Biology subjects

Moreno, J.

Publications and source records attributed to Moreno, J..

At least 19 recordsLinked to original sources

bioRxiv2026

Synergistic cytotoxicity with Chk1/Chk2-inhibitor prexasertib in small cell lung cancer following lurbinectedin-induced G2/M-checkpoint activation

Small cell lung cancer (SCLC) is an aggressive thoracic malignancy with a 5-year survival rate under 7%. Lack of meaningful improvement of survival rates despite advances in treatment highlights the need for novel therapeutic approaches to improve patient outcomes. Currently, carboplatin + etoposide chemotherapy is the backbone of treatment for most patients. Lurbinectedin is a cytotoxic drug with unique activity against small cell lung cancers in patients with extensive disease and acquired resistance to carboplatin + etoposide. Our preliminary experiments in human SCLC cell lines treated with lurbinectedin demonstrated a dose-dependent increase in Chk1 and Chk2 protein phosphorylation. A consequence of the frequent TP53 inactivation in SCLC is tumor cell reliance on G2/M cell cycle checkpoints involving Chk1/Chk2 to maintain genomic integrity and allow cell survival following DNA damage. We hypothesised that inhibition of Chk1/Chk2-dependent responses with dual-inhibitor prexasertib (ACR-368), would potentiate tumor cell killing by lurbinectedin potentially in a synergistic manner. SCLC cells underwent cell death following single agent prexasertib exposure and this further increased with prexasertib + lurbinectedin combination. Highest Single Agent (HSA) synergy score calculations based on cell viability measurements suggested synergistic action between prexasertib and lurbinectedin at select dose combinations. Western blot analysis of intracellular proteins from SCLC cells treated with both drugs demonstrate dynamic, dose-dependent effects on Chk2, Chk1 and downstream effector Wee1, with lurbinectedin increasing intracellular levels of pChk1 and pChk2, while co-treatment with prexasertib deregulates this process across multiple human-derived cell lines. Synergistic killing was associated with elevated {psi}-H2AX levels indicative of DNA double strand breaks and PARP-cleavage due to apoptotic caspase activation. Despite some heterogeneity among treated SCLC cells, the increased phosphorylation of Chk1 was noted at several kinase-activating sites including Serine 296, 317, and 345 while Chk2 Tyrosine 68 phosphorylation was consistently upregulated by lurbinectedin. The results provide a preclinical mechanistic rationale for overcoming a pro-survival, drug resistance-promoting checkpoint pathway to enhance the unique efficacy of single-agent lurbinectedin in patients with SCLC.

cancer biology↗

bioRxiv2026

Toward Large-Scale Numerical Modeling of the Cardiovascular System with up to 34 Billion Vessels

Cardiovascular diseases, such as stroke and heart attacks, are the leading cause of death worldwide. Computational models like cardiovascular digital twins (CVDTs) offer a promising path for research and intervention but are challenged by the complexity of simulating the full human vasculature. This study evaluates the feasibility of simulating blood flow through a vascular network containing 34 billion vessels (the estimated number in the human body) using first-principles physics and simplified geometry which is a first step towards CVDT. We synthesized 3D vasculature using a fractal model and computed blood flow rates via Poiseuille equation and steady-state fluid dynamics, implemented with high-performance computing. Simulations were conducted for networks ranging from 6 vessels to 34 billion vessels. The results demonstrated high accuracy (within 1% of bench-marks), reproducibility across platforms, and strong scalability. Simulating the full vasculature required 156 node-hours on the second-fastest supercomputer in the world, using 29 TB of memory and 84 TFLOPS. Maximum speedup factor was 80, with parallel efficiency no lower than 0.48. These findings show it is computationally feasible to simulate blood flow through a full-body vascular network at scale. The approach is well suited to parallel computing, suggesting that with continued development, CVDTs could enable whole-organism modeling for applications such as stroke, trauma, radiation injury, and cancer metastasis.

bioinformatics↗

bioRxiv2026

Semaglutide Attenuates Neuroinflammation in Mice

Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) have shown promise in preclinical models of neurodegeneration, with emerging evidence suggesting these effects may be driven by modulation of neuroinflammation. However, the cellular mechanisms underlying GLP-1 RA effects on neuroinflammation remain poorly understood. Here, using a mouse model of lipopolysaccharide-induced neuroinflammation, we investigated how semaglutide coordinates cellular responses to resolve neuroinflammation. We find that semaglutide prevents brain infiltration of neutrophils, excessive cytokine release, and suppresses neuroinflammation-associated transcriptional signatures specifically in microglia, endothelial cells, and a subset of pericytes. Mechanistically, we identify a subset of Glp1r-expressing neurons in the dorsal vagal complex that, upon semaglutide treatment, regulate genes involved in anti-inflammatory signaling. Semaglutide-modulated pathways overlap with inflammatory signatures found in human neurodegenerative diseases, including Alzheimers disease, suggesting broad relevance for conditions involving neuroinflammation. Together, these findings reveal how GLP-1R signaling orchestrates resolution of neuroinflammation through coordinated multi-cellular programs.

neuroscience↗

bioRxiv2025

Direct In-Sample Sequencing of the 3' Transcriptome Expands the Capabilities of Optical Pooled Screens

We present a platform that directly sequences single guide RNAs and endogenous 3'UTRs in fixed cells while simultaneously measuring protein abundance and cellular morphology. We demonstrate platform capability by performing optical pooled screening of CRISPR-perturbed lung cancer cells. This approach unites direct in-sample RNA sequencing with complementary phenotypic readouts, enabling comprehensive, scalable, and functional genomics analyses within a single experiment.

genomics↗

bioRxiv2025

A simplified hybrid capture approach retains high specificity and enables PCR-free workflow

Hybrid capture is a critical technology for selective enrichment of genomic regions of interest, enabling cost-effective focused sequencing in both clinical and research applications. We present a simplified hybrid capture approach that eliminates complexities typically associated with hybridization-based selection methods by directly loading the hybridization product onto the sequencing flow cell. The workflow removes bead-based steps, washes, and post-hybridization PCR, while retaining high capture specificity and library complexity. The approach is enabled by the development of a streptavidin flow cell surface, a method to circularize and amplify captured targets on the flow cell, and a fast hybridization protocol. We demonstrate application across targeted panel sizes and improvements to library complexity and variant calling. We also show how the approach can be used to create an entirely PCR-free targeted sequencing workflow that further improves variant calling and enables the detection of repeat expansions.

genomics↗

bioRxiv2025

Drug-Resistance Biomarkers in Leishmania infantum through Nanopore-Based Detection of Aneuploidy and Gene Copy Number Variations with LeishGenApp

BackgroundDrug-resistant strains of Leishmania infantum challenge the effectiveness of treatments for clinical leishmaniosis and may lead to more frequent relapses. Copy number variation (CNV) at specific genetic loci is associated with drug resistance and virulence, but information about its prevalence in endemic regions is limited. This study examines the drug resistance and virulence status of Leishmania strains in human and canine isolates from the Mediterranean region. MethodsForty-eight Leishmania infantum isolates were whole-genome sequenced with nanopore long reads, followed by de novo assembly. We analyzed chromosomal aneuploidies and gene copy number variation in loci linked to drug resistance and virulence in Leishmania, alongside the genomic structure and rearrangements responsible for these variations. ResultsComplete genomes were de novo assembled for 35 L. infantum isolates (22 from dogs and 13 from humans), revealing significant chromosomal variability. We assessed copy number variation for 22 potential biomarkers: 15 genes related to drug resistance to first-line drugs (METK for allopurinol; LdSMT for amphotericin B; AQP1 and H-locus for antimonials; LdMT, LdRos3, and MSL for miltefosine; PPM for paramomycin) and seven genes related to virulence (lipophosphoglycan and proteophosphoglycan biosynthesis, and the Lack protein). Drug-resistance biomarkers were identified in 80% of the isolates. Canine strains primarily showed resistance to allopurinol and antimonials, while human isolates exhibited a broader resistance spectrum, especially to antimonials and paromomycin. The co-occurrence of resistance biomarkers was common, especially for allopurinol and antimonial resistance. Distinct mechanisms underlie the observed copy number variations. Virulence-associated genes were less variable among isolates. ConclusionsThe prevalence of drug-resistance biomarkers in Leishmania infantum strains from the Mediterranean region, as revealed by this study, underscores the critical need for routine resistance surveillance in managing clinical leishmaniosis. These findings not only inform current clinical practice but also pave the way for more effective management strategies in the future.

genetics↗

bioRxiv2024

Microvascular preservation and cardiomyocyte hyperplasia underlie adaptive right ventricular development in congenital heart disease-associated pulmonary arterial hypertension

AbstractO_ST_ABSBackgroundC_ST_ABSRight ventricular (RV) failure is the primary cause of death among patients with pulmonary arterial hypertension (PAH). Patients with congenital heart disease- associated PAH (CHD-PAH) demonstrate improved outcomes compared to patients with other forms of PAH, which is related to the maintenance of an adaptively hypertrophied RV. In an ovine model of CHD-PAH, we aimed to elucidate the cellular, microvascular, and transcriptional adaptations to congenital pressure overload that support RV function in CHD-PAH. MethodsFetal surgery was performed on late gestation lambs to insert a large aortopulmonary graft, leading to a persistent congenital left-right shunt and RV pressure load. At 3 days and 4-6 weeks of life, shunt RV microvasculature, cardiomyocyte structure, and myocardial growth mechanisms were compared to age-matched controls and unoperated fetal RV. RNA sequencing was performed to assess differences in the RV transcriptomes. ResultsAt 4-6 weeks of age, shunt lambs demonstrate significant RV enlargement (shunt 37.1 {+/-} 2.9g vs control 15.9 {+/-} 1.0g, p<0.001) but maintain stable microvascular density (fetal 3.0 {+/-} 0.6 vs shunt 2.9 {+/-} 0.3 vs control 3.1 {+/-} 0.6 capillaries per 1000 {micro}m3, p>0.05). Shunt RV cardiomyocytes are significantly smaller by cross-sectional area and more numerous than age-matched controls (shunt 73.3 {+/-} 5.5 {micro}m2 vs control 99.2 {+/-} 4.9 {micro}m2, p=0.013). At 3 days, shunt RV cardiomyocytes show evidence of increased proliferative capacity and ongoing hyperplasia compared to controls. RNA sequencing analyses reveal a distinct gene expression profile in shunt RV consistent with a delay in terminal differentiation and metabolic adaptations to support adaptive function. ConclusionsThis study provides novel insights into the development of adaptive RV hypertrophy in CHD-PAH, demonstrating roles for preserved microvascular density and increased postnatal cardiomyocyte hyperplasia in supporting RV performance. Future investigations into the mechanisms underlying these changes could have significant implications for the development of novel therapeutic strategies for supporting RV function.

physiology↗

bioRxiv2024

Mapping atherogenesis mechanisms in smooth muscle cells by targeting genes linked to coronary artery disease

Recent genome-wide association studies (GWAS) have identified multiple vascular cell-expressed genes linked to coronary artery disease (CAD), suggesting that smooth muscle cells (SMCs) and SMC-derived metaplastic cells are promising targets for novel antiatherosclerosis therapies. However, the disease-promoting pathways of most GWAS-identified genes are unknown, hindering their translation into therapeutic targets. This study integrated public GWAS data for CAD and single-cell RNA sequencing (scRNA-seq) analyses of human atherosclerotic plaques to identify 20 GWAS risk genes with a putative mechanism of action in SMCs or SMC-derived cells. Gene perturbation experiments in SMCs coaxed to plaque-relevant phenotypes revealed that the selected risk genes, despite encoding very different types of proteins, regulated shared sets of genes associated with contractile functions, cell cycle pathways, NF{kappa}B, and type I interferon signaling. By integrating information about GWAS gene effect direction and a deep analysis of cholesterol- and stretch-induced gene modules in SMCs, we find evidence that cholesterol-induced signaling is a pro-atherogenic disease mechanism in SMCs that is upregulated by detrimental and downregulated by protective GWAS genes. Overall, our study identifies a set of candidate disease mechanisms in SMCs that are regulated by multiple GWAS genes across several SMC assays. Furthermore, it provides proof-of-concept for using GWAS gene effect directionality to predict the pathogenic effect of candidate disease mechanisms that can be extended to other GWAS genes and cell types in the future.

cell biology↗

bioRxiv2024

Free Amino Acids Accelerate the Time-Dependent Inactivation of Rat Liver Nucleotide Pyrophosphatase / Phosphodiesterase Enpp3 elicited by EDTA

Nucleotide-pyrophosphatases/phosphodiesterases (NPP/PDE) are membrane or secreted Zn2+-metallohydrolases of nucleoside-5-monophosphate derivatives. They hydrolyze, for instance, ATP and 4-nitrophenyl-dTMP, and belong to the ecto-nucleotide pyrophosphatase/phosphodiesterase (ENPP) family that contains seven members (ENPP1-ENPP7). Earlier we had shown that an NPP/PDE activity solubilized and partially purified from rat liver membranes is inactivated by EDTA in a time-dependent fashion, an effect enhanced by glycine and blocked by the 4-nitrophenyl-dTMP. Here, we extended this observation to other free amino acids. Activity assays started after different incubation lengths with EDTA provided first-order, apparent inactivation constants (ki(ap)). With the exception of cysteine (a strong inhibitor) and histidine (itself evoking a time-dependent inactivation), free amino acids themselves did not affect activity but increased ki(ap). The results are compatible with a conformational change of NPP/PDE evoked by interaction with free amino acids. The enzyme preparation was analyzed to identify what ENPP family members were present. First, the hydrolytic activity on 2,3-cGAMP was assayed because until very recently ENPP1 was the only mammalian enzyme known to display it. 2,3-cGAMP hydrolase activity was clearly detected, but mass spectrometry data obtained by LC-MS/MS gave evidence that only rat Enpp3, Enpp4 and Enpp5 were present with low abundance. This finding coincided in time with a recent publication claiming that mouse Enpp3 hydrolyzes 2,3-cGAMP, and that Enpp1 and Enpp3 account for all the 2,3-cGAMP hydrolase activity in mice. So, our results are confirmatory of Enpp3 activity towards 2,3-cGAMP. Finally, the effect of amino acids could be relevant to NPP/PDE actions dependent on protein-protein interactions, like the known insulin-related effects of ENPP1 and possibly ENPP3.

biochemistry↗

bioRxiv2024

Brain-Penetrant NF-κB and NLRP3 Targeting Nanoligomers are Therapeutic in Amyotrophic Lateral Sclerosis (ALS) and Alzheimers Disease (AD) Human Organoid and Mouse Models

Millions of people suffer worldwide from neurodegenerative diseases ranging from rapidly progressing and fatal motor neuron diseases like Amyotrophic Lateral Sclerosis (ALS) to more chronic illnesses such as frontotemporal dementia (FTD) and Alzheimers disease (AD). A growing number of studies have implicated neuroinflammation as a key and causative phenomenon and an important target for novel therapeutics for these diseases. Neuroinflammation is characterized by reactive glial cells that produce pro-inflammatory neurotoxic cytokines. Our previous studies have shown a brain-penetrant Nanoligomer cocktail (NI112) inhibiting the neuroinflammation mediators nuclear factor kappa-light-chain-enhancer of activated B cells (NF-{kappa}B) and NOD-like receptor family, pyrin domain containing 3 (NLRP3) is a safe, targeted, and effective neurotherapeutic drug. Here, we show that a four-week NI112 treatment is therapeutic using: 1) an ALS-FTD 3D human motor neuron organoid model of tar DNA binding protein 43 (TDP-43, a key contributor to ALS pathology) overexpression (knock-in); 2) an AD model of APOE4/APOE4 (AD risk allele) double mutation in human neurons comprising a 3D human prefrontal cortex (PFC) organoid; and 3) multiple in vivo (mouse models) of the same/related conditions. In 3D organoids made from healthy motor neurons (HMN negative control) and TDP-43 overexpressing (or ALS organoids), we monitored the mean firing rate using calcium signaling as a functional output, while measuring TDP-43 and other key neurodegeneration biomarkers. After 4 weeks, we observed a massive improvement in the mean firing rate of NI112-treated ALS organoids compared to untreated ALS organoids, which was more comparable to healthy HMN organoids. Similarly, we found a significant decrease in neurodegeneration markers like amyloid beta 42 (A{beta}42) in NI112-treated AD organoids compared to untreated AD organoids (A{beta}42 comparable to healthy PFC organoids). In the mouse ALS (SOD1-G93A) model, we observed behavioral improvements and restoration of motor function (e.g., grip strength) in NI112-treated mice, and in mouse AD model mice (radiation-induced accelerated neuropathology in APP/PS1, and rTg4510 phospho-tau), we observed improved cognition. In both models, we also found an accompanying reduction in neuroinflammation and reduced neuropathology. These results show the promise for further testing and development of neuroinflammation-targeting Nanoligomers to benefit patients suffering from debilitating neurodegenerative diseases like ALS, FTD, and AD.

neuroscience↗

bioRxiv2024

NF-kB/NLRP3 Translational Inhibition by Nanoligomer Therapy Mitigates Ethanol and Advanced Age-Related Neuroinflammation

Binge alcohol use is increasing among aged adults (>65 years). Alcohol-related toxicity in aged adults is associated with neurodegeneration, yet the molecular underpinnings of age-related sensitivity to alcohol are not well described. Studies utilizing rodent models of neurodegenerative disease reveal heightened activation of Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-{kappa}B) and Nod like receptor 3 (NLRP3) mediate microglia activation and associated neuronal injury. Our group, and others, have implicated hippocampal-resident microglia as key producers of inflammatory mediators, yet the link between inflammation and neurodegeneration has not been established in models of binge ethanol exposure and advanced age. Here, we report binge ethanol increased the proportion of NLRP3+ microglia in the hippocampus of aged (18-20 months) female C57BL/6N mice compared to young (3-4 months). In primary microglia, ethanol-induced expression of reactivity markers and NLRP3 inflammasome activation were more pronounced in microglia from aged mice compared to young. Making use of an NLRP3-specific inhibitor (OLT1177) and a novel brain- penetrant Nanoligomer that inhibits NF-{kappa}B and NLRP3 translation (SB_NI_112), we find ethanol- induced microglial reactivity can be attenuated by OLT1177 and SB_NI_112 in microglia from aged mice. In a model of intermittent binge ethanol exposure, SB_NI_112 prevented ethanol-mediated microglia reactivity, IL-1{beta} production, and tau hyperphosphorylation in the hippocampus of aged mice. These data suggest early indicators of neurodegeneration occurring with advanced age and binge ethanol exposure are NF-{kappa}B- and NLRP3-dependent. Further investigation is warranted to explore the use of targeted immunosuppression via Nanoligomers to attenuate neuroinflammation after alcohol consumption in the aged.

neuroscience↗

bioRxiv2024

Large- and Small-Animal Studies of Safety, Pharmacokinetics (PK), and Biodistribution of Inflammasome-Targeting Nanoligomer in the Brain and Other Target Organs

Immune malfunction or misrecognition of healthy cells and tissue, termed autoimmune disease, is implicated in more than 80 disease conditions and multiple other secondary pathologies. While pan-immunosuppressive therapies like steroids offer some relief for systemic inflammation for some organs, many patients never achieve remission and such drugs do not cross the blood-brain barrier making them ineffective for tackling neuroinflammation. Especially in the brain, unintended activation of microglia and astrocytes is hypothesized to be directly or indirectly responsible for Multiple Sclerosis (MS), Amyotrophic Lateral Sclerosis (ALS), Parkinsons Disease (PD), and Alzheimers Disease (AD). Recent studies have also shown that targeting inflammasome and specific immune targets can be beneficial for these diseases. Further, our previous studies have shown targeting NF-{kappa}B and NLRP3 through brain penetrant Nanoligomer cocktail SB_NI_112 (abbreviated to NI112) can be therapeutic for several neurodegenerative diseases. Here we show safety-toxicity studies, followed by pharmacokinetics (PK) and biodistribution in small- (mice) and large-animal (dog) studies of this inflammasome-targeting Nanoligomer cocktail NI 112. We conducted studies using four different routes of administration: intravenous (IV), subcutaneous (SQ), intraperitoneal (IP), and intranasal (IN), and identified the drug concentration over time using inductively coupled plasma mass spectrometry (ICP-MS) in the blood serum, the brain (including different brain regions), and other target organs like liver, kidney, and colon. Our results indicate the Nanoligomer cocktail has a strong safety profile, and shows high biodistribution (F [~]0.98) and delivery across multiple routes of administration. Further analysis showed high brain bioavailability with a ratio of NI112 in brain tissue to blood serum [~]30%. Our model accurately shows dose scaling, translation between different routes of administration, and interspecies scaling. These results provide an excellent platform for human clinical translation and predicting therapeutic dosage between different routes of administration.

pharmacology and toxicology↗

bioRxiv2024

Targeted-Neuroinflammation Mitigation Using Inflammasome-Inhibiting Nanoligomers is Therapeutic in Experimental Autoimmune Encephalomyelitis (EAE) Mouse Model

Multiple Sclerosis (MS) is a debilitating autoimmune disease that impacts millions of patients worldwide, disproportionately impacting women (4:1), and often presenting at highly productive stages of life. This disease affects the spinal cord and brain, and is characterized by severe neuroinflammation, demyelination, and subsequent neuronal damage, resulting in symptoms like loss of mobility. While untargeted and pan-immunosuppressive therapies have proven to be disease-modifying and manage (or prolong the time between) symptoms in many patients, a significant fraction are unable to achieve remission. Recent work has suggested more targeted neuroinflammation mitigation through selective inflammasome-inhibition can offer relief to patients, while preserving key components of immune function. Here we show a screening of potential therapeutic targets using inflammasome-inhibiting Nanoligomers (NF-{kappa}B1, TNFR1, TNF-, IL-6), that meet or far-exceed commercially available small-molecule counterparts like Ruxolitinib, MCC950, and Deucravacitinib. Using the human brain organoid model, top Nanoligomer combinations (NF-{kappa}B1+TNFR1: NI111, and NF-{kappa}B1+NLRP3: NI112) were shown to significantly reduce neuroinflammation, without any observable negative impact on organoid function. Further testing of these top Nanoligomer combinations in an aggressive Experimental Autoimmune Encephalomyelitis (EAE) mouse model for MS using intraperitoneal (IP) injections showed that NF-{kappa}B1and NLRP3 targeting Nanoligomer combination NI112 rescues mice without observable loss of mobility or disability, minimal inflammation in brain and spinal cord histology, and minimal to no immune cell infiltration of the spinal cord and no demyelination, similar to or at par with mice that received no EAE injections (negative control). Mice receiving NI111 (NF-{kappa}B1+TNFR1) also showed reduced neuroinflammation compared to saline (sham) treated EAE mice and at par/similar to other inflammasome-inhibiting small molecule treatments, although it was significantly higher than NI112 leading to subsequent worsening clinical outcomes. Furthermore, treatment with an oral formulation of NI112 at lower doses showed a significant reduction in EAE severity, albeit with higher variance owing to administration and formulation/fill-and-finish variability. Overall, these results point to the potential of further development and testing these inflammasome targeting Nanoliogmers as an effective neuroinflammation treatment for multiple neurodegenerative diseases, and potentially benefit several patients suffering from such debilitating autoimmune diseases like MS.

neuroscience↗

bioRxiv2024

Predicting the subcellular location of prokaryotic proteins with DeepLocPro

Protein subcellular location prediction is a widely explored task in bioinformatics because of its importance in proteomics research. We propose DeepLocPro, an extension to the popular method DeepLoc, tailored specifically to archaeal and bacterial organisms. DeepLocPro is a multiclass subcellular location prediction tool for prokaryotic proteins, trained on experimentally verified data curated from UniProt and PSORTdb. DeepLocPro compares favorably to the PSORTb 3.0 ensemble method, surpassing its performance across multiple metrics on our benchmark experiment. The DeepLocPro prediction tool is available online at https://ku.biolib.com/deeplocpro and https://services.healthtech.dtu.dk/services/DeepLocPro-1.0/.

bioinformatics↗

bioRxiv2023

Quantitative Three-dimensional Label-free Digital Holographic Imaging of Cardiomyocyte Size, Ploidy, and Cell Division

Cardiac regeneration in newborn rodents depends on the ability of pre-existing cardiomyocytes to proliferate and divide. This capacity is lost within the first week of postnatal development when these cells rapidly switch from hyperplasia to hypertrophy, withdraw from the cell cycle, become binucleated, and increase in size. How these dynamic changes in size and ploidy impact cardiomyocyte proliferative potential is not well understood. In this study, we innovate the application of a commercially available digital holographic imaging microscope, the Holomonitor M4, to evaluate the proliferative responses of mononucleated diploid and binucleated tetraploid cardiomyocytes. This instrument coupled with the powerful Holomonitor App Suite software enables long-term label-free quantitative three-dimensional tracking of primary cardiomyocyte dynamics in real-time with single-cell resolution. Our digital holographic imaging results provide direct evidence that mononucleated cardiomyocytes retain significant proliferative potential as most can successfully divide with high frequency. In contrast, binucleated cardiomyocytes exhibit a blunted response to a proliferative stimulus with the majority not attempting to divide at all. Nevertheless, some binucleated cardiomyocytes were capable of complete division, suggesting that these cells still do retain limited proliferative capacity. By quantitatively tracking cardiomyocyte volume dynamics during these proliferative responses, we reveal that both mononucleated and binucleated cells reach a unique size threshold prior to attempted cell division. The absolute threshold is increased by binucleation, which may limit the ability of binucleated cardiomyocytes to divide. By defining the interrelationship between cardiomyocyte size, ploidy, and cell cycle control, we will better understand the cellular mechanisms that drive the loss of mammalian cardiac regenerative capacity after birth.

cell biology↗

bioRxiv2023

Effects of a field-sprayed antibiotic on bee foraging behavior and pollination in pear orchards

Broadcast spraying of antibiotics in crops is widely used for controlling bacterial plant pathogens. The effects of antibiotics on non-target (and especially beneficial) organisms in cropping systems, however, are not well studied. Pollinators are of particular concern because in pear and apple crops, antibiotics for controlling fire blight (Erwinia amylovora) are sprayed during bloom, likely exposing pollinators. This is especially relevant as laboratory evidence suggests that antibiotics could have sublethal effects on bee foraging behavior and colony health. But to our knowledge these potential impacts have not been studied in field settings. Here, we compared the effects of two fire blight control methods, a single spray of an antibiotic (oxytetracycline) and a biological antagonist (Aureobasidium pullulans), on honey bee (Apis mellifera) foraging, pollination, and fruit set in pear orchards. Complementing these field assessments, we conducted laboratory experiments to examine the effects of these treatments on locomotion and foraging behavior of the bumble bee species, Bombus vosnesenskii. We found that honey bees visited fewer flowers and foraged longer on each flower in orchards sprayed with antibiotics than with biological product, but there were no differences in pollination and seed set. The pear cultivars we worked in, however, can self-pollinate. In the lab, we found that feeding on high doses of either the antibiotic or the biological antagonist reduced bumble bee foraging behavior relative to controls. The limited impact of antibiotics on pear pollination observed in this study suggest that antibiotics pose a low economic risk to pear growers, especially for self-compatible cultivars. Still, crops with higher pollinator dependence may be more affected by reductions in pollinator visitation. Future studies should examine the impacts of multiple antibiotic sprays within a season, which are common during warm springs, and their long-term health impacts on both individual bees and colonies. HighlightsO_LIAntibiotics are sprayed on many crops to control plant bacterial pathogens. C_LIO_LIThe impacts of antibiotics on beneficial organisms in agriculture are unknown. C_LIO_LIWe studied antibiotic impacts on bee behavior and pollination function in pears. C_LIO_LIBees exposed to antibiotics visit fewer flowers and this could impact bee fitness. C_LIO_LIDespite decreased bee visitation, we did not detect a reduction in crop pollination. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=83 HEIGHT=200 SRC="FIGDIR/small/528407v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@17c2189org.highwire.dtl.DTLVardef@7dd952org.highwire.dtl.DTLVardef@4696beorg.highwire.dtl.DTLVardef@52b7fd_HPS_FORMAT_FIGEXP M_FIG C_FIG

animal behavior and cognition↗

bioRxiv2021

Alignment of multiple metabolomics LC-MS datasets from disparate diseases to reveal fever-associated metabolites

Acute febrile illnesses are still a major cause of mortality and morbidity globally, particularly in low to middle income countries. The aim of this study was to determine any possible metabolic commonalities of patients infected with disparate pathogens that cause fever. Three liquid chromatography-mass spectrometry (LC-MS) datasets investigating the metabolic effects of malaria, leishmaniasis and Zika virus infection were used. The retention time (RT) drift between the datasets was determined using landmarks obtained from the standard reference mixtures generally used in the quality control of the LC-MS experiments. We used fitted Gaussian Process models (GPs) to perform a high level correction of the RT drift between the experiments, followed by standard peakset alignment between the samples with corrected RTs of the three LC-MS datasets. Statistical analysis, annotation and pathway analysis of the integrated peaksets were subsequently performed. Metabolic dysregulation patterns common across the datasets were identified, with kynurenine pathway being the most affected pathway between all three fever-associated datasets.

bioinformatics↗

bioRxiv2021

Complexity of genomic epidemiology of carbapenem-resistant Klebsiella pneumoniae isolates in Colombia urges the reinforcement of whole genome sequencing based surveillance programs

BackgroundCarbapenem-resistant Klebsiella pneumoniae (CRKP) is an emerging public health problem. This study explores the specifics of CRKP epidemiology in Colombia based on whole genome sequencing (WGS) of the National Reference Laboratory at Instituto Nacional de Salud (INS)s 2013-2017 sample collection. MethodsA total of 425 CRKP isolates from 21 departments were analyzed by HiSeq-X10(R)Illumina high-throughput sequencing. Bioinformatic analysis was performed, primarily using the pipelines developed collaboratively by the National Institute for Health Research Global Health Research Unit (GHRU) on Genomic Surveillance of AMR, and AGROSAVIA. ResultsOf the 425 CRKP isolates, 91.5% were carbapenemase-producing strains. The data support a recent expansion and the endemicity of CRKP in Colombia with the circulation of 7 high-risk clones, the most frequent being CG258 (48.39% of isolates). We identified genes encoding carbapenemases blaKPC-3, blaKPC-2, blaNDM-1, blaNDM-9, blaVIM-2, blaVIM-4, and blaVIM-24, and various mobile genetic elements (MGE). The virulence of CRKP isolates was low, but colibactin (clb3) was present in 25.2% of isolates, and a hypervirulent CRKP clone (CG380) was reported for the first time in Colombia. ST258, ST512, and ST4851 were characterized by low levels of diversity in the core genome (ANI> 99.9%). ConclusionsThe study outlines complex CRKP epidemiology in Colombia. CG258 expanded clonally and carries specific carbapenemases in specific MGEs, while the other high-risk clones (CG147, CG307, and CG152) present a more diverse complement of carbapenemases. The specifics of the Colombian situation stress the importance of WGS-based surveillance to monitor evolutionary trends of STs, MGE, and resistance and virulence genes. summaryIn Colombia, the dissemination of carbapenemases in carbapenem-resistant Klebsiella pneumoniae is attributed to horizontal gene transfer and successful circulation of CG258, and, to a lesser extent, other clones such as ST307, ST147, and ST152.

genomics↗