bluuebunny/crossref_metadata_2025 huggingface dataset:
A confounder debiasing method for RCT-like comparability enables Machine Learning-based personalization of survival benefit in living donor liver transplantation
Authors: Gangadhar Anirudh , Hasjim Bima J. , Zhao Xun , Sun Yingji , Chon Joseph , Sidhu Aman , Jaeckel Elmar , Selzner Nazia , Cattral Mark S. , Sayed Blayne A. , Brudno Michael , McIntosh Chris , Bhat Mamatha
Published: November 2024

Many clinical questions in medicine cannot be answered through randomized controlled trials (RCTs) due to ethical or feasibility constraints. In such cases, observational data is often the only available resource for evaluating treatment effects. To address this challenge, we have developed Decision Path Similarity Matching (DPSM), a novel machine learning (ML)-based algorithm that simulates RCT-like conditions to debias observational data. In this study, we apply DPSM to the clinical question of living donor liver transplantation (LDLT) versus deceased donor liver transplantation (DDLT), helping to identify which patients benefit most from LDLT. DPSM leverages decision paths from a Random Forest classifier to perform accurate, one-to-one matching between LDLT and DDLT recipients, minimizing confounding while retaining interpretability. Using data from the Scientific Registry of Transplant Recipients (SRTR), including 4,473 LDLT and 68,108 DDLT patients transplanted between 2002 and 2023, we trained independent Random Survival Forest (RSF) models on the matched cohorts to predict post-transplant survival. DPSM successfully reduced confounding associations between the two groups as shown by a decrease in area under the receiver operating characteristic (AUROC) from 0.82 to 0.51. Subsequently, RSF (C-indexldlt=0.67, C-indexddlt=0.74) outperformed the traditional Cox model (C-indexldlt=0.57, C-indexddlt=0.65). The predicted 10-year mean survival gain was 10.3% (SD = 5.7%). In conclusion, DPSM provides an effective approach for creating RCT-like comparability from observational data, enabling personalized survival predictions. By leveraging real-world data where RCTs are impractical, this method offers clinicians a tool for transitioning from population-level evidence to more nuanced, personalization.
In situ pressure modulation measurements by video capillaroscopy reveal capillary stiffening and reduced reactivity with age
Authors: Forst Marcus L. , Rincon Gabriela , Levy Juliette H. , Cornfield David N , Quake Stephen R
Published: November 2024

Understanding tissue degradation with age and disease requires a clear picture of capillary function, but the small size of capillaries makes them difficult to model, perturb, and image. To measure capillary reactivity and stiffness, we recorded videos of nailbed capillaries while applying increasing external pressure to the finger. Combining the velocities at each pressure into participant velocity distributions showed that participants aged fifty and over (n=19) had a significantly higher median velocity than participants under fifty (n=16), whereas the effects of sex and blood pressure were insignificant. Notably, the effects of age were independent of blood pressure, suggesting that this method probes a different aspect of vascular health. Calculating dissimilarity using the earth-mover’s distance between individual and population velocity distributions classified age groups above and below fifty with an AUC of 0.79±0.07. These findings show that pressure-applying video capillaroscopy can measure the effects of capillary stiffening with age and interrogate how individual capillaries respond to the slowing and stopping of blood flow—suggesting that this method could be used to diagnose and evaluate vascular diseases in the future.
Role of Arginine and its Metabolism in TGF-β-Induced Activation of Lung Fibroblasts
Authors: Hamanaka Robert B , Shin Kun Woo D , Atalay M Volkan , Cetin-Atalay Rengul , Shah Hardik , Houpy Szafran Jennifer C , Woods Parker S , Meliton Angelo Y , Shamaa Obada R , Tian Yufeng , Cho Takugo , Mutlu Gökhan M.
Published: November 2024

Arginine is a conditionally essential amino acid with known roles in protein production, nitric oxide synthesis, biosynthesis of proline and polyamines, and regulation of intracellular signaling pathways. Arginine biosynthesis and catabolism have been linked to TGF-β-induced activation of fibroblasts in the context of pulmonary fibrosis; however, a thorough study on the metabolic and signaling roles of arginine in the process of fibroblast activation has not been conducted. Here, we used metabolic dropouts and labeling strategies to determine how activated fibroblasts utilize arginine. We found that arginine limitation leads to activation of GCN2 while inhibiting TGF-β-induced mTORC1 activation and collagen protein production. Extracellular citrulline could rescue the effect of arginine deprivation in an ASS1-dependent manner. Using metabolic tracers of arginine and its precursors, we found little evidence of arginine synthesis or catabolism in lung fibroblasts treated with TGF-β. Extracellular ornithine or glutamine were the primary sources of ornithine and polyamines, not arginine. Our findings suggest that the major role for arginine in lung fibroblasts is for charging of arginyl-tRNAs and for promotion of mTOR signaling.

Highlights

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Arginine depletion inhibits TGF-β-induced transcription in human lung fibroblasts (HLFs).

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Arginine is not significantly catabolized in HLFs either through NOS or ARG dependent pathways.

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Extracellular glutamine and ornithine are the primary sources of polyamines in lung fibroblasts.

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The primary role of arginine in lung fibroblasts is for signaling through mTOR and GNC2.
Evidences that host genetic background more than the environment shapes the microbiota of the snailBulinus truncatus, an intermediate host ofSchistosomaspecies
Authors: Jaquet Mathilde J. , Douchet Philippe , Toulza Eve , Lefevre Thierry , Senghor Bruno , Boissier Jérôme , Lepais Olivier , Chancerel Emilie , Gourbal Benjamin , Rey Olivier
Published: November 2024

Microbiota are increasingly recognized as key players in regulating host biological functions, influencing both the ecology and evolution of organisms. However, the factors shaping microbiota diversity and structure in natural environments remain underexplored, especially the relative importance of host genetics versus environmental factors. In this study, we address this gap using the freshwater snailBulinus truncatus, an intermediate host for some human and animalSchistosomaparasites, as a model species. We developed 31 new microsatellite markers to assess the population structure ofB. truncatusacross 9 sites in Senegal. We then applied metabarcoding to characterize the diversity and structure of individual snail bacterial microbiota and environmental communities associated with each sampling site using environmental DNA. We also used molecular diagnostics to determine trematode infection status ofB. truncatusindividuals. By integrating these data through multiple regression on distance matrices (MRM) analyses, we quantified the influence ofB. truncatuspopulation genetics, spatial distribution, environmental bacterial communities, and infection status on the snail’s microbiota structure. Our results show that the genetic structure ofB. truncatuspopulations, and to a lower extent geographic distribution, are the main factors explaining the snail’s microbiota compositions. Neither the environmental bacterial communities nor trematode infection status significantly contributed to microbiota structure. A portion of the variance in microbiota composition remains unexplained, suggesting that additional ecological or intrinsic factors might be involved. These findings provide new insights into the drivers of microbiota structure in natural populations and highlight the complexity of host-microbiota-environment interactions.

Key-words:Bulinus truncatus, Microbiota, Spatial structure, Population genetics, Trematodes, Multiple regressions on distance matrices
Context-dependent NMDA receptor dysfunction predicts seizure treatment in mice with human GluN1 variant
Authors: Venkatesan Sridevi , Nazarkina Daria , Sullivan Megan T , Tan Yao-Fang , Qu Sarah , Ramsey Amy J. , Lambe Evelyn K.
Published: November 2024

Mutations in N-Methyl D-Aspartate receptors (NMDARs) cause epilepsy and profound cognitive impairment, though the underlying subunit-specific vulnerabilities remain unclear. We investigate the impact of a severe human variant in the lurcher motif of obligate GluN1 NMDAR subunit using transgenic mice, leveraging context-specific dysfunction to devise a surprising treatment. We show that the GluN1 Y647S variant significantly reduces current flow through isolated NMDARs in the mouse brain. However, this loss-of-function paradoxically extends NMDAR-dependent dendritic integration, causing prolonged circuit-wide excitation that promotes seizures. Mutant receptors fail to sufficiently engage opposing dendritic ion channels that normally prevent NMDAR overactivation. Boosting negative feedback restores normal dendritic integration and successfully treats seizuresin vivo,despite loss-of-function of isolated NMDARs. We demonstrate how seizures arise from loss-of-function NMDARs and target the interaction between a GluN1 variant’s receptor-level effects and its dendritic environment to treat them effectively.
Gene expression and DNA methylation changes in response to hypoxia in toxicant-adapted Atlantic killifish (Fundulus heteroclitus)
Authors: Aluru Neelakanteswar , Venkataraman Yaamini R. , Murray Christopher S. , DePascuale Veronica
Published: November 2024

Coastal fish populations are threatened by multiple anthropogenic impacts, including the accumulation of industrial contaminants and the increasing frequency of hypoxia. Some populations of the Atlantic killifish (Fundulus heteroclitus), like those in New Bedford Harbor (NBH), Massachusetts, have evolved a resistance to dioxin-like polychlorinated biphenyls (PCBs) that may influence their ability to cope with secondary stressors. To address this question, we compared hepatic gene expression and DNA methylation patterns in response to mild or severe hypoxia in killifish from NBH and Scorton Creek (SC), a reference population from a relatively pristine environment. We hypothesized that NBH fish would show altered responses to hypoxia due to trade-offs linked to toxicant resistance. Our results revealed substantial differences between populations. SC fish demonstrated a dose-dependent changes in gene expression in response to hypoxia, while NBH fish exhibited a muted transcriptional response to severe hypoxia. Interestingly, NBH fish showed significant DNA methylation changes in response to hypoxia, while SC fish did not exhibit notable epigenetic alterations. These findings suggest that toxicant-adapted killifish may face trade-offs in their molecular response to environmental stress, potentially impacting their ability to survive severe hypoxia in coastal habitats. Further research is needed to elucidate the functional implications of these epigenetic modifications and their role in adaptive stress responses.

Summary Statement

This study reveals how evolved resistance to toxicants in killifish may compromise their ability to respond to hypoxia, highlighting trade-offs that impact survival in stressed coastal environments.
Distinct intraspecific diversification dynamics in Neotropical montane versus lowland birds revealed by whole-genome comparative phylogeography
Authors: Wacker K. S. , Winger B. M.
Published: November 2024

Comparing phylogeographic patterns across different biogeographic regions can illuminate how different types of landscapes promote the formation of incipient species, providing insights into the evolutionary mechanisms underlying broadscale biodiversity gradients. The Neotropics are a global biodiversity hotspot, and the megadiverse Andes-Amazonia system has elevational gradients in both species richness and speciation rates. Here, we compare the phylogeographic dynamics of birds in the tropical Andes mountains versus the Amazonian lowlands using whole genomes from a congeneric set of colorful canopy frugivores (Tangaratanagers). We first show that Andean species have greater population structuring across their geographic ranges than Amazonian species. Next, we evaluated whether differences in geographic barrier efficacy and range size drive this elevated population-level divergence in the mountains. We find greater population differentiation and reduced gene flow across individual geographic barriers in the Andes. Furthermore, Andean species have consistently lower genetic diversity and smaller effective population sizes. Together, these results support a model of Neotropical diversification whereby incipient species form more readily in the mountains than the lowlands owing to more effective geographic barriers and smaller populations. These different phylogeographic processes across the Andes-Amazonia system carry implications for our understanding of the origin and maintenance of regional biodiversity gradients.
The C3-C3aR axis modulates trained immunity in alveolar macrophages
Authors: Earhart Alexander P. , Alburquerque Rafael Aponte , Starick Marick , Nallapu Aasritha , Garnica Lorena , Ozanturk Ayse Naz , Maurya Rahul Kumar , Wu Xiaobo , Haspel Jeffrey A. , Kulkarni Hrishikesh S.
Published: November 2024

Complement protein C3 is crucial for immune responses in mucosal sites such as the lung, where it aids in microbe elimination and enhances inflammation. While trained immunity – enhanced secondary responses of innate immune cells after prior exposure – is well-studied, the role of the complement system in trained immune responses remains unclear. We investigated the role of C3 in trained immunity and found thatin vivo, trained wild-type mice showed significantly elevated pro-inflammatory cytokines and increased C3a levels upon a second stimulus, whereas C3-deficient mice exhibited a blunted cytokine response and heightened evidence of lung injury.Ex vivo, C3-deficient alveolar macrophages (AMs) displayed reduced chemokine and cytokine output after training, which was restored by exogenous C3 but not by C3a. Inhibiting C3aR, both pharmacologically and with a genetic C3aR knockout, prevented this restoration, indicating the necessity of C3aR engagement. Mechanistically, trained WT AMs demonstrated enhanced glycolytic activity compared to C3-deficient AMs – a defect corrected by exogenous C3 in a C3aR-dependent manner. These findings reveal that C3 modulates trained immunity in AMs through C3aR signaling, affecting cytokine production and metabolic reprogramming, and highlight a novel role for C3 in trained immunity.
CancerFoundation: A single-cell RNA sequencing foundation model to decipher drug resistance in cancer
Authors: Theus Alexander , Barkmann Florian , Wissel David , Boeva Valentina
Published: November 2024

We present CancerFoundation, a novel single-cell RNA-seq foundation model (scFM) trained exclusively on malignant cells. Despite being trained on only one million total cells, a fraction of the data used by existing models, CancerFoundation outperforms other scFMs in key tasks such as zero-shot batch integration and drug response prediction. During training, we employ tissue and technologyaware oversampling and domain-invariant training to enhance performance on underrepresented cancer types and sequencing technologies. We propose survival prediction as a new downstream task to evaluate the generalizability of single-cell foundation models to bulk RNA data and their applicability to patient stratification. CancerFoundation demonstrates superior batch integration performance and shows significant improvements in predicting drug responses for both unseen cell lines and drugs. These results highlight the potential of focused, smaller foundation models in advancing drug discovery and our understanding of cancer biology. Our code is available here1.
Optical Interference for the Guidance of Cryogenic Focused Ion Beam Milling Beyond the Axial Diffraction Limit
Authors: Sica Anthony V. , Zaoralová Magda , Antolini Cali , Boltje Daan B. , Penzes Judit J. , Malmqvist Lilyana M. , Jensen Grant , Kaelber Jason T. , Dahlberg Peter
Published: November 2024

Using a tri-coincident cryogenic FIB-SEM-LM system, we identify an interferometric optical response that can be used for targeting lamella production to fluorescently labelled structures with accuracy beyond the diffraction limit. We demonstrate the approach using synthetic samples of fluorescent beads embedded in micron-scale droplets of water. We then apply the approach to capture virions inside host cells. Successful targeting is confirmed by cryogenic electron tomography revealing clusters of virions in intracellular vesicles.
IKAROS facilitates antigen escape in the face of CD19- and CD22-targeted therapies for B-cell acute lymphoblastic leukemia
Authors: Domizi Pablo , Sarno Jolanda , Jager Astraea , Merchant Milton , Pacheco Kaithlen Zen B , Yamada-Hunter Sean A. , Rotiroti Maria Caterina , Liu Yuxuan , Baskar Reema , Reynolds Warren D. , Sworder Brian J. , Sahaf Bita , Bendall Sean C. , Mullighan Charles G. , Alizadeh Ash A. , Leahy Allison B. , Myers Regina M. , Yates Bonnie , Wang Hao-Wei , Shah Nirali N. , Majzner Robbie G. , Mackall Crystal L. , Grupp Stephan A. , Barrett David M. , Sotillo Elena , Davis Kara L.
Published: November 2024

Relapse due to antigen escape is a major cause of treatment failure for patients with B-cell malignancies following targeted immunotherapies, including CD19- and CD22-directed chimeric antigen receptor T (CAR T) cells. To identify tumor intrinsic factors associated with antigen loss, we performed single-cell analyses on 61 primary patient samples or patient-derived xenografts from patients with B-cell acute lymphoblastic leukemia (B-ALL) treated with CAR T cells. We identified that low levels of the transcription factor IKAROS in pro-B-like B-ALL cells before CAR T treatment are associated with antigen escape. We demonstrate that IKAROSlowB-ALL cells lose features of B cell identity and resemble progenitor cells based on their epigenetic and transcriptional state, resulting in the downregulation of B-cell immunotherapy antigens, including surface expression of CD19 and CD22. We find that modulation of CD19 and CD22 protein expression is IKAROS dose-dependent and reversible. Further, we demonstrate that IKAROSlowcells are resistant to CD19- and CD22-targeted therapies. Together, we describe a novel role for IKAROS in the regulation of B-cell immunotherapy targets and the risk of antigen escape relapse, identifying it as a potential prognostic target.

Highlights

IKAROSlowpro-B-like B-ALL cells are associated with CD19negrelapse

IKAROSlowB-ALL cells resemble progenitor cells and have lower B-cell commitment

IKAROS modulates CD19 and CD22 surface expression in a dose-dependent and reversible manner

IKAROSlowB-ALL cells are more resistant to CD19- and CD22-targeted therapies

Graphical abstract

Before immunotherapy, IKAROSlowpro-B-like B-ALL cells possess chromatin and gene expression states poised for loss of B-cell identity while maintaining expression of CD19 and CD22. Under immune pressure, IKAROShighcells maintain their antigen expression, making them more susceptible to T cell-mediated killing. Conversely, IKAROSlowcells are more likely to downregulate their antigen expression, giving them a relative advantage to escape immunotherapies, resulting in antigen escape relapse.
Rare species do not disproportionately contribute to phylogenetic diversity in a subalpine plant community
Authors: Veldhuisen Leah N , Zepeda Verónica , Enquist Brian J , Dlugosch Katrina M
Published: November 2024

Premise of the study

Worldwide, 36% of all plant species are exceedingly rare, and are at the highest risk of extinction. Extinction risk has also been observed to be phylogenetically clustered, threatening the loss of lineages that contribute unique evolutionary diversity to communities. Rare plants’ contributions to phylogenetic diversity are largely unknown, however. We investigate whether rare species (as measured by local abundance and range size) contribute disproportionately to phylogenetic diversity in a subalpine plant community.

Methods

We collected abundance data at three sites in Washington Gulch near the Rocky Mountain Biological Laboratory (RMBL, Gothic, Colorado, USA) in 2021 and 2022. We then calculated range size for each species. We calculated phylogenetic signal for abundance and range size, compared community phylogenetic metrics weighted by range size and abundance to unweighted metrics, and quantified the change in phylogenetic diversity when removing single species and groups of species ranked by rarity.

Key results

We found phylogenetic signal for abundance, but not range size. There was no difference between rarity-weighted and unweighted phylogenetic diversity metrics. Finally, phylogenetic diversity did not decline more when we removed single rare species and groups of rare species than when we removed single common species and groups of common species.

Conclusions

Overall, we found that rare species do not disproportionately contribute to phylogenetic diversity. This suggests that rare species likely provide phylogenetic redundancy with common species, and losing rare species may not cause disproportionate drops in phylogenetic diversity in this system.
Simultaneous mesoscopic measurement and manipulation of mouse cortical activity
Authors: Matveev Pascha , Li Anna J , Ye Zhiwen , Bowen Anna J , Opitz-Araya Ximena , Ting Jonathan T , Steinmetz Nicholas A
Published: November 2024

Dynamics of activity across the cerebral cortex at the mesoscopic scale - coordinated fluctuations of local populations of neurons - are essential to perception and cognition and relevant to computations like sensorimotor integration and goal-directed task engagement. However, understanding direct causal links between population dynamics and behavior requires the ability to manipulate mesoscale activity and observe the effect of manipulation across multiple brain regions simultaneously. Here, we develop a novel system enabling simultaneous recording and manipulation of activity across the dorsal cortex of awake mice, compatible with large-scale electrophysiology from any region across the brain. Transgenic mice expressing the GCaMP calcium sensor are injected systemically with an adeno-associated virus driving expression of the ChrimsonR excitatory opsin. This strategy drives expression of the blue-excited calcium indicator, GCaMP, in excitatory neurons and red-excited Chrimson opsin in inhibitory neurons. We demonstrate widefield single-photon calcium imaging and simultaneous galvo-targeted laser stimulation over the entire dorsal cortical surface. The light channels of the imaging and the opsin do not interfere. We characterize the spatial and temporal resolution of the method, which is suitable for targeting specific cortical regions and specific time windows in behavioral tasks. The preparation is stable over many months and thus well-suited for long-term behavioral experiments. This technique allows for studying the effect of cortical perturbations on cortex-wide activity, on subcortical spiking activity, and on behavior, and for designing systems to control cortical activity in closed-loop.
Transcriptomic-Based Identification of FCRL6 as a Novel Diagnostic Biomarker in Diabetic Nephropathy
Authors: Marinedrive Adarsh , Ray Sonalika , Mazumder Mohit
Published: November 2024

Diabetic nephropathy (DN) is a microvascular complication that arises in a sizable portion of individuals with diabetes mellitus, representing a growing public health issue given its progressive injury of the kidney. DN is primarily driven by the upregulation of pro-inflammatory markers and pathways, which cause hyperfiltration and significant physiological changes to the renal structure. To further understand the molecular mechanisms by which DN progresses, we used bioinformatics and meta-analysis techniques to analyze two datasets from the Gene Expression Omnibus (GEO) database: GSE142025 and GSE142153. Differential gene expressions (DGEs) were identified using the Bioconductor DESeq2 library and further used in functional enrichment analysis. DGEs were also constructed into protein-protein interaction networks and used to find regulatory hub genes. In total, 911 DGEs were found, and enrichment using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) revealed strong correlations with immunoreactive responses and cascades. In our analysis, we discovered 12 protein-coding genes novel with regards to DN; we further identified Fc Receptor-Like 6 - FCRL6 - as a potential biomarker in nephropathy and described its involvement in the suppression of natural killer cell-mediated toxicity as a mechanism of poor immunoregulation in the diabetic kidney. The current study presents comparative analyses of the DN transcriptome and a possible diagnostic marker with applications in immunotherapy treatments of nephropathy.
SNP panel for non-invasive genotyping of leopard (Panthera pardus)
Authors: Mamugy Faruk , Bertola Laura D. , De Vry Amber Mertens , Dussex Nicolas , Shiffthaler Bastian , Paijmans Joanna , Hofreiter Michael , Forbes Ryan , Kerley Graham I.H. , Everatt Kris , Becker Matthew , Creel Scott , Bourgeois Stéphanie , Drouilly Marine , Spong Göran
Published: November 2024

Genetic resources for species monitoring are ideally relevant for the species’ full distribution range, feasible economically and logistically, and validated for the range of sample types collected from the field. This is particularly important for large carnivores that are elusive and wide-ranging, where individual and population processes often traverse administrative borders, and where obtaining high-quality samples can be challenging. Here we present a small species-specific SNP panel for leopards. We used whole genome data from across the global range and RAD sequence data from Zambian leopards to select markers for assay development. These were ascertained for 590 individual leopards from eight African countries and final selection was based on marker variation and performance on non-invasive samples. The final 96 marker panel holds 5 mitochondrial markers for species recognition, 3 Y-markers for determination of individual sex, 3 X-markers and 85 somatic markers, with an associated genetic baseline holding nearly 900 individuals. The selected autosomal markers hold variation across the global range with high power to identify individuals (PID=2,45×10−35) and in most cases their provenance with high assignment probability (>95%). Markers were also selected based on their performance on samples with low target DNA content, with distinct genotype separation in the output marker plots. The genotypes from this panel are thus straightforward to analyze and do not require computationally challenging bioinformatic resources, making this a low cost and accessible resource for leopard monitoring and research.
MaternalMed12safeguards trophoblast pluripotency and placental development
Authors: Halstead Michelle M. , Goad Jyoti , Khan Amina , Deshmukh Rutuja , Rajkovic Aleksandar
Published: November 2024

For a brief but critical period post-fertilization, the mammalian embryo is entirely dependent on maternal products inherited from the oocyte. Previous research showed that oocyte-specific loss ofMed12, an X-linked gene and Mediator complex subunit, leads to female sterility despite normal folliculogenesis and ovulation. Here, we show that loss of maternalMed12has minimal effect on the oocyte transcriptome and does not manifest in embryonic lethality until post-implantation. Implants derived fromMed12-null oocytes demonstrate abnormal placentation at E9.5, with an overabundance of trophoblast giant cells (TGC). This phenotype associates with downregulation of trophoblast pluripotency markers (e.g.Cdx2) and activation of drivers of TGC identity (e.g.Stra13) in the E7.5 extraembryonic ectoderm, revealing a previously undescribed role forMed12in trophoblast pluripotency maintenance. Notably, we find consistently lowMed12expression in embryos derived fromMed12-null oocytes, potentially due to programmed paternal X chromosome inactivation (XCI). To isolate the consequences of maternalMed12depletion, we introduced an autosomalMed12transgene and show that embryonic expression of the transgene rescues development ofMed12-null oocytes. We conclude that oocyte-specific deletion ofMed12produces a maternal-zygotic double knock-out in extraembryonic tissues due to paternal XCI, leading to loss of pluripotency in the trophoblast, placental malformation, and embryonic death.
Synthesis of USC-093 and comparison with its promoiety enantiomer USC-093D against adenovirus in vitro and in a Syrian hamster model
Authors: Yang Jiayi , Riemann Samantha B. , Lyu Jinglei , Feng Sudi , Bi Yang , Lentini Nicholas A. , Kang Inah , Kashemirov Boris A. , Hartline Caroll B. , James Scott H. , Tollefson Ann E. , Cline-Smith Anna , Toth Karoly , McKenna Charles E.
Published: November 2024

Adenovirus infections of immunocompromised humans are a significant source of morbidity and mortality. At present, no drug has been approved by FDA for the treatment of adenovirus infections. A current treatment of such infections is off-label use of an antiviral acyclic nucleotide phosphonate, cidofovir (CDV, (S)-HPMPC), which requires i.v. administration and has dose-limiting kidney toxicity. We recently reported that USC-093, a homoserinamide analogue of the tyrosinamide (S)- HPMPA prodrug USC-087, was orally effective at a 10 mg/kg against disseminated human adenovirus infection (HAdV-C6) in a Syrian hamster model, although their efficacy was marginal after respiratory infection. Neither prodrug manifested GI toxicity. Unlike USC-087, USC-093 showed no significant nephrotoxicity at the effective dose. Here, we describe in detail the synthesis of USC-093 and also its D-homoserinamide analogue, USC-093D, in four steps (20-40% overall yield) starting from Boc-protected L-homoserine or D-homoserine lactone, respectively. The two stereoisomeric prodrugs had EC5030-70 nM vs. Ad5 or 1-6 nM vs. Ad6 in HFF cells, with USC-093D giving the lower values. The prodrugs were 30-59x more potent vs. Ad5 and 82-332x more potent than Ad6 relative to the positive control, CDV. To ascertain whether D-chirality in the promoiety could enhance the performance of the prodrug in vivo, USC-093D and USC-093 were compared in the Syrian hamster model (treated from day 1 q.d at an experimentally determined maximum tolerated oral dose of 20 mg/kg)). In this study, the hamsters were instilled i.n. with vehicle or 4X1010PFU/kg of HAdV-C6 to promote lung infection. Oral valganciclovir (VGCV) at 200 mg/kg b.i.d. was used as the positive control. The body weights were recorded daily, and at 3 days post challenge, gross pathological observation was performed. Lung samples were collected, and the virus burden was determined by TCID50assay. The results show that altering homoserine stereochemistry did not markedly improve the efficacy of the orally administered prodrug, consistent with the premise that its mechanism of transport is likely not dependent on stereoselective pathways, such as hPEPT1-mediated uptake.
Revised diffusion law permits quantitative nanoscale characterization of membrane organization
Authors: Svobodová Barbora , Šťastný David , Blom Hans , Mikhalyov Ilya , Gretskaya Natalia , Balleková Alena , Sezgin Erdinc , Hof Martin , Šachl Radek
Published: November 2024

Formation of functional nanoscopic domains is an inherent property of plasma membranes. Stimulated emission depletion combined with fluorescence correlation spectroscopy (STED-FCS) has been used to identify such domains, however, the information obtained by STED-FCS has been limited to presence of such domains while crucial parameters have not been accessible, such as size (Rd), the fraction of occupied membrane surface (f), in-membrane lipid diffusion inside (Din) and outside (Dout) the nanodomains as well as their self-diffusion (Dd). Here, based on a revision of the ‘diffusion law’, we present an approach to retrieve these five parameters from STED-FCS data. We verify that approach on ganglioside nanodomains in giant unilamellar vesicles (GUVs), validating the Saffman-Delbrück assumption forDd. We examined STED-FCS data in both plasma membranes of living PtK2 cells and in giant plasma membrane vesicles (GPMVs) and present a quantitative framework for molecular diffusion modes in biological membranes.
Tumor-Infiltrating Lymphocytes Display Prognostic Signatures Associated with Chemotherapy Response in TNBC Patients
Authors: Banerjee Shayantan , Tiwari Vijay K. , Raman Karthik , Inayatullah Mohammad
Published: November 2024

Triple-negative breast cancer (TNBC) is an aggressive subtype often marked by resistance to neoadjuvant chemotherapy (NAC), making treatment particularly challenging. Tumor-infiltrating lymphocytes (TILs), crucial players in the immune landscape of tumors, have been associated with treatment outcomes, but the prognostic potential of TIL-derived gene markers in pre-NAC samples from TNBC patients remains understudied. In this research, we analyzed the single-cell transcriptional profiles of approximately 5,000 cells from four chemosensitive and four chemoresistant TNBC patients using publicly available datasets. Leveraging standard single-cell analysis, we identified differentially expressed gene signatures within the TIL subpopulation, highlighting significant immune activation pathways differentiating chemoresistant from chemosensitive tumors. By employing robust feature selection and repeated cross-validation across microarray and RNA-seq datasets, we developed a stable set of 30 TIL-based gene markers with notable prognostic relevance for NAC response in TNBC. These markers achieved an AUROC of 0.78 in the training set and validated with AUROCs of 0.8, 0.658, and 0.736 across five independent test datasets, demonstrating consistency across diverse platforms and sequencing technologies. Furthermore, increased expression of these gene signatures correlated with improved recurrence-free survival (RFS) in a cohort of 220 TNBC patients. This study enhances our understanding of the TIL transcriptional landscape in NAC response, identifying potential biomarkers and therapeutic targets for improving treatment outcomes in TNBC.
Calcium transients regulate epithelial integration of multiciliated cells during Xenopus skin development
Authors: Christodoulou Neophytos , Skourides Paris A.
Published: November 2024

Abstract

Integration basally located progenitors into an existing epithelium is crucial for tissue morphogenesis during embryogenesis and tissue homeostasis of epithelial organs. In this study, using Xenopus as a model system, we explore the role of intracellular calcium in multiciliated cell (MCC) progenitors’ epithelial integration during mucociliary skin epithelium development. Our findings reveal that calcium transients precede MCC apical emergence and are essential for their successful insertion into the overlying skin epithelium. Furthermore, we demonstrate that phospholipase C (PLC) activity is required for the generation of calcium transients, which in turn regulate MCC epithelial integration via the calcium-binding protein calmodulin. Last, we demonstrate that the PLC/Ca²⁺/calmodulin axis is necessary for proper formation of the apical actin network, by influencing its stability. This study advances our understanding of the molecular mechanisms governing epithelial integration of basal progenitors and highlights the importance of calcium signalling in coordinating cytoskeletal dynamics during epithelial morphogenesis.
Secretome analysis of human and rat pancreatic islets co-cultured with adipose-derived stromal cells reveals a signature with enhanced regenerative capacities
Authors: Pinheiro-Machado Erika , de Haan Bart J. , Engelse Marten A. , Smink Alexandra M.
Published: November 2024

Pancreatic islet transplantation (PIT), a promising treatment for Type 1 Diabetes (T1D), encounters challenges in the pre- and post-transplantation phases. Co-culturing or co-transplantation of islets with mesenchymal stromal cells (MSC), known for their regenerative properties, emerged as a potential solution and was shown to increase islet function and improve PIT outcomes. This study explored the changes in the islets’ secretion signature (secretome) when co-cultured with adipose-derived stromal cells (ASC), an MSC subtype. The secretome profile of islets and co-cultures under various stressors, i.e., cytokines, high glucose, hypoxia, and a combination of hypoxia and high glucose, was investigated. The results shed light on the potential mechanisms through which ASC support islets’ functional survival. Co-culturing pancreatic islets with ASC induced substantial proteomic changes, impacting pathways crucial for energy metabolism, angiogenesis, extracellular matrix organization, and immune responses. The analysis of key signaling molecules (VEGF, PDGF, bFGF, Collagen I alpha 1, IL-1α, and IL-10) revealed alterations influenced by the culturing conditions and the presence of the ASC.In vitrofunctional assays using the secretomes also demonstrated their potential to differentially influence angiogenic processes, enhance collagen deposition, and modulate the immune system based on the conditions in which they were generated. These findings offer valuable insights into the potential of ASC co-culturing to address challenges in PIT, paving the way for enhanced therapeutic interventions in T1D and regenerative medicine.

Highlights

Bothin-silicoandin-vitrodata support that co-culturing pancreatic islets with ASC enhances islet function.

Co-culturing islets with ASC induces changes in the secretome, impacting pathways related to energy metabolism, angiogenesis, extracellular matrix organization, and immune response.

Key signaling molecules, including VEGF, PDGF, bFGF, Collagen I alpha 1, IL-1α, and IL-10, are differentially affected by various co-culturing conditions.
Structural Model of Bacteriophage P22 Scaffolding Protein in a Procapsid by Magic-Angle Spinning NMR
Authors: Guo Changmiao , Whitehead Richard D. , Struppe Jochem , Porat-Dahlerbruch Gal , Hassan Alia , Gronenborn Angela M. , Alexandrescu Andrei T. , Teschke Carolyn M. , Polenova Tatyana
Published: November 2024

Icosahedral dsDNA viruses such as the tailed bacteriophages and herpesviruses have a conserved pathway to virion assembly that is initiated from a scaffolding protein driven procapsid formation. The dsDNA is actively packaged into procapsids, which undergo complex maturation reactions to form infectious virions. In bacteriophage P22, scaffolding protein (SP) directs the assembly of coat proteins into procapsids that have a T=7 icosahedral arrangement, en route to the formation of the mature P22 capsid. Other than the C-terminal helix-turn-helix involved in interaction with coat protein, the structure of the P22 303 amino acid scaffolding protein within the procapsid is not understood. Here, we present a structural model of P22 scaffolding protein encapsulated within the 23 MDa procapsid determined by magic angle spinning NMR spectroscopy. We took advantage of the 10-fold sensitivity gains afforded by the novel CPMAS CryoProbe to establish the secondary structure of P22 scaffolding protein and employed19F MAS NMR experiments to probe its oligomeric state in the procapsid. Our results indicate that the scaffolding protein has both α-helical and disordered segments and forms a trimer of dimers when bound to the procapsid lattice. This work provides the first structural information for P22 SP beyond the C-terminal helix-turn-helix and demonstrates the power of MAS NMR to understand higher-order viral protein assemblies involving structural components that are inaccessible to other structural biology techniques.
Habenula-ventral tegmental area functional coupling and risk-aversion in humans
Authors: Lin Wanjun , Xu Jiahua , Zhang Xiaoying , Dolan Raymond J
Published: November 2024

Maladaptive responses to uncertainty, including excessive risk avoidance, are linked to a range of mental disorders. One expression of these is a pro-variance bias (PVB), wherein risk-seeking manifests in a preference for choosing options with higher variances/uncertainty. Here, using a magnitude learning task, we provide a behavioural and neural account of PVB in humans. We show that individual differences in PVB are captured by a computational model that includes asymmetric learning rates, allowing differential learning from positive prediction errors (PPEs) and negative prediction errors (NPEs). Using high-resolution 7T functional magnetic resonance imaging (fMRI), we identify distinct neural responses to PPEs and NPEs in value-sensitive regions including habenula (Hb), ventral tegmental area (VTA), nucleus accumbens (NAcc), and ventral medial prefrontal cortex (vmPFC). Prediction error signals in NAcc and vmPFC were boosted for high variance options. NPEs responses in NAcc were associated with a negative bias in learning rates linked to a stronger negative Hb-VTA functional coupling during NPE encoding. A mediation analysis revealed this coupling influenced NAcc responses to NPEs via an impact on learning rates. These findings implicate Hb-VTA coupling in the emergence of risk preferences during learning, with implications for psychopathology.
Bystander activation across a TAD boundary supports a cohesin-dependent hub-model for enhancer function
Authors: Williamson Iain , Graham Katy A. , Becher Hannes , Hill Robert E. , Bickmore Wendy A. , Lettice Laura A.
Published: November 2024

Enhancers in the mammalian genome are able to control their target genes over very large genomic distances, often across intervening genes. Yet the spatial and temporal specificity of developmental gene regulation would seem to demand that enhancers are constrained so that they only activate the correct target gene. The sculpting of three-dimensional chromosome organization, especially that brought about through cohesin-dependent loop extrusion, is thought to be important for facilitating and constraining the action of enhancers. In particular, the boundaries of topologically associating domains (TADs) are thought to delimit regulatory landscapes and prevent enhancers acting on genes close in the linear genome, but located in adjacent TADs. However, there are some examples where enhancers appear to act across TAD boundaries. In these cases it was not determined whether an enhancer can simultaneously activate transcription at genes in its own TAD and in an adjacent TAD. Here, using a combination of mouse developmental genetics, and synthetic activators in stem cells, we show that some Shh enhancers can activate transcription simultaneously, not only ofShhbut also at a geneMnx1located in an adjacent TAD. This occurs in the context of a chromatin configuration that maintains both genes and the enhancers close together and is influenced by cohesin. To the best of our knowledge this is the first report of two endogenous mammalian genes transcribed simultanously under the control of the same enhancer, and across a TAD boundary. Our data have implications for understanding the design rules of gene regulatory landscapes, and are most consistent with a transcription hub model of enhancer-promoter communication.
Inhibition of H3K27M-enhanced ATM signaling increases radiation efficacy in diffuse midline glioma
Authors: Peterson Erik , Parsels Leslie A. , Parsels Joshua D. , Zhao Xinyi , Castro Maria G. , Lawrence Theodore S. , Koschmann Carl , Zhang Qiang , Wahl Daniel R. , Morgan Meredith A.
Published: November 2024

H3K27-altered diffuse midline glioma (DMG) is an aggressive and treatment-resistant form of pediatric high-grade glioma (pHGG). The disease is defined by point mutations in histone H3 that convert lysine 27 to methionine (termed H3K27M), resulting in genome-wide epigenetic changes that drive tumorigenesis. While radiation therapy is the standard of care, subsequent recurrence, often within the high dose radiation field, is universal. We found that the apical DNA damage response (DDR) kinase Ataxia Telangiectasia-Mutated (ATM) was uniquely upregulated in H3K27M-expressing patient tumor samples compared to pHGG expressing only wild-type histone H3. Using a panel of H3K27 isogenic DMG cell lines, we further found that H3K27M was associated with reduced H3K27me3within theATMpromoter, increased ATM mRNA levels, and elevated DDR signaling, even in the absence of exogenous DNA damage. Consistent with these results, AZD1390, a clinical-grade, CNS-penetrant ATM inhibitor, sensitized H3K27M neurospheres to the long-term effects of radiation on survival, in part due to attenuated repair of radiation-induced DNA damage. Finally, AZD1390 sensitized orthotopic H3K27M mutant tumors to radiotherapy and significantly extended median survival relative to vehicle, AZD1390 or radiation alone (50 days vs 31, 36 or 39 days, respectively) with minimal adverse effects. Taken together, these data provide a direct mechanistic link between the H3K27M mutation and ATM expression and support the clinical investigation of AZD1390 with radiotherapy in H3K27M-altered DMG.
The ecological memory of landscape complexity shapes diversity of freshwater communities
Authors: Tabi Andrea , Santos Edineusa Pereira , Brejão Gabriel , Siqueira Tadeu
Published: November 2024

Land use change affects the biodiversity of terrestrial landscapes as well as of the freshwater systems they surround, and thereby numerous ecosystem services that are essential for human well-being. Previous research suggests that freshwater systems show delayed responses to abrupt disturbances such as deforestation, however less is known about whether there is a legacy effect of landscape history on freshwater communities under a long history of land use change. We addressed this research gap here by first quantifying the historical complexity of landscapes, including natural and agricultural formations, in terms of their composition and configuration in 26 tropical catchment areas surrounding 101 streams over a 30-year period. We identified clear evidence for a memory effect of past events, whereby historical landscape complexity, measured as the information entropy of landscape composition, positively affected freshwater biodiversity. Finally, using a causal discovery approach, we showed that species richness was causally related to landscape complexity only when historical values were incorporated. Our results corroborate previous work on the positive effect of landscape complexity on biodiversity, and also confirm the role of historical contingencies in predicting future ecological outcomes.