Antagonists of the Receptor-G Protein Interface Block Gi-coupled Signal Transduction

The carboxyl terminus of heterotrimeric G protein alpha subunits plays an important role in receptor interaction. We demonstrate that peptides corresponding to the last 11 residues of Galphai1/2 or Galphao1 impair agonist binding to A1 adenosine receptors, whereas Galphas or Galphat peptides have no effect. Previously, by using a combinatorial library we identified a series of Galphat peptide analogs that bind rhodopsin with high affinity (Martin, E. L., Rens-Domiano, S., Schatz, P. J., and Hamm, H. E. (1996) J. Biol. Chem. 271, 361-366). Native Galphai1/2 peptide as well as several analogs were tested for their ability to modulate agonist binding or antagonist-agonist competition using cells overexpressing human A1 adenosine receptors. Three peptide analogs decreased the Ki, suggesting that they disrupt the high affinity receptor-G protein interaction and stabilize an intermediate affinity state. To study the ability of the peptides to compete with endogenous Galphai proteins and block signal transduction in a native setting, we measured activation of G protein-coupled K+ channels through A1 adenosine or gamma-aminobutyric acid, type B, receptors in hippocampal CA1 pyramidal neurons. Native Galphai1/2, peptide, and certain analog peptides inhibited receptor-mediated K+ channel gating, dependent on which receptor was activated. This differential perturbation of receptor-G protein interaction suggests that receptors that act on the same G protein can be selectively disrupted

Serum IgG antibodies from pregnant women reacting to mimotopes of simian virus 40 large T antigen, the viral oncoprotein

Simian virus 40 (SV40) large T antigen (LT) coding sequences were revealed in different human samples, whereas SV40 antibodies (Ab) were detected in human sera of cancer patients and healthy individuals, although with a lower prevalence. Previous studies carried out by the neutralization assay gave a SV40 seroprevalence, in the general population, up to 8%, although higher rates, 12%, were detected in kidney transplant children, in a group of HIV-positive patients, and in healthy females. In this study, serum samples from pregnant women, together with those from non-pregnant women, were analyzed to check the prevalence of IgG Ab reacting to SV40 LT antigens. Serum samples were collected from pregnant and non-pregnant women, with the same mean age. Women were in the range of 15-48 years old. Samples were assayed by an indirect ELISA employing specific SV40 LT mimotopes as antigens, whereas functional analysis was performed by neutralization of the viral infectivity in cell cultures. As a control, sera were analyzed for Ab against BK polyomavirus (BKPyV), which is a human polyomavirus homologous to SV40. Statistical analyses employed chi-square with Yates' correction, and Student's t tests. Indirect ELISAs indicated that pregnant women tested SV40 LT-positive with a prevalence of 17% (23/134), whereas non-pregnant women had a prevalence of 20% (36/180) (P > 0.05). Ab against BKPyV were detected with a prevalence of 80% in pregnant women and with a prevalence of 78% in non-pregnant women. These data indicate that SV40 infects at a low prevalence pregnant women. We may speculate that SV40, or a close human polyomavirus still undetected, could be transmitted from mother to fetus

Palmitate-induced lipotoxicity alters acetylation of multiple proteins in clonal β cells and human pancreatic islets

Type 2 diabetes is characterized by progressive β cell dysfunction, with lipotoxicity playing a possible pathogenetic role. Palmitate is often used to examine the direct effects of lipotoxicity and it may cause mitochondrial alterations by activating protein acetylation. However, it is unknown whether palmitate influences protein acetylation in β cells. We investigated lysine acetylation in mitochondrial proteins from INS-1E β cells (INS-1E) and in proteins from human pancreatic islets (HPI) after 24 h palmitate exposure. First, we confirmed that palmitate damages β cells and demonstrated that chemical inhibition of deacetylation also impairs INS-1E function and survival. Then, by 2-D gel electrophoresis, Western Blot and Liquid Chromatography-Mass Spectrometry we evaluated the effects of palmitate on protein acetylation. In mitochondrial preparations from palmitate-treated INS-1E, 32 acetylated spots were detected, with 13 proteins resulting over-acetylated. In HPI, 136 acetylated proteins were found, of which 11 were over-acetylated upon culture with palmitate. Interestingly, three proteins, glutamate dehydrogenase, mitochondrial superoxide dismutase, and SREBP-1, were over-acetylated in both INS-1E and HPI. Therefore, prolonged exposure to palmitate induces changes in β cell protein lysine acetylation and this modification could play a role in causing β cell damage. Dysregulated acetylation may be a target to counteract palmitate-induced β cell lipotoxicity

High availability using virtualization - 3RC

High availability has always been one of the main problems for a data center. Till now high availability was achieved by host per host redundancy, a highly expensive method in terms of hardware and human costs. A new approach to the problem can be offered by virtualization. Using virtualization, it is possible to achieve a redundancy system for all the services running on a data center. This new approach to high availability allows the running virtual machines to be distributed over a small number of servers, by exploiting the features of the virtualization layer: start, stop and move virtual machines between physical hosts. The 3RC system is based on a finite state machine, providing the possibility to restart each virtual machine over any physical host, or reinstall it from scratch. A complete infrastructure has been developed to install operating system and middleware in a few minutes. To virtualize the main servers of a data center, a new procedure has been developed to migrate physical to virtual hosts. The whole Grid data center SNS-PISA is running at the moment in virtual environment under the high availability system.Comment: 10 page

Serum IgG against Simian Virus 40 antigens are hampered by high levels of sHLA-G in patients affected by inflammatory neurological diseases, as multiple sclerosis

Background: Many investigators detected the simian polyomavirus SV40 footprints in human brain tumors and neurologic diseases and recently it has been indicated that SV40 seems to be associated with multiple sclerosis (MS) disease. Interestingly, SV40 interacts with human leukocyte antigen (HLA) class I molecules for cell entry. HLA class I antigens, in particular non-classical HLA-G molecules, characterized by an immune-regulatory function, are involved in MS disease, and the levels of these molecules are modified according with the disease status. Objective: We investigated in serum samples, from Italian patients affected by MS, other inflammatory diseases (OIND), non-inflammatory neurological diseases (NIND) and healthy subjects (HS), SV40-antibody and soluble sHLA-G and the association between SV40-prevalence and sHLA-G levels. Methods: ELISA tests were used for SV40-antibodies detection and sHLA-G quantitation in serum samples. Results: The presence of SV40 antibodies was observed in 6 % of patients affected by MS (N = 4/63), 10 % of OIND (N = 8/77) and 15 % of NIND (N = 9/59), which is suggestive of a lower prevalence in respect to HS (22 %, N = 18/83). MS patients are characterized by higher sHLA-G serum levels (13.9 \ub1 0.9 ng/ml; mean \ub1 St. Error) in comparison with OIND (6.7 \ub1 0.8 ng/ml), NIND (2.9 \ub1 0.4 ng/ml) and HS (2.6 \ub1 0.7 ng/ml) subjects. Interestingly, we observed an inverse correlation between SV40 antibody prevalence and sHLA-G serum levels in MS patients. Conclusion: The data obtained showed a low prevalence of SV40 antibodies in MS patients. These results seems to be due to a generalized status of inability to counteract SV40 infection via antibody production. In particular, we hypothesize that SV40 immune-inhibitory direct effect and the presence of high levels of the immune-inhibitory HLA-G molecules could co-operate in impairing B lymphocyte activation towards SV40 specific peptides

Inter-fractional monitoring of 12 C ions treatments: results from a clinical trial at the CNAO facility

The high dose conformity and healthy tissue sparing achievable in Particle Therapy when using C ions calls for safety factors in treatment planning, to prevent the tumor under-dosage related to the possible occurrence of inter-fractional morphological changes during a treatment. This limitation could be overcome by a range monitor, still missing in clinical routine, capable of providing on-line feedback. The Dose Profiler (DP) is a detector developed within the INnovative Solution for In-beam Dosimetry in hadronthErapy (INSIDE) collaboration for the monitoring of carbon ion treatments at the CNAO facility (Centro Nazionale di Adroterapia Oncologica) exploiting the detection of charged secondary fragments that escape from the patient. The DP capability to detect inter-fractional changes is demonstrated by comparing the obtained fragment emission maps in different fractions of the treatments enrolled in the first ever clinical trial of such a monitoring system, performed at CNAO. The case of a CNAO patient that underwent a significant morphological change is presented in detail, focusing on the implications that can be drawn for the achievable inter-fractional monitoring DP sensitivity in real clinical conditions. The results have been cross-checked against a simulation study

On the Dynamics of the Spontaneous Activity in Neuronal Networks

Most neuronal networks, even in the absence of external stimuli, produce spontaneous bursts of spikes separated by periods of reduced activity. The origin and functional role of these neuronal events are still unclear. The present work shows that the spontaneous activity of two very different networks, intact leech ganglia and dissociated cultures of rat hippocampal neurons, share several features. Indeed, in both networks: i) the inter-spike intervals distribution of the spontaneous firing of single neurons is either regular or periodic or bursting, with the fraction of bursting neurons depending on the network activity; ii) bursts of spontaneous spikes have the same broad distributions of size and duration; iii) the degree of correlated activity increases with the bin width, and the power spectrum of the network firing rate has a 1/f behavior at low frequencies, indicating the existence of long-range temporal correlations; iv) the activity of excitatory synaptic pathways mediated by NMDA receptors is necessary for the onset of the long-range correlations and for the presence of large bursts; v) blockage of inhibitory synaptic pathways mediated by GABA(A) receptors causes instead an increase in the correlation among neurons and leads to a burst distribution composed only of very small and very large bursts. These results suggest that the spontaneous electrical activity in neuronal networks with different architectures and functions can have very similar properties and common dynamics