Human Hypertension Research Articles

Identification and validation of CCL5 as a key gene in HIV infection and pulmonary arterial hypertension.

The relationship between human immunodeficiency virus (HIV) infection and pulmonary arterial hypertension (PAH) has garnered significant scrutiny. Individuals with HIV infection have a higher risk of developing PAH. However, the specific mechanism of HIV-associated PAH remains unclear. Our study aims at investigating the shared biomarkers in HIV infection and PAH and predicting the potential therapeutic target for HIV-associated PAH. Data for HIV infection and PAH were downloaded from Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) analysis was performed to detect shared genes in HIV infection and PAH. Enrichment analysis was conducted to identify the function of common DEGs. Protein-protein interaction (PPI) analysis was used to detect key genes. These crucial genes were subsequently verified by RT-qPCR. Finally, candidate drugs were identified by using the Drug Signatures Database (DSigDB). Nineteen common DEGs were identified in HIV infection and PAH. Enrichment analysis exhibited that the functions of these genes were mainly enriched in inflammatory responses, mainly including cellular immunity and interaction between viral proteins and cytokines. By constructing PPI networks, we identified the key gene CC-type chemokine ligand 5 (CCL5), and we verified that CCL5 was highly expressed in hypoxia induced human pulmonary artery endothelial cells (hPAECs) and human pulmonary artery smooth muscle cells (hPASMCs). In addition, we predicted 10 potential drugs targeting CCL5 by Autodock Vina. This study revealed that CCL5 might be a common biomarker of HIV infection and PAH and provided a new therapeutic target for HIV-associated PAH. However, further clinical validation is still indispensable.

Association between copper intake and essential hypertension: dual evidence from Mendelian randomization analysis and the NHANES database.

Although previous studies have identified an association between trace elements and essential hypertension, the specific trace elements involved and the mechanisms of their association remain unclear. This study aimed to elucidate the relationship between various human trace elements and essential hypertension, thereby addressing existing gaps in the research. This study employed two-sample, multivariate, and inverse Mendelian randomization (MR) analyses to investigate the causal relationship between 15 human trace elements as exposure factors and essential hypertension as the outcome. The analysis revealed a statistically significant association between copper intake and essential hypertension. Further validation was conducted using logistic regression models based on data from the National Health and Nutrition Examination Survey (NHANES). Eighteen trace elements were initially identified through searches in the GWAS database and PubMed. After screening, 15 trace elements were selected as potential exposure factors. MR analysis, utilizing the 2021 genome-wide dataset for essential hypertension, identified copper as a risk factor, showing a positive association with hypertension. Subsequent logistic regression analyses based on NHANES data further confirmed a significant association between dietary copper intake and the risk of essential hypertension, except for the 0.80-1.08 mg/d group in model 3 (p < 0.05). Restricted cubic spline (RCS) analysis indicated a nonlinear relationship between copper intake and the risk of developing essential hypertension. This study demonstrates a significant association between copper intake and the development of essential hypertension. The findings suggest that higher copper intake is linked to an increased risk of hypertension, underscoring the need to monitor copper intake levels in the prevention and management of this condition.

Unilateral Lung Removal in Combination with Monocrotaline or SU5416 in Rodents: A Reliable Model to Mimic the Pathology of the Human Pulmonary Hypertension.

Pulmonary hypertension (PH) is a chronic and progressive disorder characterized by elevated mean pulmonary arterial pressure, pulmonary vascular remodeling, and the development of concentric laminar intimal fibrosis with plexiform lesions. While rodent models have been developed to study PH, they have certain deficiencies and do not entirely replicate the human disease due to the heterogeneity of PH pathology. Therefore, combined models are necessary to study PH. Recent studies have shown that altered pulmonary blood flow is a significant trigger in the development of vascular remodeling and neointimal lesions. One of the most promising rodent models for increased pulmonary flow is the combination of unilateral left pneumonectomy with a "second hit" of monocrotaline (MCT) or SU5416. The removal of one lung in this model forces blood to circulate only in the other lung and induces increased and turbulent pulmonary blood flow. This increased vascular flow leads to progressive remodeling and occlusion of small pulmonary arteries. The second hit by MCT or SU5416 leads to endothelial cell dysfunction, resulting in severe PH and the development of plexiform arteriopathy.

Correlation of Atrial Natriuretic peptide (ANP) coding gene (NPPA) and Human Hypertension

Atrial natriuretic peptide (ANP) is a polypeptide hormone secreted by atrial muscle cells that has an endogenous antagonistic effect on the renin-angiotensin-aldosterone system (RAAS). It is believed to regulate salt and fluid balance and blood pressure. Blood samples (150) from males aged less than 20 to 55 years were collected from Al-Salam and Ibn-Sena Teaching Hospital in Mosul from 1/2/2022 to 1/9/2022. These samples, 5ml that include healthy and previously blood hypertension-diagnosed patients, were put into two tubes for hormonal and molecular study and divided into three classes: Control, which includes 50 samples from healthy individuals; hypertension without treatment patients 40 individuals and patients with treatment 60 individuals. Each class was divided into five groups according to their age. group (1) less than 20, group (2) 20-29, group (3) 30-39, group (4) 40-49 and group (5) 50-59 years. Serum ANP hormone concentration was measured, and the results revealed significant differences at between patients and healthy control groups. Hormone concentration had decreased in the 1,2 group while increased in groups 3,4,5 of treated and untreated patients compared with control groups with different group concentrations ranging from 28.86-23.16 pg/ml, indicating the effect of aging on hormone concentration. Also, the results revealed a different hormone concentration in untreated patient's groups that ranged between 12.97-28 pg./ml while the concentration of treated patient's groups ranged between 12.97-33.36 pg/ml, these differences in hormone concentration in all groups revealed the effect interference of aging and treatments that had a direct effect on the hormone concentration. Besides, the Amplicon copy numbers of the ANP coding gene (Nppa gene), considered an essential indicator of gene activities utilizing the real-time technique, showed a significant increase in patient groups, especially group (5) of both treated and untreated, compared with control. These results concluded that this hormone had an essential correlation with hypertension.

Effect of Omega-3 And SCFA in Human Hypertension and Cardiovascular Health

Hypertension is an urgent and increasingly prevalent issue in global public health. Its regulation through diet has become a crucial focus. Dietary Omega-3 and SCFA have demonstrated multiple health benefits, including the blood pressure management. This article delves into their effects and mechanism on hypertension in humans. Epidemiological and clinical studies provide empirical support for the assertion that the intake of PUFA is correlated with reduced cardiovascular mortality. This reduction is achieved through various mechanisms, such as control of cellular metabolic pathways, alteration of gene transcription patterns, and favorable effects on control of blood pressure and lipid. Furthermore, this article also analyzed the effects of SCFA on hypertension. The utilization of dietary fiber by intestinal flora resulting in the synthesis of several SCFAs which subsequently influence hypertension. Additionally, a correlation exists between hypertension and gut microbiota composition, with significant variations observed in the gut microbiota composition among spontaneously hypertensive groups.

Herbal medicine (Oryeongsan) for fluid and sodium balance in renal cortex of spontaneously hypertensive rats

BackgroundHerbal medicine Oryeongsan (ORS), also known as Wulingsan in Chinese, has been used for the treatment of impaired body fluid balance. However, the mechanisms involved are not clearly defined. The purpose of the present study was to identify the actions of ORS on the renal excretory function and blood pressure (BP) and to define the mechanisms involved in association with renin-angiotensin system (RAS) and natriuretic peptide system (NPS) in spontaneously hypertensive rats (SHR), an animal model of human essential hypertension. MethodsChanges in urine volume (UV), excretion of electrolytes including Na+ (urinary excretion of Na+ (UNaV)) were measured. RT-PCR was performed to trace the changes in expression of RAS, NPS and sodium (Na+)-hydrogen (H+) exchanger 3 (NHE3) in the renal cortex. ResultsIn the SHR treated with vehicle (SHR-V) group, UV and UNaV were suppressed and the Na+ balance was maintained at the higher levels leading to an increase in BP compared to WKY-V group. These were accompanied by an increase in NHE3 expression with an accentuation of angiotensin I converting enzyme-angiotensin II type 1 (ACE-AT1) receptor and concurrent suppression of angiotensin II type 2 (AT2) receptor/ACE2-Mas receptor expression in the renal cortex. Chronic treatment with ORS increased UV and UNaV, and decreased the Na+ and water balance with a decrease in BP in the ORS-treated SHR-ORS group compared to SHR-V. These were accompanied by a decrease in NHE3 expression with a suppression of ACE-AT1 receptor and concurrent accentuation of AT2/ACE2-Mas receptor. ConclusionThe present study shows that ORS reduced BP with a decrease in Na+ and water retention by a suppression of NHE3 expression via modulation of RAS and NPS in SHR. The present study provides pharmacological rationale for the treatment of hypertension with ORS in SHR.

Cyclin D-CDK4 Disulfide Bond Attenuates Pulmonary Vascular Cell Proliferation.

Pulmonary hypertension (PH) is a chronic vascular disease characterized, among other abnormalities, by hyperproliferative smooth muscle cells and a perturbed cellular redox and metabolic balance. Oxidants induce cell cycle arrest to halt proliferation; however, little is known about the redox-regulated effector proteins that mediate these processes. Here, we report a novel kinase-inhibitory disulfide bond in cyclin D-CDK4 (cyclin-dependent kinase 4) and investigate its role in cell proliferation and PH. Oxidative modifications of cyclin D-CDK4 were detected in human pulmonary arterial smooth muscle cells and human pulmonary arterial endothelial cells. Site-directed mutagenesis, tandem mass-spectrometry, cell-based experiments, in vitro kinase activity assays, in silico structural modeling, and a novel redox-dead constitutive knock-in mouse were utilized to investigate the nature and definitively establish the importance of CDK4 cysteine modification in pulmonary vascular cell proliferation. Furthermore, the cyclin D-CDK4 oxidation was assessed in vivo in the pulmonary arteries and isolated human pulmonary arterial smooth muscle cells of patients with pulmonary arterial hypertension and in 3 preclinical models of PH. Cyclin D-CDK4 forms a reversible oxidant-induced heterodimeric disulfide dimer between C7/8 and C135, respectively, in cells in vitro and in pulmonary arteries in vivo to inhibit cyclin D-CDK4 kinase activity, decrease Rb (retinoblastoma) protein phosphorylation, and induce cell cycle arrest. Mutation of CDK4 C135 causes a kinase-impaired phenotype, which decreases cell proliferation rate and alleviates disease phenotype in an experimental mouse PH model, suggesting this cysteine is indispensable for cyclin D-CDK4 kinase activity. Pulmonary arteries and human pulmonary arterial smooth muscle cells from patients with pulmonary arterial hypertension display a decreased level of CDK4 disulfide, consistent with CDK4 being hyperactive in human pulmonary arterial hypertension. Furthermore, auranofin treatment, which induces the cyclin D-CDK4 disulfide, attenuates disease severity in experimental PH models by mitigating pulmonary vascular remodeling. A novel disulfide bond in cyclin D-CDK4 acts as a rapid switch to inhibit kinase activity and halt cell proliferation. This oxidative modification forms at a critical cysteine residue, which is unique to CDK4, offering the potential for the design of a selective covalent inhibitor predicted to be beneficial in PH.

Cardiovascular and metabolic effects of MANP. a novel GC-A activator, in human hypertension with metabolic syndrome

Abstract Background/Introduction Hypertension (HTN) and metabolic syndrome (MetS) are both leading risk factors for major cardiovascular diseases, which frequently coexist as a complicated cardiometabolic disorder. The cardiac hormone atrial natriuretic peptide (ANP) plays a key role in both blood pressure and metabolic regulation through activation of the guanylyl cyclase-A receptor (GC-A) and the second messenger 3’,5’ cyclic guanosine monophosphate (cGMP). Indeed, ANP decreases blood pressure, induces lipolysis by increasing plasma levels of non-esterified fatty acids (NEFA) and improves insulin-sensitivity. MANP is a new Mayo Clinic designed natriuretic peptide consisting of the native ANP with a 12-amino-acid C-terminus extension and its biological actions are mediated by the GC-A/cGMP pathway. MANP is less susceptible to neprilysin degradation and exert more sustained cGMP dependent biological actions than ANP. Purpose In a proof-of-principle human study, for the first time we aimed to determine the cardiovascular and metabolic properties of MANP, a novel ANP analog, in the clinical setting of HTN with MetS. Methods We conducted a double-blind, placebo-controlled trial in 22 patients (17 receiving MANP) with HTN and MetS involving single subcutaneous injection of MANP (2.5 µg/kg) or placebo (NCT03781739). Blood pressure (BP) and circulating cGMP were monitored for 24 hours post treatment. Circulating metabolic parameters including insulin, glucose, non-esterified fatty acids (NEFA) and insulin resistance (HOMA-IR) and insulin sensitivity (HOMA-IS) indexes were assessed for the first 4 hours post treatment. Results Compared to baseline, MANP increased plasma cGMP at 30 minutes (delta: 4.8 ± 2.0 pmol/mL, P = 0.02) and 1 hour (delta: 2.9 ± 1.3 pmol/mL, P = 0.03) post injection. At 6 hours, systolic BP decreased by 5.7 ± 2.9 mmHg (P = 0.06) in the MANP group, whereas no reduction in BP was observed in placebo group. Among the 17 subjects receiving MANP, lower baseline cGMP levels were associated with greater mean reductions in systolic BP (ρ = 0.66, P = 0.005). After MANP injection, glucose levels decreased at 2 (delta: -4.7 ± 2.1 mg/mL, P = 0.04) and 4 hours (delta: -13.1 ± 3.9 mg/mL, P = 0.003) compared to baseline while insulin levels remained stable. During the first 4 hours post injection, we also observed a trend of increase in insulin sensitivity index (HOMA2-IS) and decrease in insulin resistance index (HOMA2-IR) with MANP, both were opposite to the trends in placebo. At 1 hour post MANP administration, NEFA increased by 108.2 ± 37.4 µM compared to baseline (P = 0.01). Conclusion(s) Our findings support the favorable cardiovascular and metabolic actions of MANP as a potential therapeutic strategy for HTN with MetS.

MyD88 in myeloid cells promotes angiotensin II-induced vascular inflammation and is associated with all-cause mortality and incident heart failure in humans with arterial hypertension

Abstract Background Angiotensin II (AngII) causes hypertension and promotes vascular accumulation of inflammatory cells. We aimed to determine how myeloid factor of differentiation 88 (MyD88) contributes to vascular dysfunction and arterial hypertension in mice and humans. Methods Male C57BL/6 as well as MyD88-/-, LysMCre/wtMyD88LSL/LSL, LysMCre/wt, TLR2-/-, TLR4-/-, TLR7-/- and TLR9-/- were investigated in the AngII-model of hypertension (1 mg/kg/d for 7days). Biodata analyses were performed in the Gutenberg-Health Study (GHS), a population based cohort. In addition, we performed computational analyses of known hypertension-related genes from genome-wide association studies (GWAS) to investigate whether they preferentially interact with MyD88 in the human protein network. Results MyD88 deficiency attenuated AngII-induced blood pressure increase and endothelial dysfunction in conductance and resistance vessels. Vascular mRNA expression levels of vcam-1, nos2, nox2 were decreased in AngII-infused MyD88-/- mice compared to C57BL/6 controls. Flow cytometric analyses revealed that AngII-induced aortic accumulation of CD11b+Ly6Chi inflammatory monocytes was significantly dampened in MyD88-/- mice. AngII increased mRNA expression of cd62L, cd68 and ccl2, which was blocked in MyD88-/- mice, indicating a role of MyD88 for myeloid cell activation and differentiation. Additionally, aortic interferon-g+ NK cells and mRNA levels of interleukin-12 and interleukin-1b were reduced in AngII-treated MyD88-/- mice. Bone marrow transfer experiments demonstrated a vasoprotective effect of MyD88 deficiency in bone marrow-derived cells, and selective expression of MyD88 in LysMCre/wtMyD88LSL/LSL mice restored AngII-induced pathology, revealing that myelomonocytic cells drives vascular dysfunction in a MyD88-dependent manner. Furthermore, data from 1,274 individuals in the GHS indicated, that MyD88 mRNA expression was associated with all cause mortality and incident chronic heart failure after a median follow-up of 11.6 years. Computational analysis of the human protein interaction network demonstrated that MyD88 is significantly connected to proteins encoded by genetic loci associated with blood pressure traits in multiple GWAS. Conclusion We provide evidence, that MyD88 expressed by myelomonocytic cells promotes AngII-induced vascular dysfunction and arterial hypertension. MyD88 might be used as an inflammatory diagnostic marker for the risk of death in hypertensive patients.

CDN1163 alleviates SERCA2 dysfunction-induced pulmonary vascular remodeling by inhibiting the phenotypic transition of pulmonary artery smooth muscle cells

ABSTRACT Background and Purpose Substitution of Cys674 (C674) in the sarcoplasmic/endoplasmic reticulum Ca2+ ATPase 2 (SERCA2) causes SERCA2 dysfunction which leads to activated inositol requiring enzyme 1 alpha (IRE1α) and spliced X-box binding protein 1 (XBP1s) pathway accelerating cell proliferation of pulmonary artery smooth muscle cells (PASMCs) followed by significant pulmonary vascular remodeling resembling human pulmonary hypertension. Based on this knowledge, we intend to investigate other potential mechanisms involved in SERCA2 dysfunction-induced pulmonary vascular remodeling. Experimental Approach Heterozygous SERCA2 C674S knock-in (SKI) mice of which half of cysteine in 674 was substituted by serine to mimic the partial irreversible oxidation of C674 were used. The lungs of SKI mice and their littermate wild-type mice were collected for PASMC culture, protein expression, and pulmonary vascular remodeling analysis. Results SERCA2 dysfunction increased intracellular Ca2+ levels, which activated Ca2+-dependent calcineurin (CaN) and promoted the nuclear translocation and protein expression of the nuclear factor of activated T-lymphocytes 4 (NFAT4) in an IRE1α/XBP1s pathway-independent manner. In SKI PASMCs, the scavenge of intracellular Ca2+ by BAPTA-AM or inhibition of CaN by cyclosporin A can prevent PASMC phenotypic transition. CDN1163, a SERCA2 agonist, suppressed the activation of CaN/NFAT4 and IRE1α/XBP1s pathways, reversed the protein expression of PASMC phenotypic transition markers and cell cycle-related proteins, and inhibited cell proliferation and migration when given to SKI PASMCs. Furthermore, CDN1163 ameliorated pulmonary vascular remodeling in SKI mice. Conclusions and Implications SERCA2 dysfunction promotes PASMC phenotypic transition and pulmonary vascular remodeling by multiple mechanisms, which could be improved by SERCA2 agonist CDN1163.

The effect of zofenopril on the cardiovascular system of spontaneously hypertensive rats treated with the ACE2 inhibitor MLN-4760

BackgroundAngiotensin converting enzyme 2 (ACE2) plays a crucial role in the infection cycle of SARS-CoV-2 responsible for formation of COVID-19 pandemic. In the cardiovascular system, the virus enters the cells by binding to the transmembrane form of ACE2 causing detrimental effects especially in individuals with developed hypertension or heart disease. Zofenopril, a H2S-releasing angiotensin-converting enzyme inhibitor (ACEI), has been shown to be effective in the treatment of patients with essential hypertension; however, in conditions of ACE2 inhibition its potential beneficial effect has not been investigated yet. Therefore, the aim of the study was to determine the effect of zofenopril on the cardiovascular system of spontaneously hypertensive rats, an animal model of human essential hypertension and heart failure, under conditions of ACE2 inhibition induced by the administration of the specific inhibitor MLN-4760 (MLN).ResultsZofenopril reduced MLN-increased visceral fat to body weight ratio although no changes in systolic blood pressure were recorded. Zofenopril administration resulted in a favorable increase in left ventricle ejection fraction and improvement of diastolic function regardless of ACE2 inhibition, which was associated with increased H2S levels in plasma and heart tissue. Similarly, the acute hypotensive responses induced by acetylcholine, L-NAME (NOsynthase inhibitor) and captopril (ACEI) were comparable after zofenopril administration independently from ACE2 inhibition. Although simultaneous treatment with zofenopril and MLN led to increased thoracic aorta vasorelaxation, zofenopril increased the NO component equally regardless of MLN treatment, which was associated with increased NO-synthase activity in aorta and left ventricle. Moreover, unlike in control rats, the endogenous H2S participated in maintaining of aortic endothelial function in MLN-treated rats and the treatment with zofenopril had no impact on this effect.ConclusionsZofenopril treatment reduced MLN-induced adiposity and improved cardiac function regardless of ACE2 inhibition. Although the concomitant MLN and zofenopril treatment increased thoracic aorta vasorelaxation capacity, zofenopril increased the participation of H2S and NO in the maintenance of endothelial function independently from ACE2 inhibition. Our results confirmed that the beneficial effects of zofenopril were not affected by ACE2 inhibition, moreover, we assume that ACE2 inhibition itself can lead to the activation of cardiovascular compensatory mechanisms associated with Mas receptor, nitrous and sulfide signaling.

OR06-04 Environmental Pollutants Perfluoroalkyl Acids Potently Stimulated Aldosterone Biosynthesis

Abstract Disclosure: B. Caroccia: None. T.M. Seccia: None. G. Pallafacchina: None. M. Piazza: None. I. Caputo: None. R. Rizzuto: None. G. Rossi: None. A large environmental contamination of drinking water by perfluoroalkyl substances (PFAS) markedly increased by 8-fold the plasma levels of pentadecafluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) in a Northern Italy population with that shows a high prevalence of arterial hypertension and cardiovascular disease. As the link between PFAS and arterial hypertension is unknown, we investigated if they could enhance the biosynthesis of aldosterone, the main mineralocorticoid hormone that has pressor effects. We exposed human adrenocortical carcinoma HAC15 cells to 1 µM or 10 µM concentration of PFOA and PFOS, alone or in combination, for 24, 48 or 72 hours and we measured changes in cell viability, aldosterone synthase (CYP11B2) mRNA, cell and mitochondrial reactive oxygen species (ROS), and aldosterone production in the absence or presence of the superoxide scavenger tempol. Experiments were also performed in the presence of 10 or 100 nM Ang II. We found that PFAS increased CYP11B2 mRNA by 3-fold and aldosterone secretion and mitochondrial ROS production by 2-fold over controls (p &amp;lt; 0.01 for all). PFAS also enhanced the effects of the aldosterone secretagogue Ang II on CYP11B2 mRNA and aldosterone secretion: 10 nM or 100 nM Ang II increased CYP11B2 mRNA by 12- and 52-fold, respectively, compared to vehicle (p &amp;lt; 0.001). Moreover, 1 µM PFAS potentiated the effect of 10 nM or 100 nM Ang II on CYP11B2 gene expression by 30- and 80-fold, respectively (p &amp;lt; 0.001). As the ROS scavenger tempol, which decreased ROS, significantly blunted the CYP11B2 gene expression induced by either PFAS alone, or by concomitant exposure to Ang II and PFAS, the effect of PFAS likely occurs via increased ROS generation. These results indicate that at concentrations mimicking those found in human plasma of exposed individuals, PFAS exert a prooxidant effect and act as potent disruptors of human adrenocortical cell function, thus implicating them as causative factors of human arterial hypertension due to aldosteronism. Presentation: Thursday, June 15, 2023

Arsenic trioxide demonstrates efficacy in a mouse model of preclinical systemic sclerosis

BackgroundUncontrolled T-cell activation plays a key role in systemic sclerosis (SSc). Arsenic trioxide (ATO) has immunological effects and has demonstrated potential in preclinical SSc models. In this study, we assessed the efficacy of ATO in Fra2 transgenic (Fra2TG) mice, which develop severe vascular remodeling of pulmonary arterioles and nonspecific interstitial pneumonia-like lung disease, closely resembling human SSc-associated pulmonary hypertension, therefore partially resembling to the SSc human disease.MethodsThe efficacy of ATO in Fra2TG mice was evaluated through histological scoring and determination of cell infiltration. Fibrotic changes in the lungs were assessed by measuring collagen content biochemically, using second harmonic generation to measure fibrillar collagen, and imaging via computed tomography. Cardiovascular effects were determined by measuring right ventricular systolic pressure and vessel remodeling. The mechanism of action of ATO was then investigated by analyzing lung cell infiltrates using flow cytometry and bulk RNA with sequencing techniques.ResultsAfter ATO treatment, the Ashcroft histological score was substantially decreased by 33% in ATO-treated mice compared to control mice. Other investigations of fibrotic markers showed a trend of reduction in various measurements of fibrosis, but the differences did not reach significance. Further cardiovascular investigations revealed convergent findings supporting a beneficial effect of ATO, with reduced right ventricular systolic pressure and medial wall thickness, and a significant decrease in the number of muscularized distal pulmonary arteries in ATO-treated Fra2TG mice compared to untreated Fra2TG mice. Additionally, inflammatory cell infiltration was also markedly reduced in lesioned lungs. A reduction in the frequency of CD4 + and T effector memory cells, and an increase in the percentage of CD4 + T naive cells in the lungs of ATO-treated Fra-2TG mice, was observed when compared to PBS group Fra-2Tg mice. RNA-seq analysis of ATO-treated mouse lungs revealed a downregulation of biological pathways associated with immune activity and inflammation, such as T-cell activation, regulation of leucocyte activation, leucocyte cell–cell adhesion, and regulation of lymphocyte activation.ConclusionsOur results suggest the clinical relevance of ATO treatment in SSc. Using the Fra2TG mouse model, we observed significant lung histological changes, a trend towards a decrease in various fibrotic makers, and a strong reduction in vascular remodeling. The mechanism of action of ATO appears to involve a marked counteraction of the immune activation characteristic of SSc, particularly T-cell involvement. These findings pave the way for further studies in SSc.

Altered Smooth Muscle Cell Histone Acetylome by the SPHK2/S1P Axis Promotes Pulmonary Hypertension.

Epigenetic regulation of vascular remodeling in pulmonary hypertension (PH) is poorly understood. Transcription regulating, histone acetylation code alters chromatin accessibility to promote transcriptional activation. Our goal was to identify upstream mechanisms that disrupt epigenetic equilibrium in PH. Human pulmonary artery smooth muscle cells (PASMCs), human idiopathic pulmonary arterial hypertension (iPAH):human PASMCs, iPAH lung tissue, failed donor lung tissue, human pulmonary microvascular endothelial cells, iPAH:PASMC and non-iPAH:PASMC RNA-seq databases, NanoString nCounter, and cleavage under targets and release using nuclease were utilized to investigate histone acetylation, hyperacetylation targets, protein and gene expression, sphingolipid activation, cell proliferation, and gene target identification. SPHK2 (sphingosine kinase 2) knockout was compared with control C57BL/6NJ mice after 3 weeks of hypoxia and assessed for indices of PH. We identified that Human PASMCs are vulnerable to the transcription-promoting epigenetic mediator histone acetylation resulting in alterations in transcription machinery and confirmed its pathological existence in PH:PASMC cells. We report that SPHK2 is elevated as much as 20-fold in iPAH lung tissue and is elevated in iPAH:PASMC cells. During PH pathogenesis, nuclear SPHK2 activates nuclear bioactive lipid S1P (sphingosine 1-phosphate) catalyzing enzyme and mediates transcription regulating histone H3K9 acetylation (acetyl histone H3 lysine 9 [Ac-H3K9]) through EMAP (endothelial monocyte activating polypeptide) II. In iPAH lungs, we identified a 4-fold elevation of the reversible epigenetic transcription modulator Ac-H3K9:H3 ratio. Loss of SPHK2 inhibited hypoxic-induced PH and Ac-H3K9 in mice. We discovered that pulmonary vascular endothelial cells are a priming factor of the EMAP II/SPHK2/S1P axis that alters the acetylome with a specificity for PASMC, through hyperacetylation of histone H3K9. Using cleavage under targets and release using nuclease, we further show that EMAP II-mediated SPHK2 has the potential to modify the local transcription machinery of pluripotency factor KLF4 (Krüppel-like factor 4) by hyperacetylating KLF4 Cis-regulatory elements while deletion and targeted inhibition of SPHK2 rescues transcription altering Ac-H3K9. SPHK2 expression and its activation of the reversible histone H3K9 acetylation in human pulmonary artery smooth muscle cell represent new therapeutic targets that could mitigate PH vascular remodeling.

Metabolic profile of blood serum in experimental arterial hypertension.

The etiology of essential hypertension is intricate, since it employs simultaneously various body systems related to the regulation of blood pressure in one way or another: the sympathetic nervous system, renin-angiotensin-aldosterone and hypothalamic-pituitary-adrenal systems, renal and endothelial mechanisms. The pathogenesis of hypertension is influenced by a variety of both genetic and environmental factors, which determines the heterogeneity of the disease in human population. Hence, there is a need to perform research on experimental models - inbred animal strains, one of them being ISIAH rat strain, which is designed to simulate inherited stress-induced arterial hypertension as close as possible to primary (or essential) hypertension in humans. To determine specific markers of diseases, various omics technologies are applied, including metabolomics, which makes it possible to evaluate the content of low-molecular compounds - amino acids, lipids, carbohydrates, nucleic acids fragments - in biological samples available for clinical analysis (blood and urine). We analyzed the metabolic profile of the blood serum of male ISIAH rats with a genetic stress-dependent form of arterial hypertension in comparison with the normotensive WAG rats. Using the method of nuclear magnetic resonance spectroscopy (NMR spectroscopy), 56 metabolites in blood serum samples were identified, 18 of which were shown to have significant interstrain differences in serum concentrations. Statistical analysis of the data obtained showed that the hypertensive status of ISIAH rats is characterized by increased concentrations of leucine, isoleucine, valine, myo-inositol, isobutyrate, glutamate, glutamine, ornithine and creatine phosphate, and reduced concentrations of 2-hydroxyisobutyrate, betaine, tyrosine and tryptophan. Such a ratio of the metabolite concentrations is associated with changes in the regulation of glucose metabolism (metabolic markers - leucine, isoleucine, valine, myo-inositol), of nitric oxide synthesis (ornithine) and catecholamine pathway (tyrosine), and with inflammatory processes (metabolic markers - betaine, tryptophan), all of these changes being typical for hypertensive status. Thus, metabolic profiling of the stress-dependent form of arterial hypertension seems to be an important result for a personalized approach to the prevention and treatment of hypertensive disease.

Eicosanoid-Regulated Myeloid ENaC and Isolevuglandin Formation in Human Salt-Sensitive Hypertension.

The mechanisms by which salt increases blood pressure in people with salt sensitivity remain unclear. Our previous studies found that high sodium enters antigen-presenting cells (APCs) via the epithelial sodium channel and leads to the production of isolevuglandins and hypertension. In the current mechanistic clinical study, we hypothesized that epithelial sodium channel-dependent isolevuglandin-adduct formation in APCs is regulated by epoxyeicosatrienoic acids (EETs) and leads to salt-sensitive hypertension in humans. Salt sensitivity was assessed in 19 hypertensive subjects using an inpatient salt loading and depletion protocol. Isolevuglandin-adduct accumulation in APCs was analyzed using flow cytometry. Gene expression in APCs was analyzed using cellular indexing of transcriptomes and epitopes by sequencing analysis of blood mononuclear cells. Plasma and urine EETs were measured using liquid chromatography-mass spectrometry. Baseline isolevuglandin+ APCs correlated with higher salt-sensitivity index. Isolevuglandin+ APCs significantly decreased from salt loading to depletion with an increasing salt-sensitivity index. We observed that human APCs express the epithelial sodium channel δ subunit, SGK1 (salt-sensing kinase serum/glucocorticoid kinase 1), and cytochrome P450 2S1. We found a direct correlation between baseline urinary 14,15 EET and salt-sensitivity index, whereas changes in urinary 14,15 EET negatively correlated with isolevuglandin+ monocytes from salt loading to depletion. Coincubation with 14,15 EET inhibited high-salt-induced increase in isolevuglandin+ APC. Isolevuglandin formation in APCs responds to acute changes in salt intake in salt-sensitive but not salt-resistant people with hypertension, and this may be regulated by renal 14,15 EET. Baseline levels of isolevuglandin+ APCs or urinary 14,15 EET may provide diagnostic tools for salt sensitivity without a protocol of salt loading.

Elevated arsenic level in fasting serum via ingestion of fish meat increased the risk of hypertension in humans and mice.

There has been a shortage of human studies to elucidate the association between serum arsenic levels and the prevalence of hypertension. This study multidirectionally investigated associations among arsenic exposure, dietary ingestion, and the risk of hypertension by combined human epidemiological and mouse experimental studies. This study focused on the total arsenic level in fasting serum, a biomarker of arsenic exposure. Associations among ingestion frequencies of 54 diet items of Japanese food separated into six categories, total arsenic level in fasting serum, and the prevalence of hypertension were investigated in 2709 general people in Japan. Logistic regression analysis demonstrated a dose-dependent association between serum arsenic level and hypertension and a positive association between the ingestion of fish meat and hypertension. Further analysis showed that the latter association was fully mediated by increased fasting serum arsenic levels in humans. Similarly, oral exposure to the putative human-equivalent dose of arsenic species mixture with the same ratios in a common fish meat in Japan increased systolic blood pressure and arsenic levels in fasting serum in mice. This interdisciplinary approach suggests that fish-meat ingestion is a potential risk factor for arsenic-mediated hypertension. Because the increased consumption of fish meat is a recent global trend, health risks of the increased ingestion of arsenic via fish meat should be further investigated.

Abstract 011: Cluster Of Differentiation 14 (CD14) Inhibits NADPH Oxidase 2 (NOX2)-Derived Oxidative Stress In Hematopoietic Cells And Modulates Dahl Salt-Sensitive Hypertension And Renal Damage

We recently discovered that genetic deletion of CD14 in Dahl SS rats (CD14-/-) amplifies the development of salt-sensitive hypertension and renal damage in females. Interestingly, the expression of CD14 in leukocytes and the level of plasma CD14 protein are increased in cardiovascular disease and hypertension in humans. An examination of macrophages from CD14-/- females revealed enhanced production of superoxide from NOX2 compared to Dahl SS. Furthermore, a double knockout Dahl SS strain lacking both CD14 and functional NOX2 (DKO) exhibits significantly reduced salt-induced hypertension and associated renal damage, indicating that free radical production from NOX2 is likely mediating the amplification of disease following CD14 deletion. The present study examined the specific effects of CD14 and NOX2 deletion in hematopoietic cells via a total body irradiation/bone marrow transfer (TBI/BMT) approach. Female Dahl SS recipients were irradiated (11Gy) then received either CD14 or DKO bone marrow transplantation via tail vein injection. Rats that received DKO hematopoietic cells had blunted mean arterial pressure (147±4 mmHg, n=3, p=0.052) versus those that received CD14-/- (179±19mmHg, n=3) in response to a 3-week high salt challenge (HS, 4.0% NaCl, AIN-76A). As an index of renal damage, albuminuria was significantly reduced in the DKO (110.3±7 mg/day, n=3, p&lt;0.01) compared to the CD14-/- (280.9±42 mg/day, n=3) at HS day 21. Flow cytometric analysis of isolated renal immune cells indicated a reduction in renal inflammation in the DKO compared to the CD14-/-. There were fewer CD45+ total leukocytes (2.8x10 6 vs 3.8x10 6 cells/kidney, p&lt;0.05) and CD11b/c+ monocytes/macrophages (2.4x10 6 vs 3.3x10 6 cells/kidney, n=3, p&lt;0.05) in DKO versus CD14-/- kidneys. Finally, to document the TBI/BMT, macrophages from CD14-/- rats produced 7-fold greater superoxide from NOX2 when stimulated with 12-phorbol myristate 13-acetate compared to DKO macrophages. Together, these data suggest a regulatory role of CD14 in hematopoietic cells to reduce oxidative stress from NOX2 in the kidney of female Dahl SS rats during salt-sensitive hypertension and associated renal disease.

Proteomic mapping reveals dysregulated angiogenesis in the cerebral arteries of rats with early-onset hypertension

Hypertension is associated with the presence of vascular abnormalities, including remodeling and rarefaction. These processes play an important role in cerebrovascular disease development; however, the mechanistic changes leading to these diseases are not well characterized. Using data-independent acquisition-based mass spectrometry analysis, here we determined the protein changes in cerebral arteries in pre- and early-onset hypertension from the spontaneously hypertensive rat (SHR), a model that resembles essential hypertension in humans. Our analysis identified 125 proteins with expression levels that were significantly upregulated or downregulated in 12-week-old spontaneously hypertensive rats compared to normotensive Wistar Kyoto rats. Using an angiogenesis enrichment analysis, we further identified a critical imbalance in angiogenic proteins that promoted an anti-angiogenic profile in cerebral arteries at early onset of hypertension. In a comparison to previously published data, we demonstrate that this angiogenic imbalance is not present in mesenteric and renal arteries from age-matched SHRs. Finally, we identified two proteins (Fbln5 and Cdh13), whose expression levels were critically altered in cerebral arteries compared to the other arterial beds. The observation of an angiogenic imbalance in cerebral arteries from the SHR reveals critical protein changes in the cerebrovasculature at the early onset of hypertension and provides novel insights into the early pathology of cerebrovascular disease.

Abstract P186: CCR10 Promotes Blood Pressure Elevations And Renal Damage In A Mouse Model Of Hypertension

Emerging evidence suggests inflammation plays a key role in hypertension and its associatedend-organ damage. Regulatory T cells (Tregs) function primarily by limiting inflammation andare considered protective in the pathogenesis of hypertension. However, Tregs also havetissue-specific functions and have been found to play pathogenic roles in some chronicdiseases by limiting angiogenesis. As such, the role of Tregs and its subsets in chronichypertension remains unclear. Recently, we identified CCR10 + Tregs as an immune populationselectively decreased in the circulation of hypertensive patients. CCR10 is a chemokinereceptor important for recruitment of immune cells to the skin. Although skin has not been wellstudied in hypertension, prior reports in humans demonstrate that loss of skin microvessels(microvascular rarefaction) occurs as a putative mechanism by which blood pressure (BP) isincreased. Thus, we hypothesized that CCR10 + Tregs promote hypertension through skininfiltration leading to enhanced microvascular rarefaction. Herein, we demonstrate that,compared to normotensive controls, mice with Angiotensin II (Ang II)-induced hypertension haveincreased abundance of CCR10 + Tregs in the skin (p=0.0004) with a corresponding decrease incirculation (p=0.003). These mice also displayed increased cutaneous levels of CCL27(p=0.0007), a ligand for CCR10. Among circulating lymphocytes in normotensive mice, wefound that Tregs express the highest frequency of CCR10 (p=0.0007). Importantly, wedemonstrate that CCR10-deficient mice (CCR10 -/- ) are more resistant to Ang II-induced systolicblood pressure elevations (p=0.004) and albuminuria (p=0.01) compared to wild type mice.CCR10 -/- mice also exhibit higher levels of cutaneous arteriolar density (p=0.02) and VEGFA(p=0.009), consistent with reduced skin microvascular rarefaction. Finally, using PrediXcan, wedemonstrate that higher genetically predicted levels of CCR10 are associated with increasedrisk of hypertension in humans. Taken together, our data suggest that CCR10 promotes Ang II-induced BP elevations and renal damage in mice, at least in part by promoting Treg recruitmentto the skin leading to microvascular rarefaction.