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Abstract To understand how oncogenes affect genome organization, it is essential to visualize fundamental processes such as DNA replication and transcription at high resolution in intact cells. At the same time, it is important to determine the progression of the cell along the cell cycle, as cell cycle regulation is crucial for the control of cell proliferation and oncogenesis. Here, we present a super-resolution imaging-based method to analyze single cell nuclei sorted according to specific phases of the cell cycle. The sorting is based on the evaluation of the number and the intensity of pixels in the replication foci image and the colocalization analysis is based on image cross-correlation spectroscopy (ICCS). We evaluate the colocalization between replication and transcription, at different cell cycle phases, in a model of PML-RARα oncogene activation. We find that colocalization between replication and transcription is higher in cells in early S phase compared to cells in middle and late S phase. When we turn on the PML-RARα oncogene, this colocalization pattern is preserved but we detect an increase of colocalization between replication and transcription in the early S phase which points to an effect of the PML-RARα oncogene on the coordination between replication and transcription.

Abstract Glioblastoma (GBM) is the most aggressive glial tumor of the adult brain, associated with invariably fatal outcome, and a deeper understanding of the underlying malignant mechanisms is necessary to address the current therapeutic failure. We previously demonstrated the role of the CXCL12/CXCR4 axis in GBM cell migration and resistance to ionizing radiation. The atypical chemokine receptor ACKR3, responsible for CXCL12 scavenging, was previously suggested as additional important player in the context of GBM. Following validation of the detection tools, we observed that ACKR3 is expressed within GBM patient tumor tissue, distributed in diverse cell types. In contrast to CXCR4, ACKR3 expression in patient-derived stem-like cells (GSCs) remains however low, while ACKR3 gene expression by tumor cells appears to be modulated by the in-vivo environment. Using overexpression models, we also showed that in vitro ACKR3 had no significant direct effect on cell proliferation or invasion. Altogether, these results suggest that in vitro ACKR3 plays a minor role in malignant GBM cell biology and that its expression is possibly regulated by in-vivo influences. The subtle and multifaceted functions ACKR3 could exert in GBM should therefore only be tackled within a comprehensive tumor microenvironment considering tumoral but also non-tumoral cells.

Abstract Performing accurate Fluorescence Correlation Spectroscopy (FCS) measurements in cells can be challenging due to cellular motion or other intracellular processes. In this respect, it has recently been shown that analysis of FCS data in short temporal segments (segmented FCS) can be very useful to increase the accuracy of FCS measurements inside cells. Here, we demonstrate that segmented FCS can be performed on a commercial laser scanning microscope (LSM), even in the absence of the dedicated FCS module. We show how data can be acquired on a Leica SP8 confocal microscope and then exported and processed with a custom software in MATLAB. The software performs segmentation of the data to extract an average ACF and measure the diffusion coefficient in specific subcellular regions. First of all, we measure the diffusion of fluorophores of different size in solution, to show that good-quality ACFs can be obtained in a commercial LSM. Next, we validate the method by measuring the diffusion coefficient of GFP in the nucleus of HeLa cells, exploiting variations of the intensity to distinguish between nucleoplasm and nucleolus. As expected, the measured diffusion coefficient of GFP is slower in the nucleolus relative to nucleoplasm. Finally, we apply the method to HeLa cells expressing a PARP1 chromobody to measure the diffusion coefficient of PARP1 in different subcellular regions. We find that PARP1 diffusion is slower in the nucleolus compared to the nucleoplasm.

Abstract We argue that the Starobinsky model of inflation, realised via an R 2 term in the Lagrangian, can originate from quantum effects due to a tower of light species. By means of two separate arguments, we show how this implies that the scale of the R 2 term must be of order of the species scale Λs, namely the energy at which gravity becomes strongly coupled. We discuss the implications and challenges of this scenario for inflation, inflationary reheating, and string theory embeddings. In this context, we collect strong evidence to conclude that Starobinsky inflation lies in the Swampland.

Abstract Localised fruit thinning strategies must be investigated to improve precision crop load management in narrow-canopy, multileader apple trees. This study aimed to determine the effects of within-leader and tree total crop load on leaders’ and trunk’s growth, fruit set, yield, and fruit quality in ‘Ruby Matilda’ apples (marketed as Pink Lady®) over three years. Different crop loads were imposed on two leaders (primary and secondary) of bi-axis trees. Leader and trunk relative growth rate, return fruit set, yield, and fruit quality parameters at harvest were measured. High within-leader crop loads led to a significant increase in yield and reductions in trunk growth, return fruit set, and deterioration of fruit quality parameters except for flesh firmness and starch index. Similar trends were observed in whole-tree relationships. High crop load in secondary leaders had moderate negative effects on trunk growth, yield, and fruit mass of primary leaders; it only marginally affected their return fruit set and had no significant effect (p > 0.05) on their fruit quality. A crop load of 6.8 fruit no. cm−2 of leader cross-sectional area was estimated to achieve a relatively consistent return fruit set within the same leader. At a whole-tree level, a similar crop load (6.9 fruit no. cm−2 of trunk cross-sectional area) produced a consistent return fruit set despite its higher variability. These crop loads produced high yields (120 and 111 t ha−1, respectively) and good quality fruit. Using individual leaders as management units is recommended to simplify operations and reduce variability.

Nowadays, developing countries have seen a reduction in male reproductive parameters, and it has been linked to the exposure of endocrine disrupting chemicals (EDCs), which are able to mimic or disrupt steroid hormone actions. Also, nanoparticles have shown effects on the male reproductive system, in particular the use of TiO2-NPs in drugs, cosmetics, and food as pigment additives, and, thanks to their small size (1–100 nm), provide themselves the opportunity to be internalized by the body and pass the blood–testis barrier (BTB). Therefore, TiO2-NPs can act on spermatogenesis and spermatozoa. In this study, we carried out an in vitro assay on human spermatozoa to evaluate the effects of TiO2-NPs at the concentrations of 500, 250, 100, and 50 ppm. Exposure did not statistically alter sperm parameters (e.g., motility and viability) but induced damage to sperm DNA and the expression of biomarkers by spermatozoa. This immunofluorescence investigation showed a positivity for biomarkers of stress (HSP70 and MTs) on the connecting piece of spermatozoa and also for sex hormone binding globulin (SHBG) biomarkers. The SHBG protein acts as a carrier of androgens and estrogens, regulating their bioavailability; therefore, its expression in the in vitro assay did not rule out the ability of TiO2-NPs to act as endocrine disruptors.

Multiple sclerosis is a chronic inflammatory disease of the central nervous system1. Astrocytes are heterogeneous glial cells that are resident in the central nervous system and participate in the pathogenesis of multiple sclerosis and its model experimental autoimmune encephalomyelitis2,3. However, few unique surface markers are available for the isolation of astrocyte subsets, preventing their analysis and the identification of candidate therapeutic targets; these limitations are further amplified by the rarity of pathogenic astrocytes. Here, to address these challenges, we developed focused interrogation of cells by nucleic acid detection and sequencing (FIND-seq), a high-throughput microfluidic cytometry method that combines encapsulation of cells in droplets, PCR-based detection of target nucleic acids and droplet sorting to enable in-depth transcriptomic analyses of cells of interest at single-cell resolution. We applied FIND-seq to study the regulation of astrocytes characterized by the splicing-driven activation of the transcription factor XBP1, which promotes disease pathology in multiple sclerosis and experimental autoimmune encephalomyelitis4. Using FIND-seq in combination with conditional-knockout mice, in vivo CRISPR-Cas9-driven genetic perturbation studies and bulk and single-cell RNA sequencing analyses of samples from mouse experimental autoimmune encephalomyelitis and humans with multiple sclerosis, we identified a new role for the nuclear receptor NR3C2 and its corepressor NCOR2 in limiting XBP1-driven pathogenic astrocyte responses. In summary, we used FIND-seq to identify a therapeutically targetable mechanism that limits XBP1-driven pathogenic astrocyte responses. FIND-seq enables the investigation of previously inaccessible cells, including rare cell subsets defined by unique gene expression signatures or other nucleic acid markers.

Abstract Multiple sclerosis is a debilitating autoimmune disease, characterized by chronic inflammation of the central nervous system. While the significance of the gut microbiome on multiple sclerosis pathogenesis is established, the underlining mechanisms are unknown. We found that serum levels of the microbial postbiotic tryptophan metabolite indole-3-carboxaldehyde (3-IAld) inversely correlated with disease duration in multiple sclerosis patients. Much like the host-derived tryptophan derivative l-Kynurenine, 3-IAld would bind and activate the Aryl hydrocarbon Receptor (AhR), which, in turn, controls endogenous tryptophan catabolic pathways. As a result, in peripheral lymph nodes, microbial 3-IAld, affected mast-cell tryptophan metabolism, forcing mast cells to produce serotonin via Tph1. We thus propose a protective role for AhR–mast-cell activation driven by the microbiome, whereby natural metabolites or postbiotics will have a physiological role in immune homeostasis and may act as therapeutic targets in autoimmune diseases.

The aim of this study was to present our preliminary experience with transarterial radioembolization (TARE) using Yttrium-90 (90Y), compare the cancer-specific survival (CSS) of patients with hepatocellular carcinoma (HCC) and colorectal cancer (CRC) liver metastases undergoing TARE, and investigate the influence of additional treatments on CSS. Our database was interrogated to retrieve patients who had undergone TARE using Yttrium-90 (90Y) glass or resin microspheres. Kaplan–Meier curves and the log-rank test were employed to conduct survival analysis for the different groups (p < 0.05). Thirty-nine patients were retrieved (sex: 27 M, 12 F; mean age: 63.59 ± 15.66 years): twenty-three with hepatocellular carcinoma (HCC) and sixteen with CRC liver metastasis. Globally, the patients with HCC demonstrated a significantly longer CSS than those with CRC liver metastasis (22.64 ± 2.7 vs. 7.21 ± 1.65 months; p = 0.014). Among the patients with CRC liver metastasis, those receiving TARE and additional concomitant treatments (n = 10) demonstrated a longer CSS than the CRC patients receiving only TARE (9.97 ± 2.21 vs. 2.59 ± 0.24 months; p = 0.06). In the HCC group, there was a trend of a longer CSS in patients (n = 8) receiving TARE and additional treatments (27.89 ± 3.1 vs. 17.69 ± 3.14 months; p = 0.15). Patients with HCC seem to achieve a longer survival after TARE compared to patients with CRC liver metastases. In patients with CRC liver metastases, the combination of TARE and additional concomitant treatments may improve survival.

La displasia cleidocraneal (OMIM #119600) es una osteocondrodisplasia de herencia autosómica dominante debida a una mutación en heterocigosis en el gen RUNX2. Su prevalencia es 1 en 1.000.000 individuos. Puede detectarse en pacientes de estatura baja asociada a alteraciones dentales, malformaciones craneofaciales y ausencia o hipoplasia de clavículas; el diagnóstico se corrobora con estudios imagenológicos y moleculares; el tratamiento es sintomático, ortopédico y dental. Objetivo: describir una enfermedad genética de baja prevalencia, sus características clínicas y la importancia de la aproximación diagnóstica. Se presenta caso de adolescente masculino con talla baja, presencia prolongada de dientes primarios y supernumerarios, macrocefalia, prominencia frontal, hipertelorismo ocular, asimetría de tórax y extremidades; antecedentes familiares positivos; estudio radiológico revela ausencia de clavículas y estudio molecular confirma la displasia cleidocraneal. El paciente asistió tardíamente a consulta de nutrición y desarrollo. La historia clínica exhaustiva, estudios complementarios adecuados y seguimiento oportuno por equipo multidisciplinario son aspectos claves para un diagnóstico precoz y pronóstico favorable.

Reseñas iberoamericanas

Abstract Dendritic cells (DCs) are essential orchestrators of immune responses and represent potential targets for immunomodulation in autoimmune diseases. Human amniotic fluid secretome is abundant in immunoregulatory factors, with extracellular vesicles (EVs) being a significant component. However, the impact of these EVs on dendritic cells subsets remain unexplored. In this study, we investigated the interaction between highly purified dendritic cell subsets and EVs derived from amniotic fluid stem cell lines (HAFSC‐EVs). Our results suggest that HAFSC‐EVs are preferentially taken up by conventional dendritic cell type 2 (cDC2) through CD29 receptor‐mediated internalization, resulting in a tolerogenic DC phenotype characterized by reduced expression and production of pro‐inflammatory mediators. Furthermore, treatment of cDC2 cells with HAFSC‐EVs in coculture systems resulted in a higher proportion of T cells expressing the regulatory T cell marker Foxp3 compared to vehicle‐treated control cells. Moreover, transfer of HAFSC‐EV‐treated cDC2s into an EAE mouse model resulted in the suppression of autoimmune responses and clinical improvement. These results suggest that HAFSC‐EVs may serve as a promising tool for reprogramming inflammatory cDC2s towards a tolerogenic phenotype and for controlling autoimmune responses in the central nervous system, representing a potential platform for the study of the effects of EVs in DC subsets.

Volume 15 of AGATHÓN follows on from its predecessors on Innovability©® | Digital Transition and Innovability©® | Ecological Transition, aware of its pressing topicality but also of the scope that the proposal of a threefold key of interpretation suggests. We have clarified the meaning of the term ‘innovability’©®, formerly in use in the economic and social sciences, which is attributed to a renewed driving force for a new paradigm of development that expresses one of the most crucial challenges of our time and the need for a ‘solidary’ convergence between the two inescapable instances of ‘innovation’ and ‘sustainability’, as if they were opposites and contrasts: beyond the term used, in a historical moment characterised by environmental, social and economic emergencies, Humanity promotes one of its prerogatives, the use of the ‘things’ that nature makes available to us to do something other than their primary function (innovation), aware that those resources are not inexhaustible (sustainability). In this context, which must always look forward, we must design our best political and systemic actions to promote the need to innovate by using the Planet’s resources well and consciously. The goals of climate neutrality by 2050 and the reduction of CO2 emissions by 55% (compared to the 1990 level) by 2030 (European Commission, 2019, 2021) pose the European Union, and even more so the rest of the world, with several complex issues, including a significant increase in ‘clean’ energy production from alternative and renewable sources, the reduction of energy poverty, greater security of energy supply and a drastic decrease in dependence on energy imports; at the same time, the aim is to foster modern economic growth decoupled from the use of non-renewable resources, the creation of new jobs, and to generate environmental and health benefits, objectives with inevitable cultural, political, economic, production, technological and social implications to be addressed both within one’s borders and in foreign policy. The energy transition is, therefore, complex and challenging to implement because it involves ‘everything’ and is needed ‘everywhere’ but also because globally, primary energy consumption has been steadily increasing for at least half a century. The theoretical and experimental framework presented in Volume 15 of AGATHÓN demonstrates how energy, ecological and digital transitions can contribute synergistically to achieving the goals of decarbonisation and climate neutrality. The contributions published in the form of essays and research papers appear consistent with the 2022 Strategic Foresight Report (European Commission, 2022) based on the JRC report entitled Towards a Green and Digital Future – Key Requirements for Successful Twin Transitions in the European Union (Muench et alii, 2022) and founded on the fundamental concepts of (a) ‘twin’ transitions, as the key to a sustainable, fair and competitive future; (b) ‘just’ transition, for widespread acceptance of green and digital solutions to mitigate consumption and improve efficiency; (c) ‘integrated approach’ to challenges, to maximise the benefits of synergies and better manage risks. From the published contributions, it emerges the need for a paradigm shift that, on the one hand, is characterised by a ‘sufficiency’ approach (regarding new land occupation and new constructions) and a circular economy (to limit the use of non-renewable resources) capable of exploiting the potential of technologies for the new services made possible by digitisation, and on the other hand, relies on new user awareness of the limits of the Planet, pursuable through ‘soft’ urgent actions that are robust, flexible and easy to implement as they require a lower financial commitment. We assume that community energy renewable energy production from hydrogen and production chains can help mitigate greenhouse gas emissions. In that case, the vast existing real estate heritage is an area in which it is possible to intervene effectively, even where it has a historical and cultural value, using tools such as digital twins or analysis methodologies capable of assessing ex-ante the impacts on the ecosystem and prefigure scenarios for cities, buildings and production processes aimed at sustainable development and compatible with the urgent objectives set for 2030 and 2050. These are some of the strategies, pathways, measures and actions that can take place by exploiting the availability of the substantial financial resources allocated by governments for transitions, stimulating the sensitivity of local administrators and enhancing the skills and transversal competencies of technicians and operators in the sector, but also and above all by raising users’ awareness of the risks posed by climate change, to activate their ‘behavioural’ response to the consumption of energy and non-renewable natural resources.

Design, understood as a modern discipline in its broadest sense, has always been inextricably linked to energy production, availability and accessibility. Every project resulting from human creativity and ingenuity, whether material or immaterial, analogue or digital, depends on it; it is necessary to break free from this atavistic dependence by finding alternative paths that lead us to the energy transition. An increase in human awareness is required to accomplish this and achieve social and environmental equity. In this perspective, this paper, starting with the concepts of ‘mitigation’, ‘sufficiency’, ‘circular economy’ and ‘innovability’, illustrates a number of highly replicable experiments carried out by the multidisciplinary Dotdotdot Studio by employing storytelling, emotive technology and the value of data to generate awareness in users on the issue of energy conservation. Article info Received: 15/05/2024; Revised: 17/05/2024; Accepted: 20/05/2024

The Gulf of Maine in the northwestern Atlantic Ocean is one of the world’s fastest warming marine regions. Changes in ocean conditions are affecting growth, survival, and distribution of American lobster (Homarus americanus), which supports iconic fisheries along the coast of Maine, USA. In this study, we analyzed 15 years of oral records from the Maine Fishermen’s Climate Roundtables to explore fishermen’s observations of and responses to social–ecological changes. Fishermen reported an overall shift in lobster biomass further east and offshore, resulting in strategic expansion of fishing seasons and areas. Biomass shifts were thought to be connected to increases in temperature, decreases in salinity, a shift in ocean currents, and a loss of predator species. Fishing strategies were categorized according to five domains of adaptive capacity, but the majority of fishers’ responses fell into two domains: “access to assets” and “diversity and flexibility”. Strategies within these domains included increased expansion into federal lobster fisheries and extension of fishing seasons. Fishermen highlighted data gaps that need to be addressed to meet the challenges of climate change. Fisheries learning exchanges, such as the Climate Roundtables, create social networks that foster knowledge sharing to support the continued viability of local livelihoods.

This study aimed to characterize the physiological and structural responses of potted one-year-old olive trees belonging to two olive cultivars—‘Nocellara del Belice’ and ‘Cerasuola’—exposed to prolonged drought under greenhouse conditions. Two irrigation treatments based on evapotranspiration (ET) were imposed for 69 days, i.e., well-watered (WW, 100% ET) and drought-stressed (DS, 10−30% ET). Leaf stomatal conductance (gs), stem water potential (Ψstem), transpiration (E), photosynthetic capacity (Amax), water use efficiency (WUE), stem (Kstem) and root (Kroot) hydraulic conductance, trunk diameter variations (TDV), and leaf patch attenuated pressure fluctuations (pp, a proxy of the inverse of leaf turgor pressure) were measured in WW and DS trees at different stages of the experiment. Leaf gs did not significantly differ between cultivars under DS, whereas differences in Ψstem only became significant at the end of prolonged drought, when ‘Nocellara del Belice’ experienced Ψstem < −4 MPa. ‘Cerasuola’ trees expressed the best WUE under drought, although they were more susceptible to photoinhibition under optimal plant water status. Both cultivars tended to increase their Kstem at the end of the drought period. A marked reduction in Kroot occurred in ‘Cerasuola’ plants after prolonged drought; however, a similar mechanism was not observed in ‘Nocellara del Belice’. The ratio between Kstem and Kroot exponentially increased towards the end of the prolonged drought period in both cultivars, but more markedly in ‘Cerasuola’. TDV and pp trends suggested that ‘Cerasuola’ plants keep better plant water status under severe drought compared to ‘Nocellara del Belice’ by maintaining high leaf turgor and reduced trunk diameter fluctuations. These responses may be related to reduced cell wall elasticity and xylem vessel size and/or wall thickness—drought avoidance mechanisms. The Kstem/Kroot ratio can serve as an indicator of drought stress avoidance mechanisms to compare genotype-specific responses to drought stress.

Abstract Early Dark Energy (EDE) is a prominent model to resolve the Hubble tension, which employs a dynamical axion with a periodic potential. In this work, we take first steps towards the embedding of this model into stable compactifications of string theory. First, we provide a pedagogical review of the EDE scenario and its main challenges. Second, we construct a simple supergravity toy model using only minimal ingredients. Already at this level, we can understand the origin of the harmonics of the EDE scalar potential in terms of a delicate balance of the leading terms from separate non-perturbative effects. Third and final, we embed the model into a KKLT-type compactification, with the EDE scalar field realized by a two-form axion. We find that a successful embedding, with all moduli stabilized, requires restrictive assumptions, which are lacking at present a known string theory implementation mechanism, both on the Pfaffians and on the exponents of the non-perturbative terms responsible for the EDE dynamics. We point out that such non-generic conditions reflect well known challenges of the EDE model and further investigation might guide us towards a conclusive resolution. An alternative viewpoint suggests that rather than serving as the foundation for a string theory embedding, these conditions might instead indicate that EDE is, in fact, incompatible with string theory, at least in the realizations we study here. Further investigation will be needed to ascertain the status of EDE in string theory.

Abstract The Laser Interferometer Space Antenna (LISA) has two scientific objectives of cosmological focus: to probe the expansion rate of the universe, and to understand stochastic gravitational-wave backgrounds and their implications for early universe and particle physics, from the MeV to the Planck scale. However, the range of potential cosmological applications of gravitational-wave observations extends well beyond these two objectives. This publication presents a summary of the state of the art in LISA cosmology, theory and methods, and identifies new opportunities to use gravitational-wave observations by LISA to probe the universe.

Plastic additives include a wide range of pollutants, added throughout the production process of plastics aiming to improve their properties. Given that they are not chemically bound to the plastic items, they can easily migrate in the marine environment allowing their uptake by marine organisms and accumulation in their tissues. Representatives of Phthalic acid esters (PAEs) and bisphenols have been characterized for their ability to impact not only marine organisms but also humans via fish and seafood consumption. In this study, a liquid-liquid method was applied to determine the concentrations of selected PAEs and Bisphenol A (BPA) in the tissues of E.encrasicolus, S.pilchardus, B.boops, and M.barbatus from two important divisions of fisheries (North Aegean and the Western Ionian Seas, thereof NAS and IOS respectively) of the Mediterranean Sea. The level of contamination varied among the different species and geographical locations. DEHP (bis(2-ethylhexyl) phthalate) had the highest mean concentrations of the examined PAEs for all four species examined. Statistically significant differences in DEHP mean concentrations were observed between the tissues of B.boops (collected from NAS) and S.pilchardus (from IOS) (p < 0.05, Wilcoxon rank test). DIDP (di-isodecyl phthalate) was the following most common PAEs, although no statistically significant differences were presented between sites, species, and tissues. DINP (di-isononyl phthalate) was recorded only in the GIT of E.encrasicolus (from NAS), while the rest of the examined PAEs were presented in a smaller subset of the samples. Noticeably, the parent diester DBP and the metabolic monoester MNBP were concurrently detected in a part of the analyzed samples. The level of risk via fish consumption, based on the Estimated Daily Intake and the Target Hazard Quotient (THQ), showed that the PAEs and BPA had a low likelihood of a negative effect occurring for every scenario that was examined, although THQ for BPA indicated there was a higher likelihood than PAEs of an adverse effect to be presented. The results of this study highlighted the need for future efforts focusing on the factors affecting plastic additives occurrences in the marine environment, especially for marine organisms intended for consumption.