A relay velocity model infers cell-dependent RNA velocity.

RNA velocity provides an approach for inferring cellular state transitions from single-cell RNA sequencing (scRNA-seq) data. Conventional RNA velocity models infer universal kinetics from all cells in an scRNA-seq experiment, resulting in unpredictable performance in experiments with multi-stage and/or multi-lineage transition of cell states where the assumption of the same kinetic rates for all cells no longer holds. Here we present cellDancer, a scalable deep neural network that locally infers velocity for each cell from its neighbors and then relays a series of local velocities to provide single-cell resolution inference of velocity kinetics. In the simulation benchmark, cellDancer shows robust performance in multiple kinetic regimes, high dropout ratio datasets and sparse datasets. We show that cellDancer overcomes the limitations of existing RNA velocity models in modeling erythroid maturation and hippocampus development. Moreover, cellDancer provides cell-specific predictions of transcription, splicing and degradation rates, which we identify as potential indicators of cell fate in the mouse pancreas.

Telomerase mRNA Enhances Human Skin Engraftment for Wound Healing.

Deep skin wounds represent a serious condition and frequently require split-thickness skin grafts (STSG) to heal. The application of autologous human-skin-cell-suspension (hSCS) requires less donor skin than STSG without compromising the healing capacity. Impaired function and replicative ability of senescent cutaneous cells in the aging skin affects healing with autologous hSCS. Major determinants of senescence are telomere erosion and DNA damage. Human telomerase reverse transcriptase (hTERT) adds telomeric repeats to the DNA and can protect against DNA damage. Herein, hTERT mRNA lipid nanoparticles (LNP) are proposed and evaluated for enhancing cellular engraftment and proliferation of hSCS. Transfection with optimized hTERT mRNA LNP system enables delivery and expression of mRNA in vitro in keratinocytes, fibroblasts, and in hSCS prepared from donors' skin. Telomerase activity in hSCS is significantly increased. hTERT mRNA LNP enhance the generation of a partial-thickness human skin equivalent in the mouse model, increasing hSCS engraftment (Lamin) and proliferation (Ki67), while reducing cellular senescence (p21) and DNA damage (53BP1).

Disrupted Stiffness Ratio Alters Nuclear Mechanosensing.

The ability of endothelial cells to sense and respond to dynamic changes in blood flow is critical for vascular homeostasis and cardiovascular health. The mechanical and geometric properties of the nuclear and cytoplasmic compartments affect mechanotransduction. We hypothesized that alterations to these parameters have resulting mechanosensory consequences. Using atomic force microscopy and mathematical modeling, we assessed how the nuclear and cytoplasmic compartment stiffnesses modulate shear stress transfer to the nucleus within aging endothelial cells. Our computational studies revealed that the critical parameter controlling shear transfer is not the individual mechanics of these compartments, but the stiffness ratio between them. Replicatively aged cells had a reduced stiffness ratio, attenuating shear transfer, while the ratio was not altered in a genetic model of accelerated aging. We provide a theoretical framework suggesting that dysregulation of the shear stress response can be uniquely imparted by relative mechanical changes in subcellular compartments.

Premature senescence and cardiovascular disease following cancer treatments: mechanistic insights.

Cardiovascular disease (CVD) is a leading cause of morbidity and mortality, especially among the aging population. The "response-to-injury" model proposed by Dr. Russell Ross in 1999 emphasizes inflammation as a critical factor in atherosclerosis development, with atherosclerotic plaques forming due to endothelial cell (EC) injury, followed by myeloid cell adhesion and invasion into the blood vessel walls. Recent evidence indicates that cancer and its treatments can lead to long-term complications, including CVD. Cellular senescence, a hallmark of aging, is implicated in CVD pathogenesis, particularly in cancer survivors. However, the precise mechanisms linking premature senescence to CVD in cancer survivors remain poorly understood. This article aims to provide mechanistic insights into this association and propose future directions to better comprehend this complex interplay.

Transflammation in tissue regeneration and response to injury: How cell-autonomous inflammatory signaling mediates cell plasticity.

Inflammation is a first responder against injury and infection and is also critical for the regeneration and repair of tissue after injury. The role of professional immune cells in tissue healing is well characterized. Professional immune cells respond to pathogens with humoral and cytotoxic responses; remove cellular debris through efferocytosis; secrete angiogenic cytokines and growth factors to repair the microvasculature and parenchyma. However, non-immune cells are also capable of responding to damage or pathogens. Non-immune somatic cells express pattern recognition receptors (PRRs) to detect pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs). The PRRs activation leads to the release of inflammatory cytokines required for tissue defense and repair. Notably, the activation of PRRs also triggers epigenetic changes that promote DNA accessibility and cellular plasticity. Thus, non-immune cells directly respond to the local inflammatory cues and can undergo phenotypic modifications or even cell lineage transitions to facilitate tissue regeneration. This review will focus on the novel role of cell-autonomous inflammatory signaling in mediating cell plasticity, a process which is termed transflammation. We will discuss the regulation of this process by changes in the functions and expression levels of epigenetic modifiers, as well as metabolic and ROS/RNS-mediated epigenetic modulation of DNA accessibility during cell fate transition. We will highlight the recent technological developments in detecting cell plasticity and potential therapeutic applications of transflammation in tissue regeneration.

Endothelial activation and fibrotic changes are impeded by laminar flow-induced CHK1-SENP2 activity through mechanisms distinct from endothelial-to-mesenchymal cell transition.

Background: The deSUMOylase sentrin-specific isopeptidase 2 (SENP2) plays a crucial role in atheroprotection. However, the phosphorylation of SENP2 at T368 under disturbed flow (D-flow) conditions hinders its nuclear function and promotes endothelial cell (EC) activation. SUMOylation has been implicated in D-flow-induced endothelial-to-mesenchymal transition (endoMT), but the precise role of SENP2 in counteracting this process remains unclear. Method: We developed a phospho-specific SENP2 S344 antibody and generated knock-in (KI) mice with a phospho-site mutation of SENP2 S344A using CRISPR/Cas9 technology. We then investigated the effects of SENP2 S344 phosphorylation under two distinct flow patterns and during hypercholesteremia (HC)-mediated EC activation. Result: Our findings demonstrate that laminar flow (L-flow) induces phosphorylation of SENP2 at S344 through the activation of checkpoint kinase 1 (CHK1), leading to the inhibition of ERK5 and p53 SUMOylation and subsequent suppression of EC activation. We observed a significant increase in lipid-laden lesions in both the aortic arch (under D-flow) and descending aorta (under L-flow) of female hypercholesterolemic SENP2 S344A KI mice. In male hypercholesterolemic SENP2 S344A KI mice, larger lipid-laden lesions were only observed in the aortic arch area, suggesting a weaker HC-mediated atherogenesis in male mice compared to females. Ionizing radiation (IR) reduced CHK1 expression and SENP2 S344 phosphorylation, attenuating the pro-atherosclerotic effects observed in female SENP2 S344A KI mice after bone marrow transplantation (BMT), particularly in L-flow areas. The phospho-site mutation SENP2 S344A upregulates processes associated with EC activation, including inflammation, migration, and proliferation. Additionally, fibrotic changes and up-regulated expression of EC marker genes were observed. Apoptosis was augmented in ECs derived from the lungs of SENP2 S344A KI mice, primarily through the inhibition of ERK5-mediated expression of DNA damage-induced apoptosis suppressor (DDIAS). Summary: In this study, we have revealed a novel mechanism underlying the suppressive effects of L-flow on EC inflammation, migration, proliferation, apoptosis, and fibrotic changes through promoting CHK1-induced SENP2 S344 phosphorylation. The phospho-site mutation SENP2 S344A responds to L-flow through a distinct mechanism, which involves the upregulation of both mesenchymal and EC marker genes.

An ERK5-NRF2 Axis Mediates Senescence-Associated Stemness and Atherosclerosis.

BACKGROUND: ERK5 (extracellular signal-regulated kinase 5) is a dual kinase transcription factor containing an N-terminal kinase domain and a C-terminal transcriptional activation domain. Many ERK5 kinase inhibitors have been developed and tested to treat cancer and inflammatory diseases. However, recent data have raised questions about the role of the catalytic activity of ERK5 in proliferation and inflammation. We aimed to investigate how ERK5 reprograms myeloid cells to the proinflammatory senescent phenotype, subsequently leading to atherosclerosis. METHODS: A ERK5 S496A (dephosphorylation mimic) knock in (KI) mouse model was generated using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeat-associated 9), and atherosclerosis was characterized by hypercholesterolemia induction. The plaque phenotyping in homozygous ERK5 S496A KI and wild type (WT) mice was studied using imaging mass cytometry. Bone marrow-derived macrophages were isolated from hypercholesterolemic mice and characterized using RNA sequencing and functional in vitro approaches, including senescence, mitochondria reactive oxygen species, and inflammation assays, as well as by metabolic extracellular flux analysis. RESULTS: We show that atherosclerosis was inhibited in ERK5 S496A KI mice. Furthermore, ERK5 S496 phosphorylation mediates both senescence-associated secretory phenotype and senescence-associated stemness by upregulating AHR (aryl hydrocarbon receptor) in plaque and bone marrow-derived macrophages isolated from hypercholesterolemic mice. We also discovered that ERK5 S496 phosphorylation could induce NRF2 (NFE2-related factor 2) SUMOylation at a novel K518 site to inhibit NRF2 transcriptional activity without altering ERK5 catalytic activity and mediates oxidized LDL (low-density lipoprotein)-induced senescence-associated secretory phenotype. Specific ERK5 kinase inhibitors (AX15836 and XMD8-92) also inhibited ERK5 S496 phosphorylation, suggesting the involvement of ERK5 S496 phosphorylation in the anti-inflammatory effects of these ERK5 kinase inhibitors. CONCLUSIONS: We discovered a novel mechanism by which the macrophage ERK5-NRF2 axis develops a unique senescence-associated secretory phenotype/stemness phenotype by upregulating AHR to engender atherogenesis. The finding of senescence-associated stemness phenotype provides a molecular explanation to resolve the paradox of senescence in proliferative plaque by permitting myeloid cells to escape the senescence-induced cell cycle arrest during atherosclerosis formation.

Possible molecular mechanisms underlying the development of atherosclerosis in cancer survivors.

Cancer survivors undergone treatment face an increased risk of developing atherosclerotic cardiovascular disease (CVD), yet the underlying mechanisms remain elusive. Recent studies have revealed that chemotherapy can drive senescent cancer cells to acquire a proliferative phenotype known as senescence-associated stemness (SAS). These SAS cells exhibit enhanced growth and resistance to cancer treatment, thereby contributing to disease progression. Endothelial cell (EC) senescence has been implicated in atherosclerosis and cancer, including among cancer survivors. Treatment modalities for cancer can induce EC senescence, leading to the development of SAS phenotype and subsequent atherosclerosis in cancer survivors. Consequently, targeting senescent ECs displaying the SAS phenotype hold promise as a therapeutic approach for managing atherosclerotic CVD in this population. This review aims to provide a mechanistic understanding of SAS induction in ECs and its contribution to atherosclerosis among cancer survivors. We delve into the mechanisms underlying EC senescence in response to disturbed flow and ionizing radiation, which play pivotal role in atherosclerosis and cancer. Key pathways, including p90RSK/TERF2IP, TGFßR1/SMAD, and BH4 signaling are explored as potential targets for cancer treatment. By comprehending the similarities and distinctions between different types of senescence and the associated pathways, we can pave the way for targeted interventions aim at enhancing the cardiovascular health of this vulnerable population. The insights gained from this review may facilitate the development of novel therapeutic strategies for managing atherosclerotic CVD in cancer survivors.

A2AR-mediated lymphangiogenesis via VEGFR2 signaling prevents salt-sensitive hypertension.

AIMS: Excess dietary sodium intake and retention lead to hypertension. Impaired dermal lymphangiogenesis and lymphatic dysfunction-mediated sodium and fluid imbalance are pathological mechanisms. The adenosine A2A receptor (A2AR) is expressed in lymphatic endothelial cells (LECs), while the roles and mechanisms of LEC-A2AR in skin lymphangiogenesis during salt-induced hypertension are not clear. METHODS AND RESULTS: The expression of LEC-A2AR correlated with lymphatic vessel density in both high-salt diet (HSD)-induced hypertensive mice and hypertensive patients. Lymphatic endothelial cell-specific A2AR knockout mice fed HSD exhibited 17 ± 2% increase in blood pressure and 17 ± 3% increase in Na+ content associated with decreased lymphatic density (-19 ± 2%) compared with HSD-WT mice. A2AR activation by agonist CGS21680 increased lymphatic capillary density and decreased blood pressure in HSD-WT mice. Furthermore, this A2AR agonist activated MSK1 directly to promote VEGFR2 activation and endocytosis independently of VEGF as assessed by phosphoprotein profiling and immunoprecipitation assays in LECs. VEGFR2 kinase activity inhibitor fruquintinib or VEGFR2 knockout in LECs but not VEGF-neutralizing antibody bevacizumab suppressed A2AR activation-mediated decrease in blood pressure. Immunostaining revealed phosphorylated VEGFR2 and MSK1 expression in the LECs were positively correlated with skin lymphatic vessel density and A2AR level in hypertensive patients. CONCLUSION: The study highlights a novel A2AR-mediated VEGF-independent activation of VEGFR2 signaling in dermal lymphangiogenesis and sodium balance, which might be a potential therapeutic target in salt-sensitive hypertension.

Role of angiogenic transdifferentiation in vascular recovery.

Tissue repair requires the orchestration of multiple processes involving a multiplicity of cellular effectors, signaling pathways, and cell-cell communication. The regeneration of the vasculature is a critical process for tissue repair and involves angiogenesis, adult vasculogenesis, and often arteriogenesis, which processes enable recovery of perfusion to deliver oxygen and nutrients to the repair or rebuild of the tissue. Endothelial cells play a major role in angiogenesis, whereas circulating angiogenic cells (primarily of hematopoietic origin) participate in adult vasculogenesis, and monocytes/macrophages have a defining role in the vascular remodeling that is necessary for arteriogenesis. Tissue fibroblasts participate in tissue repair by proliferating and generating the extracellular matrix as the structural scaffold for tissue regeneration. Heretofore, fibroblasts were not generally believed to be involved in vascular regeneration. However, we provide new data indicating that fibroblasts may undergo angiogenic transdifferentiation, to directly expand the microvasculature. Transdifferentiation of fibroblasts to endothelial cells is initiated by inflammatory signaling which increases DNA accessibility and cellular plasticity. In the environment of under-perfused tissue, the activated fibroblasts with increased DNA accessibility can now respond to angiogenic cytokines, which provide the transcriptional direction to induce fibroblasts to become endothelial cells. Periphery artery disease (PAD) involves the dysregulation of vascular repair and inflammation. Understanding the relationship between inflammation, transdifferentiation, and vascular regeneration may lead to a new therapeutic approach to PAD.

Coronary microvascular health in symptomatic patients with prior COVID-19 infection: an updated analysis.

AIMS: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with endothelial dysfunction. We aimed to determine the effects of prior coronavirus disease 2019 (COVID-19) on the coronary microvasculature accounting for time from COVID-19, disease severity, SARS-CoV-2 variants, and in subgroups of patients with diabetes and those with no known coronary artery disease. METHODS AND RESULTS: Cases consisted of patients with previous COVID-19 who had clinically indicated positron emission tomography (PET) imaging and were matched 1:3 on clinical and cardiovascular risk factors to controls having no prior infection. Myocardial flow reserve (MFR) was calculated as the ratio of stress to rest myocardial blood flow (MBF) in mL/min/g of the left ventricle. Comparisons between cases and controls were made for the odds and prevalence of impaired MFR (MFR < 2). We included 271 cases matched to 815 controls (mean ± SD age 65 ± 12 years, 52% men). The median (inter-quartile range) number of days between COVID-19 infection and PET imaging was 174 (58-338) days. Patients with prior COVID-19 had a statistically significant higher odds of MFR <2 (adjusted odds ratio 3.1, 95% confidence interval 2.8-4.25 P < 0.001). Results were similar in clinically meaningful subgroups. The proportion of cases with MFR <2 peaked 6-9 months from imaging with a statistically non-significant downtrend afterwards and was comparable across SARS-CoV-2 variants but increased with increasing severity of infection. CONCLUSION: The prevalence of impaired MFR is similar by duration of time from infection up to 1 year and SARS-CoV-2 variants, but significantly differs by severity of infection.

Epigenetic landscape reveals MECOM as an endothelial lineage regulator.

A comprehensive understanding of endothelial cell lineage specification will advance cardiovascular regenerative medicine. Recent studies found that unique epigenetic signatures preferentially regulate cell identity genes. We thus systematically investigate the epigenetic landscape of endothelial cell lineage and identify MECOM to be the leading candidate as an endothelial cell lineage regulator. Single-cell RNA-Seq analysis verifies that MECOM-positive cells are exclusively enriched in the cell cluster of bona fide endothelial cells derived from induced pluripotent stem cells. Our experiments demonstrate that MECOM depletion impairs human endothelial cell differentiation, functions, and Zebrafish angiogenesis. Through integrative analysis of Hi-C, DNase-Seq, ChIP-Seq, and RNA-Seq data, we find MECOM binds enhancers that form chromatin loops to regulate endothelial cell identity genes. Further, we identify and verify the VEGF signaling pathway to be a key target of MECOM. Our work provides important insights into epigenetic regulation of cell identity and uncovered MECOM as an endothelial cell lineage regulator.

Electrically stimulated gene expression under exogenously applied electric fields.

Introduction: Electrical stimulation, the application of an electric field to cells and tissues grown in culture to accelerate growth and tight junction formation among endothelial cells, could be impactful in cardiovascular tissue engineering, allotransplantation, and wound healing. Methods: Using Electrical Cell Stimulation And Recording Apparatus (ECSARA), the exploration of the stimulatory influences of electric fields of different magnitude and frequencies on growth and proliferation, trans endothelial electrical resistance (TEER) and gene expression of human endothelia cells (HUVECs) were explored. Results: Within the range of endogenous electrical pulses studied, frequency was found to be more significant (p = 0.05) than voltage in influencing HUVEC gene expression. Localization of Yes Associated Protein (YAP) and expression of CD-144 are shown to be consistent with temporal manifestations of TEER. Discussion: This work introduces the field of electromics, the study of cellular gene expression profiles and their implications under the influence of exogenously applied electric fields. Homology of electrobiology and mechanobiology suggests use of such exogenous cues in tissue and regenerative engineering.

Design and rationale of re-energize fontan: Randomized exercise intervention designed to maximize fitness in fontan patients.

In this manuscript, we describe the design and rationale of a randomized controlled trial in pediatric Fontan patients to test the hypothesis that a live-video-supervised exercise (aerobic+resistance) intervention will improve cardiac and physical capacity; muscle mass, strength, and function; and endothelial function. Survival of children with single ventricles beyond the neonatal period has increased dramatically with the staged Fontan palliation. Yet, long-term morbidity remains high. By age 40, 50% of Fontan patients will have died or undergone heart transplantation. Factors that contribute to onset and progression of heart failure in Fontan patients remain incompletely understood. However, it is established that Fontan patients have poor exercise capacity which is associated with a greater risk of morbidity and mortality. Furthermore, decreased muscle mass, abnormal muscle function, and endothelial dysfunction in this patient population is known to contribute to disease progression. In adult patients with 2 ventricles and heart failure, reduced exercise capacity, muscle mass, and muscle strength are powerful predictors of poor outcomes, and exercise interventions can not only improve exercise capacity and muscle mass, but also reverse endothelial dysfunction. Despite these known benefits of exercise, pediatric Fontan patients do not exercise routinely due to their chronic condition, perceived restrictions to exercise, and parental overprotection. Limited exercise interventions in children with congenital heart disease have demonstrated that exercise is safe and effective; however, these studies have been conducted in small, heterogeneous groups, and most had few Fontan patients. Critically, adherence is a major limitation in pediatric exercise interventions delivered on-site, with adherence rates as low as 10%, due to distance from site, transportation difficulties, and missed school or workdays. To overcome these challenges, we utilize live-video conferencing to deliver the supervised exercise sessions. Our multidisciplinary team of experts will assess the effectiveness of a live-video-supervised exercise intervention, rigorously designed to maximize adherence, and improve key and novel measures of health in pediatric Fontan patients associated with poor long-term outcomes. Our ultimate goal is the translation of this model to clinical application as an "exercise prescription" to intervene early in pediatric Fontan patients and decrease long-term morbidity and mortality.

Long noncoding RNA LEENE promotes angiogenesis and ischemic recovery in diabetes models.

Impaired angiogenesis in diabetes is a key process contributing to ischemic diseases such as peripheral arterial disease. Epigenetic mechanisms, including those mediated by long noncoding RNAs (lncRNAs), are crucial links connecting diabetes and the related chronic tissue ischemia. Here we identify the lncRNA that enhances endothelial nitric oxide synthase (eNOS) expression (LEENE) as a regulator of angiogenesis and ischemic response. LEENE expression was decreased in diabetic conditions in cultured endothelial cells (ECs), mouse hind limb muscles, and human arteries. Inhibition of LEENE in human microvascular ECs reduced their angiogenic capacity with a dysregulated angiogenic gene program. Diabetic mice deficient in Leene demonstrated impaired angiogenesis and perfusion following hind limb ischemia. Importantly, overexpression of human LEENE rescued the impaired ischemic response in Leene-knockout mice at tissue functional and single-cell transcriptomic levels. Mechanistically, LEENE RNA promoted transcription of proangiogenic genes in ECs, such as KDR (encoding VEGFR2) and NOS3 (encoding eNOS), potentially by interacting with LEO1, a key component of the RNA polymerase II-associated factor complex and MYC, a crucial transcription factor for angiogenesis. Taken together, our findings demonstrate an essential role for LEENE in the regulation of angiogenesis and tissue perfusion. Functional enhancement of LEENE to restore angiogenesis for tissue repair and regeneration may represent a potential strategy to tackle ischemic vascular diseases.

TNIK regulation of interferon signaling and endothelial cell response to virus infection.

Background: Traf2 and Nck-interacting kinase (TNIK) is known for its regulatory role in various processes within cancer cells. However, its role within endothelial cells (ECs) has remained relatively unexplored. Methods: Leveraging RNA-seq data and Ingenuity Pathway Analysis (IPA), we probed the potential impact of TNIK depletion on ECs. Results: Examination of RNA-seq data uncovered more than 450 Differentially Expressed Genes (DEGs) in TNIK-depleted ECs, displaying a fold change exceeding 2 with a false discovery rate (FDR) below 0.05. IPA analysis unveiled that TNIK depletion leads to the inhibition of the interferon (IFN) pathway [-log (p-value) >11], downregulation of IFN-related genes, and inhibition of Hypercytokinemia/Hyperchemokinemia [-log (p-value) >8]. The validation process encompassed qRT-PCR to evaluate mRNA expression of crucial IFN-related genes, immunoblotting to gauge STAT1 and STAT2 protein levels, and ELISA for the quantification of IFN and cytokine secretion in siTNIK-depleted ECs. These assessments consistently revealed substantial reductions upon TNIK depletion. When transducing HUVECs with replication incompetent E1-E4 deleted adenovirus expressing green fluorescent protein (Ad-GFP), it was demonstrated that TNIK depletion did not affect the uptake of Ad-GFP. Nonetheless, TNIK depletion induced cytopathic effects (CPE) in ECs transduced with wild-type human adenovirus serotype 5 (Ad-WT). Summary: Our findings suggest that TNIK plays a crucial role in regulating the EC response to virus infections through modulation of the IFN pathway.

Future Impact of mRNA Therapy on Cardiovascular Diseases.

The silver lining of the recent pandemic was that it accelerated the emergence of messenger ribonucleic acid (mRNA) therapeutics. The great promise of mRNA therapeutics was highlighted by the speed at which the vaccines were created, tested, and proven to be relatively safe and highly effective. There are a wide variety of mRNA therapeutics now under development, and dozens of these are in clinical trials. These therapeutics are generating a major paradigm shift in medical therapy, including the treatment of cardiovascular disease. Most of the cardiovascular mRNA therapies are still in preclinical development, although a phase 2a trial of mRNA therapy for myocardial ischemia has been completed with promising results.1 The application of mRNA therapies to cardiovascular diseases is virtually limitless, and ongoing work includes mRNA therapies for myocardial ischemia, heart failure, arrhythmias, hypercholesterolemia, and arterial occlusive diseases. In addition, mRNA may be used to enhance cell therapies. In the future, mRNA therapies for cardiovascular disease are destined to supplant some of our current biologics and pharmacotherapies and will be used to treat previously untreatable cardiovascular diseases. Furthermore, mRNA therapies can be personalized, and they can be rapidly generated in current Good Manufacturing Practice facilities with a modest footprint, facilitating the rise of hospital-based regional centers of RNA therapeutics.