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Aims Elevated serum immunoglobulins have been associated with experimental and human hypertension for decades but whether immunoglobulins and B cells play a causal role in hypertension pathology is unclear. In this study, we sought to determine the role of B cells and high-affinity class-switched immunoglobulins on hypertension and hypertensive end-organ damage to determine if they might represent viable therapeutic targets for this disease. Methods and results We purified serum IgG from mice exposed to vehicle or angiotensin (Ang) II to induce hypertension and adoptively transferred these to wild type (WT) recipient mice receiving a subpressor dose of Ang II. We found that transfer of IgG from hypertensive animals leads does not affect blood pressure, endothelial function, renal inflammation, albuminuria, or T cell derived cytokine production compared with transfer of IgG from vehicle infused animals. As an alternative approach to investigate the role of high affinity, class-switched immunoglobulins, we studied mice with genetic deletion of activation-induced deaminase (Aicda-/-). These mice have elevated levels of IgM but virtual absence of class-switched immunoglobulins such as IgG subclasses and IgA. Neither male nor female Aicda-/- mice were protected from Ang II-induced hypertension and renal/vascular damage. To determine if IgM or non-immunoglobulin dependent innate functions of B cells play a role in hypertension, we studied mice with severe global B cell deficiency due to deletion of the membrane exon of the IgM heavy chain (μMT-/-). μMT-/- mice were also not protected from hypertension or end-organ damage induced by Ang II infusion or deoxycorticosterone acetate (DOCA)-salt treatment. Conclusions These results suggest that B cells and serum immunoglobulins do not play a causal role in hypertension pathology. Translational perspective Our results suggest that in most cases of essential hypertension, B cells are not a causal factor in the pathophysiology of disease. Thus, elevated serum immunoglobulins seen in hypertensive animals and humans may reflect a biomarker of aberrant immune activation in hypertension and not a therapeutic target. However, autoantibodies may cause hypertension in special cases, and more work is needed to determine whether specific B cell subsets might play an important role that is masked by global B cell deficiency.

Justification: Hypertension and systemic lupus erythematosus (SLE) share similarities including elevations of blood pressure, proteinuria, inflammation and renal dysfunction and both involve formation of isoLG adducts. We hypothesized that isoLG scavenging would modulate overlapping gene pathways in inflammatory cells in these two conditions. Methods: We performed 10 X genomics single cell sequencing on splenocytes of C57Bl/6 mice with angiotensin II (Ang II)-induced hypertension, Ang II co-treatment with the isoLG scavenger 2HOBA, and 24-week-old B6.SLE123 mice with or without 6-weeks of 2HOBA. Matched C57Bl/6 females were used as controls for the B6.SLE123 mice. Results: Both models exhibited myeloid expansion and genes associated with neutrophil infiltration compared to their respective controls and 2HOBA attenuated this ( Table 1 ). Hypertension was associated with neutrophil expansion whereas SLE was associated an expansion of neutrophils, monocytes, and dendritic cells. In SLE, 2HOBA predominantly modulated gene expression in dendritic cells. Gene ontology revealed 2HOBA downregulated genes governing inflammation including Il1 β (Avglog2(fold change) = -1.6, P adj = 0.002) in both hypertension and SLE. Conclusions: In a mouse model of SLE, scavenging of isoLGs with 2HOBA downregulates inflammatory genes specifically in DCs. In models of both hypertension and SLE, scavenging of isoLG prevents neutrophil expansion. Combined these data describe a shared role of isoLGs in hypertension and SLE and suggest a specific role of DCs and neutrophil activation in the pathogenesis of both conditions. Table 1. Percentage of myeloid derived cells in all groups.

Background: Metabolic remodeling precedes most alterations during cardiac hypertrophic growth under hemodynamic stress. The elevation of glucose utilization has been recognized as a hallmark of metabolic remodeling. However, its role in cardiac hypertrophic growth and heart failure in response to pressure overload remains to be fully illustrated. Here, we aimed to dissect the role of cardiac PKM1 (pyruvate kinase muscle isozyme 1) in glucose metabolic regulation and cardiac response under pressure overload. Methods: Cardiac-specific deletion of PKM1 was achieved by crossing the floxed PKM1 mouse model with the cardiomyocyte-specific Cre transgenic mouse. PKM1 transgenic mice were generated under the control of tetracycline response elements, and cardiac-specific overexpression of PKM1 was induced by doxycycline administration in adult mice. Pressure overload was triggered by transverse aortic constriction. Primary neonatal rat ventricular myocytes were used to dissect molecular mechanisms. Moreover, metabolomics and nuclear magnetic resonance spectroscopy analyses were conducted to determine cardiac metabolic flux in response to pressure overload. Results: We found that PKM1 expression is reduced in failing human and mouse hearts. It is important to note that cardiomyocyte-specific deletion of PKM1 exacerbates cardiac dysfunction and fibrosis in response to pressure overload. Inducible overexpression of PKM1 in cardiomyocytes protects the heart against transverse aortic constriction–induced cardiomyopathy and heart failure. At the mechanistic level, PKM1 is required for the augmentation of glycolytic flux, mitochondrial respiration, and ATP production under pressure overload. Furthermore, deficiency of PKM1 causes a defect in cardiomyocyte growth and a decrease in pyruvate dehydrogenase complex activity at both in vitro and in vivo levels. Conclusions: These findings suggest that PKM1 plays an essential role in maintaining a homeostatic response in the heart under hemodynamic stress.

Elevated cardiovascular risk including stroke, heart failure, and heart attack is present even after normalization of blood pressure in patients with hypertension. Underlying immune cell activation is a likely culprit. Although immune cells are important for protection against invading pathogens, their chronic overactivation may lead to tissue damage and high blood pressure. Triggers that may initiate immune activation include viral infections, autoimmunity, and lifestyle factors such as excess dietary salt. These conditions activate the immune system either directly or through their impact on the gut microbiome, which ultimately produces chronic inflammation and hypertension. T cells are central to the immune responses contributing to hypertension. They are activated in part by binding specific antigens that are presented in major histocompatibility complex molecules on professional antigen-presenting cells, and they generate repertoires of rearranged T-cell receptors. Activated T cells infiltrate tissues and produce cytokines including interleukin 17A, which promote renal and vascular dysfunction and end-organ damage leading to hypertension. In this comprehensive review, we highlight environmental, genetic, and microbial associated mechanisms contributing to both innate and adaptive immune cell activation leading to hypertension. Targeting the underlying chronic immune cell activation in hypertension has the potential to mitigate the excess cardiovascular risk associated with this common and deadly disease.

Introduction: Inflammation and oxidative stress are linked to multiple risk factors for atrial fibrillation (AF), and to AF itself in the setting of sterile injury (e.g. after catheter ablation or cardiac surgery). However, anti-inflammatory therapies and conventional antioxidants cause adverse effects or are ineffective to prevent AF. Highly reactive mediators of lipid peroxidation such as isolevuglandins (IsoLGs) have been identified as a major component of oxidative stress-related injury. We hypothesized that during AF promoted by cardiac inflammation, a scavenger of IsoLG will decrease AF susceptibility. Methods: We studied mice with a systemic inflammatory phenotype due to deficiency in the lymphocyte adaptor protein ( Lnk -/- ), a negative regulator of cytokine signaling. At weaning, Lnk -/- mice and their wild-type (WT) littermates received either vehicle or a potent IsoLG scavenger, 2-hydroxybenzylamine (2-HOBA), by oral administration. At age 14 weeks, animals underwent transesophageal burst pacing, echocardiography, and tissue harvest or flow cytometry to measure atrial inflammation and IsoLG-adducts. Results: Cardiac histology and echocardiography revealed no major histologic or structural abnormalities in Lnk -/- mice. Nevertheless, Lnk -/- mice demonstrated a significant increase in AF burden compared to WT controls (124.8±43.3 vs 6.8±3 sec, respectively [mean±SEM, n=28, 12; P 30 sec; 48.1% vs 0%; P-/- mice, with a significant increase in CD3, CD19, NK1.1, and CD11b/MHCII positive cells, compared to atria from WT control mice. Furthermore, there was a 2 to 4-fold increase in IsoLG-adducts for Lnk -/- atrial immune cells positive for CD3, CD19, NK1.1 and CD11b/MHCII, compared to cells from WT atria. Lnk -/- mice treated with 2-HOBA had significantly reduced AF burden (4.7±4.5 sec, n=7; P Conclusions: IsoLGs play a critical role in the pathogenesis of inflammation-mediated AF, and 2-HOBA, a scavenger of IsoLGs, represents a potentially novel therapeutic strategy for AF in this clinical setting.

Deoxycorticosterone acetate (DOCA)-salt is a common hypertension model in mice and has recently been used to study heart failure with preserved ejection fraction (HFpEF) in rats. Our goal was to validate DOCA-salt as a mouse model of HFpEF and determine how DOCA-salt alters the cardiac immunological landscape to identify novel therapeutic targets for this disease. DOCA-salt mice underwent uninephrectomy, implantion of a DOCA pellet, and supplementation of the drinking water with 1% NaCl water for three weeks. Control mice underwent a sham procedure and received normal water. Compared to control mice, DOCA-salt mice exhibited elevated systolic BP, increased heart weight to body weight ratios (5.6 vs 7.1), increased lung wet to dry weight ratios (4.4 vs 4.8) indicative of pulmonary congestion, and decreased time to exhaustion upon treadmill exercise testing (23.0 vs. 18.5 seconds). On conscious echocardiography, DOCA-salt mice exhibited a preserved ejection fraction. Invasive hemodynamic studies revealed an increased tau constant (5.7 vs 8.2) and increased end-diastolic pressures in DOCA-salt mice (1.7 vs 2.6), consistent with diastolic dysfunction. CITE-seq, a novel technique to obtain transcriptomic and surface marker expression on single cells, was performed on a total of 4,359 and 7,600 cells sorted live CD45+ leukocytes from four sham and four DOCA-salt left ventricles, respectively. Unsupervised computational analysis revealed 29 clusters of immune cells. Six clusters containing natural killer, T lymphocyte and myeloid cell populations were overrepresented and five B cell clusters were underrepresented in DOCA-salt samples. Differential expression analysis of CD11b+CD64+ cardiac macrophages revealed transcriptional changes between groups with 146 significantly upregulated and 111 downregulated genes. Gene set enrichment analysis showed upregulation of leukocyte migration, response to type I interferon, and cytokine-mediated signaling pathways in DOCA-salt macrophages. In conclusion, the DOCA-salt mouse model recapitulates key features of HFpEF including diastolic dysfunction with preserved ejection fraction, cardiac hypertrophy, and pulmonary congestion and is associated with an altered cardiac immune cell profile.

Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is the cause of the global pandemic of coronavirus disease‐2019 (COVID‐19). SARS‐CoV‐2 is a zoonotic disease, but little is known about variations in species susceptibility that could identify potential reservoir species, animal models, and the risk to pets, wildlife, and livestock. Certain species, such as domestic cats and tigers, are susceptible to SARS‐CoV‐2 infection, while other species such as mice and chickens are not. Most animal species, including those in close contact with humans, have unknown susceptibility. Hence, methods to predict the infection risk of animal species are urgently needed. SARS‐CoV‐2 spike protein binding to angiotensin‐converting enzyme 2 (ACE2) is critical for viral cell entry and infection. Here we integrate species differences in susceptibility with multiple in‐depth structural analyses to identify key ACE2 amino acid positions including 30, 83, 90, 322, and 354 that distinguish susceptible from resistant species. Using differences in these residues across species, we developed a susceptibility score that predicts an elevated risk of SARS‐CoV‐2 infection for multiple species including horses and camels. We also demonstrate that SARS‐CoV‐2 is nearly optimal for binding ACE2 of humans compared to other animals, which may underlie the highly contagious transmissibility of this virus among humans. Taken together, our findings define potential ACE2 and SARS‐CoV‐2 residues for therapeutic targeting and identification of animal species on which to focus research and protection measures for environmental and public health.

In only a few months, the novel coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic, leaving physicians, scientists, and public health officials racing to understand, treat, and contain this zoonotic disease. SARS-CoV-2 has made the leap from animals to humans, but little is known about variations in species susceptibility that could identify potential reservoir species, animal models, and the risk to pets, wildlife, and livestock. While there is evidence that certain species, such as cats, are susceptible, the vast majority of animal species, including those in close contact with humans, have unknown susceptibility. Hence, methods to predict their infection risk are urgently needed. SARS-CoV-2 spike protein binding to angiotensin converting enzyme 2 (ACE2) is critical for viral cell entry and infection. Here we identified key ACE2 residues that distinguish susceptible from resistant species using in-depth sequence and structural analyses of ACE2 and its binding to SARS-CoV-2. Our findings have important implications for identification of ACE2 and SARS-CoV-2 residues for therapeutic targeting and identification of animal species with increased susceptibility for infection on which to focus research and protection measures for environmental and public health.

We and others have demonstrated that interleukin-17A (IL-17A) plays a critical role in the pathogenesis of cardiovascular disease and hypertension. Patients with psoriasis, an IL-17A mediated autoimmune disease, are at elevated risk for cardiovascular events. New biologic therapies for psoriasis have been developed that target IL-17A (secukinumab) or its receptor (brodalumab), but their impact on incident hypertension and its associated complications is unknown and remain unexplored outside of short-term safety in clinical trials. Our goal is to perform a retrospective case-control study utilizing the Synthetic Derivative (Vanderbilt’s de-identified version of the electronic health record) to determine whether incident hypertension or other major adverse cardiovascular events is reduced in patients with psoriasis receiving IL-17A targeted therapies versus other therapeutics. Patients are defined as cases by 2 or more mentions of a psoriasis ICD9 or ICD10 code and at least one documented prescription for secukinumab or brodalumab. Controls are defined by the presence of 2 or more psoriasis ICD codes, no documented exposure to the drugs of interest, and a mention of “psoriasis” in their Problem List. Using these definitions, we identified 571 cases and 4484 controls in the Synthetic Derivative. Of the 571 potential cases identified, 65 were confirmed by manual review for an initial validation study. Of the manually confirmed cases, 22 (34%) were male and 43 (66%) were female. Manually reviewed cases had a mean record length of 5.0 years with an average of 29.9 discrete ICD mentions. Twenty-three cases (35%) had an ICD code for hypertension, which predated the first documented psoriasis code in eight patients. Thus, we are able to use the Synthetic Derivative to identify patients with psoriasis receiving biologic therapies and assess temporal relationships relevant to hypertension. We plan to expand this study to the larger cohort identified and analyze rates of hypertension and a composite outcome of hypertension complications, including stroke, myocardial infarction, heart failure, and chronic kidney disease, in cases and controls matched for age, gender, race, and BMI.

T and B cells have been implicated in hypertension, but the mechanisms by which they produce a coordinated response is unknown. T follicular helper (Tfh) cells that produce interleukin 21 (IL21) promote germinal center (GC) B cell responses leading to immunoglobulin (Ig) production. Here we investigate the role of IL21 and Tfh cells in hypertension. In response to angiotensin (Ang) II-induced hypertension, T cell IL21 production is increased, and Il21-/- mice develop blunted hypertension, attenuated vascular end-organ damage, and decreased interleukin 17A (IL17A) and interferon gamma production. Tfh-like cells and GC B cells accumulate in the aorta and plasma IgG1 is increased in hypertensive WT but not Il21-/-mice. Furthermore, Tfh cell deficient mice develop blunted hypertension and vascular hypertrophy in response to Ang II infusion. Importantly, IL21 neutralization reduces blood pressure (BP) and reverses endothelial dysfunction and vascular inflammation. Moreover, recombinant IL21 impairs endothelium-dependent relaxation ex vivo and decreases nitric oxide production from cultured endothelial cells. Finally, we show in humans that peripheral blood T cell production of IL21 correlates with systolic BP and IL17A production. These data suggest that IL21 may be a novel therapeutic target for the treatment of hypertension and its micro- and macrovascular complications.