Carnosine is a natural endogenous dipeptide widely distributed in mammalian tissues, existing at particularly high concentrations in the muscles and brain and possesses well-characterized antioxidant ...and anti-inflammatory activities. In an in vitro model of macrophage activation, induced by lipopolysaccharide + interferon-gamma (LPS + IFN-γ), we here report the ability of carnosine to modulate pro-oxidant and pro-inflammatory activities of macrophages, representing the primary cell type that is activated as a part of the immune response. An ample set of parameters aimed to evaluate cytotoxicity (MTT assay), energy metabolism (HPLC), gene expressions (high-throughput real-time PCR (qRT-PCR)), protein expressions (western blot) and nitric oxide production (qRT-PCR and HPLC), was used to assess the effects of carnosine on activated macrophages challenged with a non cytotoxic LPS (100 ng/mL) + IFN-γ (600 U/mL) concentration. In our experimental model, main carnosine beneficial effects were: (1) the modulation of nitric oxide production and metabolism; (2) the amelioration of the macrophage energy state; (3) the decrease of the expressions of pro-oxidant enzymes (Nox-2, Cox-2) and of the lipid peroxidation product malondialdehyde; (4) the restoration and/or increase of the expressions of antioxidant enzymes (Gpx1, SOD-2 and Cat); (5) the increase of the transforming growth factor-β1 (TGF-β1) and the down-regulation of the expressions of interleukins 1β and 6 (IL-1β and IL-6) and 6) the increase of the expressions of Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and heme oxygenase-1 (HO-1). According to these results carnosine is worth being tested in the treatment of diseases characterized by elevated levels of oxidative stress and inflammation (atherosclerosis, cancer, depression, metabolic syndrome, and neurodegenerative diseases).
Tau is a microtubule‐associated protein exerting several physiological functions in neurons. In Alzheimer's disease (AD) misfolded tau accumulates intraneuronally and leads to axonal degeneration. ...However, tau has also been found in the extracellular medium. Recent studies indicated that extracellular tau uploaded from neurons causes synaptic dysfunction and contributes to tau pathology propagation. Here we report novel evidence that extracellular tau oligomers are abundantly and rapidly accumulated in astrocytes where they disrupt intracellular Ca2+ signaling and Ca2+‐dependent release of gliotransmitters, especially ATP. Consequently, synaptic vesicle release, the expression of pre‐ and postsynaptic proteins, and mEPSC frequency and amplitude were reduced in neighboring neurons. Notably, we found that tau uploading from astrocytes required the amyloid precursor protein, APP. Collectively, our findings suggests that astrocytes play a critical role in the synaptotoxic effects of tau via reduced gliotransmitter availability, and that astrocytes are major determinants of tau pathology in AD.
Main Points
Tau oligomers enter astrocytes more efficiently than neurons.
Tau treatment affects Ca2+‐dependent gliotransmitter release.
Tau induces ATP‐dependent inhibition of synaptic protein expression and synaptic transmission.
Deregulation of the tricarboxylic acid cycle (TCA) due to mutations in specific enzymes or defective aerobic metabolism is associated with tumour growth. Aconitase 2 (ACO2) participates in the TCA ...cycle by converting citrate to isocitrate, but no evident demonstrations of its involvement in cancer metabolism have been provided so far.
Biochemical assays coupled with molecular biology, in silico, and cellular tools were applied to circumstantiate the impact of ACO2 in the breast cancer cell line MCF-7 metabolism. Fluorescence lifetime imaging microscopy (FLIM) of NADH was used to corroborate the changes in bioenergetics.
We showed that ACO2 levels are decreased in breast cancer cell lines and human tumour biopsies. We generated ACO2- overexpressing MCF-7 cells and employed comparative analyses to identify metabolic adaptations. We found that increased ACO2 expression impairs cell proliferation and commits cells to redirect pyruvate to mitochondria, which weakens Warburg-like bioenergetic features. We also demonstrated that the enhancement of oxidative metabolism was supported by mitochondrial biogenesis and FoxO1-mediated autophagy/mitophagy that sustains the increased ROS burst.
This work identifies ACO2 as a relevant gene in cancer metabolic rewiring of MCF-7 cells, promoting a different utilisation of pyruvate and revealing the potential metabolic vulnerability of ACO2-associated malignancies.
The COVID-19 global pandemic is caused by SARS-CoV-2, and represents an urgent medical and social issue. Unfortunately, there is still not a single proven effective drug available, and therefore, ...current therapeutic guidelines recommend supportive care including oxygen administration and treatment with antibiotics. Recently, patients have been also treated with off-label therapies which comprise antiretrovirals, anti-inflammatory compounds, antiparasitic agents and plasma from convalescent patients, all with controversial results. The ubiquitin-proteasome system (UPS) is important for the maintenance of cellular homeostasis, and plays a pivotal role in viral replication processes. In this review, we discuss several aspects of the UPS and the effects of its inhibition with particular regard to the life cycle of the coronaviruses (CoVs). In fact, proteasome inhibition by various chemical compounds, such as MG132, epoxomycin and bortezomib, may reduce the virus entry into the eucariotic cell, the synthesis of RNA, and the subsequent protein expression necessary for CoVs. Importantly, since UPS inhibitors reduce the cytokine storm associated with various inflammatory conditions, it is reasonable to assume that they might be repurposed for SARS-CoV-2, thus providing an additional tool to counteract both virus replication as well as its most deleterious consequences triggered by abnormal immunological response.
Mitochondrial dynamics are regulated by a complex system of proteins representing the mitochondrial quality control (MQC). MQC balances antagonistic forces of fusion and fission determining ...mitochondrial and cell fates. In several neurological disorders, dysfunctional mitochondria show significant changes in gene and protein expression of the MQC and contribute to the pathophysiological mechanisms of cell damage. In this study, we evaluated the main gene and protein expression involved in the MQC in rats receiving traumatic brain injury (TBI) of different severities. At 6, 24, 48 and 120 hours after mild TBI (mTBI) or severe TBI (sTBI), gene and protein expressions of fusion and fission were measured in brain tissue homogenates. Compared to intact brain controls, results showed that genes and proteins inducing fusion or fission were upregulated and downregulated, respectively, in mTBI, but downregulated and upregulated, respectively, in sTBI. In particular, OPA1, regulating inner membrane dynamics, cristae remodelling, oxidative phosphorylation, was post-translationally cleaved generating differential amounts of long and short OPA1 in mTBI and sTBI. Corroborated by data referring to citrate synthase, these results confirm the transitory (mTBI) or permanent (sTBI) mitochondrial dysfunction, enhancing MQC importance to maintain cell functions and indicating in OPA1 an attractive potential therapeutic target for TBI.
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 450g from 1 (mild TBI; mTBI) or 2m height (severe TBI; sTBI). After 6, 12, 24, 48, and 120h 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 (48–120h 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 (6–24h 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.
•Metabolic, enzymatic and gene changes are involved in glucose dysmetabolism after TBI.•Activation of glycolysis in mild TBI occurs on recovery of mitochondrial functions.•Activation of glycolysis in severe TBI occurs during mitochondrial malfunctioning.•Pentosephosphate pathway changes in severe TBI are limited to NADPH supply.•Metabolic and gene mechanisms of glucose dysregulation depend on TBI severity.
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.
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.
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.
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.
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.
EudraCT 2017-005065-47.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Ozone therapy has been widely used in everyday clinical practice over the last few years, leading to significant clinical results in the treatment of herniated discs and pain management. ...Nevertheless, further studies have demonstrated its potential efficacy and safety under other clinical and experimental conditions. However, some of these studies showed controversial results regarding the safety and efficacy of ozone therapy, thus mining its potential use in an everyday clinical practice. To this regard, it should be considered that extensive literature review reported the use of ozone in a significant different dose range and with different delivery systems. The aim of the present review is to describe the various pharmacological effects of ozone in different organs and clinical conditions and to provide possible biochemical and molecular insights for ozone biological properties, thus providing a possible explanation for various controversial clinical outcomes described in the scientific literature.
Using a closed-head impact acceleration model of mild or severe traumatic brain injury (mTBI or sTBI, respectively) in rats, we evaluated the effects of graded head impacts on the gene and protein ...expressions of pyruvate dehydrogenase (PDH), as well as major enzymes of mitochondrial tricarboxylic acid cycle (TCA). TBI was induced in anaesthetized rats by dropping 450 g from 1 (mTBI) or 2 m height (sTBI). After 6 h, 12 h, 24 h, 48 h, and 120 h gene expressions of enzymes and subunits of PDH. PDH kinases and phosphatases (PDK1-4 and PDP1-2, respectively), citrate synthase (CS), isocitrate dehydrogenase (IDH), oxoglutarate dehydrogenase (OGDH), succinate dehydrogenase (SDH), succinyl-CoA synthase (SUCLG), and malate dehydrogenase (MDH) were determined in whole brain extracts (n = 6 rats at each time for both TBI levels). In the same samples, the high performance liquid chromatographic (HPLC) determination of acetyl-coenzyme A (acetyl-CoA) and free coenzyme A (CoA-SH) was performed. Sham-operated animals (
= 6) were used as controls. After mTBI, the results indicated a general transient decrease, followed by significant increases, in PDH and TCA gene expressions. Conversely, permanent PDH and TCA downregulation occurred following sTBI. The inhibitory conditions of PDH (caused by PDP1-2 downregulations and PDK1-4 overexpression) and SDH appeared to operate only after sTBI. This produced almost no change in acetyl-CoA and free CoA-SH following mTBI and a remarkable depletion of both compounds after sTBI. These results again demonstrated temporary or steady mitochondrial malfunctioning, causing minimal or profound modifications to energy-related metabolites, following mTBI or sTBI, respectively. Additionally, PDH and SDH appeared to be highly sensitive to traumatic insults and are deeply involved in mitochondrial-related energy metabolism imbalance.
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