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Background: Recent findings enlightened the pivotal role of cyclooxygenases-1 and -2 (COX-1 and COX-2) in human diseases with inflammation as the committed earliest stage, such as cancer and neurodegenerative diseases. COXs are the main targets of nonsteroidal anti-inflammatory drugs and catalyze the bis-oxygenation of arachidonic acid into prostaglandin PGH2, then converted into prostaglandins, thromboxane, and prostacyclin by tissue-specific isomerases. A remarkable amount of pure COX-1 is necessary to investigate COX-1 structure and function, as well as for in vitro disease biochemical pathway investigations. Methods: Spodoptera frugiperda cells were infected with Baculovirus that revealed to be an efficient expression system to obtain a high amount of ovine(o)COX-1. Protein solubilization time in the presence of a non-ionic detergent was modified, and a second purification step was introduced. Results and Discussion: An improvement of a previously reported method for pure recombinant oCOX-1 production and isolation has been achieved, leading to a lower starting volume of infected cells for each purification, an increased cell density, an increased number of viral particles per cell, and a shortened infection period. The protocol for the recombinant oCOX-1 expression and purification has been in-depth elaborated to obtain 1 mg/L of protein. Conclusion: The optimized procedure could be suitable for producing other membrane proteins as well, for which an improvement in the solubilization step is necessary to have the availability of high concentration proteins.

SARS-CoV-2 replication requires the synthesis of a set of structural proteins expressed through discontinuous transcription of ten subgenomic mRNAs (sgmRNAs). Here, we have fine-tuned droplet digital PCR (ddPCR) assays to accurately detect and quantify SARS-CoV-2 genomic ORF1ab and sgmRNAs for the nucleocapsid (N) and spike (S) proteins. We analyzed 166 RNA samples from anonymized SARS-CoV-2 positive subjects and we observed a recurrent and characteristic pattern of sgmRNAs expression in relation to the total viral RNA content. Additionally, expression profiles of sgmRNAs, as determined by meta-transcriptomics sequencing of a subset of 110 RNA samples, were highly correlated with those obtained by ddPCR. By providing a comprehensive and dynamic snapshot of the levels of SARS-CoV-2 sgmRNAs in infected individuals, our results may contribute a better understanding of the dynamics of transcription and expression of the genome of SARS-CoV-2 and facilitate the development of more accurate molecular diagnostic tools for the stratification of COVID-19 patients., Oranger et al use digital droplet PCR to develop an assay that provides a dynamic snapshot of the levels of SARS-CoV-2 sub-genomic messanger RNA in infected individuals. This has the potential to improve diagnostic accuracy for COVID-19

Background Choroid plexuses (CPs) have been suggested as a key gateway for inflammation in experimental autoimmune encephalitis, but in vivo evidence of their involvement in multiple sclerosis (MS) is lacking. Purpose To assess CP volumetric and inflammatory changes in patients with MS versus healthy control participants. Materials and Methods This was a secondary analysis of 97 patients (61 with relapsing-remitting MS [RRMS] and 36 with progressive MS) and 44 healthy control participants who participated in three prospective 3.0-T brain MRI studies between May 2009 and September 2017. A subgroup of 37 patients and 19 healthy control participants also underwent translocator protein fluorine 18 (

Chagas disease is caused by the protozoan parasite Trypanosoma cruzi, and in Central America, it is considered one of the four most infectious diseases. This study aimed to screen the anti-trypanosomal activity of plant species from Salvadoran flora. Plants were selected through literature search for plants ethnobotanically used for antiparasitic and Chagas disease symptomatology, and reported in Museo de Historia Natural de El Salvador (MUHNES) database. T. cruzi was incubated for 72 h with 2 different concentrations of methanolic extracts of 38 species, among which four species, Piper jacquemontianum, Piper lacunosum, Trichilia havanensis, and Peperomia pseudopereskiifolia, showed the activity (≤ 52.0% viability) at 100 µg/mL. Separation of the methanolic extract of aerial parts from Piper jacquemontianum afforded a new flavanone (4) and four known compounds, 2,2-dimethyl-6-carboxymethoxychroman-4-one (1), 2,2-dimethyl-6-carboxychroman-4-one (2), cardamomin (3), and pinocembrin (5), among which cardamomin exhibited the highest anti-trypanosomal activity (IC50 = 66 µM). Detailed analyses of the spectral data revealed that the new compound 4, named as jaqueflavanone A, was a derivative of pinocembrin having a prenylated benzoate moiety at the 8-position of the A ring. Graphic abstract

Many protected areas conduct awareness-raising activities to increase local knowledge and support conservation programmes, but the effectiveness of such activities is rarely assessed. Public awareness-raising has been carried out since the early 2000s around Bawangling National Nature Reserve, Hainan, China, to improve conservation knowledge about the Critically Endangered Hainan gibbon Nomascus hainanus, one of the rarest mammals. We conducted 207 interviews in 25 villages around Bawangling National Nature Reserve to evaluate the outcome of previous conservation education, through comparison of variation in local respondent knowledge and attitudes, and specific enquiries about sources of knowledge acquisition. Likelihood of accurate responses to most of our questions regarding the species was positively correlated with local exposure to gibbon-themed billboards and murals, and respondents exhibited greater knowledge about several key conservation indices for gibbons compared to their knowledge about sympatrically occurring rhesus macaques Macaca mulatta. Many respondents specifically reported they knew about local existence, population size, conservation status, and threats to gibbons from past awareness-raising activities, with village education sessions and billboards widely identified as key sources of information. However, other known awareness-raising approaches have had little detectable effect on shaping local conservation awareness. Although educational activities have improved awareness about gibbons and their conservation requirements in relative terms, overall levels of knowledge remain low in many important areas and ongoing improvement of local awareness is still needed, in particular around poorly-understood topics such as gibbon conservation status, rarity and threats, and for socio-demographic groups possessing less conservation knowledge.

Hereditary angioedema due to pathogenic FXII variants (HAE-FXII) is a rare dominant disease caused by increased activation of the plasma contact system. The most prevalent HAE-FXII variant, c.1032C > A p.Thr309Lys (FXII309Lys), results in a smaller FXII protein with increased sensitivity to fluid-phase activation by poorly understood mechanisms. We aimed to investigate the functionality of the FXII309Lys variant in 33 HAE-FXII patients, 25 healthy controls and 46 patients with congenital disorders of glycosylation (CDG). Activation of the plasma contact system was assessed by western blot and amidolytic assay in basal conditions or after treatment with either artificial or physiological activators. Recombinant wild-type and FXII309Lys variants were expressed in S2 insect (Drosophila) cells. Amidolytic and fibrin generation assays were performed in fresh plasma samples. FXII309Lys samples exhibited an increased electrophoretic mobility comparable with N-glycan-deficient FXII from CDG patients and asialo-FXII generated by neuraminidase treatment. They presented increased sensitivity to activation by dextran sulphate and silica which resulted in the generation of an aberrant 37-kDa heavy chain. We did not observe increased susceptibility of FXII309Lys to proteolysis by exogenous or tPA-generated plasmin. However, both exogenous and endogenous thrombin cleaved the FXII309Lys variant, releasing a 37-kDa fragment and resulting in enhanced proteolytic activation on the fluid phase. This model supports a sequential proteolytic activation process involving thrombin priming of FXII309Lys, followed by kallikrein cleavage and generation of active βFXIIa. The present results and the observation that angioedema episodes in HAE-FXII patients occur predominantly during hypercoagulable situations suggest a key role for thrombin.

Purpose The human antibody repertoire forms in response to infections, the microbiome, vaccinations, and environmental exposures. The specificity of such antibody responses was compared among a cohort of toddlers to identify differences between seropositive versus seronegative responses. Methods An assessment of the serum IgM and IgG antibody reactivities in 197 toddlers of 1- and 2-years of age was performed with a microfluidic array containing 110 distinct antigens. Longitudinal profiling was done from years 1 to 2. Seropositivity to RNA and DNA viruses; bacteria; live attenuated, inactive, and subunit vaccines; and autoantigens was compared. A stratification was developed based on quantitative variations in the IgG responses. Clinical presentations and previously known genetic risk alleles for various immune system conditions were investigated in relation to IgG responses. Results IgG reactivities stratified toddlers into low, moderate, and high responder groups. The high group (17%) had elevated IgG responses to multiple RNA and DNA viruses (e.g., respiratory syncytial virus, Epstein-Barr virus, adenovirus, Coxsackievirus) and this correlated with increased responses to live attenuated viral vaccines and certain autoantigens. This high group was more likely to be associated with gestational diabetes and an older age. Genetic analyses identified polymorphisms in the IL2RB, TNFSF4, and INS genes in two high responder individuals that were associated with their elevated cytokine levels and clinical history of eczema and asthma. Conclusion Serum IgG profiling of toddlers reveals correlations between the magnitude of the antibody responses towards viruses, live attenuated vaccines, and certain autoantigens. A low responder group had much weaker responses overall, including against vaccines. The serum antibody screen also identifies individuals with IgG responses to less common infections (West Nile virus, parvovirus, tuberculosis). The characterization of the antibody responses in combination with the identification of genetic risk alleles provides an opportunity to identify children with increased risk of clinical disease. Supplementary Information The online version contains supplementary material available at 10.1007/s10875-021-00993-w.

Objective The aim of this study was to evaluate the quality of diagnosis and treatment of COPD using Big Data methodology on the Savana Manager 2.1 clinical platform. Materials and methods A total of 59,369 patients with a diagnosis of COPD were included from a population of 1,219,749 adults over 40 years of age. Results In total, 78% were men. Spirometry data were available for only 26,453 (43.5%) subjects. Disease severity was classified in 18,172 patients: 4396 mild, 7100 moderate, and 6676 severe, although only 27%, 34%, and 28%, respectively, presented obstructive spirometry. The clinical management of COPD is mainly the responsibility of the primary care and pulmonology departments, while internal medicine and, to a lesser extent, geriatrics also participate. Drug treatment was based on bronchodilators and inhaled corticosteroids (ICS). A marked decline in the use of long-acting beta-2 agonists (LABA) in monotherapy and a slight reduction in ICS/LABA combinations, associated with a long-acting anticholinergic (LAMA) in 74% of cases, were observed. All-cause in-hospital mortality among the overall population was 5.6% compared to 1% of the general population older than 40 years. In total, 35% were admitted to hospital, with an average stay of 6.6 days and an in-hospital mortality rate in this group of 10.74%. Discussion This study identifies the main features of an unselected COPD population and the main errors made in the management of the disease.

16 p.-6 fig., Clavulanic acid (CLV) and amoxicillin, frequently administered in combination, can be independently involved in allergic reactions. Protein haptenation with β-lactams is considered necessary to activate the immune system. The aim of this study was to assess the suitability of biotinylated analogues of CLV as probes to study protein haptenation by this β-lactam. Two synthetic approaches afforded the labeling of CLV through esterification of its carboxylic group with a biotin moiety, via either direct binding (CLV-B) or tetraethylenglycol linker (CLV-TEG-B). The second analogue offered advantages as solubility in aqueous solution and potential lower steric hindrance for both intended interactions, with the protein and with avidin. NMR reactivity studies showed that both CLV and CLV-TEG-B reacts through β-lactam ring opening by aliphatic amino nitrogen, however with different stability of resulting conjugates. Unlike CLV conjugates, that promoted the decomposition of clavulanate fragment, the conjugates obtained with the CLV-TEG-B remained linked, as a whole structure including biotin, to nucleophile and showed a better stability. This was a desired key feature to allow CLV-TEG-B conjugated protein detection at great sensitivity. We have used biotin detection and mass spectrometry (MS) to detect the haptenation of human serum albumin (HSA) and human serum proteins. MS of conjugates showed that HSA could be modified by CLV-TEG-B. Remarkably, HSA preincubation with CLV excess only reduced moderately the incorporation of CLV-TEG-B, which could be attributed to different protein interferences. The CLV-TEG-B fragment with opened β-lactam was detected bound to the 404–430HSA peptide of the treated protein. Incubation of human serum with CLV-TEG-B resulted in the haptenation of several proteins that were identified by 2D-electrophoresis and peptide mass fingerprinting as HSA, haptoglobin, and heavy and light chains of immunoglobulins. Taken together, our results show that tagged-CLV keeps some of the CLV features. Moreover, although we observe a different behavior in the conjugate stability and in the site of protein modification, the similar reactivity indicates that it could constitute a valuable tool to identify protein targets for haptenation by CLV with high sensitivity to get insights into the activation of the immune system by CLV and mechanisms involved in β-lactams allergy., Work at MJT and MIM laboratory was supported by Instituto de Salud Carlos III (ISCIII) of MICINN (grants cofunded by ERDF: “Una manera de hacer Europa” (PI17/01237, PI18/00095, RETIC ARADYAL RD16/0006/0001 and Euronanomed Program AC19/00082), Miguel Servet I program (CP15/00103) and Sara Borrell program (CD17/00146)), Andalusian Regional Ministry of Health (PI-0179-2014, PE-0172-2018). Work at EP-I laboratory was supported by the Spanish Ministerio de Economía, Industria y Competitividad (CTQ 2016-75870-P), Ministerio de Ciencia y Educación (PID 2019-104293GB-I00), Ministerio de Ciencia e Innovación [Proyectos de I+D+I Programación Conjunta Internacional, EuroNanoMed 2019 (PCI 2019-111825-2)], ISCIII RETIC ARADYAL RD16/0006/0012 and Junta de Andalucía (UMA18-FEDERJA-007). Work at DP-S laboratory was supported by Grants from Agencia Estatal de Investigación, Ministerio de Ciencia e Innovación (MICINN, Spain) and European Regional Development Fund, SAF 2015-68590-R and RTI 2018-097624-B-I00, ISCIII RETIC ARADyAL RD16/0006/0021.

Herein, we explored the impact of the lysosome dysfunction during the progression of Amyotrophic Lateral Sclerosis type-1 (ALS1). We conducted the study in non-neural cells, primary fibroblasts (rFFFs), and bone marrow-mesenchymal stem cells (rBM-MSCs), isolated from the animal model ratG93A for ALS1 at two stages of the disease: Pre-symptomatic-stage (ALS1-PreS) and Terminal-stage (ALS1-EndS). We documented the storage of human mutant Superoxide Dismutase 1, SOD1G93A (SOD1*) in the lysosomes of ALS1-rFFFs and ALS1-rBM-MSCs and demonstrated the hallmarks of the disease in non-neural cells as in ratG93A-ALS1-tissues. We showed that the SOD1* storage is associated with the altered glycohydrolases and proteases levels in tissues and both cell types from ALS1-PreS to ALS1-EndS. Only in ALS1-rFFFs, the lysosomes lost homeostasis, enlarge drastically, and contribute to the cell metabolic damage. Contrariwise, in ALS1-rBM-MSCs, we found a negligible metabolic dysfunction, which makes these cells’ status similar to WT. We addressed this phenomenon to a safety mechanism perhaps associated with an enhanced lysosomal autophagic activity in ALS1-rBM-MSCs compared to ALS1-rFFFs, in which the lysosomal level of LC3-II/LC3I was comparable to that of WT-rFFFs. We suggested that the autophagic machinery could balance the storage of SOD1* aggregates and the lysosomal enzyme dysfunction even in ALS1-EndS-stem cells.

TO THE EDITOR: Factor XI (FXI) is a key element for the amplification of thrombin generation, which participates in the contact pathway. This explains why FXI deficiency has minor physiological relevance but protects against thrombotic events, supporting FXI as an excellent target for

Pediatric thromboembolism (≤18 years) is very rare (0.07-0.14/10,000/year) but may be more prevalent in children with severe thrombophilia (protein C, protein S or antithrombin deficiency). The aim of this study was to define the prevalence and clinical characteristics of pediatric thrombosis in subjects with inherited antithrombin deficiency. Our observational retrospective multicentric study from two countries recruited 968 patients of any age from 441 unrelated families with genetically, biochemically and functionally characterized antithrombin deficiency. Seventy-three subjects (7.5%) developed thrombosis before 19 years of age. Two high-risk periods for thrombosis were identified: adolescence (12-18 years, n=49) with thrombus localization (lower limb deep venous thrombosis or pulmonary embolism) and triggering factors common to adults (oral contraceptives, surgery or pregnancy); and the neonatal period (

Streptococcus uberis is one of the leading causes worldwide of mastitis in the dairy industry, with the most likely sources of infection attributed to environmental reservoirs such as contaminated bedding materials. Early detection of those cases most likely to progress to clinical disease would lead to improved animal welfare, a critical component of overall health and productivity. A multiplex PCR-based diagnostic test was developed for detection of S. uberis directly from milk and targeting two genes previously identified as important for intramammary colonisation and persistence in dairy cattle. Results indicated the threshold for detection directly from milk was 20,000 CFU/ml and this was achieved without the need for preenrichment. In addition, S. uberis could be identified from milk samples collected during intramammary challenge studies, prior to clinical signs of infection and at much lower detection limits. The PCR test developed for confirmation of the presence of S. uberis directly from infected milk has potential value as a diagnostic test to identify early infection and/or to confirm that antibiotic therapy has been successful.

Objectives The optimal dosage regimen of gentamicin in horses is still under investigation. The objectives of this study were to determine gentamicin plasma concentrations in hospitalized horses treated with 10 mg/kg gentamicin (IV, q 24 h) and to determine whether a plasma concentration to minimum inhibitory concentration (MIC) ratio of 10:1 is reached for equine pathogens using this dose. Design Prospective clinical observational study; retrospective study on MICs of 131 gram-negative bacteria isolated from horses (2012-2015). Setting University teaching hospital. Animals Ninety-eight horses >6 months old, treated with gentamicin for their primary disease, consecutive samples. Measurements and main results Plasma concentrations were measured 1 hour (C1h ) and 20 hours (C20h ) after gentamicin administration using fluorescence polarization. Presence of systemic inflammatory response syndrome (SIRS) and azotemia was recorded, as well as the reason for antimicrobial administration (primary disease) and whether administration was prophylactic or therapeutic. The target C1h of ≥20 µg/mL gentamicin was reached in 90% of horses and was sufficient to reach a plasma concentration:MIC of 10:1 in 32 of 131 (24%) of gram-negative aerobic bacteria. A C20h ≤ 2 µg/mL was reached in 97% horses. Therapeutic versus prophylactic administration, primary disease, azotemia, and systemic inflammatory response syndrome were not associated with a failure to reach a desired peak or trough. Conclusions The gentamicin dose of 10 mg/kg every 24 hours should be further investigated and safety assessed because a target gentamicin plasma concentration of ≥20 µg/mL was achieved in the majority of cases. Nephrotoxic side effects were not assessed. Individual drug monitoring should be performed because clinical factors are unreliable predictors of plasma concentrations. A gentamicin target concentration of ≥40 µg/mL does not offer additional benefits compared to ≥20 µg/mL, due to the bimodal distribution of resistance in bacterial isolates.

Abstracts of the Virtual Conference Insects to Feed the World. November 23-26, 2020.

Supplemental Digital Content is available in the text., OBJECTIVES: We aimed to assess the frequency of ICU-acquired bloodstream infections in coronavirus disease 2019 patients. DESIGN: Retrospective observational study. SETTING: The emergency expansion of an ICU from eight general beds to 30 coronavirus disease 2019 beds. PARTICIPANTS: Patients with coronavirus disease 2019 admitted to the ICU of Luigi Sacco Hospital (Milan, Italy) for greater than or equal to 48 hours between February 21, 2020, and April 30, 2020. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The frequency of bloodstream infections per 1,000 days of ICU stay was calculated in 89 coronavirus disease 2019 patients, and the cumulative probability of bloodstream infection was estimated using death and ICU discharge as competing events. Sixty patients (67.4%) experienced at least one of the 93 recorded episodes of bloodstream infection, a frequency of 87 per 1,000 days of ICU stay (95% CI, 67–112).The patients who experienced a bloodstream infection had a higher Sequential Organ Failure Assessment score upon ICU admission (9.5; interquartile range, 8–12 vs 8, interquartile range, 5–10; p = 0.042), a longer median ICU stay (15 d; interquartile range, 11–23 vs 8, interquartile range, 5–12; p < 0.001), and more frequently required invasive mechanical ventilation (98.3% vs 82.8%; p = 0.013) than those who did not. The median time from ICU admission to the first bloodstream infection episode was 10 days. Gram-positive bacteria accounted for 74 episodes (79.6%), with Enterococcus species being the most prevalent (53 episodes, 55.8%). Thirty-two isolates (27.3%) showed multidrug resistance. CONCLUSIONS: Coronavirus disease 2019 seemed to increase the frequency of bloodstream infections (particularly Enterococcus-related bloodstream infection) after ICU admission. This may have been due to enteric involvement in patients with severe coronavirus disease 2019 and/or limitations in controlling the patient-to-patient transmission of infectious agents in extremely challenging circumstances.

ObjectiveTo analyse autoantibody status in a well-defined European multicentre cohort of patients with epilepsy of unknown aetiology and to validate the recently proposed Antibody Prevalence in Epilepsy (APE2) and Response to ImmunoTherapy in Epilepsy (RITE2) scores.MethodsWe retrospectively collected clinical and paraclinical data of 92 patients referred to the Neurology Units of Verona and Salzburg between January 2014 and July 2019 with new-onset epilepsy, status epilepticus or chronic epilepsy of unknown aetiology. Fixed and live cell-based assays, tissue-based assays, immunoblot, and live rat hippocampal cell cultures were performed in paired serum/cerebrospinal fluid (CSF) to detect antineuronal and antiglial antibodies. The APE2 and RITE2 scores were then calculated and compared with clinical and laboratory data.ResultsAutoantibodies were detected in 29/92 patients (31.5%), with multiple positivity observed in 6/29 cases. The APE2 score (median 5, range 1–15) significantly correlated with antibody positivity (p=0.014), especially for the presence of neuropsychiatric symptoms (pConclusionsThis is the first study that independently validates the APE2 and RITE2 scores and includes the largest cohort of patients whose paired serum and CSF samples have been tested for autoantibodies possibly associated with autoimmune epilepsy.

A global reshaping of the immune responses occurs with ageing, indicated as immunosenescence, where mitochondria and mitochondrial metabolism play an important role. However, much less is known about the role of mitochondrial stress response in this reshaping and in particular of the molecules induced by such response, collectively indicated as mitokines. In this review, we summarize the current knowledge on the role of mitokines in modulating immune response and inflammation focusing on GDF15, FGF21 and humanin and their possible involvement in the chronic age-related low-grade inflammation dubbed inflammaging. Although many aspects of their biology are still controversial, available data suggest that these mitokines have an anti-inflammatory role and increase with age. Therefore, we hypothesize that they can be considered part of an adaptive and integrated immune-metabolic mechanism activated by mitochondrial dysfunction that acts within the framework of a larger anti-inflammatory network aimed at controlling both acute inflammation and inflammaging.

BACKGROUND This study's purposes were to characterize detection rates of several sexually transmitted infection (STI) agents and describe the effect additional specimen source and analyte screening has on STI detection within a cohort of young men who have sex with men and transgender women. METHODS Within a 16-month interval, 1966 encounters involved dual urine and rectal swab submissions assessed by commercial transcription-mediated amplification-based assays for Chlamydia trachomatis and Neisseria gonorrhoeae and by off-label transcription-mediated amplification-based Trichomonas vaginalis and Mycoplasma genitalium testing. Identification of STI carriers used algorithms involving Food and Drug Administration-cleared screening methods, laboratory-modified testing for extraurogenital C. trachomatis and N. gonorrhoeae, and laboratory-developed tests for T. vaginalis and M. genitalium. RESULTS Food and Drug Administration-indicated urine C. trachomatis and N. gonorrhoeae screening revealed 39 encounters (2.0%) yielding one or both agents. Via C. trachomatis and N. gonorrhoeae screening that included rectal swab analysis, 264 encounters (13.4%) yielded evidence of either (140 C. trachomatis, 88 N. gonorrhoeae) or both (36 participants) infections. Detection rates for C. trachomatis and N. gonorrhoeae were 1.4% and 0.6% for urine screening and 8.2% and 6.2% for rectal screening, respectively. Off-label screening identified 413 additional encounters with STI (5 T. vaginalis, 396 M. genitalium, 12 with both). Of these identifications, 82.1% were generated from analysis of rectal swabs (4 participants with T. vaginalis, 323 participants with M. genitalium, 12 with both). Overall detection rates of T. vaginalis (0.2% urine, 1.3% rectal) and M. genitalium (9.1% urine, 21.5% rectal) were variable. CONCLUSIONS Additive analyte testing, including extraurogenital collections, contributes to comprehensive STI screening within a high-risk demographic.

Twin studies of disease concordance are useful to weight the relative contribution of genetic and environmental factors to the cause of common complex disorders. In multiple sclerosis (MS) different twinning rates from geographic areas at different prevalence suggested that heritable and non-heritable factors contribute in different proportions and ways to MS risk in diverse populations. This concept prompted genome-wide association studies, and the implementation of the co-twin control design, that allows stringent experimental approaches in MS-discordant identical pairs, controlling for genetic influences and many other known and unknown factors. The co-twin control design provided important clues on MS molecular model. These studies will be reviewed, focusing on those showing significant differences between affected and healthy co-twins. In some cases, differences that emerged in non-twin patients compared to matched controls were not confirmed in identical MS-discordant pairs, suggesting an 'MS subclinical trait'. Early patterns of magnetic resonance imaging and predictive biomarkers that characterize 'healthy' co-twins may be useful for the identification of a prodromal reversible phase of the disease.

International audience; AbstractBeta‐trace protein (BTP), a low molecular weight protein of 23‐29 kDa, has been proposed as a promising biomarker to estimate residual renal function (RRF) in patients on maintenance hemodialysis (HD). Indeed, BTP is cleared by native kidney but not during conventional HD session. By contrast, the removal rate of BTP using convective processes (mainly hemodiafiltration [HDF]) and peritoneal dialysis (PD) has been little or not investigated. Therefore, an aim of this study was to evaluate the impact of dialysis procedures (high‐flux HD, on‐line post‐dilution HDF and PD) on BTP removal in comparison with beta‐2 microglobulin (B2M) and cystatin C (CYSC) removals after a single session. In addition, the ability of BTP to predict RRF in PD was assessed. This observational cross‐sectional study included a total of 82 stable chronic kidney disease patients, 53 patients were on maintenance dialysis (with n = 26 in HD and n = 27 in HDF) and 29 were on PD. Serum concentrations of BTP, B2M, and CYSC were measured (a) before and after a single dialysis session in HD and HDF anuric patients to calculate reduction percentages, (b) in serum, 24‐hour‐dialysate and 24‐hour‐urine in PD patients to compute total, peritoneal, and urinary clearance. RRF was estimated using four equations developed for dialysis patients without urine collection and compared to the mean of the urea and creatinine clearances in PD. The concentrations of the three studied molecules were significantly reduced (P < .001) after dialysis session with significantly higher reduction ratio using HDF compared to HD modality (P < .001): BTP 49.3% vs 17.5%; B2M 82.3% vs 69.7%; CYSC 77.4% vs 66% in HDF and HD, respectively. In non‐anuric PD patients, B2M and CYSC were partly removed by peritoneal clearance (72.3% and 57.6% for B2M and CYSC, respectively). By contrast, BTP removal by the peritoneum was negligible and a low bias for the BTP‐based equation to estimate RRF (−1.4 mL/min/1.73 m2) was calculated. BTP is significantly removed by high‐flux HD or HDF, thereby compromising its use to estimate RRF. By contrast, BTP appears as a promising biomarker to estimate RRF in PD patients since it is not affected by peritoneal clearance, unlike B2M and CYSC, and it is well correlated to RRF.

Background Encephalitozoon cuniculi, are obligate intracellular fungi that determine emerging diseases in humans and other animals, with economic relevance. Studies have shown that Malassezia are able to modulate the production of pro and anti-inflammatory cytokines by keratinocytes and human monocytes and has an immunomodulatory effect. In the present work, we evaluate the effects of heat-killed M. pachydermatis suspension on the modulation of the response of macrophages challenged with the microsporidia E. cuniculi. Methods Heat-killed M. pachydermatis suspension was used to treat macrophage for 1 hour, subsequently, macrophages were infected with spores of E. cuniculi (2:1) and evaluated at 15, 30, 60 minutes and 48 hours. To determine the phagocytic capacity (PC) and index (PI), cultures were stained with Calcofluor and the spores, internalized or not, were counted. Results Microbicidal and phagocytic activity was evaluated by transmission electron microscopy (TEM). The untreated macrophages and had PC, PI, and larger amounts of phagocytosed spores than those treated, but by TEM it was observed that in macrophages treated the microbicidal activity was faster, since in the phagocytic vacuoles there were few spores in different degrees of degeneration and few amounts of amorphous material. Macrophages treated with heat-killed M. pachydermatis suspension showed lower PI and PC, with the incipient presence of E. cuniculi in phagosomes inside these macrophages. Additionally, these same treated macrophages had a mixed pattern of cytokine release with a Th1, Th2 and Th17 profile, with emphasis on IL-10, IL-4, IL-17, IL-6 and IFN-g secretion, particularly greater production of anti-inflammatory cytokines. Conclusions Our results showed that the treatment with heat-killed M. pachydermatis suspension determined an increase in the release of cytokines and reduced the phagocytic activity of macrophages challenged with E. cuniculi

The aim of this work was to develop processing methods that safeguard the quality and antimicrobial properties of H. illucens and B. mori oils. We adopted a vegetable diet for both insects: leftover vegetables and fruit for H. illucens and mulberry leaves for B. mori. First, alternative techniques to obtain a good oil extraction yield from the dried biomass of H. illucens larvae were tested. Traditional pressing resulted to be the best system to maximize the oil yield and it was successfully applied to B. mori pupae. Oil quality resulted comparable to that obtained with other extraction methods described in the literature. In the case of B. mori pupae, different treatments and preservation periods were investigated to evaluate their influence on the oil composition and quality. Interestingly, agar diffusion assays demonstrated the sensitivity of Gram-positive Bacillus subtilis and Staphylococcus aureus to H. illucens and B. mori derived oils, whereas the growth of Gram-negative Pseudomonas aeruginosa and Escherichia coli was not affected. This study confirms that fat and other active compounds of the oil extracted by hot pressing could represent effective antimicrobials against bacteria, a relevant result if we consider that they are by-products of the protein extraction process in the feed industry.

Background Hereditary antithrombin deficiency is a rare autosomal-dominant disorder predisposing to recurrent venous thromboembolism (VTE). To date, only two founder mutations have been described. Objectives We investigated the antithrombin p.Thr147Ala variant, found in 12 patients of African origin. This variant is known as rs2227606 with minor allele frequency of 0.5% in Africans and absent in Europeans. A possible founder effect was investigated. Methods Phenotypical characterization was established through immunological and functional methods, both under basal and stress conditions. Recombinant antithrombin molecules were constructed by site-directed mutagenesis and expressed in HEK-293T cells. Secreted antithrombin was purified and functionally characterized. Structural modeling was performed to predict the impact of the mutation on protein structure. A novel nanopore sequencing approach was used for haplotype investigation. Results Ten patients experienced VTE, stroke, or obstetric complications. Antithrombin antigen levels and anti-IIa activity were normal or slightly reduced while anti-Xa activity was reduced with only one commercial assay. On crossed immunoelectrophoresis, an increase of antithrombin fractions with reduced heparin affinity was observed under high ionic strength conditions but not under physiological conditions. The recombinant p.Thr147Ala protein displayed a reduced anti-Xa activity. Structural modeling revealed that residue Thr147 forms three hydrogen bonds that are abolished when mutated to alanine. The investigated patients shared a common haplotype involving 13 SERPINC1 intragenic single nucleotide polymorphisms. Conclusion Antithrombin p.Thr147Ala, responsible for antithrombin type II heparin binding site deficiency, is the first founder mutation reported in people of African ancestry. This study further emphasizes the limitations of commercial methods to diagnose this specific subtype.

Background Despite their indispensability in human medicine, fluoroquinolones (FQ) are used for the treatment of bacterial infections in farm animals which increases the risk of transferring FQ-resistant bacteria into the environment and via the food chain to humans. The objectives of this observational study were to follow-up of the presence of quinolone non-susceptible Escherichia coli (QNSE) qualitatively and quantitatively in faecal samples of pigs at four time points (2 weeks old, 4 weeks old, 2 weeks post weaning and during fattening period). Moreover differences between groups of FQ-treated pigs, pigs with contact to treated pigs and control pigs were investigated. Additionally, quinolone and FQ resistance of Escherichia coli isolates of the faecal samples were investigated by determining minimum inhibitory concentrations (MICs). Results 40.9% of 621 fecal samples contained QNSE. Proportion of samples with detectable QNSE from treated and contact pigs did not differ significantly and were highest in piglets of 2 and 4 weeks of age. However, the proportions of samples with QNSE were significantly lowest in control pigs (7/90; 7.8%; CI = 3.5–14.7%) among all groups. Also, the number of colony-forming units was lowest in both weaners and fattening pigs of the control group compared to treated and contact groups. Following CLSI human breakpoints, in total, 50.4% out of 254 isolates in faecal samples were intermediate or resistant to ciprofloxacin. Conclusions QNSE were present in faeces of pigs independent of age or FQ background but significantly less were found in pigs from farms without FQ usage. Due to the long half-life of FQ, it is likely that only a prolonged absence of fluoroquinolone treatments in pig farming will lead to a reduced frequency of QNSE in the farm environment. Solutions need to be found to minimise the emergence and transfer of quinolone and FQ-resistant bacteria from treated pigs to contact pigs and to farms without FQ usage.

Global commitments prioritize protection of wildlife and improvements to human wellbeing. Local disconnects in these commitments are rarely acknowledged—or their implications assessed—preventing the development of effective solutions. National and international efforts to protect marine mammals along South America's west coast have contributed to species recovery, but also to conflict between sea lions and small‐scale fisheries. To understand the concerns ultimately motivating this conflict, we assessed how 301 coastal small‐scale fishers perceive their interactions with South American sea lions (Otaria flavescens). We then reviewed the terrestrial human–wildlife literature to identify potential management solutions to resolve the conflict. We find that fishers are chiefly concerned with increases in sea lion populations, perceive that sea lion interactions have significantly increased over the past 80 years, and report sea lion‐driven catch and income losses of ≥26%. We propose solutions to manage conflict that are sensitive to heterogeneity among fisher groups.

PURPOSE The aim of this study was to compare the Streptococcus oralis biofilm formation on titanium machined turned surfaces and sandblasted surfaces that were previously characterized for their superficial topographies. MATERIALS AND METHODS Two commercially pure titanium surfaces were analyzed and compared: machined (turned surfaces subjected to a process of decontamination that also included a double acid attack) and sandblasted (sandblasted surfaces, cleaned with purified water, enzymatic detergent, acetone, and alcohol). The characterization of the samples at the nanolevel was performed using atomic force microscopy, which permitted calculation of the superficial nanoroughness (Ra). The sessile drop method was used to measure the water contact angle in both groups and allowed information to be gained about their wetting properties. Scanning electron microscope and energy-dispersive x-ray spectroscopy analysis allowed comparison of the microtopographic geometry and the chemical composition of the samples. Then, the disks were pre-incubated with saliva in order to form an acquired pellicle. Streptococcus oralis was put on the disks, and both groups were tested at 24 and 48 hours for biofilm biomass evaluation, colony-forming units (CFUs), and live/dead staining for cell viability. RESULTS The sandblasted samples were characterized by a significantly higher level of superficial oxides, superficial roughness, and hydrophilicity, compared with the machined turned samples. Although there were topographic differences, the Streptococcus oralis biofilm formation, quantified in CFUs, and biomass formation at 24 and 48 hours were similar in both groups. With the live/dead staining, the sandblasted disks were characterized by an increased percentage of dead cells compared with the machined disks. CONCLUSION Although significant topographic differences were present between machined and sandblasted disks, the Streptococcus oralis biofilm formation seems to not be significantly affected.

The identification and characterization of structural variants (SVs) in clinical genetics have remained historically challenging as routine genetic diagnostic techniques have limited ability to evaluate repetitive regions and SVs. Long-read whole-genome sequencing (LR-WGS) has emerged as a powerful approach to resolve SVs. Here, we used LR-WGS to study 19 unrelated cases with type I Antithrombin Deficiency (ATD), the most severe thrombophilia, where routine molecular tests were either negative, ambiguous, or not fully characterized. We developed an analysis workflow to identify disease-associated SVs and resolved 10 cases. For the first time, we identified a germline complex rearrangement involved in ATD previously misclassified as a deletion. Additionally, we provided molecular diagnoses for two unresolved individuals that harbored a novel SINE-VNTR-Alu retroelement insertion that we fully characterized by de novo assembly and confirmed by PCR amplification in all affected relatives. Finally, the nucleotide-level resolution achieved for all the SVs allowed breakpoint analysis, which revealed a replication-based mechanism for most of the cases. Our study underscores the utility of LR-WGS as a complementary diagnostic method to identify, characterize, and unveil the molecular mechanism of formation of disease-causing SVs, and facilitates decision making about long-term thromboprophylaxis in ATD patients.

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Feline morbillivirus (FeMV) is a newly discovered paramyxovirus infecting domestic cats and its role in the pathogenesis of feline chronic kidney disease (CKD) has been suggested, however not confirmed. The primary aim of the study was to evaluate the renal damage associated with FeMV infection in cats. In this retrospective study, clinical and clinicopathological data were compared among 14 FeMV naturally infected, 21 CKD and 22 healthy cats. FeMV positive cats had serum chemistry analytes and main urine chemistry results similar to the healthy subjects. FeMV positive cats had significantly decreased urine specific gravity (median 1054, range 1022&ndash, 1065) and urine creatinine (median 227.23 mg/dL, range 83.02&ndash, 489.75) when compared with healthy cats (median 1067, range 1040&ndash, 1080, P &lt, 0.001, median 406.50 mg/dL, range 195.32&ndash, 575.58, P &lt, 0.001, respectively). Urine protein:creatinine ratio (UPC) results of FeMV and CKD were not different (median 0.20, range 0.08&ndash, 1.03, median 0.23, range 0.10&ndash, 0.80, respectively), however UPC results were significantly increased in both groups, if compared with healthy cats (median 0.1, range 0.04&ndash, 0.250, P &lt, 0.01). Based on clinical data, serum creatinine concentration, urine specific gravity and UPC results, CKD was suspected by clinicians in 3/14 FeMV cats. Urine protein sodium-dodecyl-sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in 10/13 (77%) FeMV cats indicated a tubular pattern, with a decrease of uromodulin and an increase in the number and intensity of low molecular weight proteins. FeMV infection can be associated with different grades of renal dysfunction ranging from mild tubular proteinuria with less concentrated urine to azotemia in cats younger than those typically affected by CKD.

Sturgeon aquaculture production has increased worldwide, particularly in Italy, yet there is little information about diseases that affect sturgeons. With this study we describe the results of screening for bacterial infections during monitoring campaigns carried out from 2014 to 2017. A total of 402 sturgeons from six sturgeon farms with one or more reared sturgeon species among five species and two hybrids were analysed. Bacteriological exam was positive in 93 individuals (23%) and negative in the remaining 309 (77%). Generally, no external or internal macroscopic lesions were present, though some specimens showed skin lesions, liver anaemia or splenomegaly, which are non‐pathognomonic signs of bacterial infection. The majority of the isolated bacteria were opportunistic. This knowledge base is important to help create a bacterial profile of sturgeons reared in Italy.

Aromatic halophytes represent an exceptional source of natural bioactive compounds for the food industry. Crithmum maritimum L., also known as sea fennel, is a halophyte plant colonizing cliffs and coastal dunes along Mediterranean and Atlantic coasts. It is well known to produce essential oils and polyphenols endowed with antioxidant and biological effects. The present work reports the phytochemical profile, as well as antioxidant, antimicrobial and antimutagenic properties of C. maritimum leaf hydro-alcoholic extract. From LC-ESI-MS analysis, eighteen phenolic compounds were depicted in sea fennel extract and the amount of total phenolic content exceeds 3% DW. Accordingly, C. maritimum extract showed strong antioxidant activities, as evidenced by in vitro (DPPH, ORAC, FRAP) and ex vivo (CAA-RBC and hemolysis) assays. An important antimicrobial activity against pathogenic strains was found as well as a strong capacity to inhibit Staphylococcus aureus (ATCC 35556) biofilm formation. Sea fennel extracts showed a significant decrease of mutagenesis induced by hydrogen peroxide (H2O2) and menadione (ME) in Saccharomyces cerevisiae D7 strain. In conclusion, our results show that C. maritimum is an exceptional source of bioactive components and exert beneficial effects against oxidative or mutagenic mechanisms, and pathogenic bacteria, making it a potential functional food.

Made available in DSpace on 2022-04-28T19:42:49Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-08-01 This study aimed to obtain a better understanding of the regulatory genes and molecules involved in the development of mastitis. For this purpose, the transcription factors (TF) and MicroRNAs (miRNA) related to differentially expressed genes previously found in extracorporeal udders infected with Streptococcus agalactiae were investigated. The Gene-TF network highlighted LOC515333, SAA3, CD14, NFKBIA, APOC2 and LOC100335608 and genes that encode the most representative transcription factors STAT3, PPARG, EGR1 and NFKB1 for infected udders. In addition, it was possible to highlight, through the analysis of the gene-miRNA network, genes that could be post-transcriptionally regulated by miRNAs, such as the relationship between the CCL5 gene and the miRNA bta-miR-363. Overall, our data demonstrated genes and regulatory elements (TF and miRNA) that can play an important role in mastitis resistance. The results provide new insights into the first functional pathways and the network of genes that orchestrate the innate immune responses to infection by Streptococcus agalactiae. Our results will increase the general knowledge about the gene networks, transcription factors and miRNAs involved in fighting intramammary infection and maintaining tissue during infection and thus enable a better understanding of the pathophysiology of mastitis. Institute of Biological Sciences Universidade Federal de Juiz de Fora Zootechnics Department Universidade Federal Dos Vales Do Jequitinhonha e Mucuri Zootechnics Department Universidade Federal Do Espírito Santo Department of Exact Sciences Universidade Estadual Paulista Júlio de Mesquita Filho Molecular Genetics Laboratory Embrapa Gado de Leite Department of Exact Sciences Universidade Estadual Paulista Júlio de Mesquita Filho

The transforming growth factor beta (TGFβ) pathway could modulate the Duchenne muscular dystrophy (DMD) phenotype. This meta-analysis aims to estimate the association of genetic variants involved in the TGFβ pathway, including the latent transforming growth factor beta binding protein 4 (LTBP4) and secreted phosphoprotein 1 (SPP1) genes, among others, with age of loss of ambulation (LoA) and cardiac function in patients with DMD. Meta-analyses were conducted for the hazard ratio (HR) of LoA for each genetic variant. A subgroup analysis was performed in patients treated exclusively with glucocorticoids. Eight studies were included in the systematic review and four in the meta-analyses. The systematic review suggests a protective effect of LTBP4 haplotype IAAM (recessive model) for LoA. It is also suggested that the SPP1 rs28357094 genotype G (dominant model) is associated with early LoA in glucocorticoids-treated patients. The meta-analysis of the LTBP4 haplotype IAAM showed a protective association with LoA, with an HR = 0.78 (95% CI: 0.67–0.90). No association with LoA was observed for the SPP1 rs28357094. The LTBP4 haplotype IAAM is associated with a later LoA, especially in the Caucasian population, while the SPP1 rs28357094 genotype G could be associated with a poor response to glucocorticoids. Future research is suggested for SPP1 rs11730582, LTBP4 rs710160, and THBS1 rs2725797.

Neoantigens are tumor-specific antigens able to induce T-cell responses, generated by mutations in protein-coding regions of expressed genes. Previous studies demonstrated that only a limited subset of mutations generates neoantigens in microsatellite stable tumors. We developed a method, called VENUS (Vaccine-Encoded Neoantigens Unrestricted Selection), to prioritize mutated peptides with high potential to be neoantigens. Our method assigns to each mutation a weighted score that combines the mutation allelic frequency, the abundance of the transcript coding for the mutation, and the likelihood to bind the patient’s class-I major histocompatibility complex alleles. By ranking mutated peptides encoded by mutations detected in nine cancer patients, VENUS was able to select in the top 60 ranked peptides, the 95% of neoantigens experimentally validated including both CD8 and CD4 T cell specificities. VENUS was evaluated in a murine model in the context of vaccination with an adeno vector encoding the top ranked mutations prioritized in the MC38 cell line. Efficacy studies demonstrated anti tumoral activity of the vaccine when used in combination with checkpoint inhibitors. The results obtained highlight the importance of a combined scoring system taking into account multiple features of each tumor mutation to improve the accuracy of neoantigen prediction.

(1) Background: The objectives of this study were to evaluate the effect of several sessions of the antibacterial protocol of complex electromagnetic fields (CMFs) on planktonic Candida albicans and fungal ability, after treatment with CMFs, to adhere and proliferate on acrylic resin materials. (2) Methods: Planktonic overnight cultures of Candida albicans were subjected to different entities of CMFs treatments. Four test groups were compared: “p1”: treated only with the first program of the antibacterial protocol, “p1–p5” subjected to the first five programs, “1 antibacterial” received one complete session of the protocol and “2 antibacterial” received two complete sessions. After the treatments, the number of colony forming units (CFUs) were recorded. Then, C. albicans broth cultures were cultivated on polyacrylic resin discs and evaluated for CFUs and subjected to scanning electron microscope (SEM) analysis. (3) Results: Microbiological analysis showed that CMFs promoted a significant reduction of C. albicans CFUs when the protocol “p1–p5” was applied. No statistically significant differences between test groups were observed if the time of exposure to CMFs was increased. SEM observations and CFUs showed that CMFs treatments have the ability to reduce C. albicans adherence and proliferation on discs. (4) Conclusions: The CMFs showed an antifungal effect as well as a decrease in C. albicans adhesion on polyacrylic resin.

The Gastrointestinal (GI) tract is composed of four main barriers: microbiological, chemical, physical and immunological. These barriers play important roles in maintaining GI tract homeostasis. In the crosstalk between these barriers, microbiota and related metabolites have been shown to influence GI tract barrier integrity, and alterations of the gut microbiome might lead to an increase in intestinal permeability. As a consequence, translocation of bacteria and their products into the circulatory system increases, reaching proximal and distal tissues, such as the liver. One of the most prevalent chronic liver diseases, Nonalcoholic Fatty Liver Disease (NAFLD), has been associated with an altered gut microbiota and barrier integrity. However, the causal link between them has not been fully elucidated yet. In this review, we aim to highlight relevant bacterial taxa and their related metabolites affecting the GI tract barriers in the context of NAFLD, discussing their implications in gut homeostasis and in disease.

Aging is characterized by dynamic changes at metabolic level that lead to modifications in the composition of the metabolome. Since the identification of biomarkers that can discriminate people of different age and health status has recently attracted a great interest, we wondered whether age-specific changes in the metabolome could be identified and serve as new and informative biomarkers of aging and longevity. In the last few years, a specific branch of metabonomics devoted to the study of volatile organic compounds (VOCs) has been developed. To date, little is known about the profile of specific VOCs in healthy aging and longevity in humans; therefore, we investigated the profile of VOCs in both urine and feces samples from 73 volunteers of different age including centenarians that represent useful “super-controls” to identify potential biomarkers of successful aging and footprints of longevity. To this purpose, we performed a discriminant analysis by which we were able to identify specific profiles of urinary and fecal VOCs. Such profiles can discriminate different age groups, from young to centenarians, and, even more interesting, centenarians’ offspring from age-matched controls. Moreover, we were able to identify VOCs that are specific for the couples “centenarians – offspring” or the trios “centenarians – offspring – spouse,” suggesting the possible existence of a familiar component also for VOCs profile. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s11357-019-00143-6) contains supplementary material, which is available to authorized users.

Background Congenital disorders of glycosylation are rare inherited diseases affecting many different proteins. The lack of glycosylation notably affects the hemostatic system and leads to deficiencies of both procoagulant and anticoagulant factors. Objective To assess the hemostatic balance in patients with multiple coagulation disorders by using a thrombin generation assay. Method We performed conventional coagulation assays and a thrombin generation assay on samples from patients with congenital disorder of glycosylation. The thrombin generation assay was performed before and after activation of the protein C system by the addition of soluble thrombomodulin. Results A total of 35 patients were included: 71% and 57% had low antithrombin and factor XI levels, respectively. Protein C and protein S levels were abnormally low in 29% and 26% of the patients, respectively, whereas only 11% displayed low factor IX levels. Under baseline conditions, the thrombin generation assay revealed a significantly higher endogenous thrombin potential and thrombin peak in patients, relative to controls. After spiking with thrombomodulin, we observed impaired involvement of the protein C system. Hence, 54% of patients displayed a hypercoagulant phenotype in vitro. All the patients with a history of stroke-like episodes or thrombosis displayed this hypercoagulant phenotype. Conclusion A thrombin generation assay revealed a hypercoagulant in vitro phenotype under baseline condition; this was accentuated by impaired involvement of the protein C system. This procoagulant phenotype may thus reflect the risk of severe vascular complications. Further research will have to determine whether the thrombin generation assay is predictive of vascular events.

Severe infectious diseases are often characterized by an overwhelming and unbalanced systemic immune response to microbial infections. Human antithrombin (hAT) is a crucial coagulation inhibitor with anti-inflammatory activities. Here we identify three hAT-binding proteins (CD13, CD300f and LRP-1) on human monocytes that are involved in blocking the activity of nuclear factor-κB. We found that the modulating effect is primarily restricted to the less abundant β-isoform (hβAT) of hAT that lacks N-glycosylation at position 135. Individuals with a mutation at this position have increased production of hβAT and analysis of their blood, which was stimulated ex vivo with lipopolysaccharide, showed a decreased inflammatory response. Similar findings were recorded when heterozygotic mice expressing hAT or hβAT were challenged with lipopolysaccharide or infected with Escherichia coli bacteria. Our results finally demonstrate that in a lethal E. coli infection model, survival rates increased when mice were treated with hβAT one hour and five hours after infection. The treatment also resulted in a reduction of the inflammatory response and less severe organ damage.

The Danish diet is characterized by a high content of sugar, fat dairy products and red meat, and a low content of fruits and vegetables. As it is considered unhealthy and environmentally unfriendly, various alternatives to the standard Danish diet have been investigated and promoted in Denmark, such as the New Nordic Diet. By using a Life Cycle Assessment (LCA), this study estimates the carbon footprint (CF) of four different diet scenarios in Denmark: standard, carnivore, vegetarian and vegan. The LCA is applied to build a dataset of the 47 most widely eaten food and beverage products, which represent the average Danish eating habits and grouped into six food categories. Unlike most past LCA-based studies, where system boundaries are limited to the farm gate, this study covers all activities and relative use of materials and energy, from the food production phase to the final consumption (namely ‘from-cradle-to-fork’). We find that the highest CF value is associated with the carnivore diet, which has the highest impact (1.83 t CO 2eq person −1 year −1). The vegan and vegetarian diets record the best profiles (0.89 and 1.37 t CO 2eq person −1 year −1, respectively), whereas the standard Danish diet has a CF value of 1.59 t CO 2eq person −1 year −1. We find that the food production phase is the most significant in terms of CF (65–85%). This study confirms that dietary preferences are a strong driver of CF. A comparison with CF associated with other diets suggests that a further research could provide a guidance to promote healthy eating patterns with adequate nutritional values and better environmental performances.

Cystidicola farionis is a nematode of the swim bladder that parasitizes Salmonidae and Osmeridae families, with a range of diffusion in Europe, Asia, and North America. During a fish health monitoring performed in July 2017 in a trout farm located in northwest Italy, a prevalence of 70% for C. farionis was recorded in rainbow trout from a unique tank. In order to clarify the first occurrence of this parasite in northwest Italy, a total of 180 specimens were analysed in different seasons to obtain prevalence, mean intensity, and mean abundance of infestation. Moreover, amphipods were also sampled during each fish sampling campaign. Only fish sampled in August 2017 were positive for C. farionis, since fish were then sold and replaced with a new batch. All isolated nematodes were identified by morphological criteria as subadults (L4), but not sexually matured. All examined amphipods (genus Gammarus) resulted negative for the presence of the parasite. The most probable explanation of this report was due to the movement of a single fish batch from northeast Italy, previously described as an endemic area for this parasite. The absence of the parasite in the amphipods and consequently in other fish batches from other tanks are due to the lifetime cycle of the parasite that had no time to infect new hosts.

Inherited antithrombin (AT) deficiency, with prevalence in the general population ranging 0.02-0.17%, is an autosomal dominant disorder associated with a high risk of venous thromboembolism. In most cases, deficiency is caused by mutations in the AT-coding gene (SERPINC1). Only 24 splicing defects have been described causing AT deficiency, all affecting exon flanking regions. The aim of the current study was to characterize the mutations underlying AT deficiency in 10 venous thromboembolism Polish patients aged 42.9 (14-63) years. Whole SERPINC1 gene sequencing was done by next generation sequencing methods. Eight cases had mutations previously described. However, we identified two new intronic mutations that might affect the correct splicing of exon 6 according to in-silico predictions: c.1154-1 G>C, which strongly disturbs the acceptor sequence and c.1219-534 A>G, a deep intronic mutation that might generate a cryptic donor sequence; both might compete with the wild-type donor sequence and explain the associated moderate AT deficiency of carriers. In conclusion, we show the molecular base of AT deficiency in 10 new Polish patients, including two novel SERPINC1 gene mutations potentially affecting splicing.

Background Persistent and/or recurrent inflammatory processes are the main factor leading to multiple sclerosis (MS) lesions. The composite ultramicronized palmitoylethanolamide, an endogenous N-acylethanolamine, combined with the flavonoid luteolin, PEALut, have been found to exert neuroprotective activities in experimental models of spinal and brain injury and Alzheimer disease, as well as a clinical improvement in human stroke patients. Furthermore, PEALut enhances the expression of different myelin proteins in oligodendrocyte progenitor cells suggesting that this composite might have protective effects in MS experimental models. Methods The mouse model of experimental autoimmune encephalomyelitis (EAE) based on active immunization with a fragment of myelin oligodendrocyte glycoprotein (MOG35-55) was used. The daily assessment of clinical score and the expression of serum amyloid A (SAA1), proinflammatory cytokines TNF-α, IL-1β, IFN-γ, and NLRP3 inflammasome, as well as TLR2, Fpr2, CD137, CD3-γ, and TCR-ζ chain, heterodimers that form T cell surface glycoprotein (TCR), and cannabinoid receptors CB1, CB2, and MBP, were evaluated in the brainstem and cerebellum at different postimmunization days (PIDs). Results Vehicle-MOG35-55-immunized (MOG35-55) mice developed ascending paralysis which peaked several days later and persisted until the end of the experiment. PEALut, given intraperitoneally daily starting on day 11 post-immunization, dose-dependently improved clinical score over the range 0.1–5 mg/kg. The mRNA expression of SAA1, TNF-α, IL-1β, IFN-γ, and NLRP3 were significantly increased in MOG35-55 mice at 14 PID. In MOG35-55 mice treated with 5 mg /kg PEALut, the increase of SAA1, TNF- α, IL-1β, and IFN-γ transcripts at 14 PID was statistically downregulated as compared to vehicle-MOG35-55 mice (p

AIM An acute phase response is induced after non-surgical periodontal treatment (SRP). The main aim of this study was to compare acute phase (24-hr) and medium-term (3-months) inflammation and clinical outcomes after SRP with or without application of enamel matrix derivative (EMD) in sites with probing pocket depth (PPD) ≥ 6 mm. METHODS Thirty-eight periodontitis-affected subjects were randomized to SRP or SRP + EMD. Periodontal parameters were recorded at baseline and 3 months. Serum samples were collected at baseline, 1 and 90 days after treatment. RESULTS Both treatments triggered an intense acute inflammation on day 1, which regressed to baseline values at 3 months. D-dimer and cystatin C levels did not show sharp increases in SRP + EMD group 24 hr after treatment, compared to SRP. Significant difference between groups was observed for D-dimer (p

Remyelination in patients with multiple sclerosis frequently fails, especially in the chronic phase of the disease promoting axonal and neuronal degeneration and progressive disease disability. Drug-based therapies able to promote endogenous remyelination capability of oligodendrocytes are thus emerging as primary approaches to multiple sclerosis. We have recently reported that the co-ultramicronized composite of palmitoylethanolamide and the flavonoid luteolin (PEALut) promotes oligodendrocyte precursor cell (OPC) maturation without affecting proliferation. Since TAM receptor signaling has been reported to be important modulator of oligodendrocyte survival, we here evaluated the eventual involvement of TAM receptors in PEALut-induced OPC maturation. The mRNAs related to TAM receptors -Tyro3, Axl, and Mertk- were all present at day 2 in vitro. However, while Tyro3 gene expression significantly increased upon cell differentiation, Axl and Mertk did not change during the first week in vitro. Tyro3 gene expression developmental pattern resembled that of MBP myelin protein. In OPCs treated with PEALut the developmental increase of Tyro3 mRNA was significantly higher as compared to vehicle while was reduced gene expression related to Axl and Mertk. Rapamycin, an inhibitor of mTOR, prevented oligodendrocyte growth differentiation and myelination. PEALut, administered to the cultures 30 min after rapamycin, prevented the alteration of mRNA basal expression of the TAM receptors as well as the expression of myelin proteins MBP and CNPase. Altogether, data obtained confirm that PEALut promotes oligodendrocyte differentiation as shown by the increase of MBP and CNPase and Tyro3 mRNAs as well as CNPase and Tyro3 immunostainings. The finding that these effects are reduced when OPCs are exposed to rapamycin suggests an involvement of mTOR signaling in PEALut effects.