Search

The aims of this study were to 1) determine whether renal localization of aliskiren and its antihypertensive and renoprotective effects persist after administration of the drug is stopped and 2) define the renal localization of aliskiren by light microscopy autoradiography. Hypertensive double transgenic rats (dTGR) overexpressing genes for human renin and angiotensinogen were treated with aliskiren (3 mg·kg-1·day-1 sc; osmotic minipumps) or enalapril (18 mg/l in drinking water). After a 2-wk treatment, dTGR were assigned to either continued treatment with aliskiren ("continued"), or to cessation of their respective treatment ("stopped") for a 3-wk washout. One week of treatment with aliskiren and enalapril reduced blood pressure and albuminuria vs. baseline. After cessation of either treatment, blood pressure had returned to pretreatment levels and albuminuria remained relatively low for 1 wk, but rose thereafter similarly in both groups. In contrast, renal mRNA for transforming growth factor-β and renal collagen IV was reduced by aliskiren (continued and stopped groups), but not after cessation of enalapril. Similar patterns were found for collagen IV protein expression. Even 3 wk after stopping aliskiren treatment, renal levels of the drug exceeded its IC50, whereas enalaprilat was not detected. To localize aliskiren accumulation, Wistar rats were treated with [³H]-aliskiren for 7 days. Autoradiography demonstrated specific labeling in glomeruli, arterioles, and afferent arterioles as well as in the distal nephron. Labeling could still be observed even after 7 days' washout. These results suggest that the renophilic properties of aliskiren are different from enalapril and could have contributed to the renoprotective mechanism of this renin inhibitor. [ABSTRACT FROM AUTHOR]

There has been a marked increase in the incidence of autoimmune diseases in the past half-century. Although the underlying genetic basis of this class of diseases has recently been elucidated, implicating predominantly immune-response genes, changes in environmental factors must ultimately be driving this increase. The newly identified population of interleukin (IL)-17-producing CD4+ helper T cells (TH17 cells) has a pivotal role in autoimmune diseases. Pathogenic IL-23-dependent TH17 cells have been shown to be critical for the development of experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis, and genetic risk factors associated with multiple sclerosis are related to the IL-23-TH17 pathway. However, little is known about the environmental factors that directly influence TH17 cells. Here we show that increased salt (sodium chloride, NaCl) concentrations found locally under physiological conditions in vivo markedly boost the induction of murine and human TH17 cells. High-salt conditions activate the p38/MAPK pathway involving nuclear factor of activated T cells 5 (NFAT5; also called TONEBP) and serum/glucocorticoid-regulated kinase 1 (SGK1) during cytokine-induced TH17 polarization. Gene silencing or chemical inhibition of p38/MAPK, NFAT5 or SGK1 abrogates the high-salt-induced TH17 cell development. The TH17 cells generated under high-salt conditions display a highly pathogenic and stable phenotype characterized by the upregulation of the pro-inflammatory cytokines GM-CSF, TNF-α and IL-2. Moreover, mice fed with a high-salt diet develop a more severe form of EAE, in line with augmented central nervous system infiltrating and peripherally induced antigen-specific TH17 cells. Thus, increased dietary salt intake might represent an environmental risk factor for the development of autoimmune diseases through the induction of pathogenic TH17 cells. [ABSTRACT FROM AUTHOR]

Background: Agonistic autoantibodies to the α1-adrenergic receptor occur in nearly half of patients with refractory hypertension; however, their relevance is uncertain. Methods/Principal Findings: We immunized Lewis rats with the second extracellular-loop peptides of the human α1A-adrenergic receptor and maintained them for one year. α1A-adrenergic antibodies (α1A-AR-AB) were monitored with a neonatal cardiomyocyte contraction assay by ELISA, and by ERK1/2 phosphorylation in human α1A-adrenergic receptor transfected Chinese hamster ovary cells. The rats were followed with radiotelemetric blood pressure measurements and echocardiography. At 12 months, the left ventricles of immunized rats had greater wall thickness than control rats. The fractional shortening and dp/dtmax demonstrated preserved systolic function. A decreased E/A ratio in immunized rats indicated a diastolic dysfunction. Invasive hemodynamics revealed increased left ventricular end-diastolic pressures and decreased dp/dtmin. Mean diameter of cardiomyocytes showed hypertrophy in immunized rats. Long-term blood pressure values and heart rates were not different. Genes encoding sarcomeric proteins, collagens, extracellular matrix proteins, calcium regulating proteins, and proteins of energy metabolism in immunized rat hearts were upregulated, compared to controls. Furthermore, fibrosis was present in immunized hearts, but not in control hearts. A subset of immunized and control rats was infused with angiotensin (Ang) II. The stressor raised blood pressure to a greater degree and led to more cardiac fibrosis in immunized, than in control rats. Conclusions/Significance: We show that α1A-AR-AB cause diastolic dysfunction independent of hypertension, and can increase the sensitivity to Ang II. We suggest that α1A-AR-AB could contribute to cardiovascular endorgan damage. [ABSTRACT FROM AUTHOR]

Background: Agonistic autoantibodies directed at the α1-adrenergic receptor (α1-AAB) have been described in patients with hypertension. We implied earlier that a1-AAB might have a mechanistic role and could represent a therapeutic target. Methodology/Principal Findings: To pursue the issue, we performed clinical and basic studies. We observed that 41 of 81 patients with refractory hypertension had α1-AAB; after immunoadsorption blood pressure was significantly reduced in these patients. Rabbits were immunized to generate a1-adrenergic receptor antibodies (α1-AB). Patient a1-AAB and rabbit α1-AB were purified using affinity chromatography and characterized both by epitope mapping and surface plasmon resonance measurements. Neonatal rat cardiomyocytes, rat vascular smooth muscle cells (VSMC), and Chinese hamster ovary cells transfected with the human α1A-adrenergic receptor were incubated with patient α1-AAB and rabbit α1-AB and the activation of signal transduction pathways was investigated by Western blot, confocal laser scanning microscopy, and gene expression. We found that phospholipase A2 group IIA (PLA2-IIA) and L-type calcium channel (Cacna1c) genes were upregulated in cardiomyocytes and VSMC after stimulation with both purified antibodies. We showed that patient α1-AAB and rabbit α1-AB result in protein kinase C alpha activation and transient extracellular-related kinase (EKR1/2) phosphorylation. Finally, we showed that the antibodies exert acute effects on intracellular Ca2+ in cardiomyocytes and induce mesentery artery segment contraction. Conclusions/Significance: Patient α1-AAB and rabbit α1-AB can induce signaling pathways important for hypertension and cardiac remodeling. Our data provide evidence for a potential clinical relevance for α1-AAB in hypertensive patients, and the notion of immunity as a possible cause of hypertension. [ABSTRACT FROM AUTHOR]

Examines the use of angiotensin II dependent model to test the ability of cerivastatin to prevent leukocyte adhesion and infiltration. Analysis on mitogen-activated protein kinase and transcription factor activation; Increase in inducible nitric oxied synthase expression in the endothelium vessels; Reduction of cell damage.

Echocardiography is the most commonly applied technique for non-invasive assessment of cardiac function in small animals. Manual tracing of endocardial borders is time consuming and varies with operator experience. Therefore, we aimed to evaluate a novel automated two-dimensional software algorithm (Auto2DE) for small animals and compare it to the standard use of manual 2D-echocardiographic assessment (2DE). We hypothesized that novel Auto2DE will provide rapid and robust data sets, which are in agreement with manually assessed data of animals.2DE and Auto2DE were carried out using a high-resolution imaging-system for small animals. First, validation cohorts of mouse and rat cine loops were used to compare Auto2DE against 2DE. These data were stratified for image quality by a blinded expert in small animal imaging. Second, we evaluated 2DE and Auto2DE in four mouse models and four rat models with different cardiac pathologies.Automated assessment of LV function by 2DE was faster than conventional 2DE analysis and independent of operator experience levels. The accuracy of Auto2DE-assessed data in healthy mice was dependent on cine loop quality, with excellent agreement between Auto2DE and 2DE in cine loops with adequate quality. Auto2DE allowed for valid detection of impaired cardiac function in animal models with pronounced cardiac phenotypes, but yielded poor performance in diabetic animal models independent of image quality.Auto2DE represents a novel automated analysis tool for rapid assessment of LV function, which is suitable for data acquisition in studies with good and very good echocardiographic image quality, but presents systematic problems in specific pathologies. [ABSTRACT FROM AUTHOR]

Objective: High-salt intake is associated with cardiovascular disease and hypertension. A substantial number of patients are sensitive to salt, but environmental factors influencing this phenomenon are poorly understood. The gut microbiome has not been considered in the context of salt and hypertension research to date. It is known to respond to fluctuations in lifestyle and diet, and is increasingly recognized as an important factor which influences host health and disease. Using next generation sequencing methods we aimed to determine the effect of a high-salt diet on gut microbiome composition.Methods and Results: Male C57BL6/J mice, 10 weeks of age, were individually housed and either subjected to a purified normal salt (0.5% sodium) or high-salt diet (4% sodium + 1% in drinking water) ad libitum for 14 days. Feces was collected on days 0 and 14 of the diets. DNA was extracted from feces, the V4 region of the 16S rRNA gene amplified with PCR. Samples were multiplexed and sequenced with MiSeq. Reads were quality controlled and operational taxonomic units (OTUs) were called by comparison to the Greengenes database.The high-salt diet created a distinct gut microbiome composition compared to the normal-salt diet (analysis of Jensen-Shannon divergence). Whereas normal diet’s gut microbiome is composed mostly of OTUs from the Bacteroidetes phylum (70% of reads), high-salt diet causes a shift in relative abundance to other phyla (increased Firmicutes:Bacteroidetes ratio). Moreover, the genus Allobaculum significantly spikes after high-salt feeding. Both phenomena have recently been implicated in obesity studies.Conclusion: An increase in dietary sodium chloride alters the gut microbiome composition. Our findings indicate new perspectives on how salt impacts the body. [ABSTRACT FROM AUTHOR]

There has been a marked increase in the incidence of autoimmune diseases in the last half-century. While the underlying genetic basis of this class of diseases has recently been elucidated implicating predominantly immune response genes1, changes in environmental factors must ultimate be driving this increase. The recently identified population of interleukin (IL)-17 producing CD4+ helper T cells (Th17 cells) play a pivotal role in autoimmune diseases2. Pathogenic IL-23 dependent Th17 cells have been shown to be critical for the development of experimental autoimmune encephalomyelitis (EAE) an animal model for multiple sclerosis (MS) and genetic risk factors associated with MS are related to the IL23/Th17 pathway1, 2. However, little is known regarding the environmental factors that directly influence Th17 cells. Here we show that increased salt (sodium chloride; NaCl) concentrations found under physiological conditions in vivo dramatically boost the induction of murine and human Th17 cells. High salt conditions activates the p38/MAPK pathway involving the tonicity-responsive enhancer binding protein (TonEBP/NFAT5) and the serum- and glucocorticoid-inducible kinase 1 (SGK1) during cytokine-induced Th17 polarization. Gene silencing or chemical inhibition of p38/MAPK, NFAT5 or SGK1 abrogates the high salt induced Th17 cell development. The Th17 cells generated under high salt display a highly pathogenic and stable phenotype as compared to cells differentiated under normal conditions characterized by the up-regulation of the pro-inflammatory cytokines GM-CSF, TNFα and IL-2. Moreover, mice fed with a high salt diet develop a very severe form of EAE in line with increased central nervous system infiltrating and peripherally induced antigen specific Th17 cells. Thus, increased dietary salt intake might represent an environmental risk factor for the development of autoimmune diseases through the induction of pathogenic Th17 cells. [ABSTRACT FROM AUTHOR]

Hypertension is associated with increased risk of Alzheimer disease (AD) that features Amyloid-β peptide (Aβ) deposition in brain (neuritic plaques) and in blood vessels (cerebral amyloid angiopathy). Recently, it was shown by others that Aβ peptides exert a chronotropic effect in neonatal cardiomyocytes, similar to α1-adrenoreceptor autoantibodies (α-AB) as we described earlier. We hypothesized that Aβ peptides elicit a signal transduction pathway in vascular cells, induced by activation of the α1-adrenoreceptor. Aβ peptides (10-35) and (25-35) induced a positive chorontropic effect in the cardiac contraction assay (CCA) after long term activation up to 4 hours (28.84±1.04 bpm and 28.96±1.08 bpm). The effect was attenuated by the α1-receptor blockers urapidil and prazosin (1.72±0.4 bpm and -0.76±1.16 bpm). β1, β2, AT1, or AT2 receptor blockers had no effect on the positive chronotropic effect of Aβ (10-35) and (25-35). Using confocal microscopy and phosphospecific immunoblotting we observed that both peptides induced ERK 1/2 phosphorylation in Chinese hamster ovary (CHO) cells, overexpressing the α1A adrenoreceptor (CHO-α1-AR), but not in wildtype CHO. pERK was also induced in vascular smooth muscle cells (VSMC) and cardiomyocytes. Both Aβ peptides induced an intracellular Ca2+ signal in VSMC. The positive chronotropic response to Aβ (25-35) in the CCA was blocked with a peptide corresponding to the 1st extracellular loop of the α1-AR, the identical epitope to which α1-AB binds. Our data show that Amyloid-β peptides induce a signal transduction cascade via the α1-adrenoreceptor in VSMC and cardiomyocytes. These data might help explain the newly discovered association between hypertension and Alzheimer disease. The role of Aβ peptides in vascular medicine deserves further study. [ABSTRACT FROM AUTHOR]

Recent advances in our understanding of cardiovascular diseases clearly show that inflammation and activation of immunity are central features in the pathogenesis of atherosclerosis, ischemic myocardial injury, and also in hypertension-induced target organ damage. However, the idea that special immune cells could regulate immune responses in these conditions in favor of minimizing disease is a novel concept. Regulatory T cells have unique immune modulatory properties that offer an attractive alternative to common immunosuppressant drugs. Their application in animal models of autoimmunity and neoplastic conditions offers exciting therapeutic avenues. Thus, with the use of regulatory T cells in hypertension-induced target organ damage enables new insights into the pathophysiologic mechanisms and widen our knowledge of the role of the immune system in cardiovascular disease. The aim of this review was to summarize and discuss some of the most recent insights and put them into a perspective based on well-known interactions between immunity and hypertensive damage. [Copyright &y& Elsevier]

Inducible NOS inhibition, eicosapentaenoic acid supplementation, and angiotensin II–induced renal damage.Background.Cytochrome P450(CYP)-dependent hydroxylation and epoxygenation metabolites of arachidonic acid (AA) influence renal vascular tone, salt excretion, and inflammation. Transgenic rats over expressing both human renin and angiotensinogen genes (dTGR) feature angiotensin II (Ang II)–induced organ damage, increased expression of inducible nitric oxide synthase (iNOS), decreased AA hydroxylation, and epoxygenation. As nitric oxide production via iNOS can inhibit CYP AA metabolism, we tested the hypothesis that by blocking iNOS or by supplementing eicosapentanoic acid (EPA), which can serve as an alternative CYP substrate, Ang II–induced vasculopathy could be ameliorated.Methods.We treated dTGR with the iNOS inhibitor L-N(6)-(1-iminoethyl) lysine (L-NIL), EPA, and the combination of both treatments from week 4 to 7.Results.Immunohistochemistry showed that L-NIL and EPA reduced glomerular iNOS toward control levels. L-NIL–treated dTGR showed cardiac hypertrophy and albuminuria similar to untreated dTGR. EPA and the combination of EPA+ L-NIL, ameliorated organ damage without lowering blood pressure. EPA and EPA+ L-NIL reduced cardiac hypertrophy, albuminuria, renal fibronectin expression, and infiltration of monocytes/macrophages, compared to L-NIL and untreated dTGR. Reactive oxygen species were detected in glomeruli of untreated and L-NIL–treated dTGR, but was reduced in the EPA groups. EPA treatment reduced activator protein-1 (AP-1) activation and partially inhibited nuclear factor-kappaB (NF-κB) activity in kidneys of dTGR.Conclusion.These results demonstrate that iNOS inhibition does not protect against Ang II–induced end-organ damage, while EPA treatment does. Our electromobility shift assay experiments revealed that EPA protection may involve inhibition of AP-1– and NF-κB–dependent pathways. [ABSTRACT FROM AUTHOR]

Gain-of-function mutations in the CACNA1H gene (encoding the T-type calcium channel CaV3.2) cause autosomal-dominant familial hyperaldosteronism type IV (FH-IV) and early-onset hypertension in humans. We used CRISPR/Cas9 to generate Cacna1hM1560V/+ knockin mice as a model of the most common FH-IV mutation, along with corresponding knockout mice (Cacna1h-/-). Adrenal morphology of both Cacna1hM1560V/+ and Cacna1h-/- mice was normal. Cacna1hM1560V/+ mice had elevated aldosterone:renin ratios (a screening parameter for primary aldosteronism). Their adrenal Cyp11b2 (aldosterone synthase) expression was increased and remained elevated on a high-salt diet (relative autonomy, characteristic of primary aldosteronism), but plasma aldosterone was only elevated in male animals. The systolic blood pressure of Cacna1hM1560V/+ mice was 8 mmHg higher than in wild-type littermates and remained elevated on a high-salt diet. Cacna1h-/- mice had elevated renal Ren1 (renin-1) expression but normal adrenal Cyp11b2 levels, suggesting that in the absence of CaV3.2, stimulation of the renin-angiotensin system activates alternative calcium entry pathways to maintain normal aldosterone production. On a cellular level, Cacna1hM1560V/+ adrenal slices showed increased baseline and peak intracellular calcium concentrations in the zona glomerulosa compared to controls, but the frequency of calcium spikes did not rise. We conclude that FH-IV, on a molecular level, is caused by elevated intracellular Ca2+ concentrations as a signal for aldosterone production in adrenal glomerulosa cells. We demonstrate that a germline Cacna1h gain-of-function mutation is sufficient to cause mild primary aldosteronism, whereas loss of CaV3.2 channel function can be compensated for in a chronic setting. [ABSTRACT FROM AUTHOR]

Rats harboring the human renin and angiotensinogen genes (dTGR) feature angiotensin (ANG) II/hypertension-induced cardiac damage and die suddenly between wk 7 and 8. We observed by electrocardiogram (ECG) telemetry that ventricular tachycardia (VT) is a common terminal event in these animals. Our aim was to investigate electrical remodeling. We used ECG telemetry, noninvasive cardiac magnetic field mapping (CMFM) at wk 5 and 7, and performed in vivo programmed electrical stimulation at wk 7. We also investigated whether or not losartan (Los; 30 mg·kg-1·day-1) would prevent electrical remodeling. Cardiac hypertrophy and systolic blood pressure progressively increased in dTGR compared with Sprague-Dawley (SD) controls. Already by wk 5, untreated dTGR showed increased perivascular and interstitial fibrosis, connective tissue growth factor expression, and monocyte infiltration compared with SD rats, differences that progressed through time. Left-ventricular mRNA expression of potassium channel subunit Kv4.3 and gap-junction protein connexin 43 were significantly reduced in dTGR compared with Los-treated dTGR and SD. CMFM showed that depolarization and repolarization were prolonged and inhomogeneous. Los ameliorated all disturbances. VT could be induced in 88% of dTGR but only in 33% of Los-treated dTGR and could not be induced in SD. Untreated dTGR show electrical remodeling and probably die from VT. Los treatment reduces myocardial remodeling and predisposition to arrhythmias. ANG II target organ damage induces VT. [ABSTRACT FROM AUTHOR]

Competing Interests: Disclosures None.

Aims: Cardiac energy metabolism is centrally involved in heart failure (HF), although the direction of the metabolic alterations is complex and likely dependent on the particular stage of HF progression. Vascular endothelial growth factor B (VEGF-B) has been shown to modulate metabolic processes and to induce physiological cardiac hypertrophy; thus, it could be cardioprotective in the failing myocardium. This study investigates the role of VEGF-B in cardiac proteomic and metabolic adaptation in HF during aldosterone and high-salt hypertensive challenges., Methods and Results: Male rats overexpressing the cardiac-specific VEGF-B transgene (VEGF-B TG) were treated for 3 or 6 weeks with deoxycorticosterone-acetate combined with a high-salt (HS) diet (DOCA + HS) to induce hypertension and cardiac damage. Extensive longitudinal echocardiographic studies of HF progression were conducted, starting at baseline. Sham-treated rats served as controls. To evaluate the metabolic alterations associated with HF, cardiac proteomics by mass spectrometry was performed. Hypertrophic non-treated VEGF-B TG hearts demonstrated high oxygen and adenosine triphosphate (ATP) demand with early onset of diastolic dysfunction. Administration of DOCA + HS to VEGF-B TG rats for 6 weeks amplified the progression from cardiac hypertrophy to HF, with a drastic drop in heart ATP concentration. Dobutamine stress echocardiographic analyses uncovered a significantly impaired systolic reserve. Mechanistically, the hallmark of the failing TG heart was an abnormal energy metabolism with decreased mitochondrial ATP, preceding the attenuated cardiac performance and leading to systolic HF., Conclusions: This study shows that the VEGF-B TG accelerates metabolic maladaptation which precedes structural cardiomyopathy in experimental hypertension and ultimately leads to systolic HF., Competing Interests: Conflict of interest: None declared., (© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Gain-of-function mutations in the CACNA1H gene (encoding the T-type calcium channel Ca V 3.2) cause autosomal-dominant familial hyperaldosteronism type IV (FH-IV) and early-onset hypertension in humans. We used CRISPR/Cas9 to generate Cacna1h M1560V/+ knockin mice as a model of the most common FH-IV mutation, along with corresponding knockout mice ( Cacna1h -/- ). Adrenal morphology of both Cacna1h M1560V/+ and Cacna1h -/- mice was normal. Cacna1h M1560V/+ mice had elevated aldosterone:renin ratios (a screening parameter for primary aldosteronism). Their adrenal Cyp11b2 (aldosterone synthase) expression was increased and remained elevated on a high-salt diet (relative autonomy, characteristic of primary aldosteronism), but plasma aldosterone was only elevated in male animals. The systolic blood pressure of Cacna1h M1560V/+ mice was 8 mmHg higher than in wild-type littermates and remained elevated on a high-salt diet. Cacna1h -/- mice had elevated renal Ren1 (renin-1) expression but normal adrenal Cyp11b2 levels, suggesting that in the absence of Ca V 3.2, stimulation of the renin-angiotensin system activates alternative calcium entry pathways to maintain normal aldosterone production. On a cellular level, Cacna1h M1560V/+ adrenal slices showed increased baseline and peak intracellular calcium concentrations in the zona glomerulosa compared to controls, but the frequency of calcium spikes did not rise. We conclude that FH-IV, on a molecular level, is caused by elevated intracellular Ca 2+ concentrations as a signal for aldosterone production in adrenal glomerulosa cells. We demonstrate that a germline Cacna1h gain-of-function mutation is sufficient to cause mild primary aldosteronism, whereas loss of Ca V 3.2 channel function can be compensated for in a chronic setting., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)