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Circadian rhythms have been shown in the subthalamic nucleus (STN) in Parkinsons disease (PD), but only a few studies have focused on the globus pallidus internus (GPi). This retrospective study investigates GPi circadian rhythms in a large cohort of subjects with PD (130 recordings from 93 subjects) with GPi activity chronically recorded in their home environment. We found a significant change in GPi activity between daytime and nighttime in most subjects (82.4%), with a reduction in GPi activity at nighttime in 56.2% of recordings and an increase in activity in 26.2%. GPi activity in higher frequency bands ( > 20 Hz) was more likely to decrease at night and in patients taking extended-release levodopa medication. Our results suggest that circadian fluctuations in the GPi vary across individuals and that increased power at night might be due to the reemergence of pathological neural activity. These findings should be considered to ensure successful implementation of adaptive neurostimulation paradigms in the real-world.

Abstract Neutrophils (PMNs) are key players of innate immune responses through the release of cytoplasmic granule content and the formation of neutrophil extracellular traps (NETs). RNASET2 is an acidic ribonuclease, recently proposed as an alarmin signal associated with inflammatory responses. Here we show that, along the neutrophil maturation cascade, RNASET2 is expressed in segmented and mature PMNs. In human PMNs, RNASET2 colocalized with primary and tertiary granules and was found to be associated with NETs following PMA or Nigericin stimulation. Similarly, activation of PMNs by soluble immune complexes, a hallmark of several autoimmune diseases, also induced RNASET2-associated NETs. Genome-wide association studies recently identified RNASET2 among a cluster of genes associated with increased susceptibility to develop autoimmune diseases, including rheumatoid arthritis (RA). RNASET2 was found expressed by PMNs and macrophages infiltrating inflamed joints in a murine model of RA (K/BxN Serum-Transfer-Induced Arthritis, STIA), by immunostaining. Similar results were found in synovial biopsies of RA patients with active disease. In addition, we demonstrate that RNASET2 circulating levels correlated with the onset and the severity of disease in two mouse models of inflammatory arthritis, STIA and CIA (Collagen-Induced Arthritis) and in serum of RA patients. These results show that PMNs are an important source of RNASET2 and that its circulating levels are associated with RA development suggesting a role for RNASET2 in the pathogenesis of immune-mediated diseases.

Abstract Background The physiological relevance of cell-to-cell communication mediated by small extracellular vesicle-encapsulated microRNAs (sEV-miRNAs) remains debated because of the limiting representativity of specific miRNAs within the extracellular pool. We hypothesize that sEV-miRNA non-canonical function consisting of the stimulation of Toll-like receptor 7 (TLR7) may rely on a global shift of the sEV cargo rather than on the induction of one or few specific miRNAs. Psoriasis represents an ideal model to test such hypothesis as it is driven by overt activation of TLR7-expressing plasmacytoid dendritic cells (pDCs) following keratinocyte damage. Methods To mimic the onset of psoriasis, keratinocytes were treated with a cocktail of psoriatic cytokines or UV-irradiated. SmallRNA sequencing was performed on sEVs released by healthy and UV-treated keratinocytes. sEV-miRNAs were analyzed for nucleotide composition as well as for the presence of putative TLR7-binding triplets. Primary human pDCs where stimulated with sEVs +/- inhibitors of TLR7 (Enpatoran), of sEV release (GW4869 + manumycin) and of TLR7-mediated pDC activation (anti-BDCA-2 antibody). Secretion of type I IFNs and activation of CD8+T cells were used as readouts. qPCR on psoriatic and healthy skin biopsies was conducted to identify induced miRNAs. Results sEV-miRNAs released by damaged keratinocytes revealed a significantly higher content of TLR7-activating sequences than healthy cells. As expected, differential expression analysis confirmed the presence of miRNAs upregulated in psoriatic skin, including miR203a. More importantly, 76.5% of induced miRNAs possessed TLR7-binding features and among these we could detect several previously demonstrated TLR7 ligands. In accordance with this in silico analysis, sEVs from damaged keratinocytes recapitulated key events of psoriatic pathogenesis by triggering pDCs to release type I interferon and activate cytotoxic CD8+T cells in a TLR7- and sEV-dependent manner. Discussion Our results demonstrate that miR203a is just one paradigmatic TLR7-activating miRNA among the hundreds released by UV-irradiated keratinocytes, which altogether trigger pDC activation in psoriatic conditions. This represents the first evidence that cell damage shifts the miRNA content of sEVs towards a TLR7-activating cargo capable to propagate inflammation and immunity, offering strong support to the physiological role of systemic miRNA-based cell-to-cell communication.

Abstract Background Prevention and treatment of metastatic breast cancer (BC) is an unmet clinical need. The retinoic acid derivative fenretinide (FeR) was previously evaluated in Phase I-III clinical trials but, despite its excellent tolerability and antitumor activity in preclinical models, showed limited therapeutic efficacy due to poor bioavailability. We recently generated a new micellar formulation of FeR, Bionanofenretinide (Bio-nFeR) showing enhanced bioavailability, low toxicity, and strong antitumor efficacy on human lung cancer, colorectal cancer, and melanoma xenografts. In the present study, we tested the effect of Bio-nFeR on a preclinical model of metastatic BC. Methods We used BC cell lines for in vitro analyses of cell viability, cell cycle and migratory capacity. For in vivo studies, we used HER2/neu transgenic mice (neuT) as a model of spontaneously metastatic BC. Mice were treated orally with Bio-nFeR and at sacrifice primary and metastatic breast tumors were analyzed by histology and immunohistochemistry. Molecular pathways activated in primary tumors were analyzed by immunoblotting. Stem cell content was assessed by flow cytometry, immunoblotting and functional assays such as colony formation ex vivo and second transplantation assay in immunocompromised mice. Results Bio-nFeR inhibited the proliferation and migration of neuT BC cells in vitro and showed significant efficacy against BC onset in neuT mice. Importantly, Bio-nFeR showed the highest effectiveness against metastatic progression, counteracting both metastasis initiation and expansion. The main mechanism of Bio-nFeR action consists of promoting tumor dormancy through a combined induction of antiproliferative signals and inhibition of the mTOR pathway. Conclusion The high effectiveness of Bio-nFeR in the neuT model of mammary carcinogenesis, coupled with its low toxicity, indicates this formulation as a potential candidate for the treatment of metastatic BC and for the adjuvant therapy of BC patients at high risk of developing metastasis.

Abstract Extracellular vesicles (EVs) are nanosized heat-stable vesicles released by virtually all cells in the body, including tumor cells and tumor-infiltrating dendritic cells (DCs). By carrying molecules from originating cells, EVs work as cell-to-cell communicators in both homeostasis and cancer but may also represent valuable therapeutic and diagnostic tools. This review focuses on the role of tumor-derived EVs (TEVs) in the modulation of DC functions and on the therapeutic potential of both tumor- and DC-derived EVs in the context of immunotherapy and DC-based vaccine design. TEVs were originally characterized for their capability to transfer tumor antigens to DCs but are currently regarded as mainly immunosuppressive because of the expression of DC-inhibiting molecules such as PD-L1, HLA-G, PGE2 and others. However, TEVs may still represent a privileged system to deliver antigenic material to DCs upon appropriate engineering to reduce their immunosuppressive cargo or increase immunogenicity. DC-derived EVs are more promising than tumor-derived EVs since they expose antigen-loaded MHC, costimulatory molecules and NK cell-activating ligands in the absence of an immunosuppressive cargo. Moreover, DC-derived EVs possess several advantages as compared to cell-based drugs such as a higher antigen/MHC concentration and ease of manipulation and a lower sensitivity to immunosuppressive microenvironments. Preclinical models showed that DC-derived EVs efficiently activate tumor-specific NK and T cell responses either directly or indirectly by transferring antigens to tumor-infiltrating DCs. By contrast, however, phase I and II trials showed a limited clinical efficacy of EV-based anticancer vaccines. We discuss that the future of EV-based therapy depends on our capability to overcome major challenges such as a still incomplete understanding of their biology and pharmacokinetic and the lack of standardized methods for high-throughput isolation and purification. Despite this, EVs remain in the limelight as candidates for cancer immunotherapy which may outmatch cell-based strategies in the fullness of their time.

New Italian data on the distribution of the Annex I Habitats are reported in this contribution. Specifically, 9 new occurrences in Natura 2000 sites are presented and 34 new cells are added in the EEA 10 km × 10 km reference grid. The new data refer to the Italian administrative regions of Abruzzo, Apulia, Calabria, Latium, Lombardy, Marche, Sardinia, Sicily, Tuscany and Umbria.

We studied the vascular flora of the isthmus of Feniglia, a Nature Reserve in southern Tuscany stretching between the Italian Peninsula and Mt. Argentario. Since the nineteenth century, the area has undergone significant environmental modifications due to intensive grazing and deforestation. Later, rehabilitation interventions were carried out, including reforestation and planting of dune species, making it an interesting protected area to study for its botanical aspects. Therefore, we aim to compile the floristic inventory of this distinctive Mediterranean area by integrating old and new data. The checklist comprises 502 specific and subspecific taxa of vascular plants. The life-form spectrum shows a predominance of therophytes, followed by hemicryptophytes. The chorological spectrum highlights the dominance of Mediterranean species, followed by Euromediterranean and Eurasian species. The presence of 15 species of regional importance, 3 species of the Italian red list and 3 Italian endemics (Ornithogalum exscapum Ten., Limonium multiforme Pignatti, Linaria purpurea (L.) Mill.) is noteworthy. Despite the environmental changes undergone in this area, our study reports the presence of remarkable species, including rare ones such as Atriplex littoralis L., Ruppia spiralis L. ex Dumort., and species at their distribution margin in the Italian Peninsula, such as Staphisagria macrosperma Spach, Juncus sorrentinoi Parl. and Maresia nana (DC.) Batt. Additionally the presence of 26 alien species, of which 15 are invasive, 9 naturalized and 2 casual, indicates a relatively low presence of alien species. Notwithstanding the significant environmental changes that the Feniglia isthmus has experienced in the last century and the high tourist pressure during the summer, the presence of the protected area suggested the effectiveness and benefits of such a form of protection for plant diversity.

Camera trap studies on ocelots in Mexico have focused on abundance, habitat use, and activity patterns, mainly within Protected Areas of tropical humid forests in southern regions. However, their ecology, including breeding effort and productivity in seasonal dry Pacific forests and areas outside Protected Areas, has received less attention. The species’ low reproductive rates make their populations vulnerable to decline. In evaluating the reproduction, the species’ breeding activity was examined for two survey periods performed during 2010–2016. We estimated the percentage of females raising young in the surveys and their associated fecundity–”productivity”. The overall results revealed a population of 26 males and 46 females that included 10 females (21%) rearing 12 young, defined as kittens, cubs, or juveniles, for an effective sample area (ESA) of 200 km2. Overall, the productivity averaged 2.05/100 km2 per season. All parameters varied between the two areas, and productivity was not constant in time in either area or site. Information on species breeding activity in combination with density estimations may facilitate determining the minimum area required for a viable ocelot population in the region. Protected Areas are required in the studied region for the ocelot in order to maintain a viable local population. This is due to the increasing fragmentation caused by agriculture, cattle ranching, and forest fires, and it is hypothesized that ocelots do not tolerate disturbance or severe fragmentation.

Abstract The COVID-19 pandemic caused by SARS-CoV-2 has highlighted the urgent need for innovative antiviral strategies to fight viral infections. Although a substantial part of the overall effort has been directed at the Spike protein to create an effective global vaccination strategy, other proteins have also been examined and identified as possible therapeutic targets. Among them, although initially underestimated, there is the SARS-CoV-2 E-protein, which turned out to be a key factor in viral pathogenesis due to its role in virus budding, assembly and spreading. The C-terminus of E-protein contains a PDZ-binding motif (PBM) that plays a key role in SARS-CoV-2 virulence as it is recognized and bound by the PDZ2 domain of the human tight junction protein ZO-1. The binding between the PDZ2 domain of ZO-1 and the C-terminal portion of SARS-CoV-2 E-protein has been extensively characterized. Our results prompted us to develop a possible adjuvant therapeutic strategy aimed at slowing down or inhibiting virus-mediated pathogenesis. Such innovation consists in the design and synthesis of externally PDZ2-ZO1 functionalized PLGA-based nanoparticles to be used as intracellular decoy. Contrary to conventional strategies, this innovative approach aims to capitalize on the E protein-PDZ2 interaction to prevent virus assembly and replication. In fact, the conjugation of the PDZ2 domain to polymeric nanoparticles increases the affinity toward the E protein effectively creating a “molecular sponge” able to sequester E proteins within the intracellular environment of infected cells. Our in vitro studies on selected cellular models, show that these nanodevices significantly reduce SARS-CoV-2-mediated virulence, emphasizing the importance of exploiting viral-host interactions for therapeutic benefit.

In this contribution, new data concerning algae, bryophytes, fungi and lichens of the Italian flora are presented. It includes new records and confirmations for the algal genera Chara and Nitella, the bryophyte genera Brachythecium, Didymodon, Fissidens, Physcomitrium, and Riccia, the fungal genera Biatoropsis, Cantharellus, Coprinellus, Dacrymyces, Inosperma, Nigropuncta, Urocystis, and Xanthoriicola, and the lichen genera Arthonia, Bellemerea, Circinaria, Lecania, Lecanora, Lecidella, Mycobilimbia, Naetrocymbe, Parmelia, Peltigera, Porpidia, Scytinium, and Usnea.

Undergraduates' first-year experience in engineering education impacts their retention and persistence. Strategic learning strategies (i.e. skill, will, and self-regulation) are important for students' academic achievement and retention. We examined to what extent (1) students' skill mediated the relationship between self-regulation and will and (2) the constructivist practices in the learning environment related to students' self-regulation. Data were collected from 293 engineering undergraduates in a redesigned first-year experience course. Path analysis showed that (1) skill mediated the relationship between self-regulation and will and (2) one dimension of the constructivist learning environment (i.e. sharing ideas with others) was correlated with self-regulation. This study helped faculty and researchers to better understand how to use integrated strategic learning strategies and create a constructivist learning environment for engineering students' academic success.

Effective conservation planning requires identifying priority hotspots to allocate resources. To preserve biodiversity, it is crucial to consider α, β and γ-diversity and protect the irreplaceable sites with high ecological uniqueness, which can host uncommon species assemblages that would be lost if only species-rich sites were protected. Coastal dunes, hosting highly specialized plant communities, are among the most threatened ecosystems worldwide. In this study, we identified conservation priority hotspots to assess the effectiveness of the network of protected areas in coastal dunes of Tuscany (central Italy), using data on plant communities collected in 506 plots. We additively partitioned γ-diversity in its α and β components, observing a significant variation at all spatial levels only for dune species. In terms of α-diversity, we found that Northern protected sites were richer in dune species, while synanthropic and alien species were equally present inside and outside protected areas of the region. By partitioning the total β-diversity into its components (replacement and richness difference), we found a prevalence of replacement for dune species, indicating the most unique sites as the ones to favor for conservation. Unique sites were identified through Local Contributions to Beta Diversity and their conservation value was determined by their species composition and the relationship with landscape variables. Unique sites with high conservation value were only partly protected, while some protected sites were altered and required restoration. Our approach proved effective for identifying the most unique sites, indicating some issues in the existing protected network, while providing valuable information on sites to prioritize for future conservation actions.

Transcatheter aortic valve replacement (TAVR) has undergone rapid expansion, emerging as a viable therapeutic option for low-risk patients in lieu of surgical aortic valve replacement. This paper aims to provide a review of the scientific evidence concerning TAVR in low-risk patients, encompassing both observational and clinical trial data. Furthermore, a substantial proportion of low-risk patients possesses a bicuspid aortic valve, necessitating careful examination of the pertinent anatomic and clinical considerations to TAVR that is highlighted in this review. Additionally, the review expands upon some of the unique challenges associated with alternate access in low-risk patients evaluated for TAVR. Last, this review outlines the pivotal role of a multidisciplinary heart team approach in the execution of all TAVR procedures and the authors’ vision of ‘minimalist TAVR’ as a new era in low-risk TAVR.

This 2-year pilot study aimed to investigate the effects of a job coaching application (i.e., Job Coaching on Demand) on the knowledge and competencies of teachers and job coaches that work with young adults with developmental disabilities. Due to the global pandemic, this research was conducted virtually over a 2-year period. During the first year, a one-group formative pretest/posttest design with key informants was used to pilot the app with three teachers and five job coaches. The anecdotal findings from this pilot year were then used to further develop the application. The intent of the application was to provide professional development support to teachers and job coaches across the categories of (a) exploring careers, (b) ready for work, (c) part of the team, and (d) advocacy. In Year 2, a total of 31 teachers and job coaches were recruited, and a pretest/posttest was conducted to compare the differences in gained competencies between the two groups. The results show that those in the treatment group showed a larger increase in gains compared to the control group. The overall results of the 2 years of data, the current application development, and practical implications of the technology are further discussed.

This research investigates the linkage between training evaluation, learning design and training transfer. A new training evaluation model, (i.e., learning-transfer evaluation model [LTEM]), was used to examine its ability to provide evaluative evidence through robust assessments in pre-, post- and delayed assessments. The model was used to improve the training design of a nursing training programme and to nudge the stakeholders to change their training practices. A mixed-methods research approach, called convergent parallel, was employed to study the influence of the LTEM model in training design improvement by assessing the perceptions of nurse trainees, trainers and a nurse educator towards the instructional design of the training programme. A quantitative approach was used to examine the extent to which the improved programme using an iterative design cycle would result in improved performance among four groups (i.e., one baseline and three treatment groups). The integrated data showed that the LTEM model influenced the stakeholders to focus more on skills practice. With this shift of mindset, the training design was enhanced based on the principles of learning design, cognitive science and multimedia principles. Results of the pre-, post- and delayed posttests showed higher mean scores in the three treatment groups compared to the baseline group. For the measure of transfer, while the treatment groups scored higher, there was no statistically significant difference among all four groups. This could be attributed to nurses' varied levels of experience, negative transfer and the use of different evaluation matrices in the study and by the hospital.