Background-Angiotensin-converting enzyme 2 (ACE2) is a pleiotropic monocarboxypeptidase capable of metabolizing several peptide substrates. We hypothesized that ACE2 is a negative regulator of angiotensin II (Ang II)-mediated signaling and its adverse effects on the cardiovascular system. Methods and Results-Ang II infusion (1.5 mg ⅐ kg Ϫ1 ⅐ d Ϫ1) for 14 days resulted in worsening cardiac fibrosis and pathological hypertrophy in ACE2 knockout (Ace2 Ϫ/y ) mice compared with wild-type (WT) mice. Daily treatment of Ang II-infused wild-type mice with recombinant human ACE2 (rhACE2; 2 mg ⅐ kg Ϫ1 ⅐ d Ϫ1 IP) blunted the hypertrophic response and expression of hypertrophy markers and reduced Ang II-induced superoxide production. Ang II-mediated myocardial fibrosis and expression of procollagen type I␣1, procollagen type III␣1, transforming growth factor-␤1, and fibronectin were also suppressed by rhACE2. Ang II-induced diastolic dysfunction was inhibited by rhACE2 in association with reduced plasma and myocardial Ang II and increased plasma Ang 1-7 levels. rhACE2 treatment inhibited Ang II-mediated activation of protein kinase C-␣ and protein kinase C-␤1 protein levels and phosphorylation of the extracellular signal-regulated 1/2, Janus kinase 2, and signal transducer and activator of transcription 3 signaling pathways in wild-type mice. A subpressor dose of Ang II (0.15 mg ⅐ kg) resulted in a milder phenotype that was strikingly attenuated by rhACE2 (2 mg ⅐ kgIn adult ventricular cardiomyocytes and cardiofibroblasts, Ang II-mediated superoxide generation, collagen production, and extracellular signal-regulated 1/2 signaling were inhibited by rhACE2 in an Ang 1-7-dependent manner. Importantly, rhACE2 partially prevented the development of dilated cardiomyopathy in pressure-overloaded wild-type mice. Conclusions-Elevated Ang II induced hypertension, myocardial hypertrophy, fibrosis, and diastolic dysfunction, which were exacerbated by ACE2 deficiency, whereas rhACE2 attenuated Ang II-and pressure-overload-induced adverse myocardial remodeling. Hence, ACE2 is an important negative regulator of Ang II-induced heart disease and suppresses adverse myocardial remodeling. (Circulation. 2010;122:717-728.)Key Words: angiotensin Ⅲ signal transduction Ⅲ hypertrophy Ⅲ remodeling Ⅲ diastole A ctivation of the renin-angiotensin system (RAS) and the subsequent generation of angiotensin (Ang) II are important mediators of myocardial fibrosis, pathological hypertrophy, and heart failure. 1-3 Pathological hypertrophy and increased myocardial interstitial fibrosis contribute to increased ventricular wall stiffness, thereby impairing cardiac diastolic function, and represent an important risk factor for heart failure in experimental models and patients. 4 -6 Drugs that target Ang II and the Ang II type 1 receptor (AT 1 ) are widely used for the treatment of cardiovascular diseases such as hypertension, myocardial infarction, and heart failure. 7 Angiotensin-converting enzyme 2 (ACE2) is a pleiotropic monocarboxypeptidase capable of metabo...

Background-Angiotensin-converting enzyme 2 (ACE2) is a monocarboxypeptidase that metabolizes Ang II into Ang 1-7, thereby functioning as a negative regulator of the renin-angiotensin system. We hypothesized that ACE2 deficiency may compromise the cardiac response to myocardial infarction (MI). Methods and Results-In response to MI (induced by left anterior descending artery ligation), there was a persistent increase in ACE2 protein in the infarct zone in wild-type mice, whereas loss of ACE2 enhanced the susceptibility to MI, with increased mortality, infarct expansion, and adverse ventricular remodeling characterized by ventricular dilation and systolic dysfunction. In ACE2-deficient hearts, elevated myocardial levels of Ang II and decreased levels of Ang 1-7 in the infarct-related zone was associated with increased production of reactive oxygen species. ACE2 deficiency leads to increased matrix metalloproteinase (MMP) 2 and MMP9 levels with MMP2 activation in the infarct and peri-infarct regions, as well as increased gelatinase activity leading to a disrupted extracellular matrix structure after MI. Loss of ACE2 also leads to increased neutrophilic infiltration in the infarct and peri-infarct regions, resulting in upregulation of inflammatory cytokines, interferon-␥, interleukin-6, and the chemokine, monocyte chemoattractant protein-1, as well as increased phosphorylation of ERK1/2 and JNK1/2 signaling pathways. Treatment of Ace2 Ϫ/y -MI mice with irbesartan, an AT1 receptor blocker, reduced nicotinamide-adenine dinucleotide phosphate oxidase activity, infarct size, MMP activation, and myocardial inflammation, ultimately resulting in improved post-MI ventricular function. Conclusions-We conclude that loss of ACE2 facilitates adverse post-MI ventricular remodeling by potentiation of Ang II effects by means of the AT1 receptors, and supplementing ACE2 can be a potential therapy for ischemic heart disease. (Circ Heart Fail. 2009;2:446-455.)

Angiotensin-converting enzyme 2 (ACE2) is a monocarboxypeptidase capable of metabolizing angiotensin (Ang) II into Ang 1 to 7. We hypothesized that ACE2 is a negative regulator of Ang II signaling and its adverse effects on the kidneys. Ang II infusion (1.5 mg/kg⁻¹/d⁻¹) for 4 days resulted in higher renal Ang II levels and increased nicotinamide adenine dinucleotide phosphate oxidase activity in ACE2 knockout (Ace2(-/y)) mice compared to wild-type mice. Expression of proinflammatory cytokines, interleukin-1β and chemokine (C-C motif) ligand 5, were increased in association with greater activation of extracellular-regulated kinase 1/2 and increase of protein kinase C-α levels. These changes were associated with increased expression of fibrosis-associated genes (α-smooth muscle actin, transforming growth factor-β, procollagen type Iα1) and increased protein levels of collagen I with histological evidence of increased tubulointerstitial fibrosis. Ang II-infused wild-type mice were then treated with recombinant human ACE2 (2 mg/kg⁻¹/d⁻¹, intraperitoneal). Daily treatment with recombinant human ACE2 reduced Ang II-induced pressor response and normalized renal Ang II levels and oxidative stress. These changes were associated with a suppression of Ang II-mediated activation of extracellular-regulated kinase 1/2 and protein kinase C pathway and Ang II-mediated renal fibrosis and T-lymphocyte-mediated inflammation. We conclude that loss of ACE2 enhances renal Ang II levels and Ang II-induced renal oxidative stress, resulting in greater renal injury, whereas recombinant human ACE2 prevents Ang II-induced hypertension, renal oxidative stress, and tubulointerstitial fibrosis. ACE2 is an important negative regulator of Ang II-induced renal disease and enhancing ACE2 action may have therapeutic potential for patients with kidney disease.

Rationale Diabetic cardiovascular complications are reaching epidemic proportions. Angiotensin-converting enzyme-2 (ACE2) is a negative regulator of the renin-angiotensin system. We hypothesize that loss of ACE2 exacerbates cardiovascular complications induced by diabetes. Objective To define the role of ACE2 in diabetic cardiovascular complications. Methods and Results We used the well-validated Akita mice, a model of human diabetes, and generated double-mutant mice using the ACE2 knockout (KO) mice (Akita/ACE2−/y). Diabetic state was associated with increased ACE2 in Akita mice, whereas additional loss of ACE2 in these mice leads to increased plasma and tissue angiotensin II levels, resulting in systolic dysfunction on a background of impaired diastolic function. Downregulation of SERCA2 and lipotoxicity were equivalent in Akita and Akita/ACE2KO hearts and are likely mediators of the diastolic dysfunction. However, greater activation of protein kinase C and loss of Akt and endothelial nitric oxide synthase phosphorylation occurred in the Akita/ACE2KO hearts. Systolic dysfunction in Akita/ACE2KO mice was linked to enhanced activation of NADPH oxidase and metalloproteinases, resulting in greater oxidative stress and degradation of the extracellular matrix. Impaired flow-mediated dilation in vivo correlated with increased vascular oxidative stress in Akita/ACE2KO mice. Treatment with the AT1 receptor blocker, irbesartan rescued the systolic dysfunction, normalized altered signaling pathways, flow-mediated dilation, and the increased oxidative stress in the cardiovascular system. Conclusions Loss of ACE2 disrupts the balance of the renin-angiotensin system in a diabetic state and leads to an angiotensin II/AT1 receptor-dependent systolic dysfunction and impaired vascular function. Our study demonstrates that ACE2 serves as a protective mechanism against diabetes-induced cardiovascular complications.

Background With the establishment of the heart-gut axis concept, accumulating studies suggest that the gut microbiome plays an important role in the pathogenesis of cardiovascular diseases. Yet, little evidence has been reported in characterizing the gut microbiota shift in atrial fibrillation. Methods We include the result of the global alterations that occur in the intestinal microbiota in a cohort of 50 patients with atrial fibrillation and 50 matched controls based on a strategy of metagenomic and metabolomic analyses. Results The alterations include a dramatic elevation in microbial diversity and a specific perturbation of gut microbiota composition. Overgrowth of Ruminococcus , Streptococcus , and Enterococcus , as well as reduction of Faecalibacterium , Alistipes , Oscillibacter , and Bilophila were detected in patients with atrial fibrillation. A gut microbial function imbalance and correlated metabolic pattern changes were observed with atrial fibrillation in both fecal and serum samples. The differential gut microbiome signatures could be used to identify patients with atrial fibrillation. Conclusions Our findings characterize the disordered gut microbiota and microbial metabolite profiles in atrial fibrillation. Further research could determine whether intervention strategies targeting intestinal microbiome composition might be useful to counteract the progression of atrial fibrillation.

Abstract-Loss of angiotensin (Ang)-converting enzyme 2 (ACE2) and inability to metabolize Ang II to Ang 1-7 perpetuate the actions of Ang II after biomechanical stress and exacerbate early adverse myocardial remodeling. Ang receptor blockers are known to antagonize the effect of Ang II by blocking Ang II type 1 receptor (AT 1 R) and also by upregulating the ACE2 expression. We directly compare the benefits of AT 1 R blockade versus enhancing Ang 1-7 action in pressure-overload-induced heart failure in ACE2 knockout mice. AT 1 R blockade and Ang 1-7 both resulted in marked recovery of systolic dysfunction in pressure-overloaded ACE2-null mice. Similarly, both therapies attenuated the increase in NADPH oxidase activation by downregulating the expression of Nox2 and p47 phox subunits and also by limiting the p47 phox phosphorylation. Biomechanical stress-induced increase in protein kinase C-␣ expression and phosphorylation of extracellular signal-regulated kinase 1/2, signal transducer and activator of transcription 3, Akt, and glycogen synthase kinase 3␤ were normalized by irbesartan and Ang 1-7. Ang receptor blocker and Ang 1-7 effectively reduced matrix metalloproteinase 2 activation and matrix metalloproteinase 9 levels. Ang II-mediated cellular effects in cultured adult cardiomyocytes and cardiofibrolasts isolated from pressure-overloaded ACE2-null hearts were inhibited to similar degree by AT 1 R blockade and stimulation with Ang 1-7. Thus, treatment with the AT 1 R blocker irbesartan and Ang 1-7 prevented the cardiac hypertrophy and improved cardiac remodeling in pressure-overloaded ACE2-null mice by suppressing NADPH oxidase and normalizing pathological signaling pathways. Key Words: renin-angiotensin system Ⅲ angiotensin 1-7 Ⅲ angiotensin-converting enzyme 2 Ⅲ NADPH oxidase Ⅲ AT 1 receptor Ⅲ heart failure Ⅲ signaling S everal lines of experimental and clinical evidence implicate a key role for the renin-angiotensin system in the pathophysiology of a number of cardiovascular diseases, such as myocardial infarction, hypertension, and heart failure. 1,2Angiotensin II (Ang II), acting via the Ang II type 1 receptor (AT 1 R) and Ang II type 2 receptor, modulates production of reactive oxygen species (ROS), impairing myocardial contractility and extracellular matrix remodeling, thereby negatively impacting on heart function.3 Angiotensin-converting enzyme 2 (ACE2), a homologue of angiotensin-converting enzyme, is a monocarboxypeptidase that metabolizes Ang II to yield angiotensin 1-7 (Ang 1-7) and lowers the Ang II/Ang 1-7 ratio.4-9 Ang II receptor blockers that selectively antagonize the AT 1 R became a valid alternative approach to interfere with the renin-angiotensin system axis and also upregulates ACE2, resulting in the generation of Ang 1-7. 10,11 Ang 1-7 acts on the Mas receptor and plays an important role in counteracting the actions of Ang II. 12-18Ang II-mediated oxidative stress, cardiac hypertrophy, contractile dysfunction, and fibrosis are exacerbated in ACE2-deficient mice, 5,7 whereas recombinant ...

Dysregulation of microRNAs has been implicated in many cardiovascular diseases including fibrosis. Here we report that miR-433 was consistently elevated in three models of heart disease with prominent cardiac fibrosis, and was enriched in fibroblasts compared to cardiomyocytes. Forced expression of miR-433 in neonatal rat cardiac fibroblasts increased proliferation and their differentiation into myofibroblasts as determined by EdU incorporation, α-SMA staining, and expression levels of fibrosis-associated genes. Conversely, inhibition of miR-433 exhibited opposite results. AZIN1 and JNK1 were identified as two target genes of miR-433. Decreased level of AZIN1 activated TGF-β1 while down-regulation of JNK1 resulted in activation of ERK and p38 kinase leading to Smad3 activation and ultimately cardiac fibrosis. Importantly, systemic neutralization of miR-433 or adeno-associated virus 9 (AAV9)-mediated cardiac transfer of a miR-433 sponge attenuated cardiac fibrosis and ventricular dysfunction following myocardial infarction. Thus, our work suggests that miR-433 is a potential target for amelioration of cardiac fibrosis.

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