Differential contribution of the guanylyl cyclase-cyclic GMP-protein kinase G pathway to the proliferation of neural stem cells stimulated by nitric oxide.

Nitric oxide (NO) is an important inflammatory mediator involved in the initial boost in the proliferation of neural stem cells following brain injury. However, the mechanisms underlying the proliferative effect of NO are still unclear. The aim of this work was to investigate whether cyclic GMP (cGMP) and the cGMP-dependent kinase (PKG) are involved in the proliferative effect triggered by NO in neural stem cells. For this purpose, cultures of neural stem cells isolated from the mouse subventricular zone (SVZ) were used. We observed that long-term exposure to the NO donor (24 h), NOC-18, increased the proliferation of SVZ cells in a cGMP-dependent manner, since the guanylate cyclase inhibitor, ODQ, prevented cell proliferation. Similarly to NOC-18, the cGMP analogue, 8-Br-cGMP, also increased cell proliferation. Interestingly, shorter exposures to NO (6 h) increased cell proliferation in a cGMP-independent manner via the ERK/MAP kinase pathway. The selective inhibitor of PKG, KT5823, prevented the proliferative effect induced by NO at 24 h but not at 6 h. In conclusion, the proliferative effect of NO is initially mediated by the ERK/MAPK pathway, and at later stages by the GC/cGMP/PKG pathway. Thus, our work shows that NO induces neural stem cell proliferation by targeting these two pathways in a biphasic manner.

How can we modulate aging through nutrition and physical exercise? An epigenetic approach.

The World Health Organization predicts that by 2050, 2.1 billion people worldwide will be over 60 years old, a drastic increase from only 1 billion in 2019. Considering these numbers, strategies to ensure an extended "healthspan" or healthy longevity are urgently needed. The present study approaches the promotion of healthspan from an epigenetic perspective. Epigenetic phenomena are modifiable in response to an individual's environmental exposures, and therefore link an individual's environment to their gene expression pattern. Epigenetic studies demonstrate that aging is associated with decondensation of the chromatin, leading to an altered heterochromatin structure, which promotes the accumulation of errors. In this review, we describe how aging impacts epigenetics and how nutrition and physical exercise can positively impact the aging process, from an epigenetic point of view. Canonical histones are replaced by histone variants, concomitant with an increase in histone post-translational modifications. A slight increase in DNA methylation at promoters has been observed, which represses transcription of previously active genes, in parallel with global genome hypomethylation. Aging is also associated with deregulation of gene expression - usually provided by non-coding RNAs - leading to both the repression of previously transcribed genes and to the transcription of previously repressed genes. Age-associated epigenetic events are less common in individuals with a healthy lifestyle, including balanced nutrition, caloric restriction and physical exercise. Healthy aging is associated with more tightly condensed chromatin, fewer PTMs and greater regulation by ncRNAs.

Leprosy in a Patient With Lymphoma: A Challenge in the Twenty-First Century.

Leprosy, or Hansen's disease, mistakenly considered a disease from the past by some, is still common nowadays, especially in tropical and subtropical regions. In the absence of appropriate medical treatment, it may progress and cause permanent damage to multiple organs. This case report illustrates the diagnostic challenge of a south-american adult man who had been living in Europe for over 14 years. He was referred to the Hematology department due to persistent lymphocytosis and a CD5+ B-cell lymphoproliferative disorder was identified. During clinical surveillance, the patient developed skin lesions in his limbs with associated hypoesthesia. A histological diagnosis of lepromatous leprosy was made, and he underwent a long-term three-drug therapeutic regimen (dapsone, rifampicin, and clofazimine). Adding to the complexity of the case, the patient progressed with splenomegaly and constitutional symptoms, more than 7 years after development of lymphocytosis. Through a comprehensive evaluation, a definitive diagnosis of mantle cell lymphoma was established and received 6-cycle R-CHOP induction, followed by maintenance rituximab. Importantly, prophylaxis for leprosy reactivation was not administered as there were no recommendations in available guidelines. Eventually, the patient experienced a leprosy relapse while on maintenance therapy, 58 months after completing the initial anti-leprous treatment. Clinical response was attained with a new treatment regimen consisting of rifampicin, clofazimine, and minocycline.  Although leprosy is primarily observed in tropical and subtropical regions, the long incubation period of this disease combined with the global flow of migrants, made us consider it. Despite being rare, leprosy relapses can occur even after a few decades. The contribution of rituximab or previously administered chemotherapeutic agents is still unknown. The question remains whether antibiotic prophylaxis should be performed in patients undergoing immunochemotherapy for malignant diseases.

Heart Failure: From Typical Clinical Manifestations to the Surprising Final Diagnosis.

The authors report a case of an 80-year-old woman with multiple cardiovascular risk factors, with exuberant acute congestive heart failure at admission. Fever, anemia, and an increase in inflammatory parameters were present, with imaging suggesting a respiratory infection as the main reason for decompensation. Empirical antibiotic therapy was instituted, with no clinical improvement even after escalation to broad-spectrum antibiotics and non-invasive ventilation with high support pressures, with no possibility of weaning. Due to maintenance of symptoms, a transthoracic echocardiogram was performed, revealing a large left atrial myxoma, obstructing the mitral valve in diastole. This case illustrates the potential severity of these benign tumors and their ability to mimic symptoms that are often evaluated in the daily life of an internist. The high clinical suspicion led to a diagnosis that was surprising due to its rarity and severity, with the patient being urgently referred for cardiac surgery.

Seronegative Celiac Disease - A Challenging Case.

Celiac disease is an inflammatory disorder of the small intestine caused by sensitivity to gluten. This enteropathy results from the interaction between genetics, autoimmunity, and an environmental trigger (gluten). It can manifest at all ages. We present a case of a 76-year-old woman with nausea and vomiting for six months. She reported asthenia, weight loss, and a brief period of diarrhea without blood or mucus. The search for evidence of infection, tumours, and endocrinopathies was negative, as well as the immunological study, including antibodies for celiac disease. Upper endoscopy with biopsies revealed villous atrophy. Capsule endoscopy showed macroscopic features suggestive of celiac/Whipple's disease. Duodenal biopsies were reviewed, and Whipple's disease was considered unlikely. The genetic analysis was positive for HLA DR7-DQ2. After one year on a gluten-free diet, there was a clinical and histological improvement. The diagnosis of seronegative celiac disease is complex and requires the exclusion of other causes of villous atrophy, as well as a histological improvement after one year of treatment. The genetic test has a high negative predictive value.

Coronal brain atlas in stereotaxic coordinates of the African spiny mouse, Acomys cahirinus.

The African spiny mouse (Acomys cahirinus) is an emerging model of mammalian epimorphic regeneration that has aroused the interest of the scientific community in the last decade. To date, studies on brain repair have been hindered by the lack of knowledge on the neuroanatomy of this species. Here, we present a coronal brain atlas in stereotaxic coordinates, which allows for three-dimensional identification and localization of the brain structures of this species. The brain of 12-week-old spiny mice was mapped in stereotaxic coordinates using cresyl violet-stained brain sections obtained from coronal cryosectioning of the brain after transcardial perfusion with fixative. The atlas is presented in 42 plates representing sections spaced 240 µm apart. Stereotaxic coordinates were validated using both a model of Parkinsonian lesion of the striatum with 6-hydroxydopamine and labeling of the corticospinal tract in the spiny mouse spinal cord using AAV1/2-GFP intracortical injections. This work presents a new tool in A. cahirinus neurobiology and opens new avenues of research for the investigation of the regenerative ability of A. cahirinus in models of brain disorders.

Rewired glycosylation activity promotes scarless regeneration and functional recovery in spiny mice after complete spinal cord transection.

Regeneration of adult mammalian central nervous system (CNS) axons is abortive, resulting in inability to recover function after CNS lesion, including spinal cord injury (SCI). Here, we show that the spiny mouse (Acomys) is an exception to other mammals, being capable of spontaneous and fast restoration of function after severe SCI, re-establishing hind limb coordination. Remarkably, Acomys assembles a scarless pro-regenerative tissue at the injury site, providing a unique structural continuity of the initial spinal cord geometry. The Acomys SCI site shows robust axon regeneration of multiple tracts, synapse formation, and electrophysiological signal propagation. Transcriptomic analysis of the spinal cord following transcriptome reconstruction revealed that Acomys rewires glycosylation biosynthetic pathways, culminating in a specific pro-regenerative proteoglycan signature at SCI site. Our work uncovers that a glycosylation switch is critical for axon regeneration after SCI and identifies ß3gnt7, a crucial enzyme of keratan sulfate biosynthesis, as an enhancer of axon growth.

Calpains and delayed calcium deregulation in excitotoxicity.

Overactivation of glutamate receptors results in neurodegeneration in a variety of brain pathologies, including ischemia, epilepsy, traumatic brain injury and slow-progressing neurodegenerative disorders. In all these pathologies, it is well accepted that the calcium-dependent cysteine proteases calpains are key players in the mechanisms of neuronal cell death. Many research groups have been actively pursuing to establish a link between the deregulation of intracellular Ca(2+) homeostasis associated with excitotoxicity and calpain activity. It is well established that these two events are connected and interact synergistically to promote neurodegeneration, but whether calpain activity depends on or contributes to Ca(2+) deregulation is still under debate.

Regulation of catecholamine release and tyrosine hydroxylase in human adrenal chromaffin cells by interleukin-1beta: role of neuropeptide Y and nitric oxide.

Adrenal chromaffin cells synthesize and secrete catecholamines and neuropeptides that may regulate hormonal and paracrine signaling in stress and also during inflammation. The aim of our work was to study the role of the cytokine interleukin-1beta (IL-1beta) on catecholamine release and synthesis from primary cell cultures of human adrenal chromaffin cells. The effect of IL-1beta on neuropeptide Y (NPY) release and the intracellular pathways involved in catecholamine release evoked by IL-1beta and NPY were also investigated. We observed that IL-1beta increases the release of NPY, norepinephrine (NE), and epinephrine (EP) from human chromaffin cells. Moreover, the immunoneutralization of released NPY inhibits catecholamine release evoked by IL-1beta. Moreover, IL-1beta regulates catecholamine synthesis as the inhibition of tyrosine hydroxylase decreases IL-1beta-evoked catecholamine release and the cytokine induces tyrosine hydroxylase Ser40 phosphorylation. Moreover, IL-1beta induces catecholamine release by a mitogen-activated protein kinase (MAPK)-dependent mechanism, and by nitric oxide synthase activation. Furthermore, MAPK, protein kinase C (PKC), protein kinase A (PKA), and nitric oxide (NO) production are involved in catecholamine release evoked by NPY. Using human chromaffin cells, our data suggest that IL-1beta, NPY, and nitric oxide (NO) may contribute to a regulatory loop between the immune and the adrenal systems, and this is relevant in pathological conditions such as infection, trauma, stress, or in hypertension.

Adenosine A2A receptor antagonists exert motor and neuroprotective effects by distinct cellular mechanisms.

OBJECTIVE: To investigate whether the motor and neuroprotective effects of adenosine A(2A) receptor (A(2A)R) antagonists are mediated by distinct cell types in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of Parkinson's disease. METHODS: We used the forebrain A(2A)R knock-out mice coupled with flow cytometric analyses and intracerebroventricular injection to determine the contribution of A(2A)Rs in forebrain neurons and glial cells to A(2A)R antagonist-mediated motor and neuroprotective effects. RESULTS: The selective deletion of A(2A)Rs in forebrain neurons abolished the motor stimulant effects of the A(2A)R antagonist KW-6002 but did not affect acute MPTP neurotoxicity. Intracerebroventricular administration of KW-6002 into forebrain A(2A)R knock-out mice reinstated protection against acute MPTP-induced dopaminergic neurotoxicity and attenuated MPTP-induced striatal microglial and astroglial activation. INTERPRETATION: A(2A)R activity in forebrain neurons is critical to the control of motor activity, whereas brain cells other than forebrain neurons (likely glial cells) are important components for protection against acute MPTP toxicity.

Leishmanicidal activity and immobilization of dermaseptin 01 antimicrobial peptides in ultrathin films for nanomedicine applications.

Antimicrobial peptides (AMPs) are essential for the innate immune system of eukaryotes, imparting protection against pathogens and their proliferation in host organisms. The recent interest in AMPs as active materials in bionanostructures is due to the properties shown by these biological molecules, such as the presence of an alpha-helix structure and distribution of positive charges along the chain. In this study the antimicrobial peptide dermaseptin 01 (DS 01), from the skin secretion of Phyllomedusa hypochondrialis frogs was immobilized in nanostructured layered films in conjunction with nickel tetrasulfonated phthalocyanines. The leishmanicidal activity of DS 01 was confirmed using kinetic essays, in which DS 01 promoted death of all metacyclic promastigote cells in 45 minutes. Surprisingly, the immobilized DS 01 molecules displayed electroactivity, as revealed by electrochemical experiments, in which an oxidation peak at about 0.61 V was observed for a DS 01 monolayer deposited on top of a conductive electrode. Such electroactivity was used to investigate the sensing abilities of the nanostructured films toward Leishmania. We observed an increase in the oxidation current as a function of number of Leishmania cells in the electrolytic solution at concentrations down to 10(3) cells/mL. The latter is indicative that the use of AMPs immobilized in electroactive nanostructured films may be of interest for applications in the pharmaceutical industry and diagnosis. FROM THE CLINICAL EDITOR: The recent interest in Antimicrobial peptides (AMPs) as active materials in bionanostructures is due to the properties shown by these biological molecules. Leishmanicidal activity of a particular AMP is demonstrated in this paper.

Extracellular electrophysiological based sensor to monitor cancer cells cooperative migration and cell-cell connections.

Herein, we describe an electrophysiological based sensor that reproducibly monitors and quantifies in real-time collective migration and the formation of cell-cell junctions by C6 glioma cells seeded on top of electrodes. The signal amplitude and frequency generated by the migrating cells changed over time and these parameters were used to accurately calculate the migration speed. Electrophysiological measurements could also distinguish individual from collective cell migration. The migration of densely packed cells generated strong signals, while dispersed cells showed weak bioelectrical activity. We propose this electrophysiological technique as a cell-based biosensor to gain insight into the mechanisms of cooperative migration of cancer cells. Possible applications include screening for anti-migratory compounds, which may lead to the development of novel strategies for antineoplastic chemotherapy.

Neuropeptide Y stimulates retinal neural cell proliferation--involvement of nitric oxide.

Neuropeptide Y (NPY) is a 36 amino acid peptide widely present in the CNS, including the retina. Previous studies have demonstrated that NPY promotes cell proliferation of rat post-natal hippocampal and olfactory epithelium precursor cells. The aim of this work was to investigate the role of NPY on cell proliferation of rat retinal neural cells. For this purpose, primary retinal cell cultures expressing NPY, and NPY Y(1), Y(2), Y(4) and Y(5) receptors [Alvaro et al., (2007) Neurochem. Int., 50, 757] were used. NPY (10-1000 nM) stimulated cell proliferation through the activation of NPY Y(1), Y(2) and Y(5) receptors. NPY also increased the number of proliferating neuronal progenitor cells (BrdU(+)/nestin(+) cells). The intracellular mechanisms coupled to NPY receptors activation that mediate the increase in cell proliferation were also investigated. The stimulatory effect of NPY on cell proliferation was reduced by L-nitroarginine-methyl-esther (L-NAME; 500 microM), a nitric oxide synthase inhibitor, 1H-[1,2,4]oxadiazolo-[4, 3-a]quinoxalin-1-one (ODQ; 20 microM), a soluble guanylyl cyclase inhibitor or U0126 (1 microM), an inhibitor of the extracellular signal-regulated kinase 1/2 (ERK 1/2). In conclusion, NPY stimulates retinal neural cell proliferation, and this effect is mediated through nitric oxide-cyclic GMP and ERK 1/2 pathways.

Calpain activation is involved in early caspase-independent neurodegeneration in the hippocampus following status epilepticus.

Evidence for increased calpain activity has been described in the hippocampus of rodent models of temporal lobe epilepsy. However, it is not known whether calpains are involved in the cell death that accompanies seizures. In this work, we characterized calpain activation by examining the proteolysis of calpain substrates and in parallel we followed cell death in the hippocampus of epileptic rats. Male Wistar rats were injected with kainic acid (10 mg/kg) intraperitoneally and killed 24 h later, after development of grade 5 seizures. We observed a strong Fluoro-Jade labeling in the CA1 and CA3 areas of the hippocampus in the rats that received kainic acid, when compared with saline-treated rats. Immunohistochemistry and western blot analysis for the calpain-derived breakdown products of spectrin showed evidence of increased calpain activity in the same regions of the hippocampus where cell death is observed. No evidence was found for caspase activation, in the same conditions. Treatment with the calpain inhibitor MDL 28170 significantly prevented the neurodegeneration observed in CA1. Taken together, our data suggest that early calpain activation, but not caspase activation, is involved in neurotoxicity in the hippocampus after status epilepticus.

Hydrothermal synthesis of bacterial cellulose-copper oxide nanocomposites and evaluation of their antimicrobial activity.

In this work, for the first time bacterial cellulose (BC) hydrogel membranes were used for the fabrication of antimicrobial cellulosic nanocomposites by hydrothermal deposition of Cu derivative nanoparticles (i.e.Cu(0) and CuxOy species). BC-Cu nanocomposites were characterized by FTIR, SEM, AFM, XRD and TGA, to study the effect of hydrothermal processing time on the final physicochemical properties of final products. XRD result show that depending on heating time (3-48h), different CuxOy phases were achieved. SEM and AFM analyses unveil the presence of the Cu(0) and copper CuxOy nanoparticles over BC fibrils while the surface of 3D network became more compact and smother for longer heating times. Furthermore, the increase of heating time placed deleterious effect on the structure of BC network leading to decrease of BC crystallinity as well as of the on-set degradation temperature. Notwithstanding, BC-Cu nanocomposites showed excellent antimicrobial activity against E. coli, S. aureus and Salmonella bacteria suggesting potential applications as bactericidal films.

Neuronal Adenosine A2A Receptors Are Critical Mediators of Neurodegeneration Triggered by Convulsions.

Neurodegeneration is a process transversal to neuropsychiatric diseases and the understanding of its mechanisms should allow devising strategies to prevent this irreversible step in brain diseases. Neurodegeneration caused by seizures is a critical step in the aggravation of temporal lobe epilepsy, but its mechanisms remain undetermined. Convulsions trigger an elevation of extracellular adenosine and upregulate adenosine A2A receptors (A2AR), which have been associated with the control of neurodegenerative diseases. Using the rat and mouse kainate model of temporal lobe epilepsy, we now tested whether A2AR control convulsions-induced hippocampal neurodegeneration. The pharmacological or genetic blockade of A2AR did not affect kainate-induced convulsions but dampened the subsequent neurotoxicity. This neurotoxicity began with a rapid A2AR upregulation within glutamatergic synapses (within 2 h), through local translation of synaptic A2AR mRNA. This bolstered A2AR-mediated facilitation of glutamate release and of long-term potentiation (LTP) in CA1 synapses (4 h), triggered a subsequent synaptotoxicity, heralded by decreased synaptic plasticity and loss of synaptic markers coupled to calpain activation (12 h), that predated overt neuronal loss (24 h). All modifications were prevented by the deletion of A2AR selectively in forebrain neurons. This shows that synaptic A2AR critically control synaptic excitotoxicity, which underlies the development of convulsions-induced neurodegeneration.

Calpastatin Overexpression Preserves Cognitive Function Following Seizures, While Maintaining Post-Injury Neurogenesis.

In the adult mammalian brain, new neurons continue to be produced throughout life in two main regions in the brain, the subgranular zone (SGZ) in the hippocampus and the subventricular zone in the walls of the lateral ventricles. Neural stem cells (NSCs) proliferate in these niches, and migrate as neuroblasts, to further differentiate in locations where new neurons are needed, either in normal or pathological conditions. However, the endogenous attempt of brain repair is not very efficient. Calpains are proteases known to be involved in neuronal damage and in cell proliferation, migration and differentiation of several cell types, though their effects on neurogenesis are not well known. Previous work by our group has shown that the absence of calpastatin (CAST), the endogenous inhibitor of calpains, impairs early stages of neurogenesis. Since the hippocampus is highly associated with learning and memory, we aimed to evaluate whether calpain inhibition would help improve cognitive recovery after lesion and efficiency of post-injury neurogenesis in this region. For that purpose, we used the kainic acid (KA) model of seizure-induced hippocampal lesion and mice overexpressing CAST. Selected cognitive tests were performed on the 3rd and 8th week after KA-induced lesion, and cell proliferation, migration and differentiation in the dentate gyrus (DG) of the hippocampus of adult mice were analyzed using specific markers. Cognitive recovery was evaluated by testing the animals for recognition, spatial and associative learning and memory. Cognitive function was preserved by CAST overexpression following seizures, while modulation of post-injury neurogenesis was similar to wild type (WT) mice. Calpain inhibition could still be potentially able to prevent the impairment in the formation of new neurons, given that the levels of calpain activity could be reduced under a certain threshold and other harmful effects from the pathological environment could also be controlled.

Ultrasensitive gold micro-structured electrodes enabling the detection of extra-cellular long-lasting potentials in astrocytes populations.

Ultra-sensitive electrodes for extracellular recordings were fabricated and electrically characterized. A signal detection limit defined by a noise level of 0.3-0.4 µV for a bandwidth of 12.5 Hz was achieved. To obtain this high sensitivity, large area (4 mm2) electrodes were used. The electrode surface is also micro-structured with an array of gold mushroom-like shapes to further enhance the active area. In comparison with a flat gold surface, the micro-structured surface increases the capacitance of the electrode/electrolyte interface by 54%. The electrode low impedance and low noise enable the detection of weak and low frequency quasi-periodic signals produced by astrocytes populations that thus far had remained inaccessible using conventional extracellular electrodes. Signals with 5 µV in amplitude and lasting for 5-10 s were measured, with a peak-to-peak signal-to-noise ratio of 16. The electrodes and the methodology developed here can be used as an ultrasensitive electrophysiological tool to reveal the synchronization dynamics of ultra-slow ionic signalling between non-electrogenic cells.

Extracellular Electrophysiological Measurements of Cooperative Signals in Astrocytes Populations.

Astrocytes are neuroglial cells that exhibit functional electrical properties sensitive to neuronal activity and capable of modulating neurotransmission. Thus, electrophysiological recordings of astroglial activity are very attractive to study the dynamics of glial signaling. This contribution reports on the use of ultra-sensitive planar electrodes combined with low noise and low frequency amplifiers that enable the detection of extracellular signals produced by primary cultures of astrocytes isolated from mouse cerebral cortex. Recorded activity is characterized by spontaneous bursts comprised of discrete signals with pronounced changes on the signal rate and amplitude. Weak and sporadic signals become synchronized and evolve with time to higher amplitude signals with a quasi-periodic behavior, revealing a cooperative signaling process. The methodology presented herewith enables the study of ionic fluctuations of population of cells, complementing the single cells observation by calcium imaging as well as by patch-clamp techniques.

Caffeine Reverts Memory But Not Mood Impairment in a Depression-Prone Mouse Strain with Up-Regulated Adenosine A2A Receptor in Hippocampal Glutamate Synapses.

Caffeine prophylactically prevents mood and memory impairments through adenosine A2A receptor (A2AR) antagonism. A2AR antagonists also therapeutically revert mood and memory impairments, but it is not known if caffeine is also therapeutically or only prophylactically effective. Since depression is accompanied by mood and memory alterations, we now explored if chronic (4 weeks) caffeine consumption (0.3 g/L) reverts mood and memory impairment in helpless mice (HM, 12 weeks old), a bred-based model of depression. HM displayed higher immobility in the tail suspension and forced swimming tests, greater anxiety in the elevated plus maze, and poorer memory performance (modified Y-maze and object recognition). HM also had reduced density of synaptic (synaptophysin, SNAP-25), namely, glutamatergic (vGluT1; -22 ± 7 %) and GABAergic (vGAT; -23 ± 8 %) markers in the hippocampus. HM displayed higher A2AR density (72 ± 6 %) in hippocampal synapses, an enhanced facilitation of hippocampal glutamate release by the A2AR agonist, CGS21680 (30 nM), and a larger LTP amplitude (54 ± 8 % vs. 21 ± 5 % in controls) that was restored to control levels (30 ± 10 %) by the A2AR antagonist, SCH58261 (50 nM). Notably, caffeine intake reverted memory deficits and reverted the loss of hippocampal synaptic markers but did not affect helpless or anxiety behavior. These results reinforce the validity of HM as an animal model of depression by showing that they also display reference memory deficits. Furthermore, caffeine intake selectively reverted memory but not mood deficits displayed by HM, which are associated with an increased density and functional impact of hippocampal A2AR controlling synaptic glutamatergic function.