Phosphorylation does not prompt, nor prevent, the formation of α-synuclein toxic species in a rat model of Parkinson's disease

Phosphorylation is involved in numerous neurodegenerative diseases. In particular, alpha-synuclein is extensively phosphorylated in aggregates in patients suffering from synucleinopathies. However, the share of this modification in the events that lead to the conversion of alpha-synuclein to aggregated toxic species needed to be clarified. The rat model that we developed through rAAV2/6-mediated expression of alpha-synuclein demonstrates a correlation between neurodegeneration and formation of small filamentous alpha-synuclein aggregates. A mutation preventing phosphorylation (S129A) significantly increases alpha-synuclein toxicity and leads to enhanced formation of beta-sheet-rich, proteinase K-resistant aggregates, increased affinity for intracellular membranes, a disarrayed network of neurofilaments and enhanced alpha-synuclein nuclear localization. The expression of a mutation mimicking phosphorylation (S129D) does not lead to dopaminergic cell loss. Nevertheless, fewer but larger aggregates are formed, and signals of apoptosis are also activated in rats expressing the phosphorylation-mimicking form of alpha-synuclein. These observations strongly suggest that phosphorylation does not play an active role in the accumulation of cytotoxic pre-inclusion aggregates. Unexpectedly, the study also demonstrates that constitutive expression of phosphorylation-mimicking forms of alpha-synuclein does not protect from neurodegeneration. The role of phosphorylation at Serine 129 in the early phase of Parkinson's disease is examined, which brings new perspective to therapeutic approaches focusing on the modulation of kinases/phosphatases activity to control alpha-synuclein toxicit

Intact satellite cells lead to remarkable protection against Smn gene defect in differentiated skeletal muscle

Deletion of murine Smn exon 7, the most frequent mutation found in spinal muscular atrophy, has been directed to either both satellite cells, the muscle progenitor cells and fused myotubes, or fused myotubes only. When satellite cells were mutated, mutant mice develop severe myopathic process, progressive motor paralysis, and early death at 1 mo of age (severe mutant). Impaired muscle regeneration of severe mutants correlated with defect of myogenic precursor cells both in vitro and in vivo. In contrast, when satellite cells remained intact, mutant mice develop similar myopathic process but exhibit mild phenotype with median survival of 8 mo and motor performance similar to that of controls (mild mutant). High proportion of regenerating myofibers expressing SMN was observed in mild mutants compensating for progressive loss of mature myofibers within the first 6 mo of age. Then, in spite of normal contractile properties of myofibers, mild mutants develop reduction of muscle force and mass. Progressive decline of muscle regeneration process was no more able to counterbalance muscle degeneration leading to dramatic loss of myofibers. These data indicate that intact satellite cells remarkably improve the survival and motor performance of mutant mice suffering from chronic myopathy, and suggest a limited potential of satellite cells to regenerate skeletal muscle

Deletion of Murine SMN Exon 7 Directed to Skeletal Muscle Leads to Severe Muscular Dystrophy

Spinal muscular atrophy (SMA) is characterized by degeneration of motor neurons of the spinal cord associated with muscle paralysis and caused by mutations of the survival motor neuron gene (SMN). To determine whether SMN gene defect in skeletal muscle might have a role in SMA pathogenesis, deletion of murine SMN exon 7, the most frequent mutation found in SMA, has been restricted to skeletal muscle by using the Cre-loxP system. Mutant mice display ongoing muscle necrosis with a dystrophic phenotype leading to muscle paralysis and death. The dystrophic phenotype is associated with elevated levels of creatine kinase activity, Evans blue dye uptake into muscle fibers, reduced amount of dystrophin and upregulation of utrophin expression suggesting a destabilization of the sarcolemma components. The mutant mice will be a valuable model for elucidating the underlying mechanism. Moreover, our results suggest a primary involvement of skeletal muscle in human SMA, which may contribute to motor defect in addition to muscle denervation caused by the motor neuron degeneration. These data may have important implications for the development of therapeutic strategies in SMA

Protein 4.1B Contributes to the Organization of Peripheral Myelinated Axons

Neurons are characterized by extremely long axons. This exceptional cell shape is likely to depend on multiple factors including interactions between the cytoskeleton and membrane proteins. In many cell types, members of the protein 4.1 family play an important role in tethering the cortical actin-spectrin cytoskeleton to the plasma membrane. Protein 4.1B is localized in myelinated axons, enriched in paranodal and juxtaparanodal regions, and also all along the internodes, but not at nodes of Ranvier where are localized the voltage-dependent sodium channels responsible for action potential propagation. To shed light on the role of protein 4.1B in the general organization of myelinated peripheral axons, we studied 4.1B knockout mice. These mice displayed a mildly impaired gait and motility. Whereas nodes were unaffected, the distribution of Caspr/paranodin, which anchors 4.1B to the membrane, was disorganized in paranodal regions and its levels were decreased. In juxtaparanodes, the enrichment of Caspr2, which also interacts with 4.1B, and of the associated TAG-1 and Kv1.1, was absent in mutant mice, whereas their levels were unaltered. Ultrastructural abnormalities were observed both at paranodes and juxtaparanodes. Axon calibers were slightly diminished in phrenic nerves and preterminal motor axons were dysmorphic in skeletal muscle. βII spectrin enrichment was decreased along the axolemma. Electrophysiological recordings at 3 post-natal weeks showed the occurrence of spontaneous and evoked repetitive activity indicating neuronal hyperexcitability, without change in conduction velocity. Thus, our results show that in myelinated axons 4.1B contributes to the stabilization of membrane proteins at paranodes, to the clustering of juxtaparanodal proteins, and to the regulation of the internodal axon caliber

Nodes of Ranvier and Paranodes in Chronic Acquired Neuropathies

Chronic acquired neuropathies of unknown origin are classified as chronic inflammatory demyelinating polyneuropathies (CIDP) and chronic idiopathic axonal polyneuropathies (CIAP). The diagnosis can be very difficult, although it has important therapeutic implications since CIDP can be improved by immunomodulating treatment. The aim of this study was to examine the possible abnormalities of nodal and paranodal regions in these two types of neuropathies. Longitudinal sections of superficial peroneal nerves were obtained from biopsy material from 12 patients with CIDP and 10 patients with CIAP and studied by immunofluorescence and in some cases electron microscopy. Electron microscopy revealed multiple alterations in the nodal and paranodal regions which predominated in Schwann cells in CIDP and in axons in CIAP. In CIDP paranodin/Caspr immunofluorescence was more widespread than in control nerves, extending along the axon in internodes where it appeared intense. Nodal channels Nav and KCNQ2 were less altered but were also detected in the internodes. In CIAP paranodes, paranodin labeling was irregular and/or decreased. To test the consequences of acquired primary Schwann cells alteration on axonal proteins, we used a mouse model based on induced deletion of the transcription factor Krox-20 gene. In the demyelinated sciatic nerves of these mice we observed alterations similar to those found in CIDP by immunofluorescence, and immunoblotting demonstrated increased levels of paranodin. Finally we examined whether the alterations in paranodin immunoreactivity could have a diagnosis value. In a sample of 16 biopsies, the study of paranodin immunofluorescence by blind evaluators led to correct diagnosis in 70±4% of the cases. This study characterizes for the first time the abnormalities of nodes of Ranvier in CIAP and CIDP, and the altered expression and distribution of nodal and paranodal proteins. Marked differences were observed between CIDP and CIAP and the alterations in paranodin immunofluorescence may be an interesting tool for their differential diagnosis

Strategies to reengage patients lost to follow up in HIV care in high income countries, a scoping review

Background: Despite remarkable achievements in antiretroviral therapy (ART), losses to follow-up (LTFU) might prevent the long-term success of HIV treatment and might delay the achievement of the 90-90-90 objectives. This scoping review is aimed at the description and analysis of the strategies used in high-income countries to reengage LTFU in HIV care, their implementation and impact. Methods: A scoping review was done following Arksey & O'Malley's methodological framework and recommendations from Joanna Briggs Institute. Peer reviewed articles were searched for in Pubmed, Scopus and Web of Science; and grey literature was searched for in Google and other sources of information. Documents were charted according to the information presented on LTFU, the reengagement procedures used in HIV units in high-income countries, published during the last 15 years. In addition, bibliographies of chosen articles were reviewed for additional articles. Results: Twenty-eight documents were finally included, over 80% of them published in the United States later than 2015. Database searches, phone calls and/or mail contacts were the most common strategies used to locate and track LTFU, while motivational interviews and strengths-based techniques were used most often during reengagement visits. Outcomes like tracing activities efficacy, rates of reengagement and viral load reduction were reported as outcome measures. Conclusions: This review shows a recent and growing trend in developing and implementing patient reengagement strategies in HIV care. However, most of these strategies have been implemented in the United States and little information is available for other high-income countries. The procedures used to trace and contact LTFU are similar across reviewed studies, but their impact and sustainability are widely different depending on the country studied

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

MUSiC : a model-unspecific search for new physics in proton-proton collisions at root s=13TeV

Results of the Model Unspecific Search in CMS (MUSiC), using proton-proton collision data recorded at the LHC at a centre-of-mass energy of 13 TeV, corresponding to an integrated luminosity of 35.9 fb(-1), are presented. The MUSiC analysis searches for anomalies that could be signatures of physics beyond the standard model. The analysis is based on the comparison of observed data with the standard model prediction, as determined from simulation, in several hundred final states and multiple kinematic distributions. Events containing at least one electron or muon are classified based on their final state topology, and an automated search algorithm surveys the observed data for deviations from the prediction. The sensitivity of the search is validated using multiple methods. No significant deviations from the predictions have been observed. For a wide range of final state topologies, agreement is found between the data and the standard model simulation. This analysis complements dedicated search analyses by significantly expanding the range of final states covered using a model independent approach with the largest data set to date to probe phase space regions beyond the reach of previous general searches.Peer reviewe

Measurement of prompt open-charm production cross sections in proton-proton collisions at root s=13 TeV

The production cross sections for prompt open-charm mesons in proton-proton collisions at a center-of-mass energy of 13TeV are reported. The measurement is performed using a data sample collected by the CMS experiment corresponding to an integrated luminosity of 29 nb(-1). The differential production cross sections of the D*(+/-), D-+/-, and D-0 ((D) over bar (0)) mesons are presented in ranges of transverse momentum and pseudorapidity 4 < p(T) < 100 GeV and vertical bar eta vertical bar < 2.1, respectively. The results are compared to several theoretical calculations and to previous measurements.Peer reviewe