Role of Dickkopf-3 in Blood Pressure Regulation in Mice and Hypertensive Rats.

BACKGROUND: Dkk3 (Dickkopf-3) is a secreted glycoprotein known for its proapoptotic and angiogenic activity. The role of Dkk3 in cardiovascular homeostasis is largely unknown. Remarkably, the Dkk3 gene maps within a chromosome segment linked to the hypertensive phenotype in spontaneously hypertensive rats (SHR). METHODS: We used Dkk3-/- mice or stroke-resistant (sr) and stroke-prone (sp) SHR to examine the role of Dkk3 in the central and peripheral regulation of blood pressure (BP). We used lentiviral expression vector to rescue Dkk3 in knockout mice or to induce Dkk3 overexpression or silencing in SHR. RESULTS: Genetic deletion of Dkk3 in mice enhanced BP and impaired endothelium-dependent acetylcholine-induced relaxation of resistance arteries. These alterations were rescued by restoring Dkk3 expression either in the periphery or in the central nervous system (CNS). Dkk3 was required for the constitutive expression of VEGF (vascular endothelium growth factor), and the action of Dkk3 on BP and endothelium-dependent vasorelaxation was mediated by VEGF-stimulated phosphatidylinositol-3-kinase pathway, leading to eNOS (endothelial NO synthase) activation both in resistance arteries and the CNS. The regulatory function of Dkk3 on BP was confirmed in SHR stroke-resistant and SHR stroke-prone in which was blunted in both resistance arteries and brainstem. In SHR stroke-resistant, lentiviral expression vector-induced Dkk3 expression in the CNS largely reduced BP, whereas Dkk3 knock-down further enhanced BP. In SHR stroke-prone challenged with a hypersodic diet, lentiviral expression vector-induced Dkk3 expression in the CNS displayed a substantial antihypertensive effect and delayed the occurrence of stroke. CONCLUSIONS: These findings demonstrate that Dkk3 acts as peripheral and central regulator of BP by promoting VEGF expression and activating a VEGF/Akt (protein kinase B)/eNOS hypotensive axis.

1,5-disubstituted-1,2,3-triazoles counteract mitochondrial dysfunction acting on F1FO-ATPase in models of cardiovascular diseases.

The compromised viability and function of cardiovascular cells are rescued by small molecules of triazole derivatives (Tzs), identified as 3a and 3b, by preventing mitochondrial dysfunction. The oxidative phosphorylation improves the respiratory control rate in the presence of Tzs independently of the substrates that energize the mitochondria. The F1FO-ATPase, the main candidate in mitochondrial permeability transition pore (mPTP) formation, is the biological target of Tzs and hydrophilic F1 domain of the enzyme is depicted as the binding region of Tzs. The protective effect of Tz molecules on isolated mitochondria was corroborated by immortalized cardiomyocytes results. Indeed, mPTP opening was attenuated in response to ionomycin. Consequently, increased mitochondrial roundness and reduction of both length and interconnections between mitochondria. In in-vitro and ex-vivo models of cardiovascular pathologies (i.e., hypoxia-reoxygenation and hypertension) were used to evaluate the Tzs cardioprotective action. Key parameters of porcine aortic endothelial cells (pAECs) oxidative metabolism and cell viability were not affected by Tzs. However, in the presence of either 1 µM 3a or 0.5 µM 3b the impaired cell metabolism of pAECs injured by hypoxia-reoxygenation was restored to control respiratory profile. Moreover, endothelial cells isolated from SHRSP exposed to high-salt treatment rescued the Complex I activity and the endothelial capability to form vessel-like tubes and vascular function in presence of Tzs. As a result, the specific biochemical mechanism of Tzs to block Ca2+-activated F1FO-ATPase protected cell viability and preserved the pAECs bioenergetic metabolism upon hypoxia-reoxygenation injury. Moreover, SHRSP improved vascular dysfunction in response to a high-salt treatment.

Impact of a NDUFC2 Variant on the Occurrence of Acute Coronary Syndromes.

Background: Among several potential mechanisms, mitochondrial dysfunction has been proposed to be involved in the pathogenesis of coronary artery disease (CAD). A mitochondrial complex I deficiency severely impairs cardiovascular health and contributes to CAD development. Previous evidence highlighted a key role of NDUFC2, a subunit of complex I, deficiency in the increased occurrence of renal and cerebrovascular damage in an animal model of hypertension, and of juvenile ischemic stroke occurrence in humans. Furthermore, a significant decrease of NDUFC2 mRNA was detected in peripheral blood mononuclear cells from patients experiencing acute coronary syndrome (ACS). The T allele at NDUFC2/rs23117379 variant is known to associate with reduced gene expression and mitochondrial dysfunction. Objective: In the present study we tested the impact of the T/C NDUFC2/rs23117379 variant on occurrence of ACS in a prospective cohort of CAD patients (n = 260). Results: Hypertension, smoking habit, diabetes and hypercholesterolemia were present in a large proportion of patients. Non-ST-elevation myocardial infarction (NSTEMI) represented the most frequent type of ACS (44%, n = 115), followed by ST-elevation myocardial infarction (STEMI) (34%, n = 88) and unstable angina (22%, n = 57). The alleles/genotypes distribution for T/C at NDUFC2/rs23117379 revealed that the TT genotype was associated with a trend toward the development of ACS at an earlier age (TT 61 ± 12, CT 65 ± 12 and CC 66 ± 11 years; p = 0.051 after adjustment for gender, hypertension, smoking habit, diabetes and hypercholesterolemia) and with a significant predictive role for ACS recurrence (hazard ratio [HR]1.671; 95% confidence interval [CI], 1.138-2.472; p = 0.009). Conclusions: Our findings are consistent with a deleterious effect of NDUFC2 deficiency on acute coronary events predisposition and further support a role of the NDUFC2/rs23117379 variant as a genetic cardiovascular risk factor.

Role of Uncoupling Protein 2 Gene Polymorphisms on the Risk of Ischemic Stroke in a Sardinian Population.

The mitochondrial uncoupling protein 2 (UCP2) acts as an anion transporter and as an antioxidant factor able to reduce the reactive oxygen species level. Based on its effects, UCP2 prevents the membrane lipids, proteins, and DNA damage while preserving normal cellular functions. Many variants have been identified within the human UCP2. Some of them were associated with a higher risk of obesity, diabetes and cardiovascular diseases in different populations. UCP2 appears a suitable candidate also for the risk of ischemic stroke. In the current study, we investigated the possible association between few variants of UCP2 (rs659366, rs660339, rs1554995310) and the risk of ischemic stroke in a genetically homogenous cohort of cases and controls selected in Sardinia Island. This population has been previously analysed for other candidate genes. A total of 250 cases of ischemic stroke and 241 controls were enrolled in the study. The allelic/genotypic distribution of the 3 UCP2 variants was characterized and compared among cases and controls. The results of our study confirmed known risk factors for ischemic stroke: age, history of smoking, hypertension, hypercholesterolemia, and atrial fibrillation. No association was found between the 3 UCP2 variants and the risk of ischemic stroke in our Sardinian cohort.

Relevance of stromal interaction molecule 1 (STIM1) in experimental and human stroke.

Stroke represents a main cause of death and permanent disability worldwide. In the attempt to develop targeted preventive and therapeutic strategies, several efforts were performed over the last decades to identify the specific molecular abnormalities preceding cerebral ischemia and neuronal death. In this regard, mitochondrial dysfunction, autophagy, and intracellular calcium homeostasis appear important contributors to stroke development, as underscored by recent pre-clinical evidence. Intracellular calcium (Ca2+) homeostasis is regulated, among other mechanisms, by the calcium sensor stromal interaction molecule 1 (STIM1) and calcium release-activated calcium modulator (ORAI) members, which mediate the store-operated Ca2+ entry (SOCE). The activity of SOCE is deregulated in animal models of ischemic stroke, leading to ischemic injury exacerbation. We found a different pattern of expression of few SOCE components, dependent from a STIM1 mutation, in cerebral endothelial cells isolated from the stroke-prone spontaneously hypertensive rat (SHRSP), compared to the stroke-resistant (SHRSR) strain, suggesting a potential involvement of this mechanism into the stroke predisposition of SHRSP. In this article, we discuss the relevant role of STIM1 in experimental stroke, as highlighted by the current literature and by our recent experimental findings, and the available evidence in the human disease. We also provide a glance on future perspectives and clinical implications of STIM1.

An interplay between UCP2 and ROS protects cells from high-salt-induced injury through autophagy stimulation.

The mitochondrial uncoupling protein 2 (UCP2) plays a protective function in the vascular disease of both animal models and humans. UCP2 downregulation upon high-salt feeding favors vascular dysfunction in knock-out mice, and accelerates cerebrovascular and renal damage in the stroke-prone spontaneously hypertensive rat. Overexpression of UCP2 counteracts the negative effects of high-salt feeding in both animal models. We tested in vitro the ability of UCP2 to stimulate autophagy and mitophagy as a mechanism mediating its protective effects upon high-salt exposure in endothelial and renal tubular cells. UCP2 silencing reduced autophagy and mitophagy, whereas the opposite was true upon UCP2 overexpression. High-salt exposure increased level of reactive oxygen species (ROS), UCP2, autophagy and autophagic flux in both endothelial and renal tubular cells. In contrast, high-salt was unable to induce autophagy and autophagic flux in UCP2-silenced cells, concomitantly with excessive ROS accumulation. The addition of an autophagy inducer, Tat-Beclin 1, rescued the viability of UCP2-silenced cells even when exposed to high-salt. In summary, UCP2 mediated the interaction between high-salt-induced oxidative stress and autophagy to preserve viability of both endothelial and renal tubular cells. In the presence of excessive ROS accumulation (achieved upon UCP2 silencing and high-salt exposure of silenced cells) autophagy was turned off. In this condition, an exogenous autophagy inducer rescued the cellular damage induced by excess ROS level. Our data confirm the protective role of UCP2 toward high-salt-induced vascular and renal injury, and they underscore the role of autophagy/mitophagy as a mechanism counteracting the high-salt-induced oxidative stress damage.

Trehalose, a natural disaccharide, reduces stroke occurrence in the stroke-prone spontaneously hypertensive rat.

Cerebrovascular disease, a frequent complication of hypertension, is a major public health issue for which novel therapeutic and preventive approaches are needed. Autophagy activation is emerging as a potential therapeutic and preventive strategy toward stroke. Among usual activators of autophagy, the natural disaccharide trehalose (TRE) has been reported to be beneficial in preclinical models of neurodegenerative diseases, atherosclerosis and myocardial infarction. In this study, we tested for the first time the effects of TRE in the stroke-prone spontaneously hypertensive rat (SHRSP) fed with a high-salt stroke permissive diet (JD). We found that TRE reduced stroke occurrence and renal damage in high salt-fed SHRSP. TRE was also able to decrease systolic blood pressure. Through ex-vivo studies, we assessed the beneficial effect of TRE on the vascular function of high salt-fed SHRSP. At the molecular level, TRE restored brain autophagy and reduced mitochondrial mass, along with the improvement of mitochondrial function. The beneficial effects of TRE were associated with increased nuclear translocation of TFEB, a transcriptional activator of autophagy. Our results suggest that TRE may be considered as a natural compound efficacious for the prevention of hypertension-related target organ damage, with particular regard to stroke and renal damage.

Inhibition of miR-155 Attenuates Detrimental Vascular Effects of Tobacco Cigarette Smoking.

Background The role of microRNAs dysregulation in tobacco cigarette smoking-induced vascular damage still needs to be clarified. We assessed the acute effects of tobacco cigarette smoking on endothelial cell-related circulating microRNAs in healthy subjects. In addition, we investigated the potential role of microRNAs in smoking-dependent endothelial cell damage. Methods and Results A panel of endothelial-related microRNAs was quantified in healthy subjects before and after smoking 1 tobacco cigarette. Serum levels of miR-155 were found to be significantly increased shortly after smoking. We also observed a progressive and significant miR-155 accumulation in culture media of human endothelial cells after 30 minutes and up to 4 hours of cigarette smoke condensate treatment in vitro without evidence of cell death, indicating that miR-155 can be released by endothelial cells in response to smoking stress. Cigarette smoke condensate appeared to enhance oxidative stress and impair cell survival, angiogenesis, and NO metabolism in human endothelial cells. Notably, these effects were abrogated by miR-155 inhibition. We also observed that miR-155 inhibition rescued the deleterious effects of cigarette smoke condensate on endothelial-mediated vascular relaxation and oxidative stress in isolated mouse mesenteric arteries. Finally, we found that exogenous miR-155 overexpression mimics the effects of smoking stress by inducing the upregulation of inflammatory markers, impairing angiogenesis and reducing cell survival. These deleterious effects were associated with downregulation of vascular endothelial growth factor and endothelial NO synthetase. Conclusions Our results suggest that miR-155 dysregulation may contribute to the deleterious vascular effects of tobacco smoking.

Pathogenesis of Ischemic Stroke: Role of Epigenetic Mechanisms.

Epigenetics is the branch of molecular biology that studies modifications able to change gene expression without altering the DNA sequence. Epigenetic modulations include DNA methylation, histone modifications, and noncoding RNAs. These gene modifications are heritable and modifiable and can be triggered by lifestyle and nutritional factors. In recent years, epigenetic changes have been associated with the pathogenesis of several diseases such as diabetes, obesity, renal pathology, and different types of cancer. They have also been related with the pathogenesis of cardiovascular diseases including ischemic stroke. Importantly, since epigenetic modifications are reversible processes they could assist with the development of new therapeutic approaches for the treatment of human diseases. In the present review article, we aim to collect the most recent evidence concerning the impact of epigenetic modifications on the pathogenesis of ischemic stroke in both animal models and humans.

Effects of dual angiotensin type 1 receptor/neprilysin inhibition vs. angiotensin type 1 receptor inhibition on target organ injury in the stroke-prone spontaneously hypertensive rat.

OBJECTIVES: The combination of AT1 blocker/neutroendopeptidase neprilysin inhibition (ARNi) represents an interesting approach to reduce cardiovascular risk in hypertension. We assessed the efficacy of ARNi, compared with angiotensin II type 1 receptor blockade alone, on blood pressure (BP) and on protection from target organ damage development in the stroke-prone spontaneously hypertensive rat (SHRSP). METHODS: In high-salt fed SHRSP, we assessed plasma and tissue natriuretic peptides, urinary volume, BP and body weight over a short-term treatment (6 weeks) with either ARNi (sacubitril/valsartan 68 mg/kg per day) or valsartan (30 mg/kg per day), protection from stroke and renal damage (as documented by proteinuria) over 4 months of treatment with either sacubitril/valsartan or valsartan; the ability of either treatment to reduce progression of cerebrovascular and renal damage after 2 weeks of high-salt diet. RESULTS: Higher levels of plasma and tissue atrial natriuretic peptide, of urinary cyclic guanosine 3'5'monophosphate and urine volumes, along with lower BP levels, were found upon sacubitril/valsartan as compared with valsartan over the short-term treatment. Sacubitril/valsartan caused a significant reduction of both BP and proteinuria levels and complete prevention of stroke over the long-term treatment. Once organ damage was established, a significant delay of its progression was observed with sacubitril/valsartan. CONCLUSION: The dual angiotensin II type 1 receptor/neutroendopeptidase inhibition significantly increased atrial natriuretic peptide level and reduced BP. Complete prevention of stroke was achieved in this model. The ability of sacubitril/valsartan to reduce organ damage progression was superior to that of valsartan alone. ARNi may represent a highly effective therapeutic agent to protect from target organ damage development in hypertension.

In vitro characterization of mitochondrial function and structure in rat and human cells with a deficiency of the NADH: ubiquinone oxidoreductase Ndufc2 subunit.

Ndufc2, a subunit of the NADH: ubiquinone oxidoreductase, plays a key role in the assembly and activity of complex I within the mitochondrial OXPHOS chain. Its deficiency has been shown to be involved in diabetes, cancer and stroke. To improve our knowledge on the mechanisms underlying the increased disease risk due to Ndufc2 reduction, we performed the present in vitro study aimed at the fine characterization of the derangements in mitochondrial structure and function consequent to Ndufc2 deficiency. We found that both fibroblasts obtained from skin of heterozygous Ndufc2 knock-out rat model showed marked mitochondrial dysfunction and PBMC obtained from subjects homozygous for the TT genotype of the rs11237379/NDUFC2 variant, previously shown to associate with reduced gene expression, demonstrated increased generation of reactive oxygen species and mitochondrial damage. The latter was associated with increased oxidative stress and significant ultrastructural impairment of mitochondrial morphology with a loss of internal cristae. In both models the exposure to stress stimuli, such as high-NaCl concentration or LPS, exacerbated the mitochondrial damage and dysfunction. Resveratrol significantly counteracted the ROS generation. These findings provide additional insights on the role of an altered pattern of mitochondrial structure-function as a cause of human diseases. In particular, they contribute to underscore a potential genetic risk factor for cardiovascular diseases, including stroke.

Ndufc2 Gene Inhibition Is Associated With Mitochondrial Dysfunction and Increased Stroke Susceptibility in an Animal Model of Complex Human Disease.

BACKGROUND: The genetic basis of stroke susceptibility remains to be elucidated. STR1 quantitative trait locus (STR1/QTL) was identified on rat chromosome 1 of stroke-prone spontaneously hypertensive rat (SHRSP) upon Japanese-style stroke-permissive diet (JD), and it contributes to 20% of the stroke phenotype variance. METHODS AND RESULTS: Nine hundred eighty-six probe sets mapping on STR1 were selected from the Rat RAE230A array and screened through a microarray differential expression analysis in brains of SHRSP and stroke-resistant SHR (SHRSR) fed with either regular diet or JD. The gene encoding Ndufc2 (NADH dehydrogenase [ubiquinone] 1 subunit), mapping 8 Mb apart from STR1/QTL Lod score peak, was found significantly down-regulated under JD in SHRSP compared to SHRSR. Ndufc2 disruption altered complex I assembly and activity, reduced mitochondrial membrane potential and ATP levels, and increased reactive oxygen species production and inflammation both in vitro and in vivo. SHRSR carrying heterozygous Ndufc2 deletion showed renal abnormalities and stroke occurrence under JD, similarly to SHRSP. In humans, T allele variant at NDUFC2/rs11237379 was associated with significant reduction in gene expression and with increased occurrence of early-onset ischemic stroke by recessive mode of transmission (odds ratio [OR], 1.39; CI, 1.07-1.80; P=0.012). Subjects carrying TT/rs11237379 and A allele variant at NDUFC2/rs641836 had further increased risk of stroke (OR=1.56; CI, 1.14-2.13; P=0.006). CONCLUSIONS: A significant reduction of Ndufc2 expression causes complex I dysfunction and contributes to stroke susceptibility in SHRSP. Moreover, our current evidence may suggest that Ndufc2 can contribute to an increased occurrence of early-onset ischemic stroke in humans.

Protective effects of Brassica oleracea sprouts extract toward renal damage in high-salt-fed SHRSP: role of AMPK/PPARα/UCP2 axis.

OBJECTIVES: Renal damage precedes occurrence of stroke in high-sodium/low-potassium-fed stroke-prone spontaneously hypertensive rat (SHRSP). We previously reported a marked suppression of uncoupling protein-2 (UCP2) upon high-salt Japanese-style diet in SHRSP kidneys. Vegetable compounds are known to exert protective effects in cardiovascular diseases. We aimed at evaluating the impact of Brassica oleracea sprouts juice toward renal damage in Japanese diet-fed SHRSP and exploring the role of 5'-adenosine monophosphate-activated protein kinase (AMPK)/NAD-dependent deacetylase sirtuin-1 (SIRT1)/peroxisome proliferator-activated receptor-γ coactivator-1α (PGC1α)/peroxisome proliferator-activated receptor-α (PPARα)/UCP2 axis. METHODS: SHRSP received Japanese diet for 4 weeks. A group of SHRSP received Japanese diet and B. oleracea. A third group received Japanese diet, B. oleracea, and PPARα inhibitor (GW6471). A group of SHRSP fed with regular diet served as control. RESULTS: Japanese diet induced marked increases of oxidative stress, inflammation, and proteinuria, along with glomerular and tubular damage, as compared with regular diet. A significant suppression of AMPK/UCP2 pathway was observed. Despite Japanese diet feeding, concomitant administration of B. oleracea prevented oxidative stress accumulation, inflammation, renal damage, and proteinuria. All components of the UCP2 regulatory pathway were significantly increased by B. oleracea. Superoxide dismutase 2 and phosphoendothelial nitric oxide synthase were also stimulated. Addition of PPARα inhibitor to B. oleracea and Japanese diet significantly reduced the B. oleracea beneficial effects. SBP levels were comparable among the different groups of rats.In vitro, UCP2 inhibition by genipin offset the antioxidant effect of B. oleracea in renal mesangial and proximal tubular cells. CONCLUSION: B. oleracea administration prevented renal damage in salt-loaded SHRSP, independently from SBP, with parallel stimulation of AMPK/SIRT1/PGC1α/PPARα/UCP2 axis. Stimulation of the latter mechanism may provide relevant renal protective effect and play a therapeutic role in target organ damage progression in hypertension.

The C2238/αANP variant is a negative modulator of both viability and function of coronary artery smooth muscle cells.

BACKGROUND: Abnormalities of vascular smooth muscle cells (VSMCs) contribute to development of vascular disease. Atrial natriuretic peptide (ANP) exerts important effects on VSMCs. A common ANP molecular variant (T2238C/αANP) has recently emerged as a novel vascular risk factor. OBJECTIVES: We aimed at identifying effects of CC2238/αANP on viability, migration and motility in coronary artery SMCs, and the underlying signaling pathways. METHODS AND RESULTS: Cells were exposed to either TT2238/αANP or CC2238/αANP. At the end of treatment, cell viability, migration and motility were evaluated, along with changes in oxidative stress pathway (ROS levels, NADPH and eNOS expression), on Akt phosphorylation and miR21 expression levels. CC2238/αANP reduced cell vitality, increased apoptosis and necrosis, increased oxidative stress levels, suppressed miR21 expression along with consistent changes of its molecular targets (PDCD4, PTEN, Bcl2) and of phosphorylated Akt levels. As a result of increased oxidative stress, CC2238/αANP markedly stimulated cell migration and increased cell contraction. NPR-C gene silencing with specific siRNAs restored cell viability, miR21 expression, and reduced oxidative stress induced by CC2238/αANP. The cAMP/PKA/CREB pathway, driven by NPR-C activation, significantly contributed to both miR21 and phosphoAkt reduction upon CC2238/αANP. miR21 overexpression by mimic-hsa-miR21 rescued the cellular damage dependent on CC2238/αANP. CONCLUSIONS: CC2238/αANP negatively modulates viability through NPR-C/cAMP/PKA/CREB/miR21 signaling pathway, and it augments oxidative stress leading to increased migratory and vasoconstrictor effects in coronary artery SMCs. These novel findings further support a damaging role of this common αANP variant on vessel wall and its potential contribution to acute coronary events.

C2238 atrial natriuretic peptide molecular variant is associated with endothelial damage and dysfunction through natriuretic peptide receptor C signaling.

RATIONALE: C2238 atrial natriuretic peptide (ANP) minor allele (substitution of thymidine with cytosine in position 2238) associates with increased risk of cardiovascular events. OBJECTIVE: We investigated the mechanisms underlying the vascular effects of C2238-αANP. METHODS AND RESULTS: In vitro, human umbilical vein endothelial cell were exposed to either wild-type (T2238)- or mutant (C2238)-αANP. Cell survival and apoptosis were tested by Trypan blue, annexin V, and cleaved caspase-3 assays. C2238-αANP significantly reduced human umbilical vein endothelial cell survival and increased apoptosis. In addition, C2238-αANP reduced endothelial tube formation, as assessed by matrigel. C2238-αANP did not differentially modulate natriuretic peptide receptor (NPR)-A/B activity with respect to T2238-αANP, as evaluated by intracellular cGMP levels. In contrast, C2238-αANP, but not T2238-αANP, markedly reduced intracellular cAMP levels in an NPR-C-dependent manner. Accordingly, C2238-αANP showed higher affinity binding to NPR-C, than T2238-αANP. Either NPR-C inhibition by antisense oligonucleotide or NPR-C gene silencing by small interfering RNA rescued survival and tube formation of human umbilical vein endothelial cell exposed to C2238-αANP. Similar data were obtained in human aortic endothelial cell with NPR-C knockdown. NPR-C activation by C2238-αANP inhibited the protein kinase A/Akt1 pathway and increased reactive oxygen species. Adenovirus-mediated Akt1 reactivation rescued the detrimental effects of C2238-αANP. Overall, these data indicate that C2238-αANP affects endothelial cell integrity through NPR-C-dependent inhibition of the cAMP/protein kinase A/Akt1 pathway and increased reactive oxygen species production. Accordingly, C2238-αANP caused impairment of acetylcholine-dependent vasorelaxation ex vivo, which was rescued by NPR-C pharmacological inhibition. Finally, subjects carrying C2238 minor allele showed early endothelial dysfunction, which highlights the clinical relevance of our results. CONCLUSIONS: C2238-αANP reduces endothelial cell survival and impairs endothelial function through NPR-C signaling. NPR-C targeting represents a potential strategy to reduce cardiovascular risk in C2238 minor-allele carriers.