Characterization of a fully active N-terminal 37-kDa polypeptide obtained by limited tryptic cleavage of pig kidney D-amino acid oxidase.

In order to obtain further information on the structure of D-amino acid oxidase (EC 1.4.3.3), limited proteolysis experiments have been carried out on its apo-, holo-, and holoenzyme-benzoate forms. The enzyme is unsensitive to 10% (w/w) chymotrypsin, while incubation with 10% (w/w) trypsin, under nondenaturating conditions, produces inactivation and proteolysis patterns which are different for the three forms of enzyme analyzed. These results confirm the previously reported conformational changes which occur upon binding of coenzyme to the apoprotein, and of benzoate to holoenzyme. The stable 37.0-kDa polypeptide, obtained from the apo- and holoenzyme-benzoate complex upon cleavage of a C-terminal 2.0-kDa fragment, retains full catalytic activity with unaltered kinetic parameters, and the coenzyme binding properties of the native enzyme. These results are in agreement with the tentative localization of the FAD-binding domain in the N-terminal region of the enzyme, and with the hypothesis that the function of the C-terminal region of D-amino acid oxidase could be related to the import of the enzyme into the peroxisomes, as suggested by Gould et al. (Gould, S. J., Keller, G. A., and Subramani, S. (1988) J. Cell. Biol. 107, 897-905)

Single-Walled Carbon Nanotubes as Enhancing Substrates for PNA-Based Amperometric Genosensors

A new amperometric sandwich-format genosensor has been implemented on single-walled carbon nanotubes screen printed electrodes (SWCNT-SPEs) and compared in terms of performance with analogous genoassays developed using the same methodology on non-nanostructured glassy carbon platforms (GC-SPE). The working principle of the genosensors is based on the covalent immobilization of Peptide Nucleic Acid (PNA) capture probes (CP) on the electrode surface, carried out through the carboxylic functions present on SWCNT-SPEs (carboxylated SWCNT) or electrochemically induced on GC-SPEs. The sequence of the CP was complementary to a 20-mer portion of the target DNA; a second biotin-tagged PNA signalling probe (SP), with sequence complementary to a different contiguous portion of the target DNA, was used to obtain a sandwich hybrid with an Alkaline Phosphatase-streptavidin conjugate (ALP-Strp). Comparison of the responses obtained from the SWCNT-SPEs with those produced from the non-nanostructured substrates evidenced the remarkable enhancement effect given by the nanostructured electrode platforms, achieved both in terms of loading capability of PNA probes and amplification of the electron transfer phenomena exploited for the signal transduction, giving rise to more than four-fold higher sensitivity when using SWCNT-SPEs. The nanostructured substrate allowed to reach limit of detection (LOD) of 71 pM and limit of quantitation (LOQ) of 256 pM, while the corresponding values obtained with GC-SPEs were 430 pM and 1.43 nM, respectively

A Folding-Based Electrochemical Aptasensor for the Single-Step Detection of the SARS-CoV-2 Spike Protein

Efficient and timely testing has taken center stage in the management, control, and monitoring of the current COVID-19 pandemic. Simple, rapid, cost-effective diagnostics are needed that can complement current polymerase chain reaction-based methods and lateral flow immunoassays. Here, we report the development of an electrochemical sensing platform based on single-walled carbon nanotube screen-printed electrodes (SWCNT-SPEs) functionalized with a redox-tagged DNA aptamer that specifically binds to the receptor binding domain of the SARS-CoV-2 spike protein S1 subunit. Single-step, reagentless detection of the S1 protein is achieved through a binding-induced, concentration-dependent folding of the DNA aptamer that reduces the efficiency of the electron transfer process between the redox tag and the electrode surface and causes a suppression of the resulting amperometric signal. This aptasensor is specific for the target S1 protein with a dissociation constant (K-D) value of 43 +/- 4 nM and a limit of detection of 7 nM. We demonstrate that the target S1 protein can be detected both in a buffer solution and in an artificial viral transport medium widely used for the collection of nasopharyngeal swabs, and that no cross-reactivity is observed in the presence of different, non-target viral proteins. We expect that this SWCNT-SPE-based format of electrochemical aptasensor will prove useful for the detection of other protein targets for which nucleic acid aptamer ligands are made available

Unlocking Access to Enantiopure Fused Uracils by Chemodivergent [4+2] Cross‐Cycloadditions: DFT‐Supported Homo‐Synergistic Organocatalytic Approach

The discovery of chemical methods enabling the construction of carbocycle‐fused uracils which embody a three‐dimensional and functional‐group‐rich architecture is a useful tool in medicinal chemistry oriented synthesis. In this work, an unprecedented amine‐catalyzed [4+2] cross‐cycloaddition is documented; it involves remotely enolizable 6‐methyluracil‐5‐carbaldehydes and β‐aryl enals, and chemoselectively produces two novel bicyclic and tricyclic fused uracil chemotypes in good yields with a maximum level of enantiocontrol. In‐depth mechanistic investigations and control experiments support an intriguing homo‐synergistic organocatalytic approach, where the same amine organocatalyst concomitantly engages both aldehyde partners in a stepwise eliminative [4+2] cycloaddition, whose vinylogous iminium ion intermediate product may diverge—depending upon conditions—to either bicyclic targets by hydrolysis or tricyclic products by a second homo‐synergistic trienamine‐mediated stepwise [4+2] cycloaddition

Generalized cutting force model for peripheral milling of wood, based on the effect of density, uncut chip cross section, grain orientation and tool helix angle

Article publié en open-access sous accord avec CRUI-CARE (Université de Florence)The influence of the grain angle on the cutting force when milling wood is not yet detailed, apart from particular cases (end-grain, parallel to the grain, or in some rare cases 45°-cut). Thus, setting-up wood machining operations with complex paths still relies mainly on the experience of the operators because of the lack of scientific knowledge easily transferable to the industry. The aim of the present work is to propose an empirical model based on specific cutting coefficients for the assessment of cutting force when peripheral milling of wood based on the following input: uncut chip thickness and width, grain angle (angle between the tool velocity vector and the grain direction of the wood), density and tool helix angle. The specific cutting coefficients were determined by peripheral milling with different depths of cut wood disks issued from different wood species on a dynamometric platform to record the forces. Milling a sample into a round shape (a disk) allows to measure the cutting forces toward every grain angle into a sole basic diameter reduction operation. Force signals are then post-processed to carefully clean the natural vibrations of the system without impacting their magnitudes. The experiment is repeated on five species with a large range of densities, machining two disks per species for five depths of cut in up- and down milling conditions for three different tool helix angles. Finally, a simple cutting force model, based on the previously cited parameters, is proposed, and its robustness analysed.- This work was made possible thanks to the funding of the ANISOTROPEE project from the University of Florence (IT) and the support of the Région Bourgogne Franche-Comté (FR). Authors also want to acknowledge Gianluca Fantacci (G3 Fantacci Corp) for designing and providing the three cutting tools used during the core experimental campaign of this study. - Giacomo Goli would like to acknowledge the Art et Metiers Institute of Technology (ENSAM, Cluny campus, France) for financing 2 months in the laboratory LaBoMaP as invited researcher. - Rémi Curti would like to acknowledge the European COST Action FP1407 for funding a Short Term Scientific Mission of 1 week at DAGRI during his Ph.D to work with Giacomo Goli

Strontium doping in mullite-type bismuth aluminate: A vacancy investigation using neutrons, photons and electrons

We report on strontium doped dibismuth-nonaoxoaluminate(III) produced at 1023 K. Partial substitution of bismuth by strontium in the structure yields oxygen vacancies for charge balance. Introducing oxygen vacancies rearranged the associated Al2O7 double-tetrahedra forming “Al3O10” tri-clusters which were identified by multi-quantum 27Al MAS NMR. Both STEM-EDX and XPS showed homogeneous distribution of strontium in the bulk and on the surface, respectively. Moreover, XPS confirms the valence state of bismuth after doping. The orientations of bismuth 6s2 lone electron pairs were calculated using DFT methods. The amount of strontium in the crystal structure was further confirmed from the decomposition product SrAl12O19 formed during the temperature-dependent X-ray powder diffraction. The structural proof was carried out by refining the structure of (Bi0.94Sr0.06)2Al4O8.94 from powder neutron and X-ray diffraction data. Rietveld refinements clearly showed the under occupation of one oxygen site and the shift of two aluminum atoms from the double-tetrahedra to two tri-cluster sites

Prevalence and follow-up of occult HCV infection in an Italian population free of clinically detectable infectious liver disease.

BACKGROUND: Occult hepatitis C virus infection (OCI) is a recently described phenomenon characterized by undetectable levels of HCV-RNA in serum/plasma by current laboratory assays, with identifiable levels in peripheral blood mononuclear cells (PBMCs) and/or liver tissue by molecular tests with enhanced sensitivity. Previous results from our group showed an OCI prevalence of 3.3% in a population unselected for hepatic disease. The present study aimed to evaluate OCI prevalence in a larger cohort of infectious liver disease-free (ILDF) subjects. Clinical follow-up of OCI subjects was performed to investigate the natural history of the infection. METHODS AND FINDINGS: 439 subjects referred to a Turin Blood Bank for phlebotomy therapy were recruited. They included 314 ILDF subjects, 40 HCV-positive subjects and 85 HBV-positive subjects, of whom 7 were active HBV carriers. Six subjects (4/314 ILDF subjects [1.27%] and 2/7 active HBV carriers [28%]) were positive for HCV-RNA in PBMCs, but negative for serological and virological markers of HCV, indicating OCI. HCV genotypes were determined in the PBMCs of 3/6 OCI subjects two had type 1b; the other had type 2a/2c. OCI subjects were followed up for at least 2 years. After 12 months only one OCI persisted, showing a low HCV viral load (3.73×10(1) UI/ml). By the end of follow-up all OCI subjects were negative for HCV. No seroconversion, alteration of liver enzyme levels, or reduction of liver synthesis occurred during follow-up. CONCLUSIONS: This study demonstrated the existence of OCI in ILDF subjects, and suggested a high OCI prevalence among active HBV carriers. Follow-up suggested that OCI could be transient, with a trend toward the decrease of HCV viral load to levels undetectable by conventional methods after 12-18 months. Confirmation studies with a longer follow-up period are needed for identification of the OCI clearance or recurrence rates, and to characterize the viruses involved

Capitolo 2. La radiazione solare e il bilancio energetico di un edificio Capitolo 4. La valutazione delle prestazioni energetiche degli schermi solari in relazione all'edificio Appendice A. Determinazione delle componenti dell'irradiazione solare (CD) Appendice D. Quadro legislativo e normativo (CD)

Negli ultimi decenni i paesi industrializzati hanno maturato la consapevolezza che il modello di sviluppo attuale non è più sostenibile e che occorre controllare gli effetti dell’azione antropica sul pianeta. Dal Protocollo di Kyoto nel 1997 a oggi, si sono susseguite iniziative per ridurre le emissioni inquinanti, per mezzo di politiche rivolte alla promozione dell’efficienza energetica, allo sviluppo di fonti di energia rinnovabili, allo sviluppo di agricoltura sostenibile, nella ricerca di un maggiore equilibrio con l’ambiente. Tra tutte le azioni avviate, l’efficienza energetica delle costruzioni è sicuramente tra quelle più realizzabili: le tecnologie disponibili consentono ai progettisti di raggiungere risultati di risparmio energetico con investimenti contenuti, a condizione che la progettazione consideri tali aspetti fin dalle prime fasi, per le nuove costruzioni, o provveda all’adeguamento di quelle esistenti. Un gruppo di docenti e professionisti ha raccolto e riportato le proprie esperienze per dare strumenti di analisi e valutazione nella progettazione di nuovi elementi costruttivi. In un panorama di prodotti sempre più specializzati, le scelte richiedono motivazioni. Soluzioni comprovate che sembravano superate, dallo stimolo modernista e non considerate da quello postmoderno, ritornano attuali con la responsabilità rispetto ai nuovi obiettivi della contemporaneità. Si presenta un insieme di contributi che invitano a riprendere la progettazione degli schermi per il ruolo che possono avere nell’involucro dell’edificio (chiusura) rispetto alle nuove necessità ambientali. Una pratica poco seguita a giudicare dalla propensione per la preferenza delle grandi superfici a tutto vetro, per la chiusura, indipendentemente dal contesto climatico delle costruzioni. Dall’analisi, del primo capitolo, di alcune esperienze storiche sugli schermi e delle loro trasformazioni per raggiungere un punto di vista dell’insieme e inquadrare l’ambito di studio si passa, con il secondo e il terzo, a contributi specifici che forniscono strumenti conoscitivi e applicativi per controllare i principali fenomeni fisici che devono essere considerati nella progettazione. Con il quarto capitolo sono dati strumenti per la valutazione delle prestazioni degli schermi e quindi criteri per la scelta di prodotti. Segue un’ampia rassegna di schede, strutturate secondo un’impostazione linguistica prestazionale, che riporta il panorama delle soluzioni più diffuse. Le schede evidenziano requisiti ed esperienze progettuali implicite alle diverse articolazioni e possono essere per il progettista una memoria di argomenti da non dimenticare. Il panorama delle soluzioni è completato con un contributo specifico relativo alle schermature vegetali di cui si evidenzia il possibile contributo prestazionale oltre l’implicita visibilità paesaggistica. A conclusione due utili appendici di carattere applicativo ed esemplificativo con i programmi più diffusi e utilizzati. L’insieme dei contributi nelle intenzioni degli autori intende supportare la progettazione sia delle costruzioni edilizie sia dei prodotti per le costruzioni: conseguente l’intenzione di evidenziare le possibilità di motivate innovazioni che il settore può offrire. Un invito a riprendere in considerazione la progettazione in senso ampio, olistico, per la novità delle condizioni nelle quali ci si ritrova a operare: per affrontare progetti di nuovi elementi e non solo l’utilizzo di quelli esistenti. Il lavoro è stato impostato con la raccolta e successiva analisi di molti casi per interpretare le esperienze del passato e non trascurare esperienze rispetto ai nuovi obiettivi. Il controllo della radiazione solare attraverso l’utilizzo delle schermature rientra tra le azioni di risparmio energetico che consente di migliorare le condizioni degli spazi interni senza impiego di energia. A interpretazione della proposta di questo libro emergono due motivazioni nuove: 1. la consapevolezza della necessità del contenimento dei consumi, supportata da leggi, richiede una progettazione più responsabile e controllata. Non ci sono soluzioni disponibili e comprovate, si apre un periodo di studio e di sperimentazione; 2. si viene da lunghi periodi di costruzioni con spreco del territorio. Le costruzioni non utilizzate non sono poche, dovrebbe avviarsi un’inversione di tendenza: le attività edilizie dovrebbero essere indirizzate all’adeguamento dell’esistente, che rappresenta un patrimonio anche culturale da valorizzare, piuttosto che su nuove costruzioni. La proposta di un libro sugli schermi porta, in modo implicito, una proposta di contenimento dei costi per la climatizzazione estiva e il controllo delle dispersioni per quella invernale. Gli schermi dovrebbero avere il ruolo, oltre a quello più consueto del controllo della luce, anche della riduzione del carico solare che incide direttamente sull’involucro dell’edificio e, di conseguenza, un miglioramento delle condizioni microclimatiche degli ambienti interni senza impiego di energia. Il raggiungimento delle condizioni di comfort negli ambienti interni con un’ottimizzazione delle prestazioni dell’involucro dell’edificio può ridurre in modo significativo il contributo impiantistico, ed in alcuni casi addirittura escluderne la climatizzazione, contenendo le emissioni di inquinanti e i costi di gestione. La progettazione dell’adeguamento dell’esistente può rappresentare l’opportunità di nuove prospettive professionali, impegnative: sia per la dimensione del grande patrimonio edilizio del paese, sia per le difficoltà oggettive degli interventi. L’approccio non può essere solo di adeguamento normativo: l’esistente è espressione di tante situazioni ambientali e culturali e l’adeguamento deve avvenire all’interno di un programma di valorizzazione complessiva

Clinical effectiveness of different natalizumab interval dosing schedules in a large Italian population of patients with multiple sclerosis

Introduction Natalizumab (NTZ) is one of the most effective treatment options for multiple sclerosis (MS) treatment. Our study aimed to evaluate the effectiveness of NTZ when administered according to the extended dosing strategy compared with standard 4-weekly administration in a large Italian MS population. Materials and methods This retrospective multicentre study included patients with relapsing-remitting MS (RR-MS) who received NTZ administrations between the 1 June 2012 and the 15 May 2018 and were followed by the ’Italian MS Register’. All patients with MS were stratified into two groups based on NTZ administration schedule: standard interval dosing (SID) patients who received infusions on average from 28 to 32 days (median 30) and extended interval dosing (EID) including patients who have been infused with interval between 33 and 49 days (median 43). Clinical data were assessed at baseline (before starting NTZ), after 12 (T1) and 24 months (T2) of treatment. Results Out of 5231 patients with RR-MS screened, 2092 (mean age 43.2±12.0, 60.6% women) were enrolled. A total of 1254 (59.9%) received NTZ according to SID, and 838 (40.1%) according to EID. At 12 and 24 months, no differences in terms of annualised relapse rate and disability status were found between the two groups. Progression index and confirmed disability worsening were similar between the two groups. Discussion The use of NTZ with an extended interval schedule showed similar effectiveness compared with SID. Unchanged clinical efficacy of EID schedule may raise the question of a possible advantage in terms of tolerability and safety

Incidence rates of in-hospital carpal tunnel syndrome in the general population and possible associations with marital status

<p>Abstract</p> <p>Background</p> <p>Carpal tunnel syndrome (CTS) is a socially relevant condition associated with biomechanical risk factors. We evaluated age-sex-specific incidence rates of in-hospital cases of CTS in central/northern Italy and explored relations with marital status.</p> <p>Methods</p> <p>Seven regions were considered (overall population, 14.9 million) over 3–6-year periods between 1997 and 2002 (when out-of-hospital CTS surgery was extremely rare). Incidence rates of in-hospital cases of CTS were estimated based on 1) codified demographic, diagnostic and intervention data in obligatory discharge records from all Italian public/private hospitals, archived (according to residence) on regional databases; 2) demographic general population data for each region. We compared (using the χ_scoretest) age-sex-specific rates between married, unmarried, divorced and widowed subsets of the general population. We calculated standardized incidence ratios (SIRs) for married/unmarried men and women.</p> <p>Results</p> <p>Age-standardized incidence rates (per 100,000 person-years) of in-hospital cases of CTS were 166 in women and 44 in men (106 overall). Married subjects of both sexes showed higher age-specific rates with respect to unmarried men/women. SIRs were calculated comparing married vs unmarried rates of both sexes: 1.59 (95% confidence interval [95% CI], 1.57–1.60) in women, and 1.42 (95% CI, 1.40–1.45) in men. As compared with married women/men, widows/widowers both showed 2–3-fold higher incidence peaks during the fourth decade of life (beyond 50 years of age, widowed subjects showed similar trends to unmarried counterparts).</p> <p>Conclusion</p> <p>This large population-based study illustrates distinct age-related trends in men and women, and also raises the question whether marital status could be associated with CTS in the general population.</p