Ischemia-reperfusion model in rat spinal cord: cell viability and apoptosis signaling study

This work aimed at determining the ideal ischemia time in an in vitro ischemia-reperfusion model of spinal cord injury. Rat spinal cord slices were prepared and then exposed or not to oxygen deprivation and low glucose (ODLG) for 30, 45, 60, 75 and 90 minutes. Cell viability was assessed by triphenyltetrazolium (TTC), lactate dehydrogenase (LDH) release, and fluorochrome dyes specific for cell dead (ethidium homodimer) using the apotome system. Glutamate release was enzymatically measured by a fluorescent method. Gene expression of apoptotic factors was assessed by real time RT-PCR. Whereas spinal cord slices exposed to ODLG exhibited mild increase in fluorescence for 30 minutes after the insult, the 45, 60, 75 and 90 minutes caused a 2-fold increase. ODLG exposure for 45, 60, 75 or 90 minutes, glutamate and LDH release were significantly elevated. nNOS mRNA expression was overexpressed for 45 minutes and moderately increased for 60 minutes in ODLG groups. Bax/bcl-xl ratio, caspase 9 and caspase 3 mRNA expressions were significantly increased for 45 minutes of ODLG, but not for 30, 60, 75 and 90 minutes. Results showed that cell viability reduction in the spinal cord was dependent on ischemic time, resulting in glutamate and LDH release. ODLG for 45 minutes was adequate for gene expression evaluation of proteins and proteases involved in apoptosis pathways.     

Characterization of a Novel Putative Xer-Dependent Integrative Mobile Element Carrying the blaNMC-A Carbapenemase Gene,Inserted into the Chromosome of Members of the Enterobacter cloacae Complex

An Enterobacter ludwigii strain was isolated during routine screening of a Japanese patient for carriage of carbapenem-resistant Enterobacteriaceae. PCR analysis revealed the bla_NMC-A carbapenemase gene. Whole-genome sequencing revealed that bla_NMC-A was inserted in the chromosome and associated with a novel 29.1-kb putative Xer-dependent integrative mobile element, named EludIMEX-1. Bioinformatic analysis identified similar elements in the genomes of an Enterobacter asburiae strain and of other Enterobacter cloacae complex strains, confirming the mobile nature of this element.     

Antileishmanial Activity of Disulfiram and Thiuram Disulfide Analogs in an Ex Vivo Model System Is Selectively Enhanced by the Addition of Divalent Metal Ions

Current treatments for cutaneous and visceral leishmaniasis are toxic, expensive, difficult to administer, and limited in efficacy and availability. Disulfiram has primarily been used to treat alcoholism. More recently, it has shown some efficacy as therapy against protozoan pathogens and certain cancers, suggesting a wide range of biological activities. We used an ex vivo system to screen several thiuram disulfide compounds for antileishmanial activity. We found five compounds (compound identifier [CID] 7188, 5455, 95876, 12892, and 3117 [disulfiram]) with anti-Leishmania activity at nanomolar concentrations. We further evaluated these compounds with the addition of divalent metal salts based on studies that indicated these salts could potentiate the action of disulfiram. In addition, clinical studies suggested that zinc has some efficacy in treating cutaneous leishmaniasis. Several divalent metal salts were evaluated at 1 μM, which is lower than the normal levels of copper and zinc in plasma of healthy individuals. The leishmanicidal activity of disulfiram and CID 7188 were enhanced by several divalent metal salts at 1 μM. The in vitro therapeutic index (IVTI) of disulfiram and CID 7188 increased 12- and 2.3-fold, respectively, against L. major when combined with ZnCl₂. The combination of disulfiram with ZnSO₄ resulted in a 1.8-fold increase in IVTI against L. donovani. This novel combination of thiuram disulfides and divalent metal ions salts could have application as topical and/or oral therapies for treatment of cutaneous and visceral leishmaniasis.     

Centronuclear myopathies: genotype–phenotype correlation and frequency of defined genetic forms in an Italian cohort

Centronuclear myopathies (CNMs) are a group of clinically and genetically heterogeneous muscle disorders. To date, mutation in 7 different genes has been reported to cause CNMs but 30 % of cases still remain genetically undefined. Genetic investigations are often expensive and time consuming. Clinical and morphological clues are needed to facilitate genetic tests and to choose the best approach for genetic screening. We aimed to describe genotype–phenotype correlation in an Italian cohort of patients affected by CNMs, to define the relative frequencies of its defined genetic forms and to draw a diagnostic algorithm to address genetic investigations. We recruited patients with CNMs from all the Italian tertiary neuromuscular centers following clinical and histological criteria. All selected patients were screened for the four ‘canonical’ genes related to CNMs: MTM1, DNM2, RYR1 and BIN1. Pathogenetic mutations were found in 38 of the 54 screened patients (70 %), mostly in patients with congenital onset (25 of 30 patients, 83 %): 15 in MTM1, 6 in DNM2, 3 in RYR1 and one in TTN. Among the 13 patients with a childhood–adolescence onset, mutations were found in 6 patients (46 %), all in DNM2. In the group of the 11 patients with adult onset, mutations were identified in 7 patients (63 %), again in DNM2, confirming that variants in this gene are relatively more common in late-onset phenotypes. The present study provides the relative molecular frequency of centronuclear myopathy and of its genetically defined forms in Italy and also proposes a diagnostic algorithm to be used in clinical practice to address genetic investigations.     

Magnetic Resonance Fiber Tracking in a Neonate with Hemimegalencephaly

A magnetic resonance diffusion fiber tracking study in neonate diagnosed with left hemisphere hemimegalencephaly is presented. Despite diffuse morphologic deformities identified in conventional imaging, all major pathways were identifiable bilaterally with minor aberrations in vicinity of morphologic lesions.     

Protective variant for hippocampal atrophy identified by whole exome sequencing

We used whole‐exome sequencing to identify variants other than APOE associated with the rate of hippocampal atrophy in amnestic mild cognitive impairment. An in‐silico predicted missense variant in REST (rs3796529) was found exclusively in subjects with slow hippocampal volume loss and validated using unbiased whole‐brain analysis and meta‐analysis across 5 independent cohorts. REST is a master regulator of neurogenesis and neuronal differentiation that has not been previously implicated in Alzheimer's disease. These findings nominate REST and its functional pathways as protective and illustrate the potential of combining next‐generation sequencing with neuroimaging to discover novel disease mechanisms and potential therapeutic targets. Ann Neurol 2015;77:547–552     

The cerebral correlates of subliminal emotions: an electroencephalographic study with emotional hybrid faces

In a high‐resolution electroencephalographic study, participants evaluated the friendliness level of upright and inverted ‘hybrid faces’, i.e. facial photos containing a subliminal emotional core in the low spatial frequencies (< 6 cycles/image), superimposed on a neutral expression in the rest of the spatial frequencies. Upright happy and angry faces were judged as more friendly or less friendly than neutral faces, respectively. We observed the time course of cerebral correlates of these stimuli with event‐related potentials (ERPs), confirming that hybrid faces elicited the posterior emotion‐related and face‐related components (P1, N170 and P2), previously shown to be engaged by non‐subliminal emotional stimuli. In addition, these components were stronger in the right hemisphere and were both enhanced and delayed by face inversion. A frontal positivity (210–300 ms) was stronger for emotional than for neutral faces, and for upright than for inverted faces. Hence, hybrid faces represent an original approach in the study of subliminal emotions, which appears promising for investigating their electrophysiological correlates.     

Transcranial direct current stimulation of the posterior parietal cortex modulates arithmetic learning

The successful acquisition of arithmetic skills is an essential step in the development of mathematical competencies and has been associated with neural activity in the left posterior parietal cortex (PPC). It is unclear, however, whether this brain region plays a causal role in arithmetic skill acquisition and whether arithmetic learning can be modulated by means of non‐invasive brain stimulation of this key region. In the present study we addressed these questions by applying transcranial direct current stimulation (tDCS) over the left PPC during a short‐term training that simulates the typical path of arithmetic skill acquisition (specifically the transition from effortful procedural to memory‐based problem‐solving strategies). Sixty participants received either anodal, cathodal or sham tDCS while practising complex multiplication and subtraction problems. The stability of the stimulation‐induced learning effects was assessed in a follow‐up test 24 h after the training. Learning progress was modulated by tDCS. Cathodal tDCS (compared with sham) decreased learning rates during training and resulted in poorer performance which lasted over 24 h after stimulation. Anodal tDCS showed an operation‐specific improvement for subtraction learning. Our findings extend previous studies by demonstrating that the left PPC is causally involved in arithmetic learning (and not only in arithmetic performance) and that even a short‐term tDCS application can modulate the success of arithmetic knowledge acquisition. Moreover, our finding of operation‐specific anodal stimulation effects suggests that the enhancing effects of tDCS on learning can selectively affect just one of several cognitive processes mediated by the stimulated area.