metabolism that are measurable in serum and attributable only to DS. These findings may be of high relevance for better strategies (pharmacological, nutritional) aiming to specifically target the dysmetabolism and decreased antioxidant defenses that are associated with DS. [ABSTRACT FROM AUTHOR]

Introduction: Amyotrophic lateral sclerosis (ALS) is an invariably lethal progressive disease, causing degeneration of neurons and muscle. No current treatment halts or reverses disease advance. This single arm, open label, clinical trial in patients with ALS investigated the safety and tolerability of a novel modified low molecular weight dextran sulphate (LMW-DS, named ILB®) previously proven safe for use in healthy volunteers and shown to exert potent neurotrophic effects in pre-clinical studies. Secondary endpoints relate to efficacy and exploratory biomarkers. Methods: Thirteen patients with ALS were treated with 5 weekly subcutaneous injections of ILB®. Safety and efficacy outcome measures were recorded weekly during treatment and at regular intervals for a further 70 days. Functional and laboratory biomarkers were assessed before, during and after treatment. Results: No deaths, serious adverse events or participant withdrawals occurred during or after ILB® treatment and no significant drug-related changes in blood safety markers were evident, demonstrating safety and tolerability of the drug in this cohort of patients with ALS. The PK of ILB® in patients with ALS was similar to that seen in healthy controls. The ILB® injection elicited a transient elevation of plasma Hepatocyte Growth Factor, a neurotrophic and myogenic growth factor. Following the ILB® injections patients reported increased vitality, decreased spasticity and increased mobility. The ALSFRS-R rating improved from 36.31 ± 6.66 to 38.77 ± 6.44 and the Norris rating also improved from 70.61 ± 13.91 to 77.85 ± 14.24 by Day 36. The improvement of functions was associated with a decrease in muscle atrophy biomarkers. These therapeutic benefits decreased 3–4 weeks after the last dosage. Conclusions: This pilot clinical study demonstrates safety and tolerability of ILB® in patients with ALS. The exploratory biomarker and functional measures must be cautiously interpreted but suggest clinical benefit and have a bearing on the mechanism of action of ILB®. The results support the drug's potential as the first disease modifying treatment for patients with ALS. Trial registration: EudraCT 2017-005065-47. [ABSTRACT FROM AUTHOR]

The combined derangements in mitochondria network, function and dynamics can affect metabolism and ATP production, redox homeostasis and apoptosis triggering, contributing to cancer development in many different complex ways. In hematological malignancies, there is a strong relationship between cellular metabolism, mitochondrial bioenergetics, interconnections with supportive microenvironment and drug resistance. Lymphoma and chronic lymphocytic leukemia cells, e.g., adapt to intrinsic oxidative stress by increasing mitochondrial biogenesis. In other hematological disorders such as myeloma, on the contrary, bioenergetics changes, associated to increased mitochondrial fitness, derive from the adaptive response to drug-induced stress. In the bone marrow niche, a reverse Warburg effect has been recently described, consisting in metabolic changes occurring in stromal cells in the attempt to metabolically support adjacent cancer cells. Moreover, a physiological dynamic, based on mitochondria transfer, between tumor cells and their supporting stromal microenvironment has been described to sustain oxidative stress associated to proteostasis maintenance in multiple myeloma and leukemia. Increased mitochondrial biogenesis of tumor cells associated to acquisition of new mitochondria transferred by mesenchymal stromal cells results in augmented ATP production through increased oxidative phosphorylation (OX-PHOS), higher drug resistance, and resurgence after treatment. Accordingly, targeting mitochondrial biogenesis, electron transfer, mitochondrial DNA replication, or mitochondrial fatty acid transport increases therapy efficacy. In this review, we summarize selected examples of the mitochondrial derangements in hematological malignancies, which provide metabolic adaptation and apoptosis resistance, also supported by the crosstalk with tumor microenvironment. This field promises a rational design to improve target-therapy including the metabolic phenotype. [ABSTRACT FROM AUTHOR]

The human consumption of food animal products is the main topic of an important debate among professionals in this sector: dietologists, dietitians and nutritional biologists. The red meat provides all the essential amino acids, bioavailable iron, zinc, calcium, lipids and B-group vitamins. A valid alternative to beef could be the buffalo meat. Italy is the largest European producer of buffalo meat and derivatives. The high nutritional characteristics of buffalo meat make it suitable to be included in the Mediterranean diet to customize it in relation to the needs and conditions of the population. Polyunsaturated/saturated fatty acids ratio can be influenced by diet, breed and type of breeding, but muscle tissue fat percentage is the main factor in determining a favorable fatty acid composition. This review focuses on the biochemical and nutritional characteristics of the buffalo meat (content of fats, cholesterol, amino acids, vitamins and minerals), explaining their variability depending on the different breeds, and the favorable implications on the human health. These results suggest that buffalo meat can be a healthier alternative to beef, not only for healthy people in particular physiological conditions (i.e. pregnancy), but also for persons at risk for cardiovascular and cerebrovascular diseases, thus achieving the goal of a personalized nutrition. [ABSTRACT FROM AUTHOR]

Recent studies have revealed that treatment with N-acetyl-l-cysteine (NAC) improved airway function in Chronic Obstructive Pulmonary Disease (COPD) patients and lowered the frequency of acute exacerbations. In this study, we investigated the biochemical mechanisms through which NAC exerts its protective effects against bronchial epithelium cells exposed to cigarette smoke. To evaluate the efficacy of NAC to counteract the deleterious effects induced by cigarette smoke, we treated human bronchial epithelial cells (16 HBE) with NAC alone and in combination with cigarette smoke extract (CSE). After treatment, we measured reactive oxygen species (ROS) formation, effect on cell viabilty, glutathione resources (contents and genes), inflammatory cytokines expression (IL-1 β, IL-6 and TNF-α) and antioxidant genes expression SOD1, SOD2, iNOS, HO-1, NRF2, IRE-1α and PGC-1α. NAC treatment was able to counteract free radicals (ROS) formed and reduce viability after CSE induced oxidative damage. Moreover, NAC increased the GSH's resources and gluathione metabolism gene such as GPX, GCLC and GT, restored the gene expression of SOD1, SOD2, reduced expression of pro-inflammatory cytokines, and was able to positively regulate expression of iNOS HO-1, NRF2, IRE-1α and PGC-1α. Our results demonstrated that NAC treatment restores endogenous antioxidant defenses in human bronchial epithelial cells exposed to cigarette smoke extract suggesting its use in the therapeutic strategy of patients with COPD. [ABSTRACT FROM AUTHOR]

Study Question: Is the determination of antioxidants, oxidative/nitrosative stress-related compounds, purines, pyrimidines and energy-related metabolites in human seminal plasma of utility to evidence biomarkers related to male infertility?Summary Answer: The determination of 26 metabolites in seminal plasma allowed to evidence that 21/26 of them are biomarkers of male infertility, as well as to calculate a cumulative index, named Biomarker Score, that fully discriminates fertile controls from infertile patients and partially differentiates infertile without from infertile with spermiogram anomalies.What Is Known Already: Epidemiological studies indicated that a male factor is involved in ~50% of cases of pregnancy failure, with a significant percentage of infertile males having no alterations in the spermiogram. Further laboratory analyses of male infertility are mainly dedicated only to gross evaluations of oxidative stress or total antioxidant capacity.Study Design, Size, Duration: Seminal plasma of 48 fertile controls and 96 infertile patients (master group), were collected from September 2016 to February 2018. A second group of 44 infertile patients (validation group) was recruited in a second, independent centre from September 2017 to March 2018. Samples were analysed in blind using a 'Redox Energy Test' to determine various low-molecular weight compounds, with the aim of finding metabolic profiles and biomarkers related to male infertility.Participants/materials, Setting, Methods: In all seminal plasma, 26 water- and fat-soluble compounds (related to antioxidant defences, oxidative/nitrosative stress, purine, pyrimidine and energy metabolism) were analysed using high-performance liquid chromatographic methods. According to spermiogram, infertile patients of both groups were also categorized into normozoospermic (N, no anomalies in the spermiogram), or into the subgroup including all patients with anomalies in the spermiogram (asthenoteratooligozoospermic ATO + asthenozoospermic A + teratozoospermic T + oligozoospermic O).Main Results and the Role Of Chance: In the master group, results indicated that 21/26 compounds assayed in seminal plasma of infertile males were significantly different from corresponding values determined in fertile controls. These 21 compounds constituted the male infertility biomarkers. Similar results were recorded in patients of the validation group. Using an index cumulating the biochemical seminal plasma anomalies (Biomarker Score), we found that fertile controls had mean Biomarker Score values of 2.01 ± 1.42, whilst infertile patients of the master and of the validation group had mean values of 12.27 ± 3.15 and of 11.41 ± 4.09, respectively (P < 0.001 compared to controls). The lack of statistical differences between the master and the validation groups, in both the metabolic profiles and the Biomarker Score values, allowed to pool patients into a single cohort of infertile males. The Biomarker Score values showed that fertile controls and infertile males clustered into two distinct groups. Infertile patients without (N, n = 42) or with (ATO + A + T + O, n = 98) spermiogram anomalies differed in some biomarkers (ascorbic acid, all-trans retinol, α-tocopherol, cytidine, uridine, guanine). These differences were reinforced by distribution frequencies and posterior probability curves of the Biomarker Score in the three groups.Limitations, Reasons For Caution: Results were obtained in relatively limited number of human seminal plasma samples. Using the 'Redox Energy Test' it was possible to associate specific metabolic profiles and values of the Biomarker Score to fertile controls or infertile males. However, it was not possible to evaluate whether the different anomalies of the spermiogram are associated with specific metabolic profiles and values of the Biomarker Score.Wider Implications Of the Findings: The 'Redox Energy Test', coupled with the Biomarker Score that cumulates the biochemical characteristics of seminal plasma into a single index, evidenced a set of low-molecular weight biomarkers potentially useful in the laboratory management of male infertility.Study Funding/competing Interest(s): The study was partly funded with research grants from the University of Catania. None of the authors have any conflicting interests to declare. [ABSTRACT FROM AUTHOR]

Long‑term exposure to ultraviolet (UV) radiation is associated with pathological alterations of the retinal pigment epithelium (RPE). It has been indicated that Cortistatin (CST) and somatostatin (SST) are able to inhibit the neurodegeneration of the RPE associated with diabetic retinopathy and retinal ischemia via activation of SST receptors (SSTRs). To the best of our knowledge, the present study indicated for the first time that treatment with UV‑A (30 and 60 min) causes an increase of CST expression, rather than SST, which was linked with the upregulation of STTR3,4,5 subtype receptor gene expression levels. The study revealed that: i) SST and CST mRNA expression were both detected under basal conditions in a human retinal pigment epithelial cell line (Arpe‑19); ii) SST expression remained constant from baseline to 1 h of UV‑A treatment; iii) CST mRNA expression levels were 80 times increased compared with time 0 and after 30 min of exposition to ultraviolet irradiation; iv) SSTR1, SSTR2 mRNA and low levels of SSTR4 were expressed in basal conditions, whereas SSTR3 and SSTR5 mRNA were not detected under the same conditions; and v) only SSTR3, SSTR4 and SSTR5 were overexpressed after UV‑A treatment, although in a different way. In conclusion, the findings provide reasonable evidence to support the pathophysiological role of the CST/SST/SSTRs system in the adaptive response of the RPE exposed to UV‑A radiation. [ABSTRACT FROM AUTHOR]

Multiple sclerosis (MS) is characterized by primary inflammation, demyelination, and progressive neurodegeneration. A biochemical MS feature is neuronal mitochondrial dysfunction, compensated by anaerobic metabolism increase, likely aggravating progression of neurodegeneration. Here, we characterized a pragmatic serum profile of compounds related to mitochondrial energy metabolism of potential clinical use. Blood samples of 518 well characterized (disability, disease course) MS patients and 167 healthy controls were analyzed for serum purines, pyrimidines, creatinine, and lactate. Nine of the 15 compounds assayed, hypoxanthine, xanthine, uric acid, inosine, uracil, β-pseudouridine, uridine, creatinine, and lactate, differed significantly between MS patients and controls ( p < 0.0001). Using these nine compounds, a unifying Biomarker Score was calculated. Controls and MS patients had mean Biomarker Scores of 0.4 ± 0.7 and 4.4 ± 1.9, respectively ( p < 0.00001). The Biomarker Score was higher in patients with progressive (6.0 ± 1.8 than with relapsing remitting disease course (3.6 ± 1.5, p < 0.00001). High association between the Biomarker Score and increase in disability (EDSS) was also observed. Additionally, in 50 patients who underwent magnetic resonance imaging (MRI), increase in the Biomarker Score correlated to neuroanatomical alterations. These results, obtained in a large cohort of MS patients evaluated for serum metabolic compounds connected to energy metabolism, demonstrated that the Biomarker Score might represent a pragmatic, resource saving, easy to obtain, laboratory tool useful to monitor MS patients and predict at an early stage who will switch from an RR to a progressive disease course. For the first time, it was also clearly shown a link between mitochondrial dysfunction and MRI lesions characteristic of MS. [ABSTRACT FROM AUTHOR]

Background: Adenylosuccinate lyase (ADSL) deficiency is a defect of purine metabolism affecting purinosome assembly and reducing metabolite fluxes through purine de novo synthesis and purine nucleotide recycling pathways. The disorder shows a wide spectrum of symptoms from slowly to rapidly progressing forms. The most severe form is characterized by neonatal encephalopathy, absence of spontaneous movement, respiratory failure, intractable seizures, and early death within the first weeks of life. More commonly, ADSL presents purely neurologic clinical picture characterized by severe psychomotor retardation, microcephaly, early onset of seizures, and autistic features (type I) or a more slowly progressing form with later onset, and major features including slight to moderate psychomotor retardation, and transient contact disturbances (type II). Diagnostic markers are the presence of succinylaminoimidazole carboxamide riboside (SAICAr) and succinyladenosine (SAdo) in extracellular fluids. ADSL is a rare disorder, although its prevalence remains unknown. Of note, the wide range of essentially nonspecific manifestations and lack of awareness of the condition often prevent diagnosis. Case presentation: We present here the case of particularly mild, late onset ADSL that has been unsuccessfully investigated until whole exome sequencing (WES) was performed. Conclusions: Besides emphasizing the valuable diagnostic value of WES, this report provides new data further documenting the relatively wide clinical manifestation of ADSL. [ABSTRACT FROM AUTHOR]

In this study, the metabolic, enzymatic and gene changes causing cerebral glucose dysmetabolism following graded diffuse traumatic brain injury (TBI) were evaluated. TBI was induced in rats by dropping 450 g from 1 (mild TBI; mTBI) or 2 m height (severe TBI; sTBI). After 6, 12, 24, 48, and 120 h gene expressions and enzymatic activities of glycolysis and pentose phosphate pathway (PPP) enzymes, and levels of lactate, ATP, ADP, ATP/ADP (indexing mitochondrial phosphorylating capacity), NADP + , NADPH and GSH were determined in whole brain extracts (n = 9 rats at each time for both TBI levels). Sham-operated animals (n = 9) were used as controls. Results demonstrated that mTBI caused a late increase (48–120 h post injury) of glycolytic gene expression and enzymatic activities, concomitantly with mitochondrial functional recovery (ATP and ATP/ADP normalization). No changes in lactate and PPP genes and enzymes, were accompanied by transient decrease in GSH, NADP + , NADPH and NADPH/NADP + . Animals following sTBI showed early increase (6–24 h post injury) of glycolytic gene expression and enzymatic activities, occurring during mitochondrial malfunctioning (50% decrease in ATP and ATP/ADP). Higher lactate and lower GSH, NADP + , NADPH, NADPH/NADP + than controls were recorded at anytime post injury (p < 0.01). Both TBI levels caused metabolic and gene changes affecting glucose metabolism. Following mTBI, increased glucose flux through glycolysis is coupled to mitochondrial glucose oxidation. “True” hyperglycolysis occurs only after sTBI, where metabolic changes, caused by depressed mitochondrial phosphorylating capacity, act on genes causing net glycolytic flux increase uncoupled from mitochondrial glucose oxidation. [ABSTRACT FROM AUTHOR]

Traumatic brain injury (TBI) is a leading and rising cause of death and disability worldwide. There is great interest in S100B and Glial Fibrillary Acid Protein (GFAP) as candidate biomarkers of TBI for diagnosis, triage, prognostication and drug development. However, conflicting results especially on S100B hamper their routine application in clinical practice. To try to address this question, we mimicked TBI damage utilizing a well-validated, simplified in vitro model of graded stretch injury induced in rat organotypic hippocampal slice cultures (OHSC). Different severities of trauma, from mild to severe, have been tested by using an equi-biaxial stretch of the OHSCs at a specified Lagrangian strain of 0 (controls), 5, 10, 20 and 50 %. OHSC were analysed at 3, 6, 18, 24, 48 and 96 h post-injury. Cell death, gene expressions and release into the culture medium of S100B and GFAP were determined at each time point. Gene expression and release of S100B slightly increased only in 20 and 50 % stretched OHSC. GFAP over-expression occurred in 10, 20 and 50 % and was inversely correlated with time post-injury. GFAP release significantly increased with time at any level of injury ( p < 0.01 with respect to controls). Consequently, the total amount of GFAP released showed a strong linear relationship with the severity of injury (R = 0.7662; p < 0.001). Under these experimental conditions, S100B seems to be useful in diagnosing only moderate to severe TBI-like injuries. Differently, GFAP demonstrates adequate biomarker requisites since its cellular release is affected by all grades of injury severity. [ABSTRACT FROM AUTHOR]

Objectives To test for structural and functional contribution of mitochondrial dysfunction to neurodegeneration in multiple sclerosis ( MS). A visual pathway model void of MS lesions was chosen in order to exclude neurodegeneration secondary to lesion related axonotmesis. Methods A single-centre cohort study (230 MS patients, 63 controls). Spectral domain optical coherence tomography of the retina, 3T magnetic resonance imaging of the brain, spectrophotometric assessment of serum lactate levels. Postmortem immunohistochemistry. Results The visual pathway was void of MS lesions in 31 patients and 31 age-matched controls. Serum lactate was higher in MS compared to controls ( P = 0.029). High serum lactate was structurally related to atrophy of the retinal nerve fiber layer at the optic disc ( P = 0.041), macula ( P = 0.025), and the macular ganglion cell complex ( P = 0.041). High serum lactate was functionally related to poor color vision ( P < 0.01), Expanded Disability Status Scale score (R = 0.37, P = 0.041), Guy's Neurological disability score (R = 0.38, P = 0.037), MS walking scale (R = 0.50, P = 0.009), upper limb motor function ( R = 0.53, P = 0.002). Immunohistochemistry demonstrated increased astrocytic expression of a key lactate generating enzyme in MS lesions as well as profound vascular expression of monocarboxylate transporter-1, which is involved in lactate transport. Interpretation This study provides structural, functional, and translational evidence for visual pathway neurodegeneration in MS related to mitochondrial dysfunction. [ABSTRACT FROM AUTHOR]

Breast cancer is the most frequent tumor and the leading cause of cancer deaths in women. In recent years, lactate metabolism and, in particular, its receptor GPR81 have been shown to play a vital role in cancer biology. GPR81 is upregulated in breast cancer and promotes tumor growth by tumor cell-derived lactate. Therefore, the search for possible crosstalk and the involvement of new molecules capable of generating this pathology is always in continuous development. In this study, the relationship between GPR81 and IGFBP6 protein in tumor growth and oxidative stress in the human breast cancer cell line MDA-MB-231 was studied. Cells were treated with lactate or the GPR81 receptor agonist and antagonist 3,5-DHBA and 3-OBA, respectively. In addition, oxidative stress and proliferation were also evaluated in cells challenged with the recombinant IGFBP6 protein. Our data showed that lactate induced cell proliferation and wound healing of the MDA-231 breast cancer cell through the overexpression of both the lactate receptor GPR81 and IGFBP6. The increase in IGFBP6 was able, in turn, to improve the mitochondrial fitness and redox state, as suggested by the reduced levels of mitochondrial ROS production after IGFBP6 treatment, presumably mediated by the increase in the ROS detoxifying genes HMOX1, GSTK1 and NQO1. In conclusion, our data highlight a novel axis between GPR81 and IGFBP6 in MDA-231 cells able to modulate lactate metabolism and oxidative stress. This complex signaling may represent a new therapeutic target for breast cancer. [ABSTRACT FROM AUTHOR]

In this study, we investigated the hypothesis that mild traumatic brain injury (mTBI) triggers a controlled gene program as an adaptive response finalized to neuroprotection, similar to that found in hibernators and in ischemic preconditioning. A stretch injury device was used to produce an equi-biaxial strain field in rat organotypic hippocampal slice cultures at a specified Lagrangian strain of 10 % and a constant strain rate of 20 s. After 24 h from injury, propidium iodide staining, HPLC analysis of metabolites and microarray analysis of cDNA were performed to evaluate cell viability, cell energy state and gene expression, respectively. Compared to control cultures, 10 % stretch injured cultures showed no change in viability, but demonstrated a hypometabolic state (decreased ATP, ATP/ADP, and nicotinic coenzymes) and a peculiar pattern of gene modulation. The latter was characterized by downregulation of genes encoding for proteins of complexes I, III, and IV of the mitochondrial electron transport chain and of ATP synthase; downregulation of transcriptional and translational genes; downregulation and upregulation of genes controlling the synthesis of glutamate and GABA receptors, upregulation of calmodulin and calmodulin-binding proteins; proper modulation of genes encoding for proapoptotic and antiapoptotic proteins. These results support the hypothesis that, following mTBI, a hibernation-type response is activated in non-hibernating species. Unlike in hibernators and ischemic preconditioning, this adaptive gene programme, aimed at achieving maximal neuroprotection, is not triggered by decrease in oxygen availability. It seems rather activated to avoid increase in oxidative/nitrosative stress and apoptosis during a transient period of mitochondrial malfunctioning. [ABSTRACT FROM AUTHOR]

Nearly 40–50% of infertility problems are estimated to be of female origin. Previous studies dedicated to the analysis of metabolites in follicular fluid (FF) produced contrasting results, although some valuable indexes capable to discriminate control groups (CTRL) from infertile females (IF) and correlate with outcome measures of assisted reproduction techniques were in some instances found. In this study, we analyzed in blind FF of 35 control subjects (CTRL = patients in which inability to obtain pregnancy was exclusively due to a male factor) and 145 IF (affected by: endometriosis, n = 19; polycystic ovary syndrome, n = 14; age-related reduced ovarian reserve, n = 58; reduced ovarian reserve, n = 29; unexplained infertility, n = 14; genetic infertility, n = 11) to determine concentrations of 55 water- and fat-soluble low molecular weight compounds (antioxidants, oxidative/nitrosative stress-related compounds, purines, pyrimidines, energy-related metabolites, and amino acids). Results evidenced that 27/55 of them had significantly different values in IF with respect to those measured in CTRL. The metabolic pattern of these potential biomarkers of infertility was cumulated (in both CTRL and IF) into a Biomarker Score index (incorporating the metabolic anomalies of FF), that fully discriminated CTRL (mean Biomarker Score value = 4.00 ± 2.30) from IF (mean Biomarker Score value = 14.88 ± 3.09, p < 0.001). The Biomarker Score values were significantly higher than those of CTRL in each of the six subgroups of IF. Posterior probability curves and ROC curve indicated that values of the Biomarker Score clustered CTRL and IF into two distinct groups, based on the individual FF metabolic profile. Furthermore, Biomarker Score values correlated with outcome measures of ovarian stimulation, in vitro fertilization, number and quality of blastocysts, clinical pregnancy, and healthy offspring. These results strongly suggest that the biochemical quality of FF deeply influences not only the effectiveness of IVF procedures but also the following embryonic development up to healthy newborns. The targeted metabolomic analysis of FF (using empowered Redox Energy Test) and the subsequent calculation of the Biomarker Score evidenced a set of 27 low molecular weight infertility biomarkers potentially useful in the laboratory managing of female infertility and to predict the success of assisted reproduction techniques. [ABSTRACT FROM AUTHOR]

Oxidative/nitrosative stress and mitochondrial dysfunction is a hallmark of amyotrophic lateral sclerosis (ALS), an invariably fatal progressive neurodegenerative disease. Here, as an exploratory arm of a phase II clinical trial (EudraCT Number 2017-005065-47), we used high performance liquid chromatography(HPLC) to investigate changes in the metabolic profiles of serum from ALS patients treated weekly for 4 weeks with a repeated sub-cutaneous dose of 1 mg/kg of a proprietary low molecular weight dextran sulphate, called ILB®. A significant normalization of the serum levels of several key metabolites was observed over the treatment period, including N-acetylaspartate (NAA), oxypurines, biomarkers of oxidative/nitrosative stress and antioxidants. An improved serum metabolic profile was accompanied by significant amelioration of the patients' clinical conditions, indicating a response to ILB® treatment that appears to be mediated by improvement of tissue bioenergetics, decrease of oxidative/nitrosative stress and attenuation of (neuro)inflammatory processes. [ABSTRACT FROM AUTHOR]

Carbon-based nanomaterials are nowadays attracting lots of attention, in particular in the biomedical field, where they find a wide spectrum of applications, including, just to name a few, the drug delivery to specific tumor cells and the improvement of non-invasive imaging methods. Nanoparticles inhaled during breathing accumulate in the lung alveoli, where they interact and are covered with lung surfactants. We recently demonstrated that an apparently non-toxic concentration of engineered carbon nanodiamonds (ECNs) is able to induce oxidative/nitrosative stress, imbalance of energy metabolism, and mitochondrial dysfunction in microglial and alveolar basal epithelial cells. Therefore, the complete understanding of their "real" biosafety, along with their possible combination with other molecules mimicking the in vivo milieu, possibly allowing the modulation of their side effects becomes of utmost importance. Based on the above, the focus of the present work was to investigate whether the cellular alterations induced by an apparently non-toxic concentration of ECNs could be counteracted by their incorporation into a synthetic lung surfactant (DPPC:POPG in 7:3 molar ratio). By using two different cell lines (alveolar (A549) and microglial (BV-2)), we were able to show that the presence of lung surfactant decreased the production of ECNs-induced nitric oxide, total reactive oxygen species, and malondialdehyde, as well as counteracted reduced glutathione depletion (A549 cells only), ameliorated cell energy status (ATP and total pool of nicotinic coenzymes), and improved mitochondrial phosphorylating capacity. Overall, our results on alveolar basal epithelial and microglial cell lines clearly depict the benefits coming from the incorporation of carbon nanoparticles into a lung surfactant (mimicking its in vivo lipid composition), creating the basis for the investigation of this combination in vivo. [ABSTRACT FROM AUTHOR]