Aging is a major risk factor for impaired cardiovascular health. Because the aging myocardium is characterized by microcirculatory dysfunction, and because nerves align with vessels, we assessed the impact of aging on the cardiac neurovascular interface. We report that aging reduces nerve density in the ventricle and dysregulates vascular-derived neuroregulatory genes. Aging down-regulates microRNA 145 (miR-145) and derepresses the neurorepulsive factor semaphorin-3A. miR-145 deletion, which increased Sema3a expression or endothelial Sema3a overexpression, reduced axon density, mimicking the aged-heart phenotype. Removal of senescent cells, which accumulated with chronological age in parallel to the decline in nerve density, rescued age-induced denervation, reversed Sema3a expression, preserved heart rate patterns, and reduced electrical instability. These data suggest that senescence-mediated regulation of nerve density contributes to age-associated cardiac dysfunction.

Background: Ischemic heart diseases are leading causes of death and reduced life quality worldwide. Although revascularization strategies significantly reduce mortality after acute myocardial infarction (MI), a large number of patients with MI develop chronic heart failure over time. We previously reported that a fragment of the extracellular matrix protein agrin promotes cardiac regeneration after MI in adult mice. Methods: To test the therapeutic potential of agrin in a preclinical porcine model, we performed ischemia–reperfusion injuries using balloon occlusion for 60 minutes followed by a 3-, 7-, or 28-day reperfusion period. Results: We demonstrated that local (antegrade) delivery of recombinant human agrin to the infarcted pig heart can target the affected regions in an efficient and clinically relevant manner. A single dose of recombinant human agrin improved heart function, infarct size, fibrosis, and adverse remodeling parameters 28 days after MI. Short-term MI experiments along with complementary murine studies revealed myocardial protection, improved angiogenesis, inflammatory suppression, and cell cycle reentry as agrin’s mechanisms of action. Conclusions: A single dose of agrin is capable of reducing ischemia–reperfusion injury and improving heart function, demonstrating that agrin could serve as a therapy for patients with acute MI and potentially heart failure.

Significance: Genetically modified animals are crucial for functional studies and translational biomedical research. However, the generation of genome-edited large animals is time consuming and inefficient. Here, we report the generation of transgenic pigs and chickens that ubiquitously express Cas9 nuclease. The functionality of Cas9 was demonstrated for different target genes, a variety of cell types, and in vivo for the heart and the developing brain. Genome editing can now easily be carried out in vivo in specific organs or tissues without the need to generate germline-modified animals. These Cas9 transgenic animals provide an innovative and efficient resource for in vivo genome editing in biomedical and agricultural sciences.

Elucidation of the molecular mechanisms leading to autonomous aldosterone secretion is a prerequisite to define potential targets and biomarkers in the context of primary aldosteronism. After a genome-wide association study with subjects from the population-based Cooperative Health Research in the Region of Augsburg F4 survey, we observed a highly significant association (P=6.78×10^–11) between the aldosterone to renin ratio and a locus at 5q32. Hypothesizing that this locus may contain genes of relevance for the pathogenesis of primary aldosteronism, we investigated solute carrier family 26 member 2 (SLC26A2), a protein with known transport activity for sulfate and other cations. Within murine tissues, adrenal glands showed the highest expression levels for SLC26A2, which was significantly downregulated on in vivo stimulation with angiotensin II and potassium. SLC26A2 expression was found to be significantly lower in aldosterone-producing adenomas in comparison with normal adrenal glands. In adrenocortical NCI-H295R cells, specific knockdown of SLC26A2 resulted in a highly significant increase in aldosterone secretion. Concomitantly, expression of steroidogenic enzymes, as well as upstream effectors including transcription factors such as NR4A1, CAMK1, and intracellular Ca²⁺ content, was upregulated in knockdown cells. To substantiate further these findings in an SLC26A2 mutant mouse model, aldosterone output proved to be increased in a sex-specific manner. In summary, these findings point toward a possible effect of SLC26A2 in the regulation of aldosterone secretion potentially involved in the pathogenesis of primary aldosteronism.

The clinical translation of cell- and gene-based therapies is limited by the lack of non-invasive, quantitative and specific whole-body imaging tools. Here we present a positron emission tomography reporter system based on a membrane-anchored anticalin protein that binds a fluorine-18-labelled lanthanide complex with picomolar affinity via a bio-orthogonal interaction. The reporter was introduced into therapeutic cells, including CAR T cells and adeno-associated virus-transduced cells. In vitro, reporter expression conferred >800-fold higher radioligand binding versus controls. In mice, the radioligand demonstrated rapid renal clearance, showed no off-target accumulation and enabled high-contrast detection of as few as 1,200 CAR T cells in the bone marrow. Longitudinal positron emission tomography imaging over 4 weeks revealed precise tracking of CAR T cell expansion and migration, with signal intensity correlating linearly with flow cytometry data. The system also enabled the quantitative imaging of in vivo gene transfer using an adeno-associated viral vector. This depth-independent whole-body imaging platform offers a powerful tool for monitoring therapeutic cell dynamics and gene delivery in preclinical and potentially clinical settings.

BACKGROUND: Endothelial cells (ECs) play pivotal roles in maintaining cardiac blood supply and regulating inflammation by acting as gatekeepers for immune cell activity. This study unveils a novel immunomodulatory function of cardiac ECs following myocardial infarction. METHODS: We used single-cell RNA sequencing and spatial transcriptomics to identify EC states after acute myocardial infarction in mice. Subsequently, we mimicked the cytokine environment that was predicted to induce EC activation in cell culture studies and confirmed the results in an endothelial-specific deletion mouse model. RESULTS: Single-cell RNA sequencing analysis identified a transient myeloid CD45 ⁺ CD11b ⁺ Cdh5 ⁺ immunomodulatory EC phenotype (IMEC) emerging between days 1 and 3 after myocardial infarction. IMECs derived from Cdh5 ⁺ tissue resident cells as shown by bone marrow transplantation and lineage tracing experiment. Ligand-receptor interaction predictions indicated a cytokine-mediated activation of IMECs, which we validated through in vitro experiments in cultured ECs. Notably, while cytokine treatment with IL-1β and TGF-β (transforming growth factor β) induced mesenchymal gene expression, the addition of IFN-γ (interferon γ) facilitated the transition into the immunomodulatory phenotype. IMECs exhibited an upregulation of MHC-II (major histocompatibility complex class II) genes, along with the expression of RUNX1 (runt-related transcription factor-1) and proinflammatory cytokines, such as IL-6 and IL-12. IMECs induced T-cell activation through paracrine signaling and were colocalized with T cells in vivo. Inhibition of endothelial-specific IFN-γ-signaling in mice by IFN-γ receptor 1 deletion improved the recovery after myocardial infarction. CONCLUSIONS: These findings provide insight into the role of ECs regulating adaptive immune responses following myocardial infarction, offering potential insights into therapeutic interventions for postinfarction immunomodulation.