Oligomeric assembly of the C-terminal and transmembrane region of SARS-CoV-2 nsp3.

As for all single-stranded, positive-sense RNA (+RNA) viruses, intracellular RNA synthesis relies on extensive remodeling of host cell membranes that leads to the formation of specialized structures. In the case of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus causing COVID-19, endoplasmic reticulum membranes are modified, resulting in the formation of double-membrane vesicles (DMVs), which contain the viral dsRNA intermediate and constitute membrane-bound replication organelles. The non-structural and transmembrane protein nsp3 is a key player in the biogenesis of DMVs and, therefore, represents an interesting antiviral target. However, as an integral transmembrane protein, it is challenging to express for structural biology. The C-terminus of nsp3 encompasses all the membrane-spanning, -interacting, and -remodeling elements. By using a cell-free expression system, we successfully produced the C-terminal region of nsp3 (nsp3C) and reconstituted purified nsp3C into phospholipid nanodiscs, opening the way for structural studies. Negative-stain transmission electron microscopy revealed the presence of nsp3C oligomers very similar to the region abutting and spanning the membrane on the cytosolic side of DMVs in a recent subtomogram average of the SARS-CoV-2 nsp3-4 pore (1). AlphaFold-predicted structural models fit particularly well with our experimental data and support a pore-forming hexameric assembly. Altogether, our data give unprecedented clues to understand the structural organization of nsp3, the principal component that shapes the molecular pore that spans the DMVs and is required for the export of RNA in vivo. IMPORTANCE: Membrane remodeling is at the heart of intracellular replication for single-stranded, positive-sense RNA viruses. In the case of coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), this leads to the formation of a network of double-membrane vesicles (DMVs). Targeting DMV biogenesis offers promising prospects for antiviral therapies. This requires a better understanding of the molecular mechanisms and proteins involved. Three non-structural proteins (nsp3, nsp4, and nsp6) direct the intracellular membrane rearrangements upon SARS-CoV-2 infection. All of them contain transmembrane helices. The nsp3 component, the largest and multi-functional protein of the virus, plays an essential role in this process. Aiming to understand its structural organization, we used a cell-free protein synthesis assay to produce and reconstitute the C-terminal part of nsp3 (nsp3C) including transmembrane domains into phospholipid nanodiscs. Our work reveals the oligomeric organization of one key player in the biogenesis of SARS-CoV-2 DMVs, providing basis for the design of future antiviral strategies.

Atomic structure of Lanreotide nanotubes revealed by cryo-EM.

Functional and versatile nano- and microassemblies formed by biological molecules are found at all levels of life, from cell organelles to full organisms. Understanding the chemical and physicochemical determinants guiding the formation of these assemblies is crucial not only to understand the biological processes they carry out but also to mimic nature. Among the synthetic peptides forming well-defined nanostructures, the octapeptide Lanreotide has been considered one of the best characterized, in terms of both the atomic structure and its self-assembly process. In the present work, we determined the atomic structure of Lanreotide nanotubes at 2.5-Å resolution by cryoelectron microscopy (cryo-EM). Surprisingly, the asymmetric unit in the nanotube contains eight copies of the peptide, forming two tetramers. There are thus eight different environments for the peptide, and eight different conformations in the nanotube. The structure built from the cryo-EM map is strikingly different from the molecular model, largely based on X-ray fiber diffraction, proposed 20 y ago. Comparison of the nanotube with a crystal structure at 0.83-Å resolution of a Lanreotide derivative highlights the polymorphism for this peptide family. This work shows once again that higher-order assemblies formed by even well-characterized small peptides are very difficult to predict.

Structure and conformational variability of the HER2-trastuzumab-pertuzumab complex.

Single particle analysis from cryogenic transmission electron microscopy (cryo-EM) is particularly attractive for complexes for which structure prediction remains intractable, such as antibody-antigen complexes. Here we obtain the detailed structure of a particularly difficult complex between human epidermal growth factor receptor 2 (HER2) and the antigen-binding fragments from two distinct therapeutic antibodies binding to distant parts of the flexible HER2, pertuzumab and trastuzumab (HTP). We highlight the strengths and limitations of current data processing software in dealing with various kinds of heterogeneities, particularly continuous conformational heterogeneity, and in describing the motions that can be extracted from our dataset. Our HTP structure provides a more detailed view than the one previously available for this ternary complex. This allowed us to pinpoint a previously overlooked loop in domain IV that may be involved both in binding of trastuzumab and in HER2 dimerization. This finding may contribute to explain the synergistic anticancer effect of the two antibodies. We further propose that the flexibility of the HTP complex, beyond the difficulties it causes for cryo-EM analysis, actually reflects regulation of HER2 signaling and its inhibition by therapeutic antibodies. Notably we obtain our best data with ultra-thin continuous carbon grids, showing that with current cameras their use to alleviate particle misdistribution is compatible with a protein complex of only 162 kDa. Perhaps most importantly, we provide here a dataset for such a smallish protein complex for further development of software accounting for continuous conformational heterogeneity in cryo-EM images.

Clustering of Tau fibrils impairs the synaptic composition of α3-Na+/K+-ATPase and AMPA receptors.

Tau assemblies have prion-like properties: they propagate from one neuron to another and amplify by seeding the aggregation of endogenous Tau. Although key in prion-like propagation, the binding of exogenous Tau assemblies to the plasma membrane of naïve neurons is not understood. We report that fibrillar Tau forms clusters at the plasma membrane following lateral diffusion. We found that the fibrils interact with the Na+/K+-ATPase (NKA) and AMPA receptors. The consequence of the clustering is a reduction in the amount of α3-NKA and an increase in the amount of GluA2-AMPA receptor at synapses. Furthermore, fibrillar Tau destabilizes functional NKA complexes. Tau and α-synuclein aggregates often co-exist in patients' brains. We now show evidences for cross-talk between these pathogenic aggregates with α-synuclein fibrils dramatically enhancing fibrillar Tau clustering and synaptic localization. Our results suggest that fibrillar α-synuclein and Tau cross-talk at the plasma membrane imbalance neuronal homeostasis.

Novel Management of Granuloma Formation Secondary to Dermal Filler with Intralesional 1444 nm Nd:YAG Laser Technique.

Background: Dermal fillers for soft tissue for the treatment of face sagging, volume loss, and wrinkles have become popular among patients of all ages and ethnicities, and their use is becoming increasingly widespread. Aim: the goal of this study was to evaluate the effectiveness and safety of a micro-pulsed, 1444 nm Nd:YAG laser on dermal filler complications, in particular on granuloma management. Methods: A subcutaneous, 1444 nm Nd:YAG laser was used on five female patients (range age 52-68 years) with hyaluronic filler granulomas located on the face (two on the cheek area and three on the lips); three patients had self-injected the filler, buying it online. Before and after the therapy, the patients received a skin ultrasound to determine the form and location of the granulomas and to determine if there had been a full or partial resolution. During this study, all possible adverse effects at the treatment site were monitored. The 5-point Global Aesthetic Improvement Scale (GAIS) (0 point-no change; 1 point-25%, mild improvement; 2 points-50%, moderate improvement; 3 points-75%, good improvement; 4 points-100%, excellent improvement) was recorded at a 3-month follow-up. Results: good results were obtained in the treatment of filler granulomas with the intralesional 1444 nm laser, even if just a single treatment was performed (one intervention was effective for curing granulomas up to 5 mm in diameter). Three patients were satisfied with excellent improvement, and two patients experienced good improvement. The results are functional and aesthetically satisfying, as shown by photographic assessment. At the last follow-up, the granuloma had reduced or completely disappeared in all cases, and no infections, burns, scarring or fibrosis, episodes of severe bleeding, or other serious adverse effects had been reported. All subjects tolerated the post-treatment period well. Conclusions: Our findings showed that granuloma treatment with an intralesional 1444 nm Nd:YAG laser is a minimally invasive, easy, fast, efficient, and low-risk procedure.

Ex Vivo Human Histology Fractional Treatment with a New CO2 Scanner: A Potential Application on Deep Scarring.

Background and Objectives: For many years, fully ablative laser treatments, particularly those performed with a carbon dioxide (CO2) laser, were regarded as the gold standard for resurfacing. This study's goal is to assess the depth that can be reached by a new CO2 scanner system, through a skin model with greater dermal thickness, to use in the treatment of deep scarring. Materials and Methods: Male human skin tissue was laser-treated using a CO2 fractional laser and a new scanning system, and all samples were fixed in 10% neutral buffered formalin, dehydrated using a series of crescent alcohol, embedded in paraffin, sectioned in series (4-5 µm thick), stained with haematoxylin and eosin (H&E), and then analysed under an optical microscope. Results: From the epidermis through the underlying papillary and reticular dermis to various depths of the dermis, microablation columns of damage and coagulated microcolumns of collagen were observed. The reticular dermis was fully penetrated up to 6 mm at higher energy levels (210 mJ/DOT), resulting in deeper tissue injury. Although the laser might penetrate further, the skin stops there, leaving just the fat and muscular tissue. Conclusions: The deep layers of the dermis can be penetrated by the CO2 laser system throughout the entire dermal thickness when using the new scanning system, suggesting that this laser's potential impact, at the selected settings, covers all skin targets required to perform superficial or deep treatments on any dermatological issue. Finally, patients who have problems, such as morbid scar-deep complications, which affect their quality of life, are more likely to profit from this innovative technique.

Synergistic Sequential Emission of Fractional 10.600 and 1540 nm Lasers for Skin Resurfacing: An Ex Vivo Histological Evaluation.

Background: Fractional ablative and non-ablative lasers are useful treatments for skin rejuvenation. A procedure that provides the sequential application of fractional ablative followed by non-ablative laser treatment may reduce patients' downtime and deliver better cosmetic results than with either laser alone. Objective: The purpose of the current study was to demonstrate the ameliorative and therapeutic effects in skin remodeling of the synergistic use of the two laser wavelengths (fractional ablative CO2 and non-ablative 1540 nm) with three different types of pulse shapes, S-Pulse (SP), D-Pulse (DP) and H-Pulse (HP), through which the CO2 laser can emit, performing an ex vivo histological evaluation. Methods: In this prospective study, ex vivo sheep inner thigh skin was chosen due to its similarity to human skin tissue, and a histological evaluation was performed. Three irradiation conditions, using all of the three CO2 pulse shapes (alone or averaged), were investigated: (1) 10.600 nm alone, the sequential irradiation of the two wavelengths in the same perfectly controlled energy pulses (DOT) for the entire scan area; ((2) 10.600 nm followed immediately by 1540 nm; and (3) 1540 nm followed immediately by 10.600 nm). Results: When comparing ablative to sequential irradiations, the synergy of the two wavelengths did not alter the typical ablative pulse shape of the 10.600 nm laser alone. With the same CO2 pulse shape, the lesion depth did not vary with the synergy of the two wavelengths, while thermal lesion width increased compared to CO2 alone. The ablation rate was achieved, while the total thermal lesion coverage in the scanning area of CO2 - 1540 lasers was greater than when using CO2 alone and then the other sequential irradiation. Conclusions: This study provides important preclinical data for new and early uses of the novel 10.600/1540 nm dual-wavelength non-ablative fractional laser. The synergy of the two wavelengths enhanced all the benefits already available when using CO2 laser systems both in terms of tone strengthening, thanks to a greater shrinking effect, and in terms of stimulation and collagen remodeling thanks to a greater volumetric thermal effect.

Tunneling nanotubes spread fibrillar α-synuclein by intercellular trafficking of lysosomes.

Synucleinopathies such as Parkinson's disease are characterized by the pathological deposition of misfolded α-synuclein aggregates into inclusions throughout the central and peripheral nervous system. Mounting evidence suggests that intercellular propagation of α-synuclein aggregates may contribute to the neuropathology; however, the mechanism by which spread occurs is not fully understood. By using quantitative fluorescence microscopy with co-cultured neurons, here we show that α-synuclein fibrils efficiently transfer from donor to acceptor cells through tunneling nanotubes (TNTs) inside lysosomal vesicles. Following transfer through TNTs, α-synuclein fibrils are able to seed soluble α-synuclein aggregation in the cytosol of acceptor cells. We propose that donor cells overloaded with α-synuclein aggregates in lysosomes dispose of this material by hijacking TNT-mediated intercellular trafficking. Our findings thus reveal a possible novel role of TNTs and lysosomes in the progression of synucleinopathies.

Increased Immune Activation by Pathologic α-Synuclein in Parkinson's Disease.

OBJECTIVE: Excessive inflammation in the central nervous system (CNS) and the periphery can result in neurodegeneration and parkinsonism. Recent evidence suggests that immune responses in Parkinson disease patients are dysregulated, leading to an increased inflammatory reaction to unspecific triggers. Although α-synuclein pathology is the hallmark of Parkinson disease, it has not been investigated whether pathologic α-synuclein is a specific trigger for excessive inflammatory responses in Parkinson disease. METHODS: We investigated the immune response of primary human monocytes and a microglial cell line to pathologic forms of α-synuclein by assessing cytokine release upon exposure. RESULTS: We show that pathologic α-synuclein (mutations, aggregation) results in a robust inflammatory activation of human monocytes and microglial BV2 cells. The activation is conformation- dependent, with increasing fibrillation and early onset mutations having the strongest effect on immune activation. We also found that activation of immune cells by extracellular α-synuclein is potentiated by extracellular vesicles, possibly by facilitating the uptake of α-synuclein. Blood extracellular vesicles from Parkinson disease patients induce a stronger activation of monocytes than blood extracellular vesicles from healthy controls. Most importantly, monocytes from Parkinson disease patients are dysregulated and hyperactive in response to stimulation with pathologic α-synuclein. Furthermore, we demonstrate that α-synuclein pathology in the CNS is sufficient to induce the monocyte dysregulation in the periphery of a mouse model. INTERPRETATION: Taken together, our data suggest that α-synuclein pathology and dysregulation of monocytes in Parkinson disease can act together to induce excessive inflammatory responses to α-synuclein. ANN NEUROL 2019;86:593-606.

α-synuclein assemblies sequester neuronal α3-Na+/K+-ATPase and impair Na+ gradient.

Extracellular α-synuclein (α-syn) assemblies can be up-taken by neurons; however, their interaction with the plasma membrane and proteins has not been studied specifically. Here we demonstrate that α-syn assemblies form clusters within the plasma membrane of neurons. Using a proteomic-based approach, we identify the α3-subunit of Na+/K+-ATPase (NKA) as a cell surface partner of α-syn assemblies. The interaction strength depended on the state of α-syn, fibrils being the strongest, oligomers weak, and monomers none. Mutations within the neuron-specific α3-subunit are linked to rapid-onset dystonia Parkinsonism (RDP) and alternating hemiplegia of childhood (AHC). We show that freely diffusing α3-NKA are trapped within α-syn clusters resulting in α3-NKA redistribution and formation of larger nanoclusters. This creates regions within the plasma membrane with reduced local densities of α3-NKA, thereby decreasing the efficiency of Na+ extrusion following stimulus. Thus, interactions of α3-NKA with extracellular α-syn assemblies reduce its pumping activity as its mutations in RDP/AHC.

Ridge Preservation Using an Innovative Enzyme-deantigenic Equine Bone Paste: A Case Report with 36-month Follow-up.

AIM: This study aimed to report a successful clinical, histological, and histomorphometric outcome of a novel equine-derived bone paste for a ridge preservation surgery involving a single post-extractive socket. BACKGROUND: After tooth avulsion, unless the implant position is not carried out straightforwardly, the alveolar process undergoes resorption: to limit it, post-extractive sockets may be grafted according to the ridge preservation principles. Grafting materials should display proper biological properties and optimal handling characteristics. Bone pastes may facilitate grafting operations, avoid granules' dispersion, and maximize the contact of the graft with the surrounding bone. An innovative equine-derived bone paste has been recently introduced on the market, but its use has never been documented in the medical literature. CASE DESCRIPTION: This report describes the treatment of a patient who received the equine-derived bone paste in a post-extractive socket to allow the preservation of the alveolar ridge and was later rehabilitated with a crown supported by a single implant. CONCLUSION: The handling properties of the equine-derived bone paste were excellent. At the 36-month follow-up, the peri-implant bone levels had been maintained, with the implant being successful according to the Albrektsson and Zarb criteria. Histologic outcome showed that the bone paste was fully biocompatible; histomorphometric analysis showed that a significant amount of newly formed bone could be observed in the grafted socket. CLINICAL SIGNIFICANCE: Alveolar ridge preservation using bone grafts is a well-known approach, yet there is still no agreement about which bone graft might be considered the most suitable for this indication. The novel equine-derived bone paste used in the present study appears a promising option for effective socket preservation and may promote secondary intention healing. How to cite this article: Di Stefano DA, Arosio P, Cinci L, et al. Ridge Preservation Using an Innovative Enzyme-deantigenic Equine Bone Paste: A Case Report with 36-month Follow-up. J Contemp Dent Pract 2019;20(10):1229-1234.

Cellular response of human neuroblastoma cells to α-synuclein fibrils, the main constituent of Lewy bodies.

BACKGROUND: α-Synuclein (α-Syn) fibrils are the main constituent of Lewy bodies and a neuropathological hallmark of Parkinson's disease (PD). The propagation of α-Syn assemblies from cell to cell suggests that they are involved in PD progression. We previously showed that α-Syn fibrils are toxic because of their ability to bind and permeabilize cell membranes. Here, we document the cellular response in terms of proteome changes of SH-SY5Y cells exposed to exogenous α-Syn fibrils. METHODS: We compare the proteomes of cells of neuronal origin exposed or not either to oligomeric or fibrillar α-Syn using two dimensional differential in-gel electrophoresis (2D-DIGE) and mass spectrometry. RESULTS: Only α-Syn fibrils induce significant changes in the proteome of SH-SY5Y cells. In addition to proteins associated to apoptosis and toxicity, or proteins previously linked to neurodegenerative diseases, we report an overexpression of proteins involved in intracellular vesicle trafficking. We also report a remarkable increase in fibrillar α-Syn heterogeneity, mainly due to C-terminal truncations. CONCLUSIONS: Our results show that cells of neuronal origin adapt their proteome to exogenous α-Syn fibrils and actively modify those assemblies. GENERAL SIGNIFICANCE: Cells of neuronal origin adapt their proteome to exogenous toxic α-Syn fibrils and actively modify those assemblies. Our results bring insights into the cellular response and clearance events the cells implement to face the propagation of α-Syn assemblies associated to pathology.

Overexpression of the calpain-specific inhibitor calpastatin reduces human alpha-Synuclein processing, aggregation and synaptic impairment in [A30P]αSyn transgenic mice.

Lewy bodies, a pathological hallmark of Parkinson's disease (PD), contain aggregated alpha-synuclein (αSyn), which is found in several modified forms and can be discovered phosphorylated, ubiquitinated and truncated. Aggregation-prone truncated species of αSyn caused by aberrant cleavage of this fibrillogenic protein are hypothesized to participate in its sequestration into inclusions subsequently leading to synaptic dysfunction and neuronal death. Here, we investigated the role of calpain cleavage of αSyn in vivo by generating two opposing mouse models. We crossed into human [A30P]αSyn transgenic (i) mice deficient for calpastatin, a calpain-specific inhibitor, thus enhancing calpain activity (SynCAST(-)) and (ii) mice overexpressing human calpastatin leading to reduced calpain activity (SynCAST(+)). As anticipated, a reduced calpain activity led to a decreased number of αSyn-positive aggregates, whereas loss of calpastatin led to increased truncation of αSyn in SynCAST(-). Furthermore, overexpression of calpastatin decreased astrogliosis and the calpain-dependent degradation of synaptic proteins, potentially ameliorating the observed neuropathology in [A30P]αSyn and SynCAST(+) mice. Overall, our data further support a crucial role of calpains, particularly of calpain 1, in the pathogenesis of PD and in disease-associated aggregation of αSyn, indicating a therapeutic potential of calpain inhibition in PD.

Socket Preservation using Enzyme-treated Equine Bone Granules and an Equine Collagen Matrix: A Case Report with Histological and Histomorphometrical Assessment.

AIM: To histologically assess the effectiveness of a socket-preservation technique using enzyme-treated equine bone granules as a bone-graft material in combination with an equine collagen matrix as a scaffold for soft-tissue regeneration. BACKGROUND: Enzyme-treated equine bone granules and equine collagen matrix recently have been developed to help overcome alveolar bone deficiencies that develop in the wake of edentulism. CASE REPORT: The patient had one mandibular molar extracted and the socket grafted with equine bone granules. The graft was covered with the equine collagen matrix, placed in a double layer. No flap was prepared, and the gingival margins were stabilized with a single stitch, leaving the matrix partially exposed and the site to heal by secondary intention. The adjacent molar was extracted 1 month later, and that socket was left to heal by secondary intention without any further treatment. Three months after each surgery, an implant was placed and a biopsy was collected. The two biopsies underwent histological processing and qualitative evaluation. Histomorphometric analysis was also performed to calculate the percentage of newly formed bone (NFB) in the two cores. Healing at both sites was uneventful, and no inflammation or other adverse reactions were observed in the samples. Soft-tissue healing by secondary intention appeared to occur faster at the grafted site. The corresponding core showed a marked separation between soft and hard tissue that was not observed in the core from the nongrafted site, where soft-tissue hypertrophy could be observed. Newly formed bone at the grafted and nongrafted sites was not significantly different (27.2 ± 7.1 and 29.4 ± 6.2% respectively, p = 0.45). CONCLUSION: The surgical technique employed in this case appeared to facilitate postextraction soft-tissue healing by second intention and simplify soft-tissue management. CLINICAL SIGNIFICANCE: Using a collagen-based matrix to cover a postextraction grafted site may facilitate second intention soft-tissue healing and proper soft-tissue growth.

Horizontal-guided Bone Regeneration using a Titanium Mesh and an Equine Bone Graft.

AIM: The present work describes a horizontal ridge augmentation in which a titanium mesh was preshaped by adapting it to a stereolithographic model of the patient's jaw that was fabricated from CT scans. BACKGROUND: Guided bone regeneration (GBR) involves covering the augmentation site with a long-lasting barrier to protect it from the invasion of surrounding soft tissues. Among barriers, titanium meshes may provide a successful outcome, but the intraoperatory time needed to shape them is a disadvantage. CASE DESCRIPTION: The 54-year-old patient, missing the right mandibular second bicuspid, first molar, and second molar, had her atrophic ridge augmented with a 30:70 mixture of autogenous bone and equine, enzyme-deantigenic collagen-preserved bone substitute. Two conical implants were inserted concomitantly in the second bicuspid and first molar positions, and the site was protected with the preshaped mesh. Four months later, the titanium mesh was retrieved, a bone sample was collected, and histological and histomorphometric analyses were performed. Provisional and definitive prostheses were then delivered, and follow-up controls were performed for up to 24 months. CONCLUSION: Preshaping the mesh on a model of the patient's mandible shortened the surgical time and enabled faster mesh placement. Two years after surgery, the implants were perfectly functional, and the bone width was stable over time as shown by radiographic controls. Histological analysis of the bone sample showed the heterologous biomaterial to be biocompatible and undergoing advanced remodeling and replacement with newly formed bone. CLINICAL SIGNIFICANCE: Preshaping a titanium mesh over a stereolithographic model of the patient's jaw allowed for a significant reduction of the intraoperative time and may be therefore, advisable in routine practice.

G51D α-synuclein mutation causes a novel parkinsonian-pyramidal syndrome.

OBJECTIVE: To date, 3 rare missense mutations in the SNCA (α-synuclein) gene and the more frequent duplications or triplications of the wild-type gene are known to cause a broad array of clinical and pathological symptoms in familial Parkinson disease (PD). Here, we describe a French family with a parkinsonian-pyramidal syndrome harboring a novel heterozygous SNCA mutation. METHODS: Whole exome sequencing of DNA from 3 patients in a 3-generation pedigree was used to identify a new PD-associated mutation in SNCA. Clinical and pathological features of the patients were analyzed. The cytotoxic effects of the mutant and wild-type proteins were assessed by analytical ultracentrifugation, thioflavin T binding, transmission electron microscopy, cell viability assay, and caspase-3 activation. RESULTS: We identified a novel SNCA G51D (c.152 G>A) mutation that cosegregated with the disease and was absent from controls. G51D was associated with an unusual PD phenotype characterized by early disease onset, moderate response to levodopa, rapid progression leading to loss of autonomy and death within a few years, marked pyramidal signs including bilateral extensor plantar reflexes, occasionally spasticity, and frequently psychiatric symptoms. Pathological lesions predominated in the basal ganglia and the pyramidal tracts and included fine, diffuse cytoplasmic inclusions containing phospho-α-synuclein in superficial layers of the cerebral cortex, including the entorhinal cortex. Functional studies showed that G51D α-synuclein oligomerizes more slowly and its fibrils are more toxic than those of the wild-type protein. INTERPRETATION: We have identified a novel SNCA G51D mutation that causes a form of PD with unusual clinical, neuropathological, and biochemical features.

Novel emerging therapy for erectile dysfunction: efficacy and safety of flat magnetic stimulation.

BACKGROUND: The erectile dysfunction (ED), which is the inability to achieve and/or sustain a penile erection sufficient to result in a satisfying sexual performance, represents a very common complaint. for men over forty years old. The aim of the study was to evaluate if Flat Magnetic Stimulation (FMS) technology could help individuals with symptomatic erectile dysfunction. METHODS: Twenty patients with erectile dysfunction, underwent eight sessions of about 30 minutes each in a twice a week frequency with the study device. During treatments, every potential side effect was assessed. The International Index of Erectile Function (IIEF) was compiled by all patients at the beginning, after the eighth treatment and at 1 month from the end of the last treatment. The questionnaire scores were presented as median values along with the interquartile range (IQR) and we set the significance threshold at 0.01. RESULTS: After the treatment and at 1-month follow-up, the increase in questionnaire scores was statistically significant compared to the baseline, thus supporting the clinical usefulness of this treatment. In particular, the result of the study indicates a statistically significant difference between IIEF score before treatment (Median = 34) and IIEF score after the end of treatment (Median = 45) and between IIEF score before treatment and IIEF score at 1-month follow-up (Median = 54). CONCLUSIONS: The study findings showed that FMS represents a promising treatment option to individuals affected by symptomatic erectile dysfunction.

Genome sequence of PSonyx, a singleton bacteriophage infecting Corynebacterium glutamicum.

PSonyx is a newly isolated phage that infects Corynebacterium glutamicum. This siphovirus was isolated from a French pond in the south of Paris by students from Paris-Saclay University. Its 80,277-bp singleton genome carries 136 protein-coding genes and 5 tRNAs.

Dendritic cells with lymphocyte-stimulating activity differentiate from human CD133 positive precursors.

CD133 is a hallmark of primitive myeloid progenitors. We have addressed whether human cord blood cells selected for CD133 can generate dendritic cells, and Langerhans cells in particular, in conditions that promote that generation from CD34(+) progenitors. Transforming growth factor-ß1 (TGF-ß1) and anti-TGF-ß1 antibody, respectively, were added in some experiments. With TGF-ß, monocytoid cells were recognized after 7 days. Immunophenotypically immature dendritic cells were present at day 14. After 4 more days, the cells expressed CD54, CD80, CD83, and CD86 and were potent stimulators in mixed lymphocyte reaction; part of the cells expressed CD1a and langerin, but not Birbeck granules. Without TGF-ß, only a small fraction of cells acquired a dendritic shape and expressed the maturation-related antigens, and lymphocytes were poorly stimulated. With anti-TGF-ß, the cell growth was greatly hampered, CD54 and langerin were never expressed, and lymphocytes were stimulated weakly. In conclusion, CD133(+) progenitors can give rise in vitro, through definite steps, to mature, immunostimulatory dendritic cells with molecular features of Langerhans cells, although without Birbeck granules. Addition of TGF-ß1 helps to stimulate cell growth and promotes the acquisition of mature immunophenotypical and functional features. Neither langerin nor Birbeck granules proved indispensable for lymphocyte stimulation.