Cancer EV stimulate endothelial glycolysis to fuel protein synthesis via mTOR and AMPKα activation.

Hypoxia is a common feature of solid tumours and activates adaptation mechanisms in cancer cells that induce therapy resistance and has profound effects on cellular metabolism. As such, hypoxia is an important contributor to cancer progression and is associated with a poor prognosis. Metabolic alterations in cells within the tumour microenvironment support tumour growth via, amongst others, the suppression of immune reactions and the induction of angiogenesis. Recently, extracellular vesicles (EV) have emerged as important mediators of intercellular communication in support of cancer progression. Previously, we demonstrated the pro-angiogenic properties of hypoxic cancer cell derived EV. In this study, we investigate how (hypoxic) cancer cell derived EV mediate their effects. We demonstrate that cancer derived EV regulate cellular metabolism and protein synthesis in acceptor cells through increased activation of mTOR and AMPKα. Using metabolic tracer experiments, we demonstrate that EV stimulate glucose uptake in endothelial cells to fuel amino acid synthesis and stimulate amino acid uptake to increase protein synthesis. Despite alterations in cargo, we show that the effect of cancer derived EV on recipient cells is primarily determined by the EV producing cancer cell type rather than its oxygenation status.

Peripheral blood immune parameters, response, and adverse events after neoadjuvant chemotherapy plus durvalumab in early-stage triple-negative breast cancer.

PURPOSE: We evaluated T- and B-cell receptor (TCR and BCR) repertoire diversity and 38 serum cytokines in pre- and post-treatment peripheral blood of 66 patients with triple-negative breast cancer (TNBC) who received neoadjuvant chemotherapy plus durvalumab and assessed associations with pathologic response and immune-related adverse events (irAEs) during treatment. METHODS: Genomic DNA was isolated from buffy coat for TCR and BCR clonotype profiling using the Immunoseq platform and diversity was quantified with Pielou's evenness index. MILLIPLEX MAP Human Cytokine/Chemokine Magnetic Bead Panel was used to measure serum cytokine levels, which were compared between groups using moderated t-statistic with Benjamini-Hochberg correction for multiple testing. RESULTS: TCR and BCR diversity was high (Pielou's index > 0.75) in all samples. Baseline receptor diversities and change in diversity pre- and post-treatment were not associated with pathologic response or irAE status, except for BCR diversity that was significantly lower post-treatment in patients who developed irAE (unadjusted p = 0.0321). Five cytokines increased after treatment in patients with pathologic complete response (pCR) but decreased in patients with RD, most prominently IL-8. IFNγ, IL-7, and GM-CSF levels were higher in pre-treatment than in post-treatment samples of patients who developed irAEs but were lower in those without irAEs. CONCLUSION: Baseline peripheral blood cytokine levels may predict irAEs in patients treated with immune checkpoint inhibitors and chemotherapy, and increased post-treatment B-cell clonal expansion might mediate irAEs.

A climatic suitability indicator to support Leishmania infantum surveillance in Europe: a modelling study.

Background: Leishmaniases are neglected diseases transmitted by sand flies. They disproportionately affect vulnerable groups globally. Understanding the relationship between climate and disease transmission allows the development of relevant decision-support tools for public health policy and surveillance. The aim of this modelling study was to develop an indicator that tracks climatic suitability for Leishmania infantum transmission in Europe at the subnational level. Methods: Historical records of sand fly vectors, human leishmaniasis, bioclimatic indicators, and environmental variables were integrated in a machine learning framework (XGBoost) to predict suitability in two past periods (2001-2010 and 2011-2020). We further assessed if predictions were associated with human and animal disease data from selected countries (France, Greece, Italy, Portugal, and Spain). Findings: An increase in the number of climatically suitable regions for leishmaniasis was detected, especially in southern and eastern countries, coupled with a northward expansion towards central Europe. The final model had excellent predictive ability (AUC = 0.970 [0.947-0.993]), and the suitability predictions were positively associated with human leishmaniasis incidence and canine seroprevalence for Leishmania. Interpretation: This study demonstrates how key epidemiological data can be combined with open-source climatic and environmental information to develop an indicator that effectively tracks spatiotemporal changes in climatic suitability and disease risk. The positive association between the model predictions and human disease incidence demonstrates that this indicator could help target leishmaniasis surveillance to transmission hotspots. Funding: European Union Horizon Europe Research and Innovation Programme (European Climate-Health Cluster), United Kingdom Research and Innovation.

Median Tissue Culture Infectious Dose 50 (TCID50) Assay to Determine Infectivity of Cytopathic Viruses.

The emergence of zoonotic viruses like severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2 have significantly impacted global health and economy. The discovery of other viruses in wildlife reservoir species present a threat for future emergence in humans and animals. Therefore, assays that are less reliant on virus-specific information, such as neutralization assays, are crucial to rapidly develop diagnostics, understand virus replication and pathogenicity, and assess the efficacy of therapeutics against newly emerging viruses. Here, we describe the discontinuous median tissue culture infectious dose 50 (TCID50) assay to quantitatively determine the titer of any virus that can produce a visible cytopathic effect in infected cells.

Establishing Air-Liquid Interface (ALI) Airway Culture Models for Infectious Disease Research.

Air-liquid interface (ALI) airway culture models serve as a powerful tool to emulate the characteristic features of the respiratory tract in vitro. These models are particularly valuable for studying emerging respiratory viral and bacterial infections. Here, we describe an optimized protocol to obtain the ALI airway culture models using normal human bronchial epithelial cells (NHBECs). The protocol outlined below enables the generation of differentiated mucociliary airway epithelial cultures by day 28 following exposure to air.

Applications of Quantitative PCR (qPCR) in Studies of Virus-Host Interactions.

Emerging viruses pose significant threats to human health and the global economy. In the past two decades, three different coronaviruses have emerged to cause worldwide public health concerns. The advent of high throughput genomic and transcriptomic technologies facilitated the study of virus-host interactions, accelerating the development of diagnostics, vaccines, and therapeutics. Here, we describe quantitative PCR (qPCR) in studies of virus-host interactions to dissect host responses and viral kinetics and how these relate to one another.

Histone H1x in mouse ventral hippocampus associates with, but does not cause behavioral adaptations to stress.

Prior research has identified differential protein expression levels of linker histone H1x within the ventral hippocampus (vHipp) of stress-susceptible versus stress-resilient mice. These mice are behaviorally classified based on their divergent responses to chronic social stress. Here, we sought to determine whether elevated vHipp H1x protein levels directly contribute to these diverging behavioral adaptations to stress. First, we demonstrated that stress-susceptible mice uniquely express elevated vHipp H1x protein levels following chronic stress. Given that linker histones coordinate heterochromatin compaction, we hypothesize that elevated levels of H1x in the vHipp may impede pro-resilience transcriptional adaptations and prevent development of the resilient phenotype following social stress. To test this, 8-10-week-old male C57BL/6 J mice were randomly assigned to groups undergoing 10 days of chronic social defeat stress (CSDS) or single housing, respectively. Following CSDS, mice were classified as susceptible versus resilient based on their social interaction behaviors. We synthesized a viral overexpression (OE) vector for H1x and transduced all stressed and single housed mice with either H1x or control GFP within vHipp. Following viral delivery, we conducted social, anxiety-like, and memory-reliant behavior tests on distinct cohorts of mice. We found no behavioral adaptations following H1x OE compared to GFP controls in susceptible, resilient, or single housed mice. In sum, although we confirm elevated vHipp protein levels of H1x associate with susceptibility to social stress, we observe no significant behavioral consequence of H1x OE. Thus, we conclude elevated levels of H1x are associated with, but are not singularly sufficient to drive development of behavioral adaptations to stress.

Triblock copolymer micelles enhance solubility, permeability and activity of a quorum sensing inhibitor against Pseudomonas aeruginosa biofilms.

Antimicrobial resistance is a threat to public health for which new treatments are urgently required. The capability of bacteria to form biofilms is of particular concern as it enables high bacterial tolerance to conventional therapies by reducing drug diffusion through the dense, exopolymeric biofilm matrix and the upregulation of antimicrobial resistance machinery. Quorum sensing (QS), a process where bacteria use diffusible chemical signals to coordinate group behaviour, has been shown to be closely interconnected with biofilm formation and bacterial virulence in many top priority pathogens including Pseudomonas aeruginosa. Inhibition of QS pathways therefore pose an attractive target for new therapeutics. We have recently reported a new series of pqs quorum sensing inhibitors (QSIs) that serve as potentiators for antibiotics in P. aeruginosa infections. The impact on biofilms of some reported QSIs was however hindered by their poor penetration through the bacterial biofilm, limiting the potential for clinical translation. In this study we developed a series of poly(ß-amino ester) (PBAE) triblock copolymers and evaluated their ability to form micelles, encapsulate a QSI and enhance subsequent delivery to P. aeruginosa biofilms. We observed that the QSI could be released from polymer micelles, perturbing the pqs pathway in planktonic P. aeruginosa. In addition, one of the prepared polymer variants increased the QSIs efficacy, leading to an enhanced potentiation of ciprofloxacin (CIP) action and therefore improved reduction in biofilm viability, compared to the non-encapsulated QSI. Thus, we demonstrate QSI encapsulation in polymeric particles can enhance its efficacy through improved biofilm penetration.

Heteroptera research in Latin America and the Caribbean (Insecta, Hemiptera): status and perspectives in the 21st century.

Latin American and the Caribbean regions (LAC) harbor one of the most biodiverse areas of the world, the Neotropics. True bugs (Hemiptera: Heteroptera) are a diverse lineage of insects, with more than 45,000 species, particularly speciose in the Neotropical region. True bugs are fundamental in the dynamics of natural and modified ecosystems, with several species critical to agriculture and public health. We compiled Heteroptera research in LAC from 1998-2022 using bibliographic databases. Productivity, collaborative networks, and the main topics studied were analyzed. A total of 1,651 Heteroptera studies from LAC were found, with continuous growth being 2021 the most prolific. Four categories (Taxonomy of extant species, Faunistic inventories and new records, Pest species biology, and Community ecology) represent most of the published research. About 60 percent of the records evaluated correspond to five families (Pentatomidae, Reduviidae, Coreidae, Miridae, and Rhyparochromidae). We emphasize the need to keep working on Heteroptera taxonomy because it will allow further advances in other areas such as phylogenetic analyses, biogeography, ecology, and natural history, among others. The results of our analyses characterize the current state of heteropterology in the region, establishing a baseline for future studies and efforts to broaden the knowledge of the group.

Climate change, malaria and neglected tropical diseases: a scoping review.

To explore the effects of climate change on malaria and 20 neglected tropical diseases (NTDs), and potential effect amelioration through mitigation and adaptation, we searched for papers published from January 2010 to October 2023. We descriptively synthesised extracted data. We analysed numbers of papers meeting our inclusion criteria by country and national disease burden, healthcare access and quality index (HAQI), as well as by climate vulnerability score. From 42 693 retrieved records, 1543 full-text papers were assessed. Of 511 papers meeting the inclusion criteria, 185 studied malaria, 181 dengue and chikungunya and 53 leishmaniasis; other NTDs were relatively understudied. Mitigation was considered in 174 papers (34%) and adaption strategies in 24 (5%). Amplitude and direction of effects of climate change on malaria and NTDs are likely to vary by disease and location, be non-linear and evolve over time. Available analyses do not allow confident prediction of the overall global impact of climate change on these diseases. For dengue and chikungunya and the group of non-vector-borne NTDs, the literature privileged consideration of current low-burden countries with a high HAQI. No leishmaniasis papers considered outcomes in East Africa. Comprehensive, collaborative and standardised modelling efforts are needed to better understand how climate change will directly and indirectly affect malaria and NTDs.

Multimodal 2D and 3D microscopic mapping of growth cartilage by computational imaging techniques - a short review including new research.

Being able to image the microstructure of growth cartilage is important for understanding the onset and progression of diseases such as osteochondrosis and osteoarthritis, as well as for developing new treatments and implants. Studies of cartilage using conventional optical brightfield microscopy rely heavily on histological staining, where the added chemicals provide tissue-specific colours. Other microscopy contrast mechanisms include polarization, phase- and scattering contrast, enabling non-stained or 'label-free' imaging that significantly simplifies the sample preparation, thereby also reducing the risk of artefacts. Traditional high-performance microscopes tend to be both bulky and expensive.Computational imagingdenotes a range of techniques where computers with dedicated algorithms are used as an integral part of the image formation process. Computational imaging offers many advantages like 3D measurements, aberration correction and quantitative phase contrast, often combined with comparably cheap and compact hardware. X-ray microscopy is also progressing rapidly, in certain ways trailing the development of optical microscopy. In this study, we first briefly review the structures of growth cartilage and relevant microscopy characterization techniques, with an emphasis on Fourier ptychographic microscopy (FPM) and advanced x-ray microscopies. We next demonstrate with our own results computational imaging through FPM and compare the images with hematoxylin eosin and saffron (HES)-stained histology. Zernike phase contrast, and the nonlinear optical microscopy techniques of second harmonic generation (SHG) and two-photon excitation fluorescence (TPEF) are explored. Furthermore, X-ray attenuation-, phase- and diffraction-contrast computed tomography (CT) images of the very same sample are presented for comparisons. Future perspectives on the links to artificial intelligence, dynamic studies andin vivopossibilities conclude the article.

The DMT1 isoform lacking the iron-response element regulates normal and malignant hematopoiesis via NOTCH pathway activation.

Natural resistance-associated macrophage protein 2 (NRAMP 2; also known as DMT1 and encoded by SLC11A2) is mainly known for its iron transport activity. Recently, the DMT1 isoform lacking the iron-response element (nonIRE) was associated with aberrant NOTCH pathway activity. In this report, we investigated the function of DMT1 nonIRE in normal and malignant hematopoiesis. Knockdown of Dmt1 nonIRE in mice showed that it has non-canonical functions in hematopoietic stem cell differentiation: its knockdown suppressed development along the myeloid and lymphoid lineages, while promoting the production of platelets. These phenotypic effects on the hematopoietic system induced by Dmt1 nonIRE knockdown were linked to suppression of Notch/Myc pathway activity. Conversely, our data indicate a non-canonical function for DMT1 nonIRE overexpression in boosting NOTCH pathway activity in T-cell leukemia homeobox protein 1 (TLX1)-defective leukemia. This work sets the stage for future investigation using a multiple-hit T-cell acute lymphoblastic leukemia (T-ALL) model to further investigate the function of DMT1 nonIRE in T-ALL disease development and progression.

Effect of Various Toothpaste Tablets on Gloss and Surface Roughness of Resin-based Composite Materials.

OBJECTIVES: To evaluate the effect of various toothpaste tablets on gloss and surface roughness of resin-based composite. METHODS AND MATERIALS: Sixty-four resin-based composite specimens were divided into four groups of 16 specimens each. Gloss and roughness were measured before and after simulated brushing with three types of toothpaste tablets and one conventional toothpaste: CT: Chewtab Toothpaste Tablets; AT: Anticavity Toothpaste Tablets; HC: Charcoal Toothpaste Tablets; CP: Cavity Protection toothpaste. The Kruskal-- Wallis procedure was performed to compare the differences by groups. Post-hoc comparisons were conducted with Bonferroni corrections (α=0.05). RESULTS: There was a significant drop in gloss for all groups. CT and AT maintained the highest gloss with means of 81.6 GU and 74.1 GU, respectively. The lowest gloss of 24.5 GU was observed for HC. There was a significant increase in roughness for all groups except for CT. CT had the lowest roughness with a mean of 0.034 µm, while HC had the highest roughness with a mean of 0.074 µm. There was a significant correlation between post-brushing gloss and post-brushing roughness (p<0.001, r=-0.884). CONCLUSION: Chewtab Toothpaste Tablets had the least effect on gloss and roughness, while Charcoal Toothpaste Tablets had the most negative effect on the surface properties of resin-based composites.

Multisectoral interventions for urban health in Africa: a mixed-methods systematic review.

Increasing evidence suggests that urban health objectives are best achieved through a multisectoral approach. This approach requires multiple sectors to consider health and well-being as a central aspect of their policy development and implementation, recognising that numerous determinants of health lie outside (or beyond the confines of) the health sector. However, collaboration across sectors remains scarce and multisectoral interventions to support health are lacking in Africa. To address this gap in research, we conducted a mixed-method systematic review of multisectoral interventions aimed at enhancing health, with a particular focus on non-communicable diseases in urban African settings. Africa is the world's fastest urbanising region, making it a critical context in which to examine the impact of multisectoral approaches to improve health. This systematic review provides a valuable overview of current knowledge on multisectoral urban health interventions and enables the identification of existing knowledge gaps, and consequently, avenues for future research. We searched four academic databases (PubMed, Scopus, Web of Science, Global Health) for evidence dated 1989-2019 and identified grey literature from expert input. We identified 53 articles (17 quantitative, 20 qualitative, 12 mixed methods) involving collaborations across 22 sectors and 16 African countries. The principle guiding the majority of the multisectoral interventions was community health equity (39.6%), followed by healthy cities and healthy urban governance principles (32.1%). Targeted health outcomes were diverse, spanning behaviour, environmental and active participation from communities. With only 2% of all studies focusing on health equity as an outcome and with 47% of studies published by first authors located outside Africa, this review underlines the need for future research to prioritise equity both in terms of research outcomes and processes. A synthesised framework of seven interconnected components showcases an ecosystem on multisectoral interventions for urban health that can be examined in the future research in African urban settings that can benefit the health of people and the planet.Paper ContextMain findings: Multisectoral interventions were identified in 27.8% of African countries in the African Union, targeted at major cities with five sectors present at all intervention stages: academia or research, agriculture, government, health, and non-governmental.Added knowledge: We propose a synthesised framework showcasing an ecosystem on multisectoral interventions for urban health that can guide future research in African urban settings.Global health impact for policy and action: This study reveals a crucial gap in evidence on evaluating the long-term impact of multisectoral interventions and calls for partnerships involving various sectors and robust community engagement to effectively deliver and sustain health-promoting policies and actions.

Simulated gastric leachate of 3D printer metal-fill filaments induces cytotoxic effects in rat and human intestinal models.

Metals are used in 3-dimensional (3D) printer filaments in the manufacture of 3D printed objects. Exposure to the filaments, printed objects and emissions from printing may pose health risks from release of toxic metals. This study investigated the cytotoxicity of extruded 3D printer filament leachates in rat and human intestinal cells. Copper-, bronze-, and steel-fill extruded filaments were incubated in acidic media for 2 h. Leachates were adjusted to pH 7 and cells exposed for 4 or 24 h. Concentration- and time-dependent decreases in rat and human cell viability were observed using a colorimetric assay and confirmed by microscopy. Copper- and bronze-fill leachates were more cytotoxic than steel. Copper-fill leachates had the highest copper concentrations by ICP-MS. Exposure to CuSO4 resulted in concentration-dependent cytotoxicity in rat cells. The copper chelator bathocuproine disulphonate alleviated cytotoxicity of CuSO4 and copper-fill leachate, suggesting that copper ions have a role in the cytotoxicity. Hydrogen peroxide increased and glutathione decreased in rat cells exposed to copper-fill leachate, suggesting the formation of reactive oxygen species. Overall, our data indicate that metals released from the acidic exposure of print objects using metal-fill filaments, especially copper, are toxic to rat and human intestinal cells and additional studies are needed.

Training pathologists to assess stromal tumour-infiltrating lymphocytes in breast cancer synergises efforts in clinical care and scientific research.

A growing body of research supports stromal tumour-infiltrating lymphocyte (TIL) density in breast cancer to be a robust prognostic and predicive biomarker. The gold standard for stromal TIL density quantitation in breast cancer is pathologist visual assessment using haematoxylin and eosin-stained slides. Artificial intelligence/machine-learning algorithms are in development to automate the stromal TIL scoring process, and must be validated against a reference standard such as pathologist visual assessment. Visual TIL assessment may suffer from significant interobserver variability. To improve interobserver agreement, regulatory science experts at the US Food and Drug Administration partnered with academic pathologists internationally to create a freely available online continuing medical education (CME) course to train pathologists in assessing breast cancer stromal TILs using an interactive format with expert commentary. Here we describe and provide a user guide to this CME course, whose content was designed to improve pathologist accuracy in scoring breast cancer TILs. We also suggest subsequent steps to translate knowledge into clinical practice with proficiency testing.