Erratum: Altered gene expression linked to germline dysfunction following exposure to DEET.

[This corrects the article DOI: 10.1016/j.isci.2023.108699.].

Altered gene expression linked to germline dysfunction following exposure of Caenorhabditis elegans to DEET.

N,N-diethyl-meta-toluamide (DEET) is a commonly used synthetic insect repellent. Although the neurological effects of DEET have been widely investigated, its effects on the germline are less understood. Here, we show that exposure of the nematode Caenorhabditis elegans, which is highly predictive of mammalian reprotoxicity, resulting in internal DEET levels within the range detected in human biological samples, causes activation of p53/CEP-1-dependent germ cell apoptosis, altered meiotic recombination, chromosome abnormalities, and missegregation. RNA-sequencing analysis links DEET-induced alterations in the expression of genes related to redox processes and chromatin structure to reduced mitochondrial function, impaired DNA double-strand break repair progression, and defects during early embryogenesis. We propose that Caenorhabditis elegans exposure to DEET interferes with gene expression, leading to increased oxidative stress and altered chromatin structure, resulting in germline effects that pose a risk to reproductive health.

Evolution of Plasmodium vivax and resistance patterns for infection based on Duffy genotype and phenotype.

The Duffy protein, a transmembrane molecule, acts as a receptor for various chemokines and facilitates binding between reticulocytes and the Plasmodium Duffy antigen binding protein. Duffy expression is associated with the Duffy chemokine receptor antigen genotype on chromosome 1 and exhibits variation across different geographic regions. Traditionally, the Duffy negative genotype and phenotype have been described to confer a certain level of protection against infection and symptom development. However, recent data suggest a shift in this behavior, with significantly higher prevalence observed in individuals with Duffy negative genotype or phenotype. Given that malaria is an endemic vector-borne disease in regions of Asia, Africa, and Latin America, posing a substantial global burden of disease and prioritizing public and global health, identifying evolutionary changes in infection and resistance patterns holds great importance for the design of strategies and reevaluation of conventional interventions. Hence, the aim of this review was to analyze the evolution of Plasmodium vivax and infection resistance patterns based on Duffy genotype and phenotype. The distribution of genotypes, phenotypes, and polymorphisms of P. vivax ligands and erythrocyte receptors varies geographically, notably resistance patterns of this microorganism in individuals with Duffy negative genotype and phenotype have significantly changed compared to studies conducted 30 years ago. The prevalence of vivax malaria in individuals with a Duffy negative status can reach up to 100%. Consequently, prioritizing research on this topic is essential for public health.

Skin-type-dependent development of murine mechanosensory neurons.

Mechanosensory neurons innervating the skin underlie our sense of touch. Fast-conducting, rapidly adapting mechanoreceptors innervating glabrous (non-hairy) skin form Meissner corpuscles, while in hairy skin, they associate with hair follicles, forming longitudinal lanceolate endings. How mechanoreceptors develop axonal endings appropriate for their skin targets is unknown. We report that mechanoreceptor morphologies across different skin regions are indistinguishable during early development but diverge post-natally, in parallel with skin maturation. Neurons terminating along the glabrous and hairy skin border exhibit hybrid morphologies, forming both Meissner corpuscles and lanceolate endings. Additionally, molecular profiles of neonatal glabrous and hairy skin-innervating neurons largely overlap. In mouse mutants with ectopic glabrous skin, mechanosensory neurons form end-organs appropriate for the altered skin type. Finally, BMP5 and BMP7 are enriched in glabrous skin, and signaling through type I bone morphogenetic protein (BMP) receptors in neurons is critical for Meissner corpuscle morphology. Thus, mechanoreceptor morphogenesis is flexibly instructed by target tissues.

Can Probiotics, Particularly Limosilactobacillus fermentum UCO-979C and Lacticaseibacillus rhamnosus UCO-25A, Be Preventive Alternatives against SARS-CoV-2?

COVID-19, an infection produced by the SARS-CoV-2 virus in humans, has rapidly spread to become a high-mortality pandemic. SARS-CoV-2 is a single-stranded RNA virus characterized by infecting epithelial cells of the intestine and lungs, binding to the ACE2 receptor present on epithelial cells. COVID-19 treatment is based on antivirals and antibiotics against symptomatology in addition to a successful preventive strategy based on vaccination. At this point, several variants of the virus have emerged, altering the effectiveness of treatments and thereby attracting attention to several alternative therapies, including immunobiotics, to cope with the problem. This review, based on articles, patents, and an in silico analysis, aims to address our present knowledge of the COVID-19 disease, its symptomatology, and the possible beneficial effects for patients if probiotics with the characteristics of immunobiotics are used to confront this disease. Moreover, two probiotic strains, L. fermentum UCO-979C and L. rhamnosus UCO-25A, with different effects demonstrated at our laboratory, are emphasized. The point of view of this review highlights the possible benefits of probiotics, particularly those associated with immunomodulation as well as the production of secondary metabolites, and their potential targets during SARS-CoV-2 infection.

Low-dose interleukin-2 promotes immune regulation in face transplantation: A pilot study.

Face transplantation is a life-changing procedure for patients with severe composite facial defects. However, it is hampered by high acute rejection rates due to the immunogenicity of skin allograft and toxicity linked to high doses of immunosuppression. To reduce immunosuppression-associated complications, we, for the first time in face transplant recipients, used low-dose interleukin 2 (IL-2) therapy to expand regulatory T cells (Tregs) in vivo and to enhance immune modulation, under close immunological monitoring of peripheral blood and skin allograft. Low-dose IL-2 achieved a sustained expansion (∼4-fold to 5-fold) of circulating Tregs and a reduction (∼3.5-fold) of B cells. Post-IL-2 Tregs exhibited greater suppressive function, characterized by higher expression of TIM-3 and LAG3co-inhibitory molecules. In the skin allograft, Tregs increased after low-dose IL-2 therapy. IL-2 induced a distinct molecular signature in the allograft with reduced cytotoxicity-associated genes (granzyme B and perforin). Two complications were observed during the trial: one rejection event and an episode of autoimmune hemolytic anemia. In summary, this initial experience demonstrated that low-dose IL-2 therapy was not only able to promote immune regulation in face transplant recipients but also highlighted challenges related to its narrow therapeutic window. More specific targeted Treg expansion strategies are needed to translate this approach to the clinic.

Cambio rutinario de instrumental y guantes quirúrgicos en cirugía abdominal: ¿cuál es el impacto sobre el riesgo de infección? / Routine change of surgical instruments and gloves in abdominal surgery: what is the impact on the risk of infection?

Objetivo: Analizar la evidencia más actualizada sobre el cambio rutinario de instrumental y guantes quirúrgicos en cirugía abdominal, y su impacto en el riesgo de infecciones. Métodos: Se realizó una búsqueda bibliográfica, en las bases de datos PubMed, ScienceDirect, Web of Science, y MEDLINE. Resultados: A la fecha, la evidencia sumamente escasa sobre el potencial impacto del cambio rutinario de instrumental y guantes quirúrgicos en cirugía abdominal, y su relación con la incidencia de infección en el sitio operatorio. Sin embargo, no deja de ser un tema de interés en cirugía global. El estudio ChEETAh, ensayo realizado en siete países de bajos y medianos ingresos, que evaluó el cambio rutinario tanto de guantes como de instrumental quirúrgico en cirugía abdominal y su relación con la infección, demostró que, la frecuencia de infección en sitio operatorio fue del 16% (n=931) en el grupo intervención, comparado a un 18,9% (n=1280) en el grupo control (RR 0,87; IC 95%: 0,79 ­ 0,95; p=0,0032). Así, podría existir cierta protección adicional con el cambio rutinario de guantes e instrumental en cirugía abdominal. Conclusión: Aunque la evidencia es limitada y heterogénea, existe una tendencia respecto a un potencial beneficio frente a la incidencia de infección en sitio operatorio, en el cambio rutinario de guantes e instrumental quirúrgico en cirugía abdominal(AU)

Objective: To analyze the most recent evidence regarding the routine change of surgical instruments and gloves in abdominal surgery and its impact on the risk of infections. Methods: A literature search was conducted in the PubMed, ScienceDirect, Web of Science, and MEDLINE databases. Results: To date, the evidence regarding the potential impact of routine changes in surgical instruments and gloves in abdominal surgery and their relationship with the incidence of surgical site infections is extremely scarce. Nevertheless, it remains a topic of interest in global surgery. The ChEETAh study, conducted in seven low and middle-income countries, which assessed the routine change of both gloves and surgical instruments in abdominal surgery and its relation to infection, demonstrated that the frequency of surgical site infection was 16% (n=931) in the intervention group compared to 18.9% (n=1280) in the control group (RR 0.87; 95% CI: 0.79 ­ 0.95; p=0.0032). Thus, there may be some additional protection with the routine change of gloves and instruments in abdominal surgery. Conclusion: Although the evidence is limited and heterogeneous, there is a trend suggesting a potential benefit in reducing the incidence of surgical site infections through the routine change of gloves and surgical instruments in abdominal surgery(AU)

Ehf and Fezf2 regulate late medullary thymic epithelial cell and thymic tuft cell development.

Thymic epithelial cells are indispensable for T cell maturation and selection and the induction of central immune tolerance. The self-peptide repertoire expressed by medullary thymic epithelial cells is in part regulated by the transcriptional regulator Aire (Autoimmune regulator) and the transcription factor Fezf2. Due to the high complexity of mTEC maturation stages (i.e., post-Aire, Krt10+ mTECs, and Dclk1+ Tuft mTECs) and the heterogeneity in their gene expression profiles (i.e., mosaic expression patterns), it has been challenging to identify the additional factors complementing the transcriptional regulation. We aimed to identify the transcriptional regulators involved in the regulation of mTEC development and self-peptide expression in an unbiased and genome-wide manner. We used ATAC footprinting analysis as an indirect approach to identify transcription factors involved in the gene expression regulation in mTECs, which we validated by ChIP sequencing. This study identifies Fezf2 as a regulator of the recently described thymic Tuft cells (i.e., Tuft mTECs). Furthermore, we identify that transcriptional regulators of the ELF, ESE, ERF, and PEA3 subfamily of the ETS transcription factor family and members of the Krüppel-like family of transcription factors play a role in the transcriptional regulation of genes involved in late mTEC development and promiscuous gene expression.

Molecular convergence between Down syndrome and fragile X syndrome identified using human pluripotent stem cell models.

Down syndrome (DS), driven by an extra copy of chromosome 21 (HSA21), and fragile X syndrome (FXS), driven by loss of the RNA-binding protein FMRP, are two common genetic causes of intellectual disability and autism. Based upon the number of DS-implicated transcripts bound by FMRP, we hypothesize that DS and FXS may share underlying mechanisms. Comparing DS and FXS human pluripotent stem cell (hPSC) and glutamatergic neuron models, we identify increased protein expression of select targets and overlapping transcriptional perturbations. Moreover, acute upregulation of endogenous FMRP in DS patient cells using CRISPRa is sufficient to significantly reduce expression levels of candidate proteins and reverse 40% of global transcriptional perturbations. These results pinpoint specific molecular perturbations shared between DS and FXS that can be leveraged as a strategy for target prioritization; they also provide evidence for the functional relevance of previous associations between FMRP targets and disease-implicated genes.

Whole-genome CRISPR screening identifies genetic manipulations to reduce immune rejection of stem cell-derived islets.

Human embryonic stem cells (hESCs) provide opportunities for cell replacement therapy of insulin-dependent diabetes. Therapeutic quantities of human stem cell-derived islets (SC-islets) can be produced by directed differentiation. However, preventing allo-rejection and recurring autoimmunity, without the use of encapsulation or systemic immunosuppressants, remains a challenge. An attractive approach is to transplant SC-islets, genetically modified to reduce the impact of immune rejection. To determine the underlying forces that drive immunogenicity of SC-islets in inflammatory environments, we performed single-cell RNA sequencing (scRNA-seq) and whole-genome CRISPR screen of SC-islets under immune interaction with allogeneic peripheral blood mononuclear cells (PBMCs). Data analysis points to "alarmed" populations of SC-islets that upregulate genes in the interferon (IFN) pathway. The CRISPR screen in vivo confirms that targeting IFNγ-induced mediators has beneficial effects on SC-islet survival under immune attack. Manipulating the IFN response by depleting chemokine ligand 10 (CXCL10) in SC-islet grafts confers improved survival against allo-rejection compared with wild-type grafts in humanized mice. These results offer insights into the nature of immune destruction of SC-islets during allogeneic responses and provide targets for gene editing.

Endothelial cell-specific deletion of a microRNA accelerates atherosclerosis.

BACKGROUND AND AIMS: Chronic vascular endothelial inflammation predisposes to atherosclerosis; however, the cell-autonomous roles for endothelial-expressing microRNAs (miRNAs) are poorly understood in this process. MiR-181b is expressed in several cellular constituents relevant to lesion formation. The aim of this study is to examine the role of genetic deficiency of the miR-181b locus in endothelial cells during atherogenesis. METHODS AND RESULTS: Using a proprotein convertase subtilisin/kexin type 9 (PCSK9)-induced atherosclerosis mouse model, we demonstrated that endothelial cell (EC)-specific deletion of miR-181a2b2 significantly promoted atherosclerotic lesion formation, cell adhesion molecule expression, and the influx of lesional macrophages in the vessel wall. Yet, endothelium deletion of miR-181a2b2 did not affect body weight, lipid metabolism, anti-inflammatory Ly6Clow or the pro-inflammatory Ly6Cinterm and Ly6Chigh fractions in circulating peripheral blood mononuclear cells (PBMCs), and pro-inflammatory or anti-inflammatory mediators in both bone marrow (BM) and PBMCs. Mechanistically, bulk RNA-seq and gene set enrichment analysis of ECs enriched from the aortic arch intima, as well as single cell RNA-seq from atherosclerotic lesions, revealed that endothelial miR-181a2b2 serves as a critical regulatory hub in controlling endothelial inflammation, cell adhesion, cell cycle, and immune response during atherosclerosis. CONCLUSIONS: Our study establishes that deficiency of a miRNA specifically in the vascular endothelium is sufficient to profoundly impact atherogenesis. Endothelial miR-181a2b2 deficiency regulates multiple key pathways related to endothelial inflammation, cell adhesion, cell cycle, and immune response involved in the development of atherosclerosis.

Perivascular Fibrosis Is Mediated by a KLF10-IL-9 Signaling Axis in CD4+ T Cells.

BACKGROUND: Perivascular fibrosis, characterized by increased amount of connective tissue around vessels, is a hallmark for vascular disease. Ang II (angiotensin II) contributes to vascular disease and end-organ damage via promoting T-cell activation. Despite recent data suggesting the role of T cells in the progression of perivascular fibrosis, the underlying mechanisms are poorly understood. METHODS: TF (transcription factor) profiling was performed in peripheral blood mononuclear cells of hypertensive patients. CD4-targeted KLF10 (Kruppel like factor 10)-deficient (Klf10fl/flCD4Cre+; [TKO]) and CD4-Cre (Klf10+/+CD4Cre+; [Cre]) control mice were subjected to Ang II infusion. End point characterization included cardiac echocardiography, aortic imaging, multiorgan histology, flow cytometry, cytokine analysis, aorta and fibroblast transcriptomic analysis, and aortic single-cell RNA-sequencing. RESULTS: TF profiling identified increased KLF10 expression in hypertensive human subjects and in CD4+ T cells in Ang II-treated mice. TKO mice showed enhanced perivascular fibrosis, but not interstitial fibrosis, in aorta, heart, and kidney in response to Ang II, accompanied by alterations in global longitudinal strain, arterial stiffness, and kidney function compared with Cre control mice. However, blood pressure was unchanged between the 2 groups. Mechanistically, KLF10 bound to the IL (interleukin)-9 promoter and interacted with HDAC1 (histone deacetylase 1) inhibit IL-9 transcription. Increased IL-9 in TKO mice induced fibroblast intracellular calcium mobilization, fibroblast activation, and differentiation and increased production of collagen and extracellular matrix, thereby promoting the progression of perivascular fibrosis and impairing target organ function. Remarkably, injection of anti-IL9 antibodies reversed perivascular fibrosis in Ang II-infused TKO mice and C57BL/6 mice. Single-cell RNA-sequencing revealed fibroblast heterogeneity with activated signatures associated with robust ECM (extracellular matrix) and perivascular fibrosis in Ang II-treated TKO mice. CONCLUSIONS: CD4+ T cell deficiency of Klf10 exacerbated perivascular fibrosis and multi-organ dysfunction in response to Ang II via upregulation of IL-9. Klf10 or IL-9 in T cells might represent novel therapeutic targets for treatment of vascular or fibrotic diseases.

Hepatic mTORC1 signaling activates ATF4 as part of its metabolic response to feeding and insulin.

OBJECTIVE: The mechanistic target of rapamycin complex 1 (mTORC1) is dynamically regulated by fasting and feeding cycles in the liver to promote protein and lipid synthesis while suppressing autophagy. However, beyond these functions, the metabolic response of the liver to feeding and insulin signaling orchestrated by mTORC1 remains poorly defined. Here, we determine whether ATF4, a stress responsive transcription factor recently found to be independently regulated by mTORC1 signaling in proliferating cells, is responsive to hepatic mTORC1 signaling to alter hepatocyte metabolism. METHODS: ATF4 protein levels and expression of canonical gene targets were analyzed in the liver following fasting and physiological feeding in the presence or absence of the mTORC1 inhibitor, rapamycin. Primary hepatocytes from wild-type or liver-specific Atf4 knockout (LAtf4KO) mice were used to characterize the effects of insulin-stimulated mTORC1-ATF4 function on hepatocyte gene expression and metabolism. Both unbiased steady-state metabolomics and stable-isotope tracing methods were employed to define mTORC1 and ATF4-dependent metabolic changes. RNA-sequencing was used to determine global changes in feeding-induced transcripts in the livers of wild-type versus LAtf4KO mice. RESULTS: We demonstrate that ATF4 and its metabolic gene targets are stimulated by mTORC1 signaling in the liver, in a hepatocyte-intrinsic manner by insulin in response to feeding. While we demonstrate that de novo purine and pyrimidine synthesis is stimulated by insulin through mTORC1 signaling in primary hepatocytes, this regulation was independent of ATF4. Metabolomics and metabolite tracing studies revealed that insulin-mTORC1-ATF4 signaling stimulates pathways of nonessential amino acid synthesis in primary hepatocytes, including those of alanine, aspartate, methionine, and cysteine, but not serine. CONCLUSIONS: The results demonstrate that ATF4 is a novel metabolic effector of mTORC1 in the liver, extending the molecular consequences of feeding and insulin-induced mTORC1 signaling in this key metabolic tissue to the control of amino acid metabolism.

Immunoregulatory and lipid presentation pathways are upregulated in human face transplant rejection.

BACKGROUNDRejection is the primary barrier to broader implementation of vascularized composite allografts (VCAs), including face and limb transplants. The immunologic pathways activated in face transplant rejection have not been fully characterized.METHODSUsing skin biopsies prospectively collected over 9 years from 7 face transplant patients, we studied rejection by gene expression profiling, histology, immunostaining, and T cell receptor sequencing.RESULTSGrade 1 rejection did not differ significantly from nonrejection, suggesting that it does not represent a pathologic state. In grade 2, there was a balanced upregulation of both proinflammatory T cell activation pathways and antiinflammatory checkpoint and immunomodulatory pathways, with a net result of no tissue injury. In grade 3, IFN-γ-driven inflammation, antigen-presenting cell activation, and infiltration of the skin by proliferative T cells bearing markers of antigen-specific activation and cytotoxicity tipped the balance toward tissue injury. Rejection of VCAs and solid organ transplants had both distinct and common features. VCA rejection was uniquely associated with upregulation of immunoregulatory genes, including SOCS1; induction of lipid antigen-presenting CD1 proteins; and infiltration by T cells predicted to recognize CD1b and CD1c.CONCLUSIONOur findings suggest that the distinct features of VCA rejection reflect the unique immunobiology of skin and that enhancing cutaneous immunoregulatory networks may be a useful strategy in combatting rejection.Trial registrationClinicalTrials.gov NCT01281267.FUNDINGAssistant Secretary of Defense and Health Affairs, through Reconstructive Transplant Research (W81XWH-17-1-0278, W81XWH-16-1-0647, W81XWH-16-1-0689, W81XWH-18-1-0784, W81XWH-1-810798); American Society of Transplantation's Transplantation and Immunology Research Network Fellowship Research Grant; Plastic Surgery Foundation Fellowship from the American Society of Plastic Surgeons; Novo Nordisk Foundation (NNF15OC0014092); Lundbeck Foundation; Aage Bangs Foundation; A.P. Moller Foundation for the Advancement of Medical Science; NIH UL1 RR025758.

Drosophila PDGF/VEGF signaling from muscles to hepatocyte-like cells protects against obesity.

PDGF/VEGF ligands regulate a plethora of biological processes in multicellular organisms via autocrine, paracrine, and endocrine mechanisms. We investigated organ-specific metabolic roles of Drosophila PDGF/VEGF-like factors (Pvfs). We combine genetic approaches and single-nuclei sequencing to demonstrate that muscle-derived Pvf1 signals to the Drosophila hepatocyte-like cells/oenocytes to suppress lipid synthesis by activating the Pi3K/Akt1/TOR signaling cascade in the oenocytes. Functionally, this signaling axis regulates expansion of adipose tissue lipid stores in newly eclosed flies. Flies emerge after pupation with limited adipose tissue lipid stores and lipid level is progressively accumulated via lipid synthesis. We find that adult muscle-specific expression of pvf1 increases rapidly during this stage and that muscle-to-oenocyte Pvf1 signaling inhibits expansion of adipose tissue lipid stores as the process reaches completion. Our findings provide the first evidence in a metazoan of a PDGF/VEGF ligand acting as a myokine that regulates systemic lipid homeostasis by activating TOR in hepatocyte-like cells.

A single-cell survey of Drosophila blood.

Drosophila blood cells, called hemocytes, are classified into plasmatocytes, crystal cells, and lamellocytes based on the expression of a few marker genes and cell morphologies, which are inadequate to classify the complete hemocyte repertoire. Here, we used single-cell RNA sequencing (scRNA-seq) to map hemocytes across different inflammatory conditions in larvae. We resolved plasmatocytes into different states based on the expression of genes involved in cell cycle, antimicrobial response, and metabolism together with the identification of intermediate states. Further, we discovered rare subsets within crystal cells and lamellocytes that express fibroblast growth factor (FGF) ligand branchless and receptor breathless, respectively. We demonstrate that these FGF components are required for mediating effective immune responses against parasitoid wasp eggs, highlighting a novel role for FGF signaling in inter-hemocyte crosstalk. Our scRNA-seq analysis reveals the diversity of hemocytes and provides a rich resource of gene expression profiles for a systems-level understanding of their functions.

Unification of miRNA and isomiR research: the mirGFF3 format and the mirtop API.

MOTIVATION: MicroRNAs (miRNAs) are small RNA molecules (∼22 nucleotide long) involved in post-transcriptional gene regulation. Advances in high-throughput sequencing technologies led to the discovery of isomiRs, which are miRNA sequence variants. While many miRNA-seq analysis tools exist, the diversity of output formats hinders accurate comparisons between tools and precludes data sharing and the development of common downstream analysis methods. RESULTS: To overcome this situation, we present here a community-based project, miRNA Transcriptomic Open Project (miRTOP) working towards the optimization of miRNA analyses. The aim of miRTOP is to promote the development of downstream isomiR analysis tools that are compatible with existing detection and quantification tools. Based on the existing GFF3 format, we first created a new standard format, mirGFF3, for the output of miRNA/isomiR detection and quantification results from small RNA-seq data. Additionally, we developed a command line Python tool, mirtop, to create and manage the mirGFF3 format. Currently, mirtop can convert into mirGFF3 the outputs of commonly used pipelines, such as seqbuster, isomiR-SEA, sRNAbench, Prost! as well as BAM files. Some tools have also incorporated the mirGFF3 format directly into their code, such as, miRge2.0, IsoMIRmap and OptimiR. Its open architecture enables any tool or pipeline to output or convert results into mirGFF3. Collectively, this isomiR categorization system, along with the accompanying mirGFF3 and mirtop API, provide a comprehensive solution for the standardization of miRNA and isomiR annotation, enabling data sharing, reporting, comparative analyses and benchmarking, while promoting the development of common miRNA methods focusing on downstream steps of miRNA detection, annotation and quantification. AVAILABILITY AND IMPLEMENTATION: https://github.com/miRTop/mirGFF3/ and https://github.com/miRTop/mirtop. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Metastable Atrial State Underlies the Primary Genetic Substrate for MYL4 Mutation-Associated Atrial Fibrillation.

BACKGROUND: Atrial fibrillation (AF) is the most common clinical arrhythmia and is associated with heart failure, stroke, and increased mortality. The myocardial substrate for AF is poorly understood because of limited access to primary human tissue and mechanistic questions around existing in vitro or in vivo models. METHODS: Using an MYH6:mCherry knock-in reporter line, we developed a protocol to generate and highly purify human pluripotent stem cell-derived cardiomyocytes displaying physiological and molecular characteristics of atrial cells. We modeled human MYL4 mutants, one of the few definitive genetic causes of AF. To explore non-cell-autonomous components of AF substrate, we also created a zebrafish Myl4 knockout model, which exhibited molecular, cellular, and physiologic abnormalities that parallel those in humans bearing the cognate mutations. RESULTS: There was evidence of increased retinoic acid signaling in both human embryonic stem cells and zebrafish mutant models, as well as abnormal expression and localization of cytoskeletal proteins, and loss of intracellular nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide + hydrogen. To identify potentially druggable proximate mechanisms, we performed a chemical suppressor screen integrating multiple human cellular and zebrafish in vivo endpoints. This screen identified Cx43 (connexin 43) hemichannel blockade as a robust suppressor of the abnormal phenotypes in both models of MYL4 (myosin light chain 4)-related atrial cardiomyopathy. Immunofluorescence and coimmunoprecipitation studies revealed an interaction between MYL4 and Cx43 with altered localization of Cx43 hemichannels to the lateral membrane in MYL4 mutants, as well as in atrial biopsies from unselected forms of human AF. The membrane fraction from MYL4-/- human embryonic stem cell derived atrial cells demonstrated increased phospho-Cx43, which was further accentuated by retinoic acid treatment and by the presence of risk alleles at the Pitx2 locus. PKC (protein kinase C) was induced by retinoic acid, and PKC inhibition also rescued the abnormal phenotypes in the atrial cardiomyopathy models. CONCLUSIONS: These data establish a mechanistic link between the transcriptional, metabolic and electrical pathways previously implicated in AF substrate and suggest novel avenues for the prevention or therapy of this common arrhythmia.

Desempeño de bancos forrajeros de Cratylia argentea (Desv.) Kuntze, en suelos degradados en el departamento de Casanare / Performance of the Cratylia argentea (Desv.) Kuntze Forage Banks in Degraded Soils in the Casanare Province / Desempenho de bancos de forragem de Cratylia argentea (Desv.) Kuntze, em solos degradados no departamento de Casanare

Resumen Las épocas de sequía o lluvias extremas afectan la eficiencia en los sistemas ganaderos, los cuales basan la producción de forrajes en el monocultivo de gramíneas, estas son más vulnerables a las condiciones climáticas extremas, lo que hace necesario buscar alternativas de plantas más resistentes a estas condiciones. Se determinó el desempeño de Cratylia argentea en bancos forrajeros, establecidos en suelos degradados con alto uso de agroquímicos, bajo condiciones de bosque húmedo tropical. El estudio se realizó durante un año y se estableció en la finca Matapantano, ubicada en Yopal, Casanare (Colombia), se determinó el efecto sobre las propiedades físico-químicas y macrofauna del suelo, producción de forraje verde y materia seca, relación hoja-tallo, calidad nutricional de la planta completa y fracciones (hoja-tallo). Se utilizó un diseño al azar y el software Infostat® para estadística descriptiva y análisis de varianza no paramétrica. Se presentaron incrementos en los contenidos de minerales, materia orgánica, carbono orgánico y macrofauna del suelo, se encontraron diferencias en producción de materia seca de las fracciones de la planta (p < 0.0001), en la producción de materia seca entre cortes (p < 0.0001) y en la relación hoja-tallo (p < 0.0001), el forraje presentó buena calidad nutricional, siendo las hojas la fracción de mejor calidad, C. argentea resistió los cambios drásticos en las condiciones climáticas. Se concluye que esta especie, presenta potencial para la recuperación de suelos degradados con intenso uso de agroquímicos, puesto que produce forraje de buena calidad para la suplementación animal en sistemas ganaderos, aun en épocas de sequía.

Abstract Extreme drought or rainy seasons affect the efficiency in the stockbreeding systems whose forage production is based on the grass monoculture. These species are more vulnerable to the extreme climate conditions, which requires looking for alternative plants with higher resistance to these conditions. This work studied the Cratylia argentea performance in forage banks used in degraded soils with high amounts of agrochemicals in areas of tropical rainforest. The study took one year and was conducted in the estate Matapantano, located in Yopal, Casanare (Colombia). The effect on the physical-chemical properties and macrofauna in this soil, the green forage and dry mass production, the leaf-stalk relationship, the nutritional quality of both fractions (leaf-stalk) and the whole plant were studied. A random design and the software Infostat® were used for the descriptive statistics and the non-parametric variance analysis. The soil showed increases in the contents of minerals, organic mass, organic carbon and macrofauna. Differences were observed in the dry mass production from the plant fractions (p < 0.0001), in the dry mass production between cuts (p < 0.0001) and in the leaf-stalk relationship (p < 0.0001). The forage showed a good nutritional quality and the leaves were the fraction with best quality. C. argentea resisted the extreme changes in the climate conditions. It is concluded that the studied species has a good potential for the recovery of degraded soils with intense use of agrochemicals as it produced good quality forage to feed animals in stockbreeding systems, even in the drought season.

Resumo As secas ou chuvas extremas afetam a eficiência dos sistemas pecuários, os quais baseiam a produção de forragem na monocultura de gramíneas, são mais vulneráveis às condições climáticas extremas, o que faz necessário procurar alternativas de plantas mais resistentes a essas condições. O desempenho de Cratylia argentea foi determinado em bancos de forragem estabelecidos em solos degradados com alto uso de agroquímicos, em condições de floresta húmida tropical. O estudo foi realizado por um ano na finca Matapantano, localizada em Yopal, Casanare (Colômbia). Determinou-se o efeito nas propriedades físico-químicas e macrofauna do solo, produção de forragem verde e matéria seca, relação folha/ caule, qualidade nutricional da planta completa e frações (folha/caule). Utilizou-se delineação aleatória e software Infostat® para estatística descritiva e análise de variância não paramétrica. Houve acréscimos nos teores de minerais, matéria orgânica, carbono orgânico e macrofauna do solo, encontraram-se diferenças na produção de matéria seca das frações da planta (p < 0.0001), na produção de matéria seca entre os cortes (p < 0.0001) e na relação folha-caule (p < 0.0001), a forragem apresentou boa qualidade nutricional, sendo as folhas a fracção de melhor qualidade, C. argentea resistiu às mudanças drásticas nas condições climáticas. Conclui-se que esta espécie apresenta potencial para a recuperação de solos degradados com intenso uso de agroquímicos, uma vez que produz forragem de boa qualidade para a suplementação animal em sistemas pecuários mesmo em épocas de seca.