A Unique Collateral Artery Development Program Promotes Neonatal Heart Regeneration.

Collateral arteries are an uncommon vessel subtype that can provide alternate blood flow to preserve tissue following vascular occlusion. Some patients with heart disease develop collateral coronary arteries, and this correlates with increased survival. However, it is not known how these collaterals develop or how to stimulate them. We demonstrate that neonatal mouse hearts use a novel mechanism to build collateral arteries in response to injury. Arterial endothelial cells (ECs) migrated away from arteries along existing capillaries and reassembled into collateral arteries, which we termed "artery reassembly". Artery ECs expressed CXCR4, and following injury, capillary ECs induced its ligand, CXCL12. CXCL12 or CXCR4 deletion impaired collateral artery formation and neonatal heart regeneration. Artery reassembly was nearly absent in adults but was induced by exogenous CXCL12. Thus, understanding neonatal regenerative mechanisms can identify pathways that restore these processes in adults and identify potentially translatable therapeutic strategies for ischemic heart disease.

DACH1 stimulates shear stress-guided endothelial cell migration and coronary artery growth through the CXCL12-CXCR4 signaling axis.

Sufficient blood flow to tissues relies on arterial blood vessels, but the mechanisms regulating their development are poorly understood. Many arteries, including coronary arteries of the heart, form through remodeling of an immature vascular plexus in a process triggered and shaped by blood flow. However, little is known about how cues from fluid shear stress are translated into responses that pattern artery development. Here, we show that mice lacking endothelial Dach1 had small coronary arteries, decreased endothelial cell polarization, and reduced expression of the chemokine Cxcl12 Under shear stress in culture, Dach1 overexpression stimulated endothelial cell polarization and migration against flow, which was reversed upon CXCL12/CXCR4 inhibition. In vivo, DACH1 was expressed during early arteriogenesis but was down in mature arteries. Mature artery-type shear stress (high, uniform laminar) specifically down-regulated DACH1, while the remodeling artery-type flow (low, variable) maintained DACH1 expression. Together, our data support a model in which DACH1 stimulates coronary artery growth by activating Cxcl12 expression and endothelial cell migration against blood flow into developing arteries. This activity is suppressed once arteries reach a mature morphology and acquire high, laminar flow that down-regulates DACH1. Thus, we identified a mechanism by which blood flow quality balances artery growth and maturation.

APJ+ cells in the SHF contribute to the cells of aorta and pulmonary trunk through APJ signaling.

Outflow tract (OFT) develops from cardiac progenitor cells in the second heart field (SHF) domain. APJ, a G-Protein Coupled Receptor, is expressed by cardiac progenitors in the SHF. By lineage tracing APJ+SHF cells, we show that these cardiac progenitors contribute to the cells of OFT, which eventually give rise to aorta and pulmonary trunk/artery upon its morphogenesis. Furthermore, we show that early APJ â€‹+ â€‹cells give rise to both aorta and pulmonary cells but late APJ â€‹+ â€‹cells predominantly give rise to pulmonary cells. APJ is expressed by the outflow tract progenitors in the SHF but its role is unclear. We performed knockout studies to determine the role of APJ in SHF cell proliferation and survival. Our data suggested that APJ knockout in the SHF reduced the proliferation of SHF progenitors, while there was no significant impact on survival. In addition, we show that ectopic overexpression of WNT in these cells disrupted aorta and pulmonary morphogenesis from OFT. Overall, our study has identified APJ â€‹+ â€‹progenitor population within the SHF that give rise to aorta and pulmonary trunk/artery cells. Furthermore, we show that APJ signaling stimulates proliferation of these cells in the SHF.

Single-cell analysis of early progenitor cells that build coronary arteries.

Arteries and veins are specified by antagonistic transcriptional programs. However, during development and regeneration, new arteries can arise from pre-existing veins through a poorly understood process of cell fate conversion. Here, using single-cell RNA sequencing and mouse genetics, we show that vein cells of the developing heart undergo an early cell fate switch to create a pre-artery population that subsequently builds coronary arteries. Vein cells underwent a gradual and simultaneous switch from venous to arterial fate before a subset of cells crossed a transcriptional threshold into the pre-artery state. Before the onset of coronary blood flow, pre-artery cells appeared in the immature vessel plexus, expressed mature artery markers, and decreased cell cycling. The vein-specifying transcription factor COUP-TF2 (also known as NR2F2) prevented plexus cells from overcoming the pre-artery threshold by inducing cell cycle genes. Thus, vein-derived coronary arteries are built by pre-artery cells that can differentiate independently of blood flow upon the release of inhibition mediated by COUP-TF2 and cell cycle factors.

Dach1 Extends Artery Networks and Protects Against Cardiac Injury.

[Figure: see text].

Early sowing can boost grain production by reducing weed infestation in organic no-till wheat.

BACKGROUND: Conservative tillage techniques have several agro-ecological benefits for organic farming. The application of these techniques, however, can create quite a few challenges due to the increased weed competition. Here, we report the results of an organic field experiment in which the responses of wheat and weeds to no tillage (NT) were evaluated compared with conventional tillage (CT). We also tested the hypothesis that, under NT, moving up the sowing date, compared with using the ordinary sowing date for the study area, can result in increased competitiveness of the crop against weeds. Two wheat genotypes, a modern variety and an ancient landrace, were tested. RESULTS: Substantial reductions in grain yield and protein content were observed in wheat under NT than under CT when the ordinary sowing date was used. This was mainly due to the considerable increase in weed biomass under NT. The tillage system also altered the composition of weed flora, with some species favored under NT and others under CT. In general, early sowing mitigated the detrimental effect of NT on yield. The two genotypes responded differently to the treatments. The early sowing in the modern variety reduced but did not eliminate the advantages of CT over NT, whereas no appreciable differences in grain yield were observed between CT and NT in the landrace. CONCLUSION: Our results show clearly that, under organic management, using NT alone as a substitute for CT is not agronomically feasible. Moving up the sowing date and using a competitive genotype can help mitigate the negative effects of NT, but surely a more effective application of NT could be achieved by acting simultaneously on other factors of the cropping management system (e.g. crop rotation, fertilization strategy, type of seeder). © 2022 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

NOTCH Activation Promotes Valve Formation by Regulating the Endocardial Secretome.

The endocardium is a specialized endothelium that lines the inner surface of the heart. Functional studies in mice and zebrafish have established that the endocardium is a source of instructive signals for the development of cardiac structures, including the heart valves and chambers. Here, we characterized the NOTCH-dependent endocardial secretome by manipulating NOTCH activity in mouse embryonic endocardial cells (MEEC) followed by mass spectrometry-based proteomics. We profiled different sets of soluble factors whose secretion not only responds to NOTCH activation but also shows differential ligand specificity, suggesting that ligand-specific inputs may regulate the expression of secreted proteins involved in different cardiac development processes. NOTCH signaling activation correlates with a transforming growth factor-ß2 (TGFß2)-rich secretome and the delivery of paracrine signals involved in focal adhesion and extracellular matrix (ECM) deposition and remodeling. In contrast, NOTCH inhibition is accompanied by the up-regulation of specific semaphorins that may modulate cell migration. The secretome protein expression data showed a good correlation with gene profiling of RNA expression in embryonic endocardial cells. Additional characterization by in situ hybridization in mouse embryos revealed expression of various NOTCH candidate effector genes (Tgfß2, Loxl2, Ptx3, Timp3, Fbln2, and Dcn) in heart valve endocardium and/or mesenchyme. Validating these results, mice with conditional Dll4 or Jag1 loss-of-function mutations showed gene expression alterations similar to those observed at the protein level in vitro These results provide the first description of the NOTCH-dependent endocardial secretome and validate MEEC as a tool for assaying the endocardial secretome response to a variety of stimuli and the potential use of this system for drug screening.

Addition of high C:N crop residues to a P-limited substrate constrains the benefits of arbuscular mycorrhizal symbiosis for wheat P and N nutrition.

Many aspects concerning the role of arbuscular mycorrhizal (AM) fungi in plant nutrient uptake from organic sources remain unclear. Here, we investigated the contribution of AM symbiosis to N and P uptake by durum wheat after the addition of a high C:N biomass to a P-limited soil. Plants were grown in pots in the presence or absence of a multispecies AM inoculum, with (Org) or without (Ctr) the addition of 15N-labelled organic matter (OM). A further treatment, in which 15N was applied in mineral form (Ctr+N) in the same amount as that supplied in the Org treatment, was also included. Inoculation with AM had positive effects on plant growth in both control treatments (Ctr and Ctr+N), mainly linked to an increase in plant P uptake. The addition of OM, increasing the P available in the soil for the plants, resulted in a marked decrease in the contribution of AM symbiosis to plant growth and nutrient uptake, although the percentage of mycorrhization was higher in the Org treatment than in the controls. In addition, mycorrhization drastically reduced the recovery of 15N from the OM added to the soil whereas it slightly increased the N recovery from the mineral fertiliser. This suggests that plants and AM fungi probably exert a differential competition for different sources of N available in the soil. On the whole, our results provide a contribution to a better understanding of the conditions under which AM fungi can play an effective role in mitigating the negative effects of nutritional stresses in plants.

Identification of microRNAS differentially regulated by water deficit in relation to mycorrhizal treatment in wheat.

Arbuscular mycorrhizal fungi (AMF) are soil microrganisms that establish symbiosis with plants positively influencing their resistance to abiotic stresses. The aim of this work was to identify wheat miRNAs differentially regulated by water deficit conditions in presence or absence of AMF treatment. Small RNA libraries were constructed for both leaf and root tissues considering four conditions: control (irrigated) or water deficit in presence/absence of mycorrhizal (AMF) treatment. A total of 12 miRNAs were significantly regulated by water deficit in leaves: five in absence and seven in presence of AMF treatment. In roots, three miRNAs were water deficit-modulated in absence of mycorrhizal treatment while six were regulated in presence of it. The most represented miRNA family was miR167 that was regulated by water deficit in both leaf and root tissues. Interestingly, miR827-5p was differentially regulated in leaves in the absence of mycorrhizal treatment while it was water deficit-modulated in roots irrespective of AMF treatment. In roots, water deficit repressed miR827-5p, miR394, miR6187, miR167e-3p, and miR9666b-3p affecting transcription, RNA synthesis, protein synthesis, and protein modifications. In leaves, mycorrhizae modulated miR5384-3p and miR156e-3p affecting trafficking and cell redox homeostasis. DNA replication and transcription regulation should be targeted by the repression of miR1432-5p and miR166h-3p. This work provided interesting insights into the post-transcriptional mechanisms of wheat responses to water deficit in relation to mycorrhizal symbiosis.

Endocardial Notch Signaling in Cardiac Development and Disease.

The Notch signaling pathway is an ancient and highly conserved signaling pathway that controls cell fate specification and tissue patterning in the embryo and in the adult. Region-specific endocardial Notch activity regulates heart morphogenesis through the interaction with multiple myocardial-, epicardial-, and neural crest-derived signals. Mutations in NOTCH signaling elements cause congenital heart disease in humans and mice, demonstrating its essential role in cardiac development. Studies in model systems have provided mechanistic understanding of Notch function in cardiac development, congenital heart disease, and heart regeneration. Notch patterns the embryonic endocardium into prospective territories for valve and chamber formation, and later regulates the signaling processes leading to outflow tract and valve morphogenesis and ventricular trabeculae compaction. Alterations in NOTCH signaling in the endocardium result in congenital structural malformations that can lead to disease in the neonate and adult heart.

Sequential Ligand-Dependent Notch Signaling Activation Regulates Valve Primordium Formation and Morphogenesis.

RATIONALE: The Notch signaling pathway is crucial for primitive cardiac valve formation by epithelial-mesenchymal transition, and NOTCH1 mutations cause bicuspid aortic valve; however, the temporal requirement for the various Notch ligands and receptors during valve ontogeny is poorly understood. OBJECTIVE: The aim of this study is to determine the functional specificity of Notch in valve development. METHODS AND RESULTS: Using cardiac-specific conditional targeted mutant mice, we find that endothelial/endocardial deletion of Mib1-Dll4-Notch1 signaling, possibly favored by Manic-Fringe, is specifically required for cardiac epithelial-mesenchymal transition. Mice lacking endocardial Jag1, Notch1, or RBPJ displayed enlarged valve cusps, bicuspid aortic valve, and septal defects, indicating that endocardial Jag1 to Notch1 signaling is required for post-epithelial-mesenchymal transition valvulogenesis. Valve dysmorphology was associated with increased mesenchyme proliferation, indicating that Jag1-Notch1 signaling restricts mesenchyme cell proliferation non-cell autonomously. Gene profiling revealed upregulated Bmp signaling in Jag1-mutant valves, providing a molecular basis for the hyperproliferative phenotype. Significantly, the negative regulator of mesenchyme proliferation, Hbegf, was markedly reduced in Jag1-mutant valves. Hbegf expression in embryonic endocardial cells could be readily activated through a RBPJ-binding site, identifying Hbegf as an endocardial Notch target. Accordingly, addition of soluble heparin-binding EGF-like growth factor to Jag1-mutant outflow tract explant cultures rescued the hyperproliferative phenotype. CONCLUSIONS: During cardiac valve formation, Dll4-Notch1 signaling leads to epithelial-mesenchymal transition and cushion formation. Jag1-Notch1 signaling subsequently restrains Bmp-mediated valve mesenchyme proliferation by sustaining Hbegf-EGF receptor signaling. Our studies identify a mechanism of signaling cross talk during valve morphogenesis involved in the origin of congenital heart defects associated with reduced NOTCH function.

Stripping the catheter: A new device for an old technique.

Fibroblastic sleeve is a common pathophysiological phenomenon characterized by the formation of a mixed fibrous-collagen tissue encasing the outside of venous access devices. Although it nearly always presents asymptomatically, this catheter-related complication represents one of the leading culprits of venous catheters malfunction. Several techniques have been described for the management of dysfunctional catheters secondary to fibroblastic sleeve, including medical therapy, catheter exchange, balloon angioplasty, and percutaneous stripping. However, there is no common consensus for the treatment management in patients who present contraindications to surgical port removal. This report illustrates the case of a port catheter malfunction due to a fibroblastic sleeve in an oncological patient with a high risk of bleeding. This was effectively treated with a minimally invasive stripping technique using an off-label device for mechanical thrombectomy, namely the ClotTriever system (Inari Medical, Irvine, CA).

Is it a complication or a consequence - a new perspective on adverse outcomes in Interventional Radiology.

The aim of the article is to introduce a new term in post-procedural events related to the procedure itself. All the Societies and Councils report these events as complications and they are divided in mild, moderate and severe or immediate and delayed.On the other hand the term error is known as the application of a wrong plan, or strategy to achieve a goal.For the first time, we are trying to introduce the term "consequence"; assuming that the procedure is the only available and the best fit to clinical indication, a consequence should be seen as an expected and unavoidable occurrence of an "adverse event" despite correct technical execution.

Transcriptome changes induced by Arbuscular mycorrhizal symbiosis in leaves of durum wheat (Triticum durum Desf.) promote higher salt tolerance.

The salinity of soil is a relevant environmental problem around the world, with climate change raising its relevance, particularly in arid and semiarid areas. Arbuscular Mycorrhizal Fungi (AMF) positively affect plant growth and health by mitigating biotic and abiotic stresses, including salt stress. The mechanisms through which these benefits manifest are, however, still unclear. This work aimed to identify key genes involved in the response to salt stress induced by AMF using RNA-Seq analysis on durum wheat (Triticum turgidum L. subsp. durum Desf. Husn.). Five hundred sixty-three differentially expressed genes (DEGs), many of which involved in pathways related to plant stress responses, were identified. The expression of genes involved in trehalose metabolism, RNA processing, vesicle trafficking, cell wall organization, and signal transduction was significantly enhanced by the AMF symbiosis. A downregulation of genes involved in both enzymatic and non-enzymatic oxidative stress responses as well as amino acids, lipids, and carbohydrates metabolisms was also detected, suggesting a lower oxidative stress condition in the AMF inoculated plants. Interestingly, many transcription factor families, including WRKY, NAC, and MYB, already known for their key role in plant abiotic stress response, were found differentially expressed between treatments. This study provides valuable insights on AMF-induced gene expression modulation and the beneficial effects of plant-AMF interaction in durum wheat under salt stress.

Nitrogen fertilization and arbuscular mycorrhizal fungi do not mitigate the adverse effects of soil contamination with polypropylene microfibers on maize growth.

Soil contamination with microplastics may adversely affect soil properties and functions and consequently crop productivity. In this study, we wanted to verify whether the adverse effects of microplastics in the soil on maize plants (Zea mays L.) are due to a reduction in nitrogen (N) availability and a reduced capacity to establish symbiotic relationships with arbuscular mycorrhizal (AM) fungi. To do this, we performed a pot experiment in which a clayey soil was exposed to two environmentally relevant concentrations of polypropylene (PP; one of the most used plastic materials) microfibers (0.4% and 0.8% w/w) with or without the addition of N fertilizer and with or without inoculation with AM fungi. The experiment began after the soil had been incubated at 23 °C for 5 months. Soil contamination with PP considerably reduced maize root and shoot biomass, leaf area, N uptake, and N content in tissue. The adverse effects increased with the concentration of PP in the soil. Adding N to the soil did not alleviate the detrimental effects of PP on plant growth, which suggests that other factors besides N availability played a major role. Similarly, although the presence of PP did not inhibit root colonization by AM fungi (no differences were observed for this trait between the uncontaminated and PP-contaminated soils), the addition of the fungal inoculum to the soil failed to mitigate the negative impact of PP on maize growth. Quite the opposite: mycorrhization further reduced maize root biomass accumulation. Undoubtedly, much research remains to be done to shed light on the mechanisms involved in determining plant behavior in microplastic-contaminated soils, which are most likely complex. This research is a priority given the magnitude of this contamination and its potential implications for human and environmental health.

Endocardium-to-coronary artery differentiation during heart development and regeneration involves sequential roles of Bmp2 and Cxcl12/Cxcr4.

Endocardial cells lining the heart lumen are coronary vessel progenitors during embryogenesis. Re-igniting this developmental process in adults could regenerate blood vessels lost during cardiac injury, but this requires additional knowledge of molecular mechanisms. Here, we use mouse genetics and scRNA-seq to identify regulators of endocardial angiogenesis and precisely assess the role of CXCL12/CXCR4 signaling. Time-specific lineage tracing demonstrated that endocardial cells differentiated into coronary endothelial cells primarily at mid-gestation. A new mouse line reporting CXCR4 activity-along with cell-specific gene deletions-demonstrated it was specifically required for artery morphogenesis rather than angiogenesis. Integrating scRNA-seq data of endocardial-derived coronary vessels from mid- and late-gestation identified a Bmp2-expressing transitioning population specific to mid-gestation. Bmp2 stimulated endocardial angiogenesis in vitro and in injured neonatal mouse hearts. Our data demonstrate how understanding the molecular mechanisms underlying endocardial angiogenesis can identify new potential therapeutic targets promoting revascularization of the injured heart.

Blood flow modeling reveals improved collateral artery performance during the regenerative period in mammalian hearts.

Collateral arteries bridge opposing artery branches, forming a natural bypass that can deliver blood flow downstream of an occlusion. Inducing coronary collateral arteries could treat cardiac ischemia, but more knowledge on their developmental mechanisms and functional capabilities is required. Here we used whole-organ imaging and three-dimensional computational fluid dynamics modeling to define spatial architecture and predict blood flow through collaterals in neonate and adult mouse hearts. Neonate collaterals were more numerous, larger in diameter and more effective at restoring blood flow. Decreased blood flow restoration in adults arose because during postnatal growth coronary arteries expanded by adding branches rather than increasing diameters, altering pressure distributions. In humans, adult hearts with total coronary occlusions averaged 2 large collaterals, with predicted moderate function, while normal fetal hearts showed over 40 collaterals, likely too small to be functionally relevant. Thus, we quantify the functional impact of collateral arteries during heart regeneration and repair-a critical step toward realizing their therapeutic potential.

The Tabula Sapiens: A multiple-organ, single-cell transcriptomic atlas of humans.

Molecular characterization of cell types using single-cell transcriptome sequencing is revolutionizing cell biology and enabling new insights into the physiology of human organs. We created a human reference atlas comprising nearly 500,000 cells from 24 different tissues and organs, many from the same donor. This atlas enabled molecular characterization of more than 400 cell types, their distribution across tissues, and tissue-specific variation in gene expression. Using multiple tissues from a single donor enabled identification of the clonal distribution of T cells between tissues, identification of the tissue-specific mutation rate in B cells, and analysis of the cell cycle state and proliferative potential of shared cell types across tissues. Cell type-specific RNA splicing was discovered and analyzed across tissues within an individual.

Coronary blood vessels from distinct origins converge to equivalent states during mouse and human development.

Most cell fate trajectories during development follow a diverging, tree-like branching pattern, but the opposite can occur when distinct progenitors contribute to the same cell type. During this convergent differentiation, it is unknown if cells 'remember' their origins transcriptionally or whether this influences cell behavior. Most coronary blood vessels of the heart develop from two different progenitor sources-the endocardium (Endo) and sinus venosus (SV)-but whether transcriptional or functional differences related to origin are retained is unknown. We addressed this by combining lineage tracing with single-cell RNA sequencing (scRNAseq) in embryonic and adult mouse hearts. Shortly after coronary development begins, capillary endothelial cells (ECs) transcriptionally segregated into two states that retained progenitor-specific gene expression. Later in development, when the coronary vasculature is well established but still remodeling, capillary ECs again segregated into two populations, but transcriptional differences were primarily related to tissue localization rather than lineage. Specifically, ECs in the heart septum expressed genes indicative of increased local hypoxia and decreased blood flow. Adult capillary ECs were more homogeneous with respect to both lineage and location. In agreement, SV- and Endo-derived ECs in adult hearts displayed similar responses to injury. Finally, scRNAseq of developing human coronary vessels indicated that the human heart followed similar principles. Thus, over the course of development, transcriptional heterogeneity in coronary ECs is first influenced by lineage, then by location, until heterogeneity declines in the homeostatic adult heart. These results highlight the plasticity of ECs during development, and the validity of the mouse as a model for human coronary development.

Nitrogen Type and Availability Drive Mycorrhizal Effects on Wheat Performance, Nitrogen Uptake and Recovery, and Production Sustainability.

Plant performance is strongly dependent on nitrogen (N), and thus increasing N nutrition is of great relevance for the productivity of agroecosystems. The effects of arbuscular mycorrhizal (AM) fungi on plant N acquisition are debated because contradictory results have been reported. Using 15N-labeled fertilizers as a tracer, we evaluated the effects of AM fungi on N uptake and recovery from mineral or organic sources in durum wheat. Under sufficient N availability, AM fungi had no effects on plant biomass but increased N concentrations in plant tissue, plant N uptake, and total N recovered from the fertilizer. In N-deficient soil, AM fungi led to decreased aboveground biomass, which suggests that plants and AM fungi may have competed for N. When the organic source had a low C:N ratio, AM fungi favored both plant N uptake and N recovery. In contrast, when the organic source had a high C:N ratio, a clear reduction in N recovery from the fertilizer was observed. Overall, the results indicate an active role of arbuscular mycorrhizae in favoring plant N-related traits when N is not a limiting factor and show that these fungi help in N recovery from the fertilizer. These results hold great potential for increasing the sustainability of durum wheat production.