Endothelin-1 Induces Degeneration of Cultured Motor Neurons Through a Mechanism Mediated by Nitric Oxide and PI3K/Akt Pathway.

Endothelin-1 (ET-1) is a vasoactive peptide produced by activated astrocytes and microglia and is implicated in initiating and sustaining reactive gliosis in neurodegenerative diseases. We have previously suggested that ET-1 can play a role in the pathophysiology of amyotrophic lateral sclerosis (ALS). Indeed, we reported that this peptide is abundantly expressed in reactive astrocytes in the spinal cord of SOD1-G93A mice and ALS patients and exerts a toxic effect on motor neurons (MNs) in an in vitro model of mixed spinal cord cultures enriched with reactive astrocytes. Here, we explored the possible mechanisms underlying the toxic effect of ET-1 on cultured MNs. We show that ET-1 toxicity is not directly caused by oxidative stress or activation of cyclooxygenase-2 but requires the synthesis of nitric oxide and is mediated by a reduced activation of the phosphoinositide 3-kinase pathway. Furthermore, we observed that ET-1 is also toxic for microglia, although its effect on MNs is independent of the presence of this type of glial cells. Our study confirms that ET-1 may contribute to MN death and corroborates the view that the modulation of ET-1 signaling might be a therapeutic strategy to slow down MN degeneration in ALS.

Transcriptional landscapes at the intersection of neuronal apoptosis and substance P-induced survival: exploring pathways and drug targets.

A change in the delicate equilibrium between apoptosis and survival regulates the neurons fate during the development of nervous system and its homeostasis in adulthood. Signaling pathways promoting or protecting from apoptosis are activated by multiple signals, including those elicited by neurotrophic factors, and depend upon specific transcriptional programs. To decipher the rescue program induced by substance P (SP) in cerebellar granule neurons, we analyzed their whole-genome expression profiles after induction of apoptosis and treatment with SP. Transcriptional pathways associated with the survival effect of SP included genes encoding for proteins that may act as pharmacological targets. Inhibition of one of these, the Myc pro-oncogene by treatment with 10058-F4, reverted in a dose-dependent manner the rescue effect of SP. In addition to elucidate the transcriptional mechanisms at the intersection of neuronal apoptosis and survival, our systems biology-based perspective paves the way towards an innovative pharmacology based on targets downstream of neurotrophic factor receptors.

Cracking the code of neuronal apoptosis and survival.

Neuronal apoptosis and survival are tightly controlled processes that regulate cell fate during the development of the central nervous system and its homeostasis throughout adulthood. A new study in primary cultures of cerebellar granule neurons identified common transcriptional cascades during rescue from apoptosis by insulin-like growth factor-1 (Igf1) and pituitary adenylyl cyclase-activating polypeptide (Pacap), thus suggesting the existence of a high degree of conservation of cell survival pathways.

Note: fast neutron efficiency in CR-39 nuclear track detectors.

CR-39 samples are commonly employed for fast neutron detection in fusion reactors and in inertial confinement fusion experiments. The literature reported efficiencies are strongly depending on experimental conditions and, in some cases, highly dispersed. The present note analyses the dependence of efficiency as a function of various parameters and experimental conditions in both the radiator-assisted and the stand-alone CR-39 configurations. Comparisons of literature experimental data with Monte Carlo calculations and optimized efficiency values are shown and discussed.

The natural history of spinal neurofibromatosis: a critical review of clinical and genetic features.

Spinal neurofibromatosis (SNF) is a related form of neurofibromatosis 1 (NF1), characterized by bilateral neurofibromas (histologically proven) of all spinal roots (and, eventually, of all the major peripheral nerve branches) with or without other manifestations of classical NF1. By rigorous application of these criteria to the 98 SNF cases published, we developed: (i) a cohort of 49 SNF patients (21 males and 28 females; aged 4-74 years]: 9 SNF families (21/49), 1 mixed SNF/NF1 family (1/49) and 27 of 49 sporadic SNF patients (including 5 unpublished patients in this report); and (ii) a group of 49 non-SNF patients including: (a) 32 patients with neurofibromas of multiple but not all spinal roots (MNFSR): 4 mixed SNF/MNFSR families (6/32); (b) 14 patients with NF1 manifestations without spinal neurofibromas, belonging to SNF (8/49) or MNFSR families (6/32); (c) 3 patients with neurofibromas in one spinal root. In addition to reduced incidence of café-au-lait spots (67% in SNF vs 56% in MNFSR), other NF1 manifestations were less frequent in either cohort. Molecular testing showed common NF1 gene abnormalities in both groups. The risk of developing SNF vs NF1 was increased for missense mutations [p = 0.0001; odds ratio (OR) = 6.16; confidence interval (CI) = 3.14-13.11], which were more frequent in SNF vs MNFSR (p = 0.0271).

A very sensitive ion collection device for plasma-laser characterization.

In this paper a very sensitive ion collection device, for diagnostic of laser ablated-target plasma, is described. It allows for reducing down to few microvolts the signal threshold at digital scope input. A standard ion collector is coupled to a transimpedance amplifier, specially designed, which increases data acquisition sensitivity by a gain ≈1100 and does not introduce any significant distortion of input signal. By time integration of current intensity, an amount of charge as small as 2.7 × 10(-2) pC can be detected for photopeak events.

Monoenergetic proton emission from nuclear reaction induced by high intensity laser-generated plasma.

A 10(16) W∕cm(2) Asterix laser pulse intensity, 1315 nm at the fundamental frequency, 300 ps pulse duration, was employed at PALS laboratory of Prague, to irradiate thick and thin primary CD(2) targets placed inside a high vacuum chamber. The laser irradiation produces non-equilibrium plasma with deutons and carbon ions emission with energy of up to about 4 MeV per charge state, as measured by time-of-flight (TOF) techniques by using ion collectors and silicon carbide detectors. Accelerated deutons may induce high D-D cross section for fusion processes generating 3 MeV protons and 2.5 MeV neutrons, as measured by TOF analyses. In order to increase the mono-energetic proton yield, secondary CD(2) targets can be employed to be irradiated by the plasma-accelerated deutons. Experiments demonstrated that high intensity laser pulses can be employed to promote nuclear reactions from which characteristic ion streams may be developed. Results open new scenario for applications of laser-generated plasma to the fields of ion sources and ion accelerators.

Proton emission from a laser ion source.

At intensities of the order of 10(10) W∕cm(2), ns pulsed lasers can be employed to ablate solid bulk targets in order to produce high emission of ions at different charge state and kinetic energy. A special interest is devoted to the production of protons with controllable energy and current from a roto-translating target irradiated in repetition rate at 1-10 Hz by a Nd:Yag pulsed laser beam. Different hydrogenated targets based on polymers and hydrates were irradiated in high vacuum. Special nanostrucutres can be embedded in the polymers in order to modify the laser absorption properties and the amount of protons to be accelerated in the plasma. For example, carbon nanotubes may increase the laser absorption and the hydrogen absorption to generate high proton yields from the plasma. Metallic nanostrucutres may increase the electron density of the plasma and the kinetic energy of the accelerated protons. Ion collectors, ion energy analyzer, and mass spectrometers, used in time-of-flight configuration, were employed to characterize the ion beam properties. A comparison with traditional proton ion source is presented and discussed.

Substance P activates ADAM9 mRNA expression and induces α-secretase-mediated amyloid precursor protein cleavage.

Altered levels of Substance P (SP), a neuropeptide endowed with neuroprotective and anti-apoptotic properties, were found in brain areas and spinal fluid of Alzheimer's disease (AD) patients. One of the hallmarks of AD is the abnormal extracellular deposition of neurotoxic beta amyloid (Aß) peptides, derived from the proteolytic processing of amyloid precursor protein (APP). In the present study, we confirmed, the neurotrophic action of SP in cultured rat cerebellar granule cells (CGCs) and investigated its effects on APP metabolism. Incubation with low (5 mM) potassium induced apoptotic cell death of CGCs and amyloidogenic processing of APP, whereas treatment with SP (200 nM) reverted these effects via NK1 receptors. The non-amyloidogenic effect of SP consisted of reduction of Aß(1-42), increase of sAPPα and enhanced α-secretase activity, without a significant change in steady-state levels of cellular APP. The intracellular mechanisms whereby SP alters APP metabolism were further investigated by measuring mRNA and/or steady-state protein levels of key enzymes involved with α-, ß- and γ-secretase activity. Among them, Adam9, both at the mRNA and protein level, was the only enzyme to be significantly down-regulated following the induction of apoptosis (K5) and up-regulated after SP treatment. In addition to its neuroprotective properties, this study shows that SP is able to stimulate non-amyloidogenic APP processing, thereby reducing the possibility of generation of toxic Aß peptides in brain.

Drug target identification for neuronal apoptosis through a genome scale screening.

During normal nervous system development, physiologically appropriate neuronal apoptosis contributes to a sculpting process that removes approximately one-half of all neurons born during neurogenesis. However, neuronal apoptosis subsequent to this developmental window is physiologically inappropriate for most systems and can contribute to neurodegenerative diseases. Neuronal apoptosis is characterized by specific morphological events and requires the activation of an intrinsic transcriptional program. With the completion of genome sequencing in humans and model organisms, and the advent of DNA microarray technology, the transcriptional cascades and networks regulating neuronal apoptosis are being elucidated providing new potential pharmacological targets. This review will introduce the reader to this genomic approach and illustrate with a few examples a methodological strategy for the rational selection of pharmacological targets and the development of neuroprotective agents.

Recent results of the laser ion source facility at INFN-LNS and applications to nuclear and applied research.

A pulsed neodymium-doped yttrium aluminum garnet laser ion source has been used as proton beams generator. The laser wavelength is 1064 nm, the pulse duration is 9 ns and the intensity reaches 10(10) W/cm(2). Laser irradiates hydrogenated polymers targets located in a chamber at 10(-7) mbar. The ions are post-accelerated in a suitable chamber by 30 kV of voltage between the target, positively biased, and the following ground electrode. The extracted beams is characterized through a time-of-flight technique. Possible applications to the field of nuclear physics, such as nuclear excitation and de-excitations, nuclear reactions and nuclear fusion, will be presented and discussed.

Isospin dependence of incomplete fusion reactions at 25 MeV/nucleon.

40Ca+;{40,48}Ca,46Ti reactions at 25 MeV/nucleon have been studied using the 4pi CHIMERA detector. An isospin effect on the competition between fusionlike and binarylike reaction mechanisms has been observed. The probability of producing a heavy residue is lower in the case of N approximately Z colliding systems as compared to the case of reactions induced on the neutron rich 48Ca target. Predictions based on constrained molecular dynamics II calculations show that the competition between fusionlike and binary reactions in the selected centrality bins can constrain the parametrization of the symmetry energy and its density dependence in the nuclear equation of state.

Distribution of ADP-ribosylation factor-related protein 1 in mouse brain.

ADP-ribosylation factor (Arf)-related protein 1 (ARFRP1) is a membrane-associated GTPase, which inhibits the Arf/Sec7-dependent activation of phospholipase D and belongs to the Arf-like (Arl) GTPases. Although ARFRP1 is involved in post-Golgi membrane trafficking and its lack leads to embryonic lethality, little is known about its possible function in the central nervous system. To obtain more knowledge about ARFRP1, we have characterized its mRNA distribution in adult mouse brain by in situ hybridization and real-time PCR. We observed a widespread distribution of ARFRP1-mRNA, with the highest levels in cerebral cortex, thalamic nuclei, colliculus, substantia nigra and granule cell layer of cerebellum. Moderate levels were observed in some amygdaloid nuclei, CA2 area and dentate gyrus of hippocampus, endopiriform nuclei, globus pallidus, striatum, molecular layer of cerebellum, and locus coeruleus, whereas no expression was detected in hypothalamic nuclei, CA1 and CA3 areas of hippocampus, zona incerta. A significant decrease of ARFRP1-mRNA was observed in cerebral cortex following sleep deprivation, whereas no change was observed in cerebellar cortex, locus courelus, brainstem, hippocampus and pontine nuclei. This study provides the first detailed analysis of the regional distribution of ARFRP1 in the mouse brain and a quantitative view of its changes following sleep deprivation.

Fast ternary and quaternary breakup of the 197Au + 197Au system in collisions at 15 MeV/nucleon.

A new reaction mechanism of violent reseparation of a heavy nucleus-nucleus system, 197Au + 197Au, into three or four massive fragments in collisions at 15 MeV/nucleon has been observed. After reseparation, the fragments are almost exactly aligned, thus showing a very short time scale of the reseparation process, of about 70-80 fm/c.

Early genomics of learning and memory: a review.

The characterization of the molecular mechanisms whereby our brain codes, stores and retrieves memories remains a fundamental puzzle in neuroscience. Despite the knowledge that memory storage involves gene induction, the identification and characterization of the effector genes has remained elusive. The completion of the Human Genome Project and a variety of new technologies are revolutionizing the way these mechanisms can be explored. This review will examine how a genomic approach can be used to dissect and analyze the complex dynamic interactions involved in gene regulation during learning and memory. This innovative approach is providing information on a new class of genes associated with learning and memory in health and disease and is elucidating new molecular targets and pathways whose pharmacological modulation may allow new therapeutic approaches for improving cognition.

Adding semantics to gene expression profiles: new tools for drug discovery.

Gene expression profiles are unveiling a wealth of new potential drug targets for a wide range of diseases, offering new opportunities for drug discoveries. The emerging challenge, however, is the effective selection of the myriad of targets to identify those with the most therapeutic utility. Numerical clustering has became a commonly used method to investigate and interpret gene expression data sets but it is often inadequate to infer the genes' and proteins' role and point to candidate genes for drug development. This review illustrates how clustering methods based on semantic characteristics, such as gene ontologies, could be used to extract more knowledge from genomic data and improve drug target and discovery processes.

Age-dependent differential expression of BACE splice variants in brain regions of tg2576 mice.

Plaques found in the brains of patients suffering from Alzheimer's disease (AD) mainly consist of beta-amyloid (Abeta), which is produced by sequential cleaving of amyloid precursor protein (APP) by two proteolytic enzymes, beta- and gamma-secretases. Any change in the fine balance between these enzymes and their substrate may contribute to the etio-pathogenesis of AD. Indeed, the protein level and enzymatic activity of beta-secretase (BACE), but not its mRNA level, were found elevated in brain areas of AD patients who suffer a high load of Abeta plaque formation. Similarly, increased BACE activity but no mRNA change was observed in a transgenic mouse model of AD, tg2576, in which over expression of the Swedish mutated human APP leads to Abeta plaque formation and learning deficits. Based on the recent demonstration of four BACE splice variants with different enzymatic activity, the discrepancy between BACE activity and mRNA expression may be explained by the altered BACE alternative splicing. To test this hypothesis, we studied the expression of all BACE splice variants in different brain areas of tg2576 mice at age of 4 months and 1 year old. We found developmental and regional differences between wild-type and tg2576 mice. Our results indicate that over expression of APP in tg2576 mice leads to the altered alternative splicing of BACE and the increase of its enzymatically more active splice variant (I-501).

Down regulation of cerebellar memory related gene-1 following classical conditioning.

We have isolated and characterized the mRNA of a mouse gene named cerebellar memory related gene-1, previously found by microarray analysis to be differentially expressed following classical conditioning of the rabbit nictitating membrane response. Quantitative RT-PCR analysis showed a significant reduction in mRNA expression in cerebellar lobule HVI but not in the hippocampus of rabbits that received classical conditioning compared to control rabbits that received either unpaired stimulus presentations or were simply restrained. The mouse mRNA encodes a protein of 485 amino acids that includes different potential post-translational modification sites and five copies of the WD-repeat suggesting involvement in protein-protein interaction and regulatory function. In-situ hybridization experiments show highly localized expression of the transcript in mouse brain with the highest expression levels located in the cerebellum, hippocampus and cortex. Taken together, our results reveal a novel gene encoding a WD-repeat protein that is down-regulated in cerebellar lobule HVI as a result of learning and memory.

Pituitary adenylate cyclase-activating polypeptide modifies the electrical activity of CA1 hippocampal neurons in the rat.

The effects of pituitary adenylate cyclase-activating polypeptide (PACAP) on neuronal excitability in the CA1 region of rat hippocampus were studied using in vivo and in vitro electrophysiological techniques. Extracellularly recorded spontaneous firing of CA1 neurons was transiently (2-7 min) increased by PACAP (106+/-32% enhancement, mean+/-SEM, n=11). Using whole-cell patch clamp, PACAP was tested on the resting membrane current of CA1 pyramidal neurons: PACAP activated a slow-onset (20-30 s) and long-lasting (over 20 min) inward current with a mean amplitude of 99+/-34 pA (mean+/-SD, n=8). These results indicate that PACAP induces depolarizing effects on CA1 hippocampal neurons. PACAP-induced long-lasting facilitation in the CA1 region might modify neuronal excitability and/or modulate the effect of other neurotransmitters.

[Occupational accidents in temporary work]. / Il fenomeno infortunistico nel lavoro interinale.

In 2000 in Italy about 470,000 workers found jobs provided by Temporary Work Agencies, i.e., more than twice the number in 1999. Temporary Work is an expanding phenomenon since it well suits the flexibility requirements of business: 77% of Temporary Work is in the manufacturing industry, involving mostly unskilled workers. The young age, low level of qualification, poor training, and lack of knowledge of the working environment are factors that can favour occupational accidents. There are studies that relate the frequency of labour accidents to the level of training and job experience. There are no data in the literature about occupational accidents during temporary work. Our study concerns sixteen temporary work Agencies that employed more than 250,000 manual workers in 2000. From the occupational accident registers of these agencies, we analysed all accidents concerning manual workers employed during 2000, with reference to: the number of accidents with up to three days prognosis, age, sex, nationality, prognosis pattern and cause. The agencies supplied data regarding the number of hours worked during 2000. We considered 5259 accidents: the overall frequency index (FI) was 92.1, with a considerable difference between the three main agencies (87.5) and the others. The mean age accident was 27.8 years, mean duration of sick leave 13.7 days, and the main causes were work tools (51.5%); 76% of the accidents concerned unskilled manual workers. The above indexes are definitely higher than those provided by INAIL (italian compulsory insurances) for the metalworking (38.1), construction (47.7) and mining (58.8) sectors during 1997. There is evidence that temporary work is related to an increased risk of occupational accidents. Further studies are required to confirm this evidence.