Effects of Fitlight training on cognitive-motor performance in élite judo athletes.

Aims: The aims of this study were to verify if a 5-week cognitive-motor training (CMT) using FitlightsTM induced changes in young adult judo athletes compared to a non-intervention group. Specifically, it was verified if CMT influenced executive functions (EFs), physical fitness and brain-derived neurotrophic factor (BDNF) levels. Additionally, athletes' competitive results were compared between groups. Method: Twenty-seven athletes (14 males and 13 females; age = 19.5 ± 2.0 years) were assigned to the Fitlight (FG) and control (CG) groups which performed 5 weeks of CMT, respectively, including 25 min per day of Fitlight training or traditional judo practice. All participants performed cognitive (flanker task and forward/backward digit span) and fitness tests (counter movement jump, handgrip test, dynamic and isometric chin up). In addition, BDNF was collected by saliva sampling and competitive results after the intervention period were considered. Results: RM-ANOVA showed significant differences in FG for the accuracy of flanker (p = 0.028) and backward digit span (p < 0.001). Moreover, significant differences in FG were found for relative dynamic chin up (p = 0.027) and counter movement jump (p = 0.05). In addition, a significant difference in FG was found for competitive results after the intervention period (p < 0.01).No significant differences were found for BDNF and other cognitive and fitness measures (p > 0.05). Conclusion: A 5-week judo-specific CMT improved EFs and motor performance in élite judo athletes. It seems that CMT with Fitlight™ could be considered an additional support to coaches during the training period.

Molecular Biology Digest of Cell Mitophagy.

The homeostasis of eukaryotic cells relies on efficient mitochondrial function. The control of mitochondrial quality is framed by the combination of distinct but interdependent mechanisms spanning biogenesis, regulation of dynamic network, and finely tuned degradation either through ubiquitin-proteasome system or autophagy (mitophagy). There is continuous evolution on the pathways orchestrating the mitochondrial response to stress signals and the organelle adaptation to quality control during acute and subtle dysfunctions. Notably, it remains indeed ill-defined whether active mitophagy leads to cell survival or death by defective mitochondrial degradation. Above all, uncharted is whether and how pharmacologically tackle these mechanisms may lead to conceive novel therapeutic strategies for treating conditions associated with the defective mitochondria. Here, we attempt to provide a chronological and comprehensive overview of the determining discoveries, which have led to the current knowledge of mitophagy.

Outcome of Transplantation Using Organs From Donors Infected or Colonized With Carbapenem-Resistant Gram-Negative Bacteria.

Donor-derived infections due to multidrug-resistant bacteria are a growing problem in solid organ transplantation, and optimal management options are not clear. In a 2-year period, 30/214 (14%) recipients received an organ from 18/170 (10.5%) deceased donors with infection or colonization caused by a carbapenem-resistant gram-negative bacteria that was unknown at the time of transplantation. Among them, 14/30 recipients (47%) received a transplant from a donor with bacteremia or with infection/colonization of the transplanted organ and were considered at high risk of donor-derived infection transmission. The remaining 16/30 (53%) recipients received an organ from a nonbacteremic donor with colonization of a nontransplanted organ and were considered at low risk of infection transmission. Proven transmission occurred in 4 of the 14 high-risk recipients because donor infection was either not recognized, underestimated, or not communicated. These recipients received late, short or inappropriate posttransplant antibiotic therapy. Transmission did not occur in high-risk recipients who received appropriate and prompt antibiotic therapy for at least 7 days. The safe use of organs from donors with multidrug-resistant bacteria requires intra- and inter-institutional communication to allow appropriate management and prompt treatment of recipients in order to avoid transmission of infection.

Improving the reliability of single-subject fMRI by weighting intra-run variability.

At present, functional magnetic resonance imaging (fMRI) is one of the most useful methods of studying cognitive processes in the human brain in vivo, both for basic science and clinical goals. Although neuroscience studies often rely on group analysis, clinical applications must investigate single subjects (patients) only. Particularly for the latter, issues regarding the reliability of fMRI readings remain to be resolved. To determine the ability of intra-run variability (IRV) weighting to consistently detect active voxels, we first acquired fMRI data from a sample of healthy subjects, each of whom performed 4 runs (4 blocks each) of self-paced finger-tapping. Each subject's data was analyzed using single-run general linear model (GLM), and each block was then analyzed separately to calculate the IRV weighting. Results show that integrating IRV information into standard single-subject GLM activation maps significantly improved the reliability (p=0.007) of the single-subject fMRI data. This suggests that taking IRV into account can help identify the most constant and relevant neuronal activity at the single-subject level.

Transglutaminase 2 ablation leads to mitophagy impairment associated with a metabolic shift towards aerobic glycolysis.

Macroautophagy selectively degrades dysfunctional mitochondria by a process known as mitophagy. Here we demonstrate the involvement of transglutaminase 2 (TG2) in the turnover and degradation of damaged mitochondria. In TG2-ablated cells we observed the presence of a large number of fragmented mitochondria that display decreased membrane potential, downregulation of IF1 along with increased Drp1 and PINK1 levels, two key proteins regulating the mitochondrial fission. Of note, we demonstrate that in healthy mitochondria, TG2 interacts with the dynamic proteins Drp1 and Fis1; interestingly, their interaction is largely reduced upon induction of the fission process by carbonyl cyanide m-chlorophenyl hydrazine (CCCP). In keeping with these findings, mitochondria lacking TG2 are more susceptible to CCCP treatment. As a consequence of accumulation of damaged mitochondria, cells lacking TG2 increased their aerobic glycolysis and became sensitive to the glycolytic inhibitor 2-deoxy-D-glucose (2-DG). In contrast, TG2-proficient cells are more resistant to 2-DG-induced apoptosis as the caspase 3 is inactivated through the enzyme's crosslinking activity. The data presented in this study show that TG2 plays a key role in cellular dynamics and consequently influences the energetic metabolism.

Essential versus accessory aspects of cell death: recommendations of the NCCD 2015.

Cells exposed to extreme physicochemical or mechanical stimuli die in an uncontrollable manner, as a result of their immediate structural breakdown. Such an unavoidable variant of cellular demise is generally referred to as 'accidental cell death' (ACD). In most settings, however, cell death is initiated by a genetically encoded apparatus, correlating with the fact that its course can be altered by pharmacologic or genetic interventions. 'Regulated cell death' (RCD) can occur as part of physiologic programs or can be activated once adaptive responses to perturbations of the extracellular or intracellular microenvironment fail. The biochemical phenomena that accompany RCD may be harnessed to classify it into a few subtypes, which often (but not always) exhibit stereotyped morphologic features. Nonetheless, efficiently inhibiting the processes that are commonly thought to cause RCD, such as the activation of executioner caspases in the course of apoptosis, does not exert true cytoprotective effects in the mammalian system, but simply alters the kinetics of cellular demise as it shifts its morphologic and biochemical correlates. Conversely, bona fide cytoprotection can be achieved by inhibiting the transduction of lethal signals in the early phases of the process, when adaptive responses are still operational. Thus, the mechanisms that truly execute RCD may be less understood, less inhibitable and perhaps more homogeneous than previously thought. Here, the Nomenclature Committee on Cell Death formulates a set of recommendations to help scientists and researchers to discriminate between essential and accessory aspects of cell death.

AMBRA1 is able to induce mitophagy via LC3 binding, regardless of PARKIN and p62/SQSTM1.

Damaged mitochondria are eliminated by mitophagy, a selective form of autophagy whose dysfunction associates with neurodegenerative diseases. PINK1, PARKIN and p62/SQTMS1 have been shown to regulate mitophagy, leaving hitherto ill-defined the contribution by key players in 'general' autophagy. In basal conditions, a pool of AMBRA1 - an upstream autophagy regulator and a PARKIN interactor - is present at the mitochondria, where its pro-autophagic activity is inhibited by Bcl-2. Here we show that, upon mitophagy induction, AMBRA1 binds the autophagosome adapter LC3 through a LIR (LC3 interacting region) motif, this interaction being crucial for regulating both canonical PARKIN-dependent and -independent mitochondrial clearance. Moreover, forcing AMBRA1 localization to the outer mitochondrial membrane unleashes a massive PARKIN- and p62-independent but LC3-dependent mitophagy. These results highlight a novel role for AMBRA1 as a powerful mitophagy regulator, through both canonical or noncanonical pathways.

Listening to speech recruits specific tongue motor synergies as revealed by transcranial magnetic stimulation and tissue-Doppler ultrasound imaging.

The activation of listener's motor system during speech processing was first demonstrated by the enhancement of electromyographic tongue potentials as evoked by single-pulse transcranial magnetic stimulation (TMS) over tongue motor cortex. This technique is, however, technically challenging and enables only a rather coarse measurement of this motor mirroring. Here, we applied TMS to listeners' tongue motor area in association with ultrasound tissue Doppler imaging to describe fine-grained tongue kinematic synergies evoked by passive listening to speech. Subjects listened to syllables requiring different patterns of dorso-ventral and antero-posterior movements (/ki/, /ko/, /ti/, /to/). Results show that passive listening to speech sounds evokes a pattern of motor synergies mirroring those occurring during speech production. Moreover, mirror motor synergies were more evident in those subjects showing good performances in discriminating speech in noise demonstrating a role of the speech-related mirror system in feed-forward processing the speaker's ongoing motor plan.

AD-linked, toxic NH2 human tau affects the quality control of mitochondria in neurons.

Functional as well as structural alterations in mitochondria size, shape and distribution are precipitating, early events in progression of Alzheimer's Disease (AD). We reported that a 20-22kDa NH2-tau fragment (aka NH2htau), mapping between 26 and 230 amino acids of the longest human tau isoform, is detected in cellular and animal AD models and is neurotoxic in hippocampal neurons. The NH2htau -but not the physiological full-length protein- interacts with Aß at human AD synapses and cooperates with it in inhibiting the mitochondrial ANT-1-dependent ADP/ATP exchange. Here we show that the NH2htau also adversely affects the interplay between the mitochondria dynamics and their selective autophagic clearance. Fragmentation and perinuclear mislocalization of mitochondria with smaller size and density are early found in dying NH2htau-expressing neurons. The specific effect of NH2htau on quality control of mitochondria is accompanied by (i) net reduction in their mass in correlation with a general Parkin-mediated remodeling of membrane proteome; (ii) their extensive association with LC3 and LAMP1 autophagic markers; (iii) bioenergetic deficits and (iv) in vitro synaptic pathology. These results suggest that NH2htau can compromise the mitochondrial biology thereby contributing to AD synaptic deficits not only by ANT-1 inactivation but also, indirectly, by impairing the quality control mechanism of these organelles.

Leaf growth dynamics in four plant species of the Patagonian Monte, Argentina.

Studying plant responses to environmental variables is an elemental key to understand the functioning of arid ecosystems. We selected four dominant species of the two main life forms. The species selected were two evergreen shrubs: Larrea divaricata and Chuquiraga avellanedae and two perennial grasses: Nassella tenuis and Pappostipa speciosa. We registered leaf/shoot growth, leaf production and environmental variables (precipitation, air temperature, and volumetric soil water content at two depths) during summer-autumn and winter-spring periods. Multiple regressions were used to test the predictive power of the environmental variables. During the summer-autumn period, the strongest predictors of leaf/shoot growth and leaf production were the soil water content of the upper layer and air temperature while during the winter-spring period, the strongest predictor was air temperature. In conclusion, we found that the leaf/shoot growth and leaf production were associated with current environmental conditions, specially to soil water content and air temperature.

IF1 limits the apoptotic-signalling cascade by preventing mitochondrial remodelling.

Mitochondrial structure has a central role both in energy conversion and in the regulation of cell death. We have previously shown that IF1 protects cells from necrotic cell death and supports cristae structure by promoting the oligomerisation of the F1Fo-ATPsynthase. As IF1 is upregulated in a large proportion of human cancers, we have here explored its contribution to the progression of apoptosis and report that an increased expression of IF1, relative to the F1Fo-ATPsynthase, protects cells from apoptotic death. We show that IF1 expression serves as a checkpoint for the release of Cytochrome c (Cyt c) and hence the completion of the apoptotic program. We show that the progression of apoptosis engages an amplification pathway mediated by: (i) Cyt c-dependent release of ER Ca(2+), (ii) Ca(2+)-dependent recruitment of the GTPase Dynamin-related protein 1 (Drp1), (iii) Bax insertion into the outer mitochondrial membrane and (iv) further release of Cyt c. This pathway is accelerated by suppression of IF1 and delayed by its overexpression. IF1 overexpression is associated with the preservation of mitochondrial morphology and ultrastructure, consistent with a central role for IF1 as a determinant of the inner membrane architecture and with the role of mitochondrial ultrastructure in the regulation of Cyt c release. These data suggest that IF1 is an antiapoptotic and potentially tumorigenic factor and may be a valuable predictor of responsiveness to chemotherapy.

Changes in the properties of normal human red blood cells during in vivo aging.

The changes in red blood cells (RBC) as they age and the mechanisms for their eventual removal have been of interest for many years. Proposed age-related changes include dehydration with increased density and decreased size, increased membrane IgG, loss of membrane phospholipid asymmetry, and decreased activity of KCl cotransport. The biotin RBC label allows unambiguous identification of older cells and exploration of their properties as they age. Autologous normal human RBC were labeled ex vivo and, after reinfusion, compared with unlabeled RBC throughout their lifespan. RBC density increased with age, with most of the change in the first weeks. Near the end of their lifespan, RBC had increased surface IgG. However, there was no evidence for elevated external phosphatidylserine (PS) even though older RBC had significantly lower activity of aminophospholipid translocase (APLT). KCl cotransport activity persisted well past the reticulocyte stage, but eventually decreased as the RBC became older. These studies place limitations on the use of density fractionation for the study of older human RBC, and do not support loss of phospholipid asymmetry as a mechanism for human RBC senescence. However, increased levels of IgG were associated with older RBC, and may contribute to their removal from the circulation.

The novel NOX inhibitor 2-acetylphenothiazine impairs collagen-dependent thrombus formation in a GPVI-dependent manner.

BACKGROUND AND PURPOSE: NADPH oxidases (NOXs) contribute to platelet activation by a largely unknown mechanism. Here, we studied the effect of the novel NOX inhibitor 2-acetylphenothiazine (2-APT) on human platelet functional responses and intracellular signaling pathways. EXPERIMENTAL APPROACH: The generation of superoxide ions was assessed by single cell imaging on adhering platelets using dihydroethidium (DHE), while other reactive oxygen species (ROS) were detected with 5-(and-6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate (CM-H(2)-DCFDA). Whole blood thrombus formation, washed platelet aggregation, integrin αIIbß3 inside-out signalling, Syk phosphorylation and PKC activation were analysed to understand the functional consequences of NOX inhibition by 2-APT in platelets. KEY RESULTS: Superoxide ion generation stimulated by platelet adhesion on collagen and fibrinogen was significantly inhibited by 2-APT in concentration-dependent manner (IC(50) = 306 nM and 227 nM, respectively), whereas cumulative ROS accumulation was not affected by this pharmacological agent. 2-APT also abolished collagen-dependent whole blood thrombus formation and washed platelet aggregation in response to collagen but not thrombin. The activation of integrin αIIbß3 and PKC in response to the GPVI-specific agonist collagen-related peptide (CRP) was significantly reduced, whereas the same responses to thrombin were not significantly affected by 2-APT. Finally, Syk activation in response to collagen but not thrombin was inhibited by 2-APT. CONCLUSIONS AND IMPLICATIONS: Taken together, our results suggest that 2-APT attenuates GPVI-specific signalling and is a novel inhibitor of collagen-induced platelet responses. Therefore, NOXs could represent a novel target for the discovery of anti-thrombotic drugs.

The autophagy-associated factors DRAM1 and p62 regulate cell migration and invasion in glioblastoma stem cells.

The aggressiveness of glioblastoma multiforme (GBM) is defined by local invasion and resistance to therapy. Within established GBM, a subpopulation of tumor-initiating cells with stem-like properties (GBM stem cells, GSCs) is believed to underlie resistance to therapy. The metabolic pathway autophagy has been implicated in the regulation of survival in GBM. However, the status of autophagy in GBM and its role in the cancer stem cell fraction is currently unclear. We found that a number of autophagy regulators are highly expressed in GBM tumors carrying a mesenchymal signature, which defines aggressiveness and invasion, and are associated with components of the MAPK pathway. This autophagy signature included the autophagy-associated genes DRAM1 and SQSTM1, which encode a key regulator of selective autophagy, p62. High levels of DRAM1 were associated with shorter overall survival in GBM patients. In GSCs, DRAM1 and SQSTM1 expression correlated with activation of MAPK and expression of the mesenchymal marker c-MET. DRAM1 knockdown decreased p62 localization to autophagosomes and its autophagy-mediated degradation, thus suggesting a role for DRAM1 in p62-mediated autophagy. In contrast, autophagy induced by starvation or inhibition of mTOR/PI-3K was not affected by either DRAM1 or p62 downregulation. Functionally, DRAM1 and p62 regulate cell motility and invasion in GSCs. This was associated with alterations of energy metabolism, in particular reduced ATP and lactate levels. Taken together, these findings shed new light on the role of autophagy in GBM and reveal a novel function of the autophagy regulators DRAM1 and p62 in control of migration/invasion in cancer stem cells.

HtrA2 deficiency causes mitochondrial uncoupling through the F1F0-ATP synthase and consequent ATP depletion.

Loss of the mitochondrial protease HtrA2 (Omi) in mice leads to mitochondrial dysfunction, neurodegeneration and premature death, but the mechanism underlying this pathology remains unclear. Using primary cultures from wild-type and HtrA2-knockout mice, we find that HtrA2 deficiency significantly reduces mitochondrial membrane potential in a range of cell types. This depolarisation was found to result from mitochondrial uncoupling, as mitochondrial respiration was increased in HtrA2-deficient cells and respiratory control ratio was dramatically reduced. HtrA2-knockout cells exhibit increased proton translocation through the ATP synthase, in combination with decreased ATP production and truncation of the F1 α-subunit, suggesting the ATP synthase as the source of the proton leak. Uncoupling in the HtrA2-deficient mice is accompanied by altered breathing pattern and, on a cellular level, ATP depletion and vulnerability to chemical ischaemia. We propose that this vulnerability may ultimately cause the neurodegeneration observed in these mice.

The 18 kDa translocator protein (TSPO): a new perspective in mitochondrial biology.

In mammals, mitochondria are central in maintaining normal cell function and dissecting the pathways that govern their physiology and pathology is therefore of utmost importance. For a long time, the science world has acknowledged the Translocator Protein (TSPO), an intriguing molecule that, through its position and association with biological processes, stands as one of the hidden regulatory pathways in mitochondrial homeostasis. Here we aim to review the literature and highlight what links TSPO to mitochondrial homeostasis in order to delineate its contribution in the functioning and malfunctioning of this critical organelle. In detail, we will discuss: 1) TSPO localization and interplay with controlling phenomena of mitochondria (e.g. mPTP); 2) TSPO interaction with the prominent mitochondrial player VDAC; consider evidence on how TSPO relates to 3) mitochondrial energy production; 4) Ca2+ signalling and 5) the generation of Reactive Oxygen Species (ROS) before finally describing 6) its part in apoptotic cell death. In essence, we hope to demonstrate the intimate involvement TSPO has in the regulation of mitochondrial homeostasis and muster attention towards this molecule, which is equally central for both cellular and mitochondrial biology.

PK11195 inhibits mitophagy targeting the F1Fo-ATPsynthase in Bcl-2 knock-down cells.

The pharmacological agent 1-(2-Chlorophenyl-N-methylpropyl)-3-isoquinolinecarboxamide (PK11195) is the prototypical ligand of the 18-kDa Translocator Protein (TSPO) but at µM concentrations deactivates the oncoprotein Bcl-2 increasing the efficiency of chemotherapeutic agents and promoting the Ca2+-dependent macro-autophagy (or autophagy). In this paper, we report that PK11195, in HeLa cells, modifies the mitochondria-targeted type of autophagy--hereafter referred to as mitophagy--and the associated resizing of the mitochondrial network but does so exclusively in absence of the oncoprotein Bcl-2 (Bcl-2 Kd cells). This is consequence of a "side" targeting of the mitochondrial F1Fo-ATPsynthase enzyme, since identical outcome is mimicked by the antibiotic Oligomycin, of which PK11195 matches the effect on: i) mitochondrial membrane potential (ΔΨm), ii) ATP homeostasis and iii) Reactive Oxygen Species (ROS) generation. Taken together, these data highlight a novel TSPO-independent biological effect for PK11195 and provide evidences for a hitherto uncovered Bcl-2-dependent role of the F1Fo-ATPsynthase in mitochondrial quality control.

Albumin uptake in OK cells exposed to rotenone: a model for studying the effects of mitochondrial dysfunction on endocytosis in the proximal tubule?

BACKGROUND: The renal proximal tubule (PT) is clinically vulnerable to mitochondrial dysfunction; sub-lethal injury can lead to the Fanconi syndrome, with elevated urinary excretion of low-molecular-weight proteins. As the mechanism that couples mitochondrial dysfunction to impaired PT low-molecular weight protein uptake is unknown, we investigated the effect of respiratory chain (RC) inhibitors on endocytosis of FITC-albumin in PT-derived OK cells. METHODS: Uptake of FITC-albumin was quantified using confocal microscopy. Cytosolic ATP levels were measured in real time using both luciferin/luciferase assays and measurements of free [Mg(2+)]. Reactive oxygen species production was measured using mitosox. RESULTS: RC blockade produced only a small decrease in cytosolic ATP levels and had minimal effect on FITC-albumin uptake. Inhibition of glycolysis caused a much bigger decrease in both cytosolic ATP levels and FITC-albumin endocytosis. Rotenone led to higher rates of reactive oxygen species production than other RC inhibitors. Rotenone also caused widespread structural damage on electron microscopy, which was mimicked by colchicine and prevented by taxol; consistent with inhibition of microtubule polymerisation as the underlying mechanism. CONCLUSIONS: Endocytosis of FITC-albumin is ATP-dependent in OK cells, but the cells are very glycolytic and therefore represent a poor metabolic model of the PT. Rotenone has toxic extra-mitochondrial structural effects.

Role of band 3 in regulating metabolic flux of red blood cells.

Deoxygenation elevates glycolytic flux and lowers pentose phosphate pathway (PPP) activity in mammalian erythrocytes. The membrane anion transport protein (band 3 or AE1) is thought to facilitate this process by binding glycolytic enzymes (GEs) and inhibiting their activity in an oxygen-dependent manner. However, this regulatory mechanism has not been demonstrated under physiological conditions. In this study, we introduce a (1)H-(13)C NMR technique for measuring metabolic fluxes in intact cells. The role of band 3 in mediating the oxygenated/deoxygenated metabolic transition was examined by treating cells with pervanadate, a reagent that prevents the GE-band 3 complex from forming. We report that pervanadate suppresses oxygen-dependent changes in glycolytic and PPP fluxes. Moreover, these metabolic alterations were not attributable to modulation of bisphosphoglycerate mutase, direct inhibition of GEs by pervanadate, or oxidation, which are the major side effects of pervanadate treatment. These data provide direct evidence supporting the role of band 3 in mediating oxygen-regulated metabolic transitions.