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ABSTRACT: Objective, robust, and repeatable assessments of fear responses of poultry can lead to improvements in research techniques, the validity of test results, and ultimately bird welfare. The objective of this study was to examine the effect of 2 different holding methods on broiler fear responses during the inversion test, a standardized method of assessing fear in poultry. On D15 and D34 mixed-sex broilers (N = 80) were inverted and held either by their shanks (N = 40) or feet (N = 40) at arm's length away from the experimenter for 30 s. The frequency of wing flapping, head movement, attempts to right, and vocalizations were observed from video recordings collected during the inversion test. Vocalizations at D15 and D34 were not different when comparing holding methods. Frequency of attempts to right and wing flapping at D15 were greater (P < 0.001) for birds held by their feet (0.90 and 16.6, respectively) than when they were held by their shanks (0.20 and 0.73, respectively). Similarly, on D34 wing flapping remained greater (P < 0.001) for birds held by their feet (23.8) compared to their shanks (8.1). Conversely, head movements were greater (P = 0.05) for birds held by their shanks (9.2) compared to their feet (6.3) on D15. Within both the shank and feet holding method, vocalizations at D15 were greater (30.7 and 35.6, respectively) compared to D34 (11.93 and 15.38, respectively) (P < 0.001). There was no holding method effect on head movements within each age. These results suggest that behaviors observed during the inversion test can be influenced by the inversion holding method and that handling should be standardized while conducting behavioral tests to assess fear in poultry.

Ultrasound-assisted extraction of (poly)phenols from pumpkin pulp was optimized using response surface methodology. Time, solvent-to-sample ratio and percentage of water in ethanol were considered and a full factorial design was applied. The optimized conditions (10 min; 30 mL/g; 50% water in ethanol) were used to isolate (poly)phenols from five pumpkin varieties (Hokkaido, Lunga di Napoli, Mantovana, Moscata di Provenza, Violina rugosa), collected for two successive years. Phenolic acids were the most abundant phenolic class (gallic acid up to 413.50 μg/g), even if differences were observed among varieties and harvesting years. Generally, Violina rugosa harvested in 2021 showed the highest total phenol content (7.59 mg gallic acid equivalents/g) and antioxidant properties (16.93 mg Trolox equivalents/g). All pumpkin variety exhibited antimicrobial activity within a concentration range of 1.95 to 250 μg/mL. In conclusion, the pumpkin pulp can be considered a promising natural source of (poly)phenols for the development of health-promoting products.

Abstract Marine protected areas (MPAs) make an essential contribution to the spatial management of critical areas, the conservation of coastal species exploited by human activities, and the sustainable use of marine resources. Within MPAs, fishing closure areas are among the most used small‐scale fishery management tools, even though their effectiveness largely remains untested or controversial. To reduce the impact of small‐scale fisheries on marine resources, a seasonal fishing closure area (SFCA) was established beginning in 2022 in autumn–winter season inside the Capo Caccia–Isola Piana MPA (Sardinia, northwestern Mediterranean Sea). Here, we assessed a posteriori whether the areas of higher habitat suitability for eight species/taxa of relevant ecological value and economic interest to small‐scale fisheries were included in the established SFCA, adequately meeting the ecological objectives of the MPA. Thus, landing data (from 2019 to 2023) were used as occurrence records to develop MaxEnt distribution models for the eight target species/taxa. The model outputs allow us to draw important insights about the spatial adequacy of the SFCA established within the MPA aimed to protect the most exploited marine resources. Furthermore, the modeling exercises were useful for understanding the local processes influencing species' habitat selection and to identify essential areas for the target species that could remain unrevealed in larger‐scale investigations.

Abstract While cardiovascular disease (CVD) is one of the major co-morbidities in patients with rheumatoid arthritis (RA), the mechanism(s) that contribute to CVD in patients with RA remain to be fully elucidated. Herein, we observe that plasma concentrations of 13-series resolvin (RvT)4 negatively correlate with vascular lipid load in mouse inflammatory arthritis. Administration of RvT4 to male arthritic mice fed an atherogenic diet significantly reduces atherosclerosis. Assessment of the mechanisms elicited by this mediator demonstrates that RvT4 activates cholesterol efflux in lipid laden macrophages via a Scavenger Receptor class B type 1 (SR-BI)-Neutral Cholesterol Ester Hydrolase-dependent pathway. This leads to the reprogramming of lipid laden macrophages yielding tissue protection. Pharmacological inhibition or knockdown of macrophage SR-BI reverses the vasculo-protective activities of RvT4 in vitro and in male mice in vivo. Together these findings elucidate a RvT4-SR-BI centered mechanism that orchestrates macrophage responses to limit atherosclerosis during inflammatory arthritis.

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Aims: The cardio-protective and immuno-regulatory properties of RTP-026, a synthetic peptide that spans the Annexin-A1 (AnxA1) N-terminal region, were tested in rat acute myocardial infarction. Methods and results: In vitro, selective activation of formyl-peptide receptor type 2 (FPR2) by RTP-026 occurred with apparent EC50 in the 10–30 nM range. With human primary cells, RTP-026 counteracted extension of neutrophil life-span and augmented phagocytosis of fluorescent E.coli by blood myeloid cells. An in vivo model of rat acute infarction was used to quantify tissue injury and phenotype immune cells in myocardium and blood. The rat left anterior descending coronary artery was occluded and then reopened for 2-hour or 24-hour reperfusion. For the 2-hour reperfusion protocol, RTP-026 (25–500 µg/kg; given i.v. at the start of reperfusion) significantly reduced infarct size by ∼50 %, with maximal efficacy at 50 µg/kg. Analyses of cardiac immune cells showed that RTP-026 reduced neutrophil and classical monocyte recruitment to the damaged heart. In the blood, RTP-026 (50 µg/kg) attenuated activation of neutrophils and monocytes monitored through CD62L and CD54 expression. Modulation of vascular inflammation by RTP-026 was demonstrated by reduction in plasma levels of mediators like TNF-α, IL-1β, KC, PGE2 and PGF2α⊡ For the 24-hour reperfusion protocol, RTP-026 (30 µg/kg given i.v. at 0, 3 and 6 h reperfusion) reduced necrotic myocardium by ∼40 %. Conclusions: RTP-026 modulate immune cell responses and decreases infarct size of the heart in preclinical settings. Tempering over-exuberant immune cell activation by RTP-026 is a suitable approach to translate the biology of AnxA1 for therapeutic purposes.

Introdução: Na doença falciforme (DF), uma mutação pontual no gene da beta-globina resulta na produção de hemoglobina S (HbS), que pode polimerizar-se após desoxigenação. A polimerização distorce a membrana dos glóbulos vermelhos e gera eritrócitos falciformes, contribuindo para oclusões microvasculares que podem manifestar-se através de crises vaso-oclusivas agudas e dolorosas (CVOs). Para pacientes (pts) com DF 70% das visitas às emergências e 90% das hospitalizações estão relacionadas a CVOs. Apesar de tratamentos existentes para prevenção de CVO, morbidade e mortalidade permanecem consideráveis; o tratamento da CVO aguda é limitado a cuidados de suporte. Dados não clínicos relacionam a desregulação do complemento na fisiopatologia da DF, incluindo vasco-oclusão, hemólise, inflamação, trombogenicidade, ativação endotelial e lesões de órgãos. Crovalimabe (Crova) é um anticorpo monoclonal anti complemento C5 que permite adm SC de baixo volume após dose IV inicial. Descrevemos 2 estudos randomizados, controlados por placebo avaliando Crova em pts com DF: CROSSWALK-a (fase 1b; NCT04912869) e CROSSWALK-c (fase 2a; NCT05075824) com centros abertos atualmente no Brasil. Objetivos: CROSSWALK-a avalia segurança de Crova no manejo de CVOs agudas não-complicadas. CROSSWALK-c avalia eficácia e segurança de Crova como terapia adjuvante na prevenção de CVOs. Métodos: Pts 12-55 anos, pesando ≥40 kg, diagnosticados com DF (HbSS homozigótica ou talassemia heterozigótica HbS/β0), vacinados contra N. meningitidis, H. influenzae tipo B e S. pneumoniae são elegíveis. Terapias concomitantes para DF são permitidas. CROSSWALK-a: Pts com CVO aguda não complicada necessitando hospitalização e opióides parenterais, histórico de ≤10 CVOs nos 12 meses pré-randomização. Pts elegíveis serão randomizados 2:1 para dose única IV peso-dependente de Crova ou placebo, monitorados na hospitalização e durante observação por 84 dias, seguido de acompanhamento de segurança por 322 dias (10,5 meses). O objetivo princial do CROSSWALK-a é avaliar a segurança até Dia 84, incluindo: Incidência e gravidade de eventos adversos, Incidência e gravidade de reações relacionadas à infusão e hipersensibilidade, Alterações em sinais vitais e resultados de exames laboratoriais específicos. Serão avaliados também eficácia, farmacocinética (PK), farmacodinâmica (PD), imunogenicidade e biomarcador exploratório. CROSSWALK-c: São elegíveis Pts com 2-10 CVOs nos 12 meses pré-randomização e serão randomizados 1:1 para receber doses diferenciadas, peso-dependentes de Crova ou placebo, consistindo em doses iniciais (IV: Dia 1; SC: Dia 2 e semanalmente nas Semanas 2-4) e doses mensais de manutenção SC nas Semanas 5-49 em 48 semanas de tratamento. O objetivo principal do CROSSWALK-c é avaliar eficácia de Crova vs placebo até 48 semanas, com base na Taxa anual de CVOs manejadas em ambientes hospitalares. Principais objetivos secundários de eficácia são: Taxas anuais de CVOs domiciliares, síndrome torácica aguda, dias de hospitalização para manejo de CVOs, Fadiga relatada por pacientes adultos. Parâmetros de segurança, farmacocinética, farmacodinâmica, imunogenicidade e biomarcador exploratório também serão avaliados. Resultados iniciais serão publicados após o readout do estudo.

Background Sustainable management requires spatial mapping of both species distribution and human activities to identify potential risk of conflict. The common bottlenose dolphin (Tursiops truncatus) is a priority species of the European Union Habitat Directive, thus, to promote its conservation, the understanding of habitat use and distribution, as well as the identification and spatial trend of the human activities which may directly affect populations traits, is pivotal. Methods A MaxEnt modeling approach was applied to predict the seasonal (from April to September) habitat use of a small population of bottlenose dolphins in the north-western Sardinia (Mediterranean Sea) in relation to environmental variables and the likelihoods of boat and fishing net presence. Then, the overlapping areas between dolphin, fishing net and boat presence were identified to provide insights for the marine spatial management of this area. Results Three of the main factors influencing the seasonal distribution of bottlenose dolphins in the area are directly (boating and fishing) or indirectly (ocean warming) related to human activities. Furthermore, almost half of the most suitable area for dolphins overlapped with areas used by fishing and boating. Finally, relying on fishing distribution models, we also shed light on the potential impact of fishing on the Posidonia oceanica beds, a protected habitat, which received higher fishing efforts than other habitat types. Discussion Modelling the spatial patterns of anthropogenic activities was fundamental to understand the ecological impacts both on cetacean habitat use and protected habitats. A greater research effort is suggested to detect potential changes in dolphin habitat suitability, also in relation to ocean warming, to assess dolphin bycatch and the status of target fish species, and to evaluate sensitive habitats conditions, such as the Posidonia oceanica meadow.

Background: Many genes associated with asthma explain only a fraction of its heritability. Most genome-wide association studies (GWASs) used a broad definition of ‘doctor-diagnosed asthma’, thereby diluting genetic signals by not considering asthma heterogeneity. The objective of our study was to identify genetic associates of childhood wheezing phenotypes. Methods: We conducted a novel multivariate GWAS meta-analysis of wheezing phenotypes jointly derived using unbiased analysis of data collected from birth to 18 years in 9568 individuals from five UK birth cohorts. Results: Forty-four independent SNPs were associated with early-onset persistent, 25 with pre-school remitting, 33 with mid-childhood remitting, and 32 with late-onset wheeze. We identified a novel locus on chr9q21.13 (close to annexin 1 [ANXA1], p

Using a global formyl-peptide receptor (Fpr)2 knockout mouse colony, we have reported the modulatory properties of this pro-resolving receptor in polymicrobial sepsis. Herein, we have used a humanized FPR2 (hFPR2) mouse colony bearing an intact or a selective receptor deficiency in myeloid cells to dwell on the cellular mechanisms. hFPR2 mice and myeloid cell-specific hFPR2 KO (abbreviated to KO) mice were subjected to cecal ligation and puncture (CLP)-induced polymicrobial sepsis. Compared with hFPR2 mice, CLP caused exacerbated cardiac dysfunction (assessed by echocardiography), worsened clinical outcome and impaired bacteria clearance in KO mice. This pathological scenario was paralleled by increased recruitment of pro-inflammatory monocytes and reduced M2-like macrophages within the KO hearts. In peritoneal exudates of KO mice, we quantified increased neutrophil and MHC II+ macrophage numbers, but decreased monocyte/macrophage and MHC II- macrophage recruitment. hFPR2 up-regulation was absent in myeloid cells and local production of lipoxin A4 was reduced in septic KO mice. Administration of the FPR2 agonist Annexin A1 (AnxA1) improved cardiac function in hFPR2 septic mice, but had limited beneficial effects in KO mice, in which the FPR2 ligand failed to polarize macrophages towards an MHC II- phenotype. In conclusion, FPR2 deficiency in myeloid cells exacerbates cardiac dysfunction and worsens clinical outcome in polymicrobial sepsis. The improvement of cardiac function and the host immune response by AnxA1 is more effective in hFPR2 competent septic mice.

Abstract One of the most studied aspects of animal communication is the acoustic repertoire difference between populations of the same species. While numerous studies have investigated the variability of bottlenose dolphin whistles between populations, very few studies have focused on the signature whistles alone and the factors underlying differentiation of signature whistles are still poorly understood. Here we describe the signature whistles produced by six distinct geographical units of the common bottlenose dolphin (Tursiops truncatus) in the Mediterranean Sea and identify the main determinants of their variability. Particularly, the influence of the region (proxy of genetic distance), the geographic site, and the environmental (sea bottom-related) and demographical (population-related) conditions on the acoustic structure of signature whistles was evaluated. The study provides the first evidence that the genetic structure, which distinguishes the eastern and western Mediterranean bottlenose dolphin populations has no strong influence on the acoustic structure of their signature whistles, and that the geographical isolation between populations only partially affected whistle variability. The environmental conditions of the areas where the whistles developed and the demographic characteristics of the belonging populations strongly influenced signature whistles, in accordance with the “acoustic adaptation hypothesis” and the theory of signature whistle determination mediated by learning.

Background and Purpose: Spinal cord (SC) cross‐sectional areas (CSAs) assessed with MRI have proven to be extremely valuable imaging markers in several diseases. Among the challenges is the delineation of vertebral levels to determine level‐dependent changes in cord atrophy. With this study, we aimed to (1) test the hypothesis that there is proportionality in the position of the first six intervertebral discs and the length of the upper portion of the SC and (2) show that a proportionality approach can simplify the CSA assessment across vertebrae offering good reliability. Methods: Forty‐six volunteers underwent standard T2‐weighted and T1‐weighted cervical SC MRI acquisitions. The distance between the obex and the intervertebral discs (from C2‐C3 to T1‐T2) was measured on the T2‐weighted acquisitions of the entire cohort. In a test‐retest experiment on 12 subjects, the % disc position values were used to define vertebral levels, and a comparison was performed with manual vertebrae assignment in terms of mean CSA and its coefficient of variation. Results: The mean upper cord length for the cohort was 144.0 ± 13.1 mm. The discs' level % position in the upper cord was found to be fairly consistent, with standard deviations of 0.8%‐1.7%. The mean vertebral CSA obtained with the proportionality method was substantially equivalent to the manual approach in terms of mean CSA values and test‐retest reliability. Conclusions: With this study, we propose a proportionality method for the assignment of cervical SC vertebral levels that can simplify the processing of MRI datasets in the context of CSA measurements. [ABSTRACT FROM AUTHOR]

Marine protected areas (MPAs) make an essential contribution to the spatial management of critical areas, the conservation of coastal species exploited by human activities, and the sustainable use of marine resources. Within MPAs, fishing closure areas are among the most used small‐scale fishery management tools, even though their effectiveness largely remains untested or controversial. To reduce the impact of small‐scale fisheries on marine resources, a seasonal fishing closure area (SFCA) was established beginning in 2022 in autumn–winter season inside the Capo Caccia–Isola Piana MPA (Sardinia, northwestern Mediterranean Sea). Here, we assessed a posteriori whether the areas of higher habitat suitability for eight species/taxa of relevant ecological value and economic interest to small‐scale fisheries were included in the established SFCA, adequately meeting the ecological objectives of the MPA. Thus, landing data (from 2019 to 2023) were used as occurrence records to develop MaxEnt distribution models for the eight target species/taxa. The model outputs allow us to draw important insights about the spatial adequacy of the SFCA established within the MPA aimed to protect the most exploited marine resources. Furthermore, the modeling exercises were useful for understanding the local processes influencing species' habitat selection and to identify essential areas for the target species that could remain unrevealed in larger‐scale investigations. [ABSTRACT FROM AUTHOR]

BackgroundMelanocortins are peptides endowed with anti-inflammatory and pro-resolving activities. Many of these effects are mediated by the Melanocortin receptor 1 (MC1) as reported in several experimental settings. As such, MC1 can be a viable target for the development of new therapies that mimic endogenous pro-resolving mediators. The aim of this study was to assess the immunopharmacology of a selective MC1 agonist (PL8177) in vitro and in a mouse model of inflammatory arthritis.MethodsPL8177 and the natural agonist αMSH were tested for activation of mouse and human Melanocortin receptors (MC1,3,4,5), monitoring cAMP accumulation and ERK1/2 phosphorylation, using transiently transfected HEK293A cells. The anti-inflammatory and pro-resolving effects of PL8177 and αMSH were evaluated using mouse peritoneal Macrophages. Finally, a model of K/BxN serum transfer induced arthritis was used to determine the in vivo potential of PL8177.ResultsPL8177 activates mouse and human MC1 with apparent EC50 values of 0.01 and 1.49 nM, respectively, using the cAMP accumulation assay. Similar profiles were observed for the induction of ERK phosphorylation (EC50: 0.05 and 1.39 nM). PL8177 displays pro-resolving activity (enhanced Macrophage efferocytosis) and counteracts the inflammatory profile of zymosan-stimulated macrophages, reducing the release of IL-1β, IL-6, TNF-α and CCL-2. In the context of joint inflammation, PL8177 (3mg/kg i.p.) reduces clinical score, paw swelling and incidence of severe disease as well as the recruitment of immune cells into the arthritic joint.ConclusionThese results demonstrate that the MC1 agonism with PL8177 affords therapeutic effects in inflammatory conditions including arthritis.SignificanceDrugs targeting the Melanocortin system have emerged as promising therapeutics for several conditions including inflammation or obesity. Multiple candidates are under clinical development, and some have already reached approval. Here we present the characterization of a novel drug candidate, PL8177, selective for the Melanocortin 1 receptor (MC1), demonstrating its selectivity profile on cAMP and ERK1/2 phosphorylation signaling pathways, of relevance as selective drugs will translate into lesser off-target effect. PL8177 also demonstrated, not only anti-inflammatory activity, but pro-resolving actions due to its ability to enhance efferocytosis (i.e. the phagocytosis of apoptotic cells), endowing this molecule with therapeutic advantages compared to classical anti-inflammatory drugs. Using a mouse model of inflammatory arthritis, the compound demonstrated in vivo efficacy by reducing clinical score, paw swelling and overall disease severity. Taken together, these results present Melanocortin-based therapies, and specifically targeting MC1 receptor, as a promising strategy to manage chronic inflammatory diseases.