Immunomodulatory agents as potential therapeutic or preventive strategies for COVID-19.

Currently, the COVID-19 pandemic, caused by the novel SARS-CoV-2 coronavirus, represents the greatest global health threat. Most people infected by the virus present mild to moderate respiratory symptoms and recover with supportive treatments. However, certain susceptible hosts develop an acute respiratory distress syndrome (ARDS), associated with an inflammatory "cytokine storm", leading to lung damage. Despite the current availability of different COVID-19 vaccines, the new emerging SARS-CoV-2 genetic variants represent a major concern worldwide, due to their increased transmissibility and rapid spread. Indeed, it seems that some mutations or combinations of mutations might confer selective advantages to the virus, such as the ability to evade the host immune responses elicited by COVID-19 vaccines. Several therapeutic approaches have been investigated but, to date, a unique and fully effective therapeutic protocol has not yet been achieved. In addition, steroid-based therapies, aimed to reduce inflammation in patients with severe COVID-19 disease, may increase the risk of opportunistic infections, increasing the hospitalization time and mortality rate of these patients. Hence, there is an unmet need to develop more effective therapeutic options. Here, we discuss the potential use of natural immunomodulators such as Thymosin α1 (Tα1), all-trans retinoic acid (ATRA), and lactoferrin (LF), as adjunctive or preventive treatment of severe COVID-19 disease. These agents are considered to be multifunctional molecules because of their ability to enhance antiviral host immunity and restore the immune balance, depending on the host immune status. Furthermore, they are able to exert a broad-spectrum antimicrobial activity by means of direct interactions with cellular or molecular targets of pathogens or indirectly by increasing the host immune response. Thus, due to the aforementioned properties, these agents might have a great potential in a clinical setting, not only to counteract SARS-CoV-2 infection, but also to prevent opportunistic infections in critically ill COVID-19 patients.

Soft x-ray spectroscopies in liquids and at solid-liquid interface at BACH beamline at Elettra.

The beamline for advanced dichroism of the Istituto Officina dei Materiali-Consiglio Nazionale delle Ricerche, operating at the Elettra synchrotron in Trieste (Italy), works in the extreme ultraviolet-soft x-ray photon energy range with selectable light polarization, high energy resolution, brilliance, and time resolution. The beamline offers a multi-technique approach for the investigation of the electronic, chemical, structural, magnetic, and dynamical properties of materials. Recently, one of the three end stations has been dedicated to experiments based on electron transfer processes at the solid/liquid interfaces and during photocatalytic or electrochemical reactions. Suitable cells to perform soft x-ray spectroscopy in the presence of liquids and reagent gases at ambient pressure were developed. Here, we present two types of static cells working in transmission or in fluorescence yield and an electrochemical flow cell that allows us to carry out cyclic voltammetry in situ and electrodeposition on a working electrode and to study chemical reactions under operando conditions. Examples of x-ray absorption spectroscopy measurements performed under ambient conditions and during electrochemical experiments in liquids are presented.

Clinical value of anti-domain I-ß2Glycoprotein 1 antibodies in antiphospholipid antibody carriers. A single centre, prospective observational follow-up study.

BACKGROUND: There seems to be a clear correlation between antibodies against domain I (anti-DI) of ß2Glycoprotein I and severe clinical profiles in antiphospholipid syndrome (APS) patients. We investigated the clinical significance of anti-DI antibodies in a cohort of aPL carriers. METHODS: One hundred and five carriers persistently positive for IgG anti-ß2Glycoprotein 1 antibodies (a-ß2GPI) and/or IgG anticardiolipin (aCL) and/or lupus anticoagulants (LAC) were tested for the presence of anti-DI antibodies using the QUANTA Flash® Beta2GPI-Domain I chemiluminescence immunoassay. RESULTS: Anti-DI antibodies were detected in 44 aPL carriers (41.9%) and they were significantly associated to triple aPL positivity (LAC plus IgG a-ß2GPI plus IgG aCL antibodies). Isolated LAC and a-ß2GPI antibodies were significantly associated to anti-DI negative aPL carriers. During a 82.2 month mean follow-up, ten aPL carriers (9.5%) developed a first thrombotic event so becoming APS patients. Anti-DI antibodies, triple aPL positivity, thromboembolic risk factors and autoimmune disorders significantly prevailed in carriers becoming APS. Logistic regression analysis showed that anti-DI positivity was an independent risk factor for thrombosis. CONCLUSIONS: Anti-DI antibody positivity can be considered a new risk factor predictive of the first thrombotic event in aPL carriers, instead, negative anti-DI may be useful to identify low-risk aPL carriers.

Clinical predictors and microbiology of ventilator-associated pneumonia in the intensive care unit: a retrospective analysis in six Italian hospitals.

The purpose of this study was to assess the main clinical predictors and microbiological features of ventilator-associated pneumonia (VAP) in the Intensive Care Unit (ICU) environment. This work is a retrospective analysis over one year from September 2010 to September 2011. Patients' risk factors, causes of admission, comorbidities and respiratory specimens collected in six Italian ICUs were reviewed. Incidence and case fatality rate of VAP were evaluated. After stratification for VAP development, univariate and multivariate analyses were performed to assess the impact of patients' conditions on the onset of this infection. A total of 1,647 ICU patients (pts) were considered. Overall, 115 patients (6.9 %) experienced at least one episode of VAP. The incidence rate for VAP was 5.82/1,000 pts-days, with a case fatality rate of 44.3 %. Multivariate analysis showed that admission for neurological disorders (aIRR 4.12, CI 1.24-13.68, p = 0.02) and emergency referral to ICU from other hospitals (aIRR 2.11, CI 1.03-4.31, p = 0.04) were associated with higher risk of VAP, whereas a tendency to a higher risk of infection was detected for admission due to respiratory disease, cardiac disease, trauma and for having obesity or renal failure. A total of 372 microbiological isolates from respiratory specimens were collected in VAP patients. The most common species were Klebsiella pneumoniae, Acinetobacter baumannii and Pseudomonas aeruginosa, showing high resistance rates to carbapenems. Neurological disorders and emergency referral at the admission into the ICU are significantly associated with the onset of VAP. A high incidence of multi-drug resistant Gram- species was detected in the respiratory specimens.

The nature of the Fe-graphene interface at the nanometer level.

The emerging fields of graphene-based magnetic and spintronic devices require a deep understanding of the interface between graphene and ferromagnetic metals. This paper reports a detailed investigation at the nanometer level of the Fe-graphene interface carried out by angle-resolved photoemission, high-resolution photoemission from core levels, near edge X-ray absorption fine structure, scanning tunnelling microscopy and spin polarized density functional theory calculations. Quasi-free-standing graphene was grown on Pt(111), and the iron film was either deposited atop or intercalated beneath graphene. Calculations and experimental results show that iron strongly modifies the graphene band structure and lifts its π band spin degeneracy.

Optoelectrochemical biorecognition by optically transparent highly conductive graphene-modified fluorine-doped tin oxide substrates.

Both optical and electrochemical graphene-based sensors have gone through rapid development, reaching high sensitivity at low cost and with fast response time. However, the complex validating biochemical operations, needed for their consistent use, currently limits their effective application. We propose an integration strategy for optoelectrochemical detection that overcomes previous limitations of these sensors used separately. We develop an optoelectrochemical sensor for aptamer-mediated protein detection based on few-layer graphene immobilization on selectively modified fluorine-doped tin oxide (FTO) substrates. Our results show that the electrochemical properties of graphene-modified FTO samples are suitable for complex biological detection due to the stability and inertness of the engineered electrodic interface. In addition, few-layer immobilization of graphene sheets through electrostatic linkage with an electrochemically grafted FTO surface allows obtaining an optically accessible and highly conductive platform. As a proof of concept, we used insulin as the target molecule to reveal in solution. Because of its transparency and low sampling volume (a few microliters), our sensing unit can be easily integrated in lab-on-a-chip cell culture systems for effectively monitoring subnanomolar concentrations of proteins relevant for biomedical applications.

Synthesis of luminescent 3D microstructures formed by carbon quantum dots and their self-assembly properties.

We report in this communication the synthesis of star-shaped carbon quantum dots-(poly-γ-benzyl-L-glutamate) conjugates that self-assemble into microstructures and retain the characteristic emission properties of the native dots. Dots were used either as an initiator to give a daisy-like peptide-polymer structure or as capping agents towards more elaborated hybrid nanostructures.

Development of a non-viral gene delivery vector based on the dynein light chain Rp3 and the TAT peptide.

Gene therapy and DNA vaccination trials are limited by the lack of gene delivery vectors that combine efficiency and safety. Hence, the development of modular recombinant proteins able to mimic mechanisms used by viruses for intracellular trafficking and nuclear delivery is an important strategy. We designed a modular protein (named T-Rp3) composed of the recombinant human dynein light chain Rp3 fused to an N-terminal DNA-binding domain and a C-terminal membrane active peptide, TAT. The T-Rp3 protein was successfully expressed in Escherichia coli and interacted with the dynein intermediate chain in vitro. It was also proven to efficiently interact and condense plasmid DNA, forming a stable, small (∼100nm) and positively charged (+28.6mV) complex. Transfection of HeLa cells using T-Rp3 revealed that the vector is highly dependent on microtubule polarization, being 400 times more efficient than protamine, and only 13 times less efficient than Lipofectamine 2000™, but with a lower cytotoxicity. Confocal laser scanning microcopy studies revealed perinuclear accumulation of the vector, most likely as a result of transport via microtubules. This study contributes to the development of more efficient and less cytotoxic proteins for non-viral gene delivery.

Characterization of the human dynein light chain Rp3 and its use as a non-viral gene delivery vector.

Dynein light chains mediate the interaction between the cargo and the dynein motor complex during retrograde microtubule-mediated transport in eukaryotic cells. In this study, we expressed and characterized the recombinant human dynein light chain Rp3 and developed a modified variant harboring an N-terminal DNA-binding domain (Rp3-Db). Our approach aimed to explore the retrograde cell machinery based on dynein to enhance plasmid DNA (pDNA) traffic along the cytosol toward the nucleus. In the context of non-viral gene delivery, Rp3-Db is expected to simultaneously interact with DNA and dynein, thereby enabling a more rapid and efficient transport of the genetic material across the cytoplasm. We successfully purified recombinant Rp3 and obtained a low-resolution structural model using small-angle X-ray scattering. Additionally, we observed that Rp3 is a homodimer under reducing conditions and remains stable over a broad pH range. The ability of Rp3 to interact with the dynein intermediate chain in vitro was also observed, indicating that the recombinant Rp3 is correctly folded and functional. Finally, Rp3-Db was successfully expressed and purified and exhibited the ability to interact with pDNA and mediate the transfection of cultured HeLa cells. Rp3-Db was also capable of interacting in vitro with dynein intermediate chains, indicating that the addition of the N-terminal DNA-binding domain does not compromise its function. The transfection level observed for Rp3-Db is far superior than that reported for protamine and is comparable to that of the cationic lipid Lipofectamine™. This report presents an initial characterization of a non-viral delivery vector based on the dynein light chain Rp3 and demonstrates the potential use of modified human light chains as gene delivery vectors.

Electron backscatter diffraction in conservation science: phase identification of pigments in paint layers.

Electron backscatter diffraction (EBSD) was used in Conservation Science for characterization of ancient materials collected from works of art. The results demonstrate the feasibility of EBSD analysis on heterogeneous matrices as very small samples of paint layers collected from paintings. Two reference pigments were selected from those used by artists to investigate the relationship existing between EBSD pattern quality and properties of the investigated material (i.e., average atomic number, density, and Mohs hardness). The technique was also tested to investigate the pigment phases on two real samples collected from Romanino's Santa Giustina altarpiece, an oil on wood painting dated 1514 (Civic Museum, Padova, Italy). Results show for the first time the acquisition of EBSD patterns from painting samples mounted in resin, i.e., painting cross sections, opening a new powerful tool to elucidate the pigment phases avoiding large sampling on works of arts and to further study the complex mechanisms of pigment deterioration.

Low occurrence of digital ulcers in scleroderma patients treated with bosentan for pulmonary arterial hypertension: a retrospective case-control study.

Digital ulcers (DU) are one of the most common and debilitating manifestations of vasculopathy in systemic sclerosis (SSc). Their prevention is important in order to improve patients' outcome and as a result of the economic impact they have on society. Randomised controlled studies have demonstrated that bosentan, an endothelin receptor antagonist, reduces the appearance of new DU. The aim of this retrospective study was to evaluate the occurrence of DU in a group of patients receiving long-term bosentan treatment for pulmonary arterial hypertension associated with SSc (PAH-SSc). Patients with PAH-SSc and treated with bosentan for at least 6 months (n = 30) were evaluated. Thirty patients with SSc not treated with bosentan, but matched for sex, age, disease duration and cutaneous form of SSc, were considered as a control group. The occurrence of DU, defined as loss of tissue of varying degrees in the epidermis, dermis and subcutaneous tissue, was determined in the bosentan-treated and untreated groups. Mean duration of bosentan treatment was 3.6 years. DU were detected in six patients in the bosentan-treated group (20.0 %) and 16 patients (53.3 %) in the untreated group (p = 0.0015). There were no significant differences in demographic or clinical characteristics between patients with or without DU at study end. The occurrence of DU in patients with PAH-SSc receiving long-term bosentan treatment was significantly lower than in untreated patients. The results from this long-term observational study provide valuable information on management of patients with PAH-SSc.

Small-angle X-ray scattering and in silico modeling approaches for the accurate functional annotation of an LysR-type transcriptional regulator.

Xylella fastidiosa is a xylem-limited, Gram-negative phytopathogen responsible for economically relevant crop diseases. Its genome was thus sequenced in an effort to characterize and understand its metabolism and pathogenic mechanisms. However, the assignment of the proper functions to the identified open reading frames (ORFs) of this pathogen was impaired due to a lack of sequence similarity in the databases. In the present work, we used small-angle X-ray scattering and in silico modeling approaches to characterize and assign a function to a predicted LysR-type transcriptional regulator in the X. fastidiosa (XfLysRL) genome. XfLysRL was predicted to be a homologue of BenM, which is a transcriptional regulator involved in the degradation pathway of aromatic compounds. Further functional assays confirmed the structural prediction because we observed that XfLysRL interacts with benzoate and cis,cis-muconic acid (also known as 2E,4E-hexa-2,4-dienedioic acid; hereafter named muconate), both of which are co-factors of BenM. In addition, we showed that the XfLysRL protein is differentially expressed during the different stages of X. fastidiosa biofilm formation and planktonic cell growth, which indicates that its expression responds to a cellular signal that is likely related to the aromatic compound degradation pathway. The assignment of the proper function to a protein is a key step toward understanding the cellular metabolic pathways and pathogenic mechanisms. In the context of X. fastidiosa, the characterization of the predicted ORFs may lead to a better understanding of the cellular pathways that are linked to its bacterial pathogenicity.

(1-3)-ß-D-Glucan vs Galactomannan Antigen in Diagnosing Invasive Fungal Infections (IFIs).

Invasive fungal infections (IFIs) are serious and often life-threatening complications in patients with haematological malignancies. Early diagnosis and the initiation of efficacious antifungal treatments could affect the prognosis of these patients. The detection of (1-3)-ß-D-Glucan (BDG) could be a promising non-culture-based, noninvasive tool for IFI analyses in haemato-oncological patients, allowing the diagnosis of the two major IFIs, invasive aspergillosis (IA) and invasive candidiasis (IC), with a single test. The aim of this work was to evaluate and compare the use of the BDG in combination with the galactomannan antigen (GAL) assay in order to exclude or confirm suspected IFIs. Sera from 46 haemato-oncological patients (24 with proven/probable IFI and 22 without IFI symptoms) were evaluated retrospectively for the detection of GAL and BDG. In 24 patients, the serum BDG levels facilitated IFI diagnosis: 18 probable IA, 3 proven IA and 3 IC. In the remaining 22 patients, the BDG level helped exclude IFIs. The BDG was positive earlier than GAL in 5/24 cases [three of probable invasive aspergillosis (IA), one of proven IA and one case of proven invasive candidiasis (IC)] and was positive at the same time as GAL in 19/24 cases; in no case was GAL positive before BDG was. The BDG detection is useful, however, the test has a great limitation because it is a completely manual procedure.

Improved diagnosis of central venous catheter-related bloodstream infections using the HB&L UROQUATTRO™ system.

The diagnosis of catheter-related bloodstream infections (CRBSIs) in febrile patients with indwelling central venous catheters (CVCs) needs improvement. To diagnose CRBSIs more efficiently, we have developed a novel culture approach using the catheter tips removed from febrile patients. CVCs and blood cultures from 1,070 patients with only CVC-related infections were obtained over a period of 3 years (January 2009 to December 2011). The CVCs were evaluated by a semi-quantitative catheter culture method according to Maki's method and by our novel method, which is based on the use of the HB&L UROQUATTRO™ system (Alifax, Padova, Italy). Using our new method, 571 (571/1,070) of the infections were confirmed as CRBSIs. The remaining 487 patients had infections that were associated with hematologic malignancies, neutropenia, prior exposure to antibiotics, and a decreased CVC removal rate. Twelve samples were identified as false-positives. The percentage of patients with CRBSIs confirmed using the HB&L UROQUATTRO™ system was 53.36 % versus 34.95 % (p-value 0.004) using Maki's method (374/1,070 CVC Maki-positive samples). Our results indicate that our new culture method allows for an improved CRBSI diagnosis rate. A significant number of tip cultures (18.41 %) tested positive for CRBSIs using our system but were negative when tested using Maki's method. Moreover, the use of the HB&L UROQUATTRO™ system allowed us to significantly reduce diagnosis time; a negative CRBSI diagnosis could be made within 6 h and a positive diagnosis could be made within 22-28 h.

Risk-based secondary prevention of obstetric antiphospholipid syndrome.

Treatment of pregnant women with antiphospholipid syndrome (APS) should be set apart from that from thrombotic APS patients. Patients with a history of pregnancy morbidity but no vascular thrombosis are usually treated with a prophylactic dose of heparin plus low-dose aspirin; whereas, those with previous vascular thrombosis alone or associated with previous pregnancy morbidity, are commonly treated with a therapeutic dose of heparin generally combined with low-dose aspirin. However, in about 20% of pregnant APS women these regimens fail. In this context, we conducted a case-control study on a large multicentre cohort of conventionally treated pregnancies to verify whether specific laboratory profiles and/or clinical characteristics are predictive of unsuccessful pregnancy outcome during conventional treatments. Multivariate analysis showed that pregnancy failure during conventional therapies was independently associated with a history of both thrombosis and pregnancy morbidity, the presence of systemic lupus erythematosus (SLE) or other systemic autoimmune diseases and triple antiphospholipid antibody positivity. With the aim to discover the most effective and safe treatments in high-risk pregnant APS women a large-scale multicentre study focusing on the effect of treatments on pregnancy outcome in women with APS and further risk factors for pregnancy failure has been designed.

Characterization of lapis lazuli and corresponding purified pigments for a provenance study of ultramarine pigments used in works of art.

In this paper, we propose an analytical methodology for attributing provenance to natural lapis lazuli pigments employed in works of art, and for distinguishing whether they are of natural or synthetic origin. A multitechnique characterization of lazurite and accessory phases in lapis lazuli stones from Afghan, Siberian and Chilean quarries, on the pigments obtained by their purification, and on synthetic ultramarine pigments was performed. According to the results obtained, infrared spectroscopy is not a suitable technique for distinguishing the provenance of lapis lazuli, but a particular absorbance band makes it relatively easy to determine whether it is of natural or synthetic origin. On the other hand, EDS elemental composition and XRD patterns show the presence of specific mineral phases associated with specific lapis lazuli sources, and can be used to distinguish the provenance of the stones as well as-albeit to a lesser extent-the corresponding purified blue pigments. In contrast, FEG-SEM observations clearly show different stone textures depending on their provenance, although these distinctive features do not persist in the corresponding pigments. PCA analyses of EDS data allow Afghan lapis lazuli stone to be distinguished from Chilean and Siberian ones, and can distinguish between the pigments resulting from their purification as well as synthetic blue ones. Although this methodology was developed using a limited number of samples, it was tested on lapis lazuli pigments collected from three paintings (from the fourteenth to sixteenth centuries) in order to perform a preliminary validation of the technique, and based on the results, the provenance of the blue pigments employed in those artworks is proposed. Finally, upon analytically monitoring the process of purifying lapis lazuli to obtain the corresponding pigments, it was found that ion-exchange reactions occur between the alkali modifiers of silicate/aluminosilicate phases and free carboxylic acids present in the doughy mixture of natural terpenes and ground stone, namely pastello. These reactions favor (i) the retention of silicate phases in the organic mixture and (ii) the selective extraction of lazurite due to the formation of Brønsted acidic sites [Al(OH)Si], which are responsible for its high hydrophilicity in comparison to the one of the other species present in the lapis lazuli stone.

Superhydrophilic and tribological improvements of polymeric surfaces via plasma enhanced chemical vapor deposition ceramic coatings.

The main object of this study is the treatment of polymeric (PVC, PC) surfaces with the aim of inducing enhanced superhydrophilic characteristics together with nanohardness features; this would allow polymeric surfaces to have longer durability and prevent the accumulation of dirt on the surface which could disable the proper use of these polymeric surfaces. Indeed plastic surfaces are difficult substrates to be covered effectively and functionalized, mainly due to their high sensitivity to heat treatments and irradiation in the UV-Vis range together with their inert behavior. Their functionalization is achieved through the deposition of ceramic coatings such as titania (TiO2), on the polymeric surfaces via PECVD (Plasma Enhanced Chemical Vapor Deposition) at low temperatures. Characterizations are carried out by contact angle analysis for the superhydrophilic characteristics, and by nanoindentation analysis for the tribological features. A cold PECVD discontinuous method allowed us to improve nanohardness, reaching a value of 1.39 GPa which is nearly ten times higher than that of the uncoated polymeric substrate, and seems a promising solution for improving uniformity of the coatings. Superhydrophilic behavior of the activated TiO2 surfaces showed contact angle values lower than 10 degrees.

Adjusted prophylactic doses of nadroparin plus low dose aspirin therapy in obstetric antiphospholipid syndrome. A prospective cohort management study.

OBJECTIVES: Current guidelines for the treatment of patients with obstetric antiphospholipid syndrome (APS) recommend low dose aspirin (LDA) and prophylactic doses of low molecular weight heparin (LMWH). Most clinicians use a fixed dosage of LMWH in pregnant APS women despite the fact that there are no clinical trials establishing that fixed doses are more efficacious than adjusted ones in preventing pregnancy complications. The efficacy and safety of adjusted single daily doses of LMWH (nadroparin) combined with LDA have thus been evaluated in 33 consecutive pregnancies in women with diagnosed obstetric APS. METHODS: LMWH doses were augmented as the pregnancies progressed and maternal/foetal weight increased. 70-80-90 U/Kg doses ranging between 3800 and 6650 U were administered daily during the first, second and third trimesters, respectively. LDA (100 mg/day) was also prescribed. RESULTS: Pregnancy outcome was successful in 97% of the patients studied, who delivered, between the 29th and 41st weeks of gestation (mean 37.4 ±2.1 SD), 32 infants with a mean birth weight of 3084 g ± 514 SD. One woman (3%) experienced a spontaneous abortion at the 8th week of gestation. CONCLUSIONS: The high live birth rate, the satisfactory mean gestational age and weight at birth and the absence of major pregnancy/neonatal-associated complications indicate that adjusted, once daily doses of LMWH together with LDA could be an efficacious treatment option for pregnant APS patients with no history of thrombosis.

Characterization of an oxidative stress response regulator, homologous to Escherichia coli OxyR, from the phytopathogen Xylella fastidiosa.

The OxyR oxidative stress transcriptional regulator is a DNA-binding protein that belongs to the LysR-type transcriptional regulators (LTTR) family. It has the ability to sense oxidative species inside the cell and to trigger the cell's response, activating the transcription of genes involved in scavenging oxidative species. In the present study, we have overexpressed, purified and characterized the predicted OxyR homologue (orf xf1273) of the phytopathogen Xylella fastidiosa. This bacterium is the causal agent of citrus variegated chlorosis (CVC) disease caused by the 9a5c strain, resulting in economic and social losses. The secondary structure of the recombinant protein was analyzed by circular dichroism. Gel filtration showed that XfoxyR is a dimer in solution. Gel shift assays indicated that it does bind to its own predicted promoter under in vitro conditions. However, considering our control experiment we cannot state that this interaction occurs in vivo. Functional complementation assays indicated that xfoxyR is able to restore the oxidative stress response in an oxyr knockout Escherichia coli strain. These results show that the predicted orfxf1273 codes for a transcriptional regulator, homologous to E. coli OxyR, involved in the oxidative stress response. This may be important for X. fastidiosa to overcome the defense mechanisms of its host during the infection and colonization processes.