RESUMEN
Changes in the renin-angiotensin system, known for its critical role in the regulation of blood pressure and sodium homeostasis, may contribute to aging and age-related diseases. While the renin-angiotensin system is suppressed during aging, little is known about its regulation and activity within tissues. However, this knowledge is required to successively treat or prevent renal disease in the elderly. Ercc1 is involved in important DNA repair pathways, and when mutated causes accelerated aging phenotypes in humans and mice. In this study, we hypothesized that unrepaired DNA damage contributes to accelerated kidney failure. We tested the use of the renin-activatable near-infrared fluorescent probe ReninSense680™ in progeroid Ercc1d/- mice and compared renin activity levels in vivo to wild-type mice. First, we validated the specificity of the probe by detecting increased intrarenal activity after losartan treatment and the virtual absence of fluorescence in renin knock-out mice. Second, age-related kidney pathology, tubular anisokaryosis, glomerulosclerosis and increased apoptosis were confirmed in the kidneys of 24-week-old Ercc1d/- mice, while initial renal development was normal. Next, we examined the in vivo renin activity in these Ercc1d/- mice. Interestingly, increased intrarenal renin activity was detected by ReninSense in Ercc1d/- compared to WT mice, while their plasma renin concentrations were lower. Hence, this study demonstrates that intrarenal RAS activity does not necessarily run in parallel with circulating renin in the aging mouse. In addition, our study supports the use of this probe for longitudinal imaging of altered RAS signaling in aging.
Asunto(s)
Envejecimiento/genética , Angiotensina II/genética , Proteínas de Unión al ADN/genética , Endonucleasas/genética , Glomeruloesclerosis Focal y Segmentaria/genética , Progeria/genética , Insuficiencia Renal Crónica/genética , Renina/genética , Envejecimiento/metabolismo , Envejecimiento/patología , Angiotensina II/metabolismo , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Animales , Daño del ADN , Reparación del ADN , Proteínas de Unión al ADN/deficiencia , Modelos Animales de Enfermedad , Endonucleasas/deficiencia , Femenino , Regulación de la Expresión Génica , Tasa de Filtración Glomerular , Glomeruloesclerosis Focal y Segmentaria/metabolismo , Glomeruloesclerosis Focal y Segmentaria/patología , Humanos , Riñón/metabolismo , Riñón/patología , Losartán/farmacología , Masculino , Ratones , Ratones Noqueados , Progeria/metabolismo , Progeria/patología , Insuficiencia Renal Crónica/metabolismo , Insuficiencia Renal Crónica/patología , Renina/metabolismo , Sistema Renina-Angiotensina/genética , Transducción de SeñalRESUMEN
DNA damage is an important contributor to endothelial dysfunction and age-related vascular disease. Recently, we demonstrated in a DNA repair-deficient, prematurely aging mouse model (Ercc1Δ/- mice) that dietary restriction (DR) strongly increases life- and health span, including ameliorating endothelial dysfunction, by preserving genomic integrity. In this mouse mutant displaying prominent accelerated, age-dependent endothelial dysfunction we investigated the signaling pathways involved in improved endothelium-mediated vasodilation by DR, and explore the potential role of the renin-angiotensin system (RAS). Ercc1Δ/- mice showed increased blood pressure and decreased aortic relaxations to acetylcholine (ACh) in organ bath experiments. Nitric oxide (NO) signaling and phospho-Ser1177-eNOS were compromised in Ercc1Δ/- DR improved relaxations by increasing prostaglandin-mediated responses. Increase of cyclo-oxygenase 2 and decrease of phosphodiesterase 4B were identified as potential mechanisms. DR also prevented loss of NO signaling in vascular smooth muscle cells and normalized angiotensin II (Ang II) vasoconstrictions, which were increased in Ercc1Δ/- mice. Ercc1Δ/- mutants showed a loss of Ang II type 2 receptor-mediated counter-regulation of Ang II type 1 receptor-induced vasoconstrictions. Chronic losartan treatment effectively decreased blood pressure, but did not improve endothelium-dependent relaxations. This result might relate to the aging-associated loss of treatment efficacy of RAS blockade with respect to endothelial function improvement. In summary, DR effectively prevents endothelium-dependent vasodilator dysfunction by augmenting prostaglandin-mediated responses, whereas chronic Ang II type 1 receptor blockade is ineffective.
Asunto(s)
Envejecimiento/metabolismo , Daño del ADN , Receptor de Angiotensina Tipo 1/metabolismo , Enfermedades Vasculares/dietoterapia , Envejecimiento/genética , Angiotensina II/metabolismo , Animales , Presión Sanguínea , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Dieta , Endonucleasas/genética , Endonucleasas/metabolismo , Femenino , Humanos , Masculino , Ratones , Ratones Noqueados , Óxido Nítrico/metabolismo , Receptor de Angiotensina Tipo 1/genética , Enfermedades Vasculares/genética , Enfermedades Vasculares/metabolismo , Enfermedades Vasculares/fisiopatología , VasodilataciónRESUMEN
The need for noninvasive imaging to distinguish stable from vulnerable atherosclerotic plaques is evident. Activated macrophages play a role in atherosclerosis and express folate receptor folate receptor ß (FR-ß). The feasibility of folate targeting to detect atherosclerosis was demonstrated in human and mouse plaques, and it was suggested that molecular imaging of FR-ß through folate conjugates might be a specific marker for plaque vulnerability. However, these studies did not allow differentiation between stable and vulnerable atherosclerotic plaques. We investigated the feasibility of a folate-based radiopharmaceutical (111)In-EC0800) with high-resolution animal single-photon emission computed tomography/computed tomography (SPECT/CT) to differentiate between stable and vulnerable atherosclerotic plaques in apolipoprotein E(−/−) mice in which we can induce plaques with the characteristics of stable and vulnerable plaques by placing a flow-modifying cast around the common carotid artery. Both plaques showed (111)In-EC0800 uptake, with higher uptake in the vulnerable plaque. However, the vulnerable plaque was larger than the stable plaque. Therefore, we determined tracer uptake per plaque volume and demonstrated higher accumulation of (111)In-EC0800 in the stable plaque normalized to plaque volume. Our data show that (111)In-EC0800 is not a clear-cut marker for the detection of vulnerable plaques but detects both stable and vulnerable atherosclerotic plaques in a mouse model of atherosclerosis.
Asunto(s)
Apolipoproteínas E/genética , Aterosclerosis/diagnóstico por imagen , Complejos de Coordinación , Receptor 2 de Folato/metabolismo , Ácido Fólico/análogos & derivados , Activación de Macrófagos/efectos de la radiación , Macrófagos/metabolismo , Placa Aterosclerótica/diagnóstico por imagen , Radiofármacos , Tomografía Computarizada de Emisión de Fotón Único/métodos , Animales , Aterosclerosis/patología , Modelos Animales de Enfermedad , Femenino , Humanos , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Cardiovascular diseases are the number one cause of death globally. The most important determinant of cardiovascular health is a person's age. Aging results in structural changes and functional decline of the cardiovascular system. DNA damage is an important contributor to the aging process, and mice with a DNA repair defect caused by Ercc1 deficiency display hypertension, vascular stiffening, and loss of vasomotor control. To determine the underlying cause, we compared important hallmarks of vascular aging in aortas of both Ercc1Δ/- and age-matched wildtype mice. Additionally, we investigated vascular aging in 104 week old wildtype mice. Ercc1Δ/- aortas displayed arterial thickening, a loss of cells, and a discontinuous endothelial layer. Aortas of 24 week old Ercc1Δ/- mice showed phenotypical switching of vascular smooth muscle cells (VSMCs), characterized by a decrease in contractile markers and a decrease in synthetic markers at the RNA level. As well as an increase in osteogenic markers, microcalcification, and an increase in markers for damage induced stress response. This suggests that Ercc1Δ/- VSMCs undergo a stress-induced contractile-to-osteogenic phenotype switch. Ercc1Δ/- aortas showed increased MMP activity, elastin fragmentation, and proteoglycan deposition, characteristic of vascular aging and indicative of age-related extracellular matrix remodeling. The 104 week old WT mice showed loss of cells, VSMC dedifferentiation, and senescence. In conclusion, Ercc1Δ/- aortas rapidly display many characteristics of vascular aging, and thus the Ercc1Δ/- mouse is an excellent model to evaluate drugs that prevent vascular aging in a short time span at the functional, histological, and cellular level.
Asunto(s)
Envejecimiento , Reparación del ADN , Endonucleasas , Matriz Extracelular , Músculo Liso Vascular , Fenotipo , Animales , Ratones , Envejecimiento/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/deficiencia , Endonucleasas/metabolismo , Endonucleasas/deficiencia , Endonucleasas/genética , Matriz Extracelular/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/metabolismoRESUMEN
PURPOSE: In this study, we explored the role of apoptosis as a potential biomarker for cardiac failure using functional micro-CT and fluorescence molecular tomography (FMT) imaging techniques in Ercc1 mutant mice. Ercc1 is involved in multiple DNA repair pathways, and its mutations contribute to accelerated aging phenotypes in both humans and mice, due to the accumulation of DNA lesions that impair vital DNA functions. We previously found that systemic mutations and cardiomyocyte-restricted deletion of Ercc1 in mice results in left ventricular (LV) dysfunction at older age. PROCEDURES AND RESULTS: Here we report that combined functional micro-CT and FMT imaging allowed us to detect apoptosis in systemic Ercc1 mutant mice prior to the development of overt LV dysfunction, suggesting its potential as an early indicator and contributing factor of cardiac impairment. The detection of apoptosis in vivo was feasible as early as 12 weeks of age, even when global LV function appeared normal, underscoring the potential of apoptosis as an early predictor of LV dysfunction, which subsequently manifested at 24 weeks. CONCLUSIONS: This study highlights the utility of combined functional micro-CT and FMT imaging in assessing cardiac function and detecting apoptosis, providing valuable insights into the potential of apoptosis as an early biomarker for cardiac failure.
Asunto(s)
Apoptosis , Proteínas de Unión al ADN , Endonucleasas , Insuficiencia Cardíaca , Miocardio , Microtomografía por Rayos X , Animales , Insuficiencia Cardíaca/diagnóstico por imagen , Insuficiencia Cardíaca/patología , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genética , Endonucleasas/metabolismo , Endonucleasas/genética , Ratones , Miocardio/patologíaRESUMEN
Heart failure has reached epidemic proportions in a progressively ageing population. The molecular mechanisms underlying heart failure remain elusive, but evidence indicates that DNA damage is enhanced in failing hearts. Here, we tested the hypothesis that endogenous DNA repair in cardiomyocytes is critical for maintaining normal cardiac function, so that perturbed repair of spontaneous DNA damage drives early onset of heart failure. To increase the burden of spontaneous DNA damage, we knocked out the DNA repair endonucleases xeroderma pigmentosum complementation group G (XPG) and excision repair cross-complementation group 1 (ERCC1), either systemically or cardiomyocyte-restricted, and studied the effects on cardiac function and structure. Loss of DNA repair permitted normal heart development but subsequently caused progressive deterioration of cardiac function, resulting in overt congestive heart failure and premature death within 6 months. Cardiac biopsies revealed increased oxidative stress associated with increased fibrosis and apoptosis. Moreover, gene set enrichment analysis showed enrichment of pathways associated with impaired DNA repair and apoptosis, and identified TP53 as one of the top active upstream transcription regulators. In support of the observed cardiac phenotype in mutant mice, several genetic variants in the ERCC1 and XPG gene in human GWAS data were found to be associated with cardiac remodelling and dysfunction. In conclusion, unrepaired spontaneous DNA damage in differentiated cardiomyocytes drives early onset of cardiac failure. These observations implicate DNA damage as a potential novel therapeutic target and highlight systemic and cardiomyocyte-restricted DNA repair-deficient mouse mutants as bona fide models of heart failure.
Asunto(s)
Proteínas de Unión al ADN , Insuficiencia Cardíaca , Ratones , Animales , Humanos , Proteínas de Unión al ADN/metabolismo , Miocitos Cardíacos/metabolismo , Reparación del ADN/genética , Daño del ADN/genética , Insuficiencia Cardíaca/genética , EndonucleasasRESUMEN
Because of the presence of the blood-brain barrier, brain renin-angiotensin system activity should depend on local (pro)renin synthesis. Indeed, an intracellular form of renin has been described in the brain, but whether it displays angiotensin (Ang) I-generating activity (AGA) is unknown. Here, we quantified brain (pro)renin, before and after buffer perfusion of the brain, in wild-type mice, renin knockout mice, deoxycorticosterone acetate salt-treated mice, and Ang II-infused mice. Brain regions were homogenized and incubated with excess angiotensinogen to detect AGA, before and after prorenin activation, using a renin inhibitor to correct for nonrenin-mediated AGA. Renin-dependent AGA was readily detectable in brain regions, the highest AGA being present in brain stem (>thalamus=cerebellum=striatum=midbrain>hippocampus=cortex). Brain AGA increased marginally after prorenin activation, suggesting that brain prorenin is low. Buffer perfusion reduced AGA in all brain areas by >60%. Plasma renin (per mL) was 40× to 800× higher than brain renin (per gram). Renin was undetectable in plasma and brain of renin knockout mice. Deoxycorticosterone acetate salt and Ang II suppressed plasma renin and brain renin in parallel, without upregulating brain prorenin. Finally, Ang I was undetectable in brains of spontaneously hypertensive rats, while their brain/plasma Ang II concentration ratio decreased by 80% after Ang II type 1 receptor blockade. In conclusion, brain renin levels (per gram) correspond with the amount of renin present in 1 to 20 µL of plasma. Brain renin disappears after buffer perfusion and varies in association with plasma renin. This indicates that brain renin represents trapped plasma renin. Brain Ang II represents Ang II taken up from blood rather than locally synthesized Ang II.
Asunto(s)
Amidas/farmacología , Angiotensina II/farmacología , Barrera Hematoencefálica/efectos de los fármacos , Encéfalo/metabolismo , Fumaratos/farmacología , Hipertensión/tratamiento farmacológico , Sistema Renina-Angiotensina/efectos de los fármacos , Angiotensinógeno/metabolismo , Animales , Presión Sanguínea/efectos de los fármacos , Barrera Hematoencefálica/metabolismo , Encéfalo/efectos de los fármacos , Acetato de Desoxicorticosterona/farmacología , Modelos Animales de Enfermedad , Ratones , Ratones Noqueados , Distribución Aleatoria , Ratas , Ratas Endogámicas SHR , Valores de ReferenciaRESUMEN
BACKGROUND: Renin inhibition with aliskiren induced the largest increases in renal plasma flow (RPF) in salt-depleted healthy volunteers of all renin-angiotensin system (RAS) blockers. However, given its side-effects at doses higher than 300âmg, no maximum effect of renin inhibition could be established. We hypothesized that VTP-27999, a novel renin inhibitor without major side-effects at high doses, would allow us to establish this. METHODS AND RESULTS: The effects of escalating VTP-27999 doses (75-600âmg) on RPF, glomerular filtration rate (GFR), and plasma RAS components were compared with those of 300âmg aliskiren in 22 normal volunteers on a low-sodium diet. VTP-27999 dose-dependently increased RPF and GFR; its effects on both parameters at 600âmg (increases of 18â±â4 and 20â±â4%, respectively) were equivalent to those at 300âmg, indicating that a maximum had been reached. The effects of 300âmg aliskiren (increases of 13â±â5 and 8â±â6%, respectively; Pâ<â0.01 vs. 300 and 600âmg VTP-27999) resembled those of 150âmg VTP-27999. VTP-27999 dose-dependently increased renin, and lowered plasma renin activity and angiotensin II to detection limit levels. The effects of aliskiren on RAS components were best comparable to those of 150âmg VTP-27999. CONCLUSION: Maximum renal renin blockade in healthy, salt-depleted volunteers, requires aliskiren doses higher than 300âmg, but can be established with 300âmg VTP-27999. To what degree such maximal effects (exceeding those of angiotensin-converting enzyme inhibitors and AT1-receptor blockers) are required in patients with renal disease, given the potential detrimental effects of excessive RAS blockade, remains to be determined.
Asunto(s)
Amidas/farmacología , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Antihipertensivos/farmacología , Carbamatos/farmacología , Fumaratos/farmacología , Piperidinas/farmacología , Sistema Renina-Angiotensina/efectos de los fármacos , Renina/antagonistas & inhibidores , Adolescente , Adulto , Anciano , Amidas/administración & dosificación , Inhibidores de la Enzima Convertidora de Angiotensina/administración & dosificación , Antihipertensivos/administración & dosificación , Presión Sanguínea/efectos de los fármacos , Carbamatos/administración & dosificación , Método Doble Ciego , Femenino , Fumaratos/administración & dosificación , Tasa de Filtración Glomerular/efectos de los fármacos , Voluntarios Sanos , Humanos , Masculino , Persona de Mediana Edad , Piperidinas/administración & dosificación , Estudios Prospectivos , Renina/sangre , Adulto JovenRESUMEN
AIMS: Increasing evidence supports a role for the angiotensin II-AT1-receptor axis in aneurysm development. Here, we studied whether counteracting this axis via stimulation of AT2 receptors is beneficial. Such stimulation occurs naturally during AT1-receptor blockade with losartan, but not during renin inhibition with aliskiren. METHODS AND RESULTS: Aneurysmal homozygous fibulin-4 mice, displaying a four-fold reduced fibulin-4 expression, were treated with placebo, losartan, aliskiren, or the ß-blocker propranolol from day 35 to 100. Their phenotype includes cystic media degeneration, aortic regurgitation, left ventricular dilation, reduced ejection fraction, and fractional shortening. Although losartan and aliskiren reduced hemodynamic stress and increased renin similarly, only losartan increased survival. Propranolol had no effect. No drug rescued elastic fiber fragmentation in established aneurysms, although losartan did reduce aneurysm size. Losartan also increased ejection fraction, decreased LV diameter, and reduced cardiac pSmad2 signaling. None of these effects were seen with aliskiren or propranolol. Longitudinal micro-CT measurements, a novel method in which each mouse serves as its own control, revealed that losartan reduced LV growth more than aneurysm growth, presumably because the heart profits both from the local (cardiac) effects of losartan and its effects on aortic root remodeling. CONCLUSION: Losartan, but not aliskiren or propranolol, improved survival in fibulin-4 mice. This most likely relates to its capacity to improve structure and function of both aorta and heart. The absence of this effect during aliskiren treatment, despite a similar degree of blood pressure reduction and renin-angiotensin system blockade, suggests that it might be because of AT2-receptor stimulation.
Asunto(s)
Aneurisma/fisiopatología , Bloqueadores del Receptor Tipo 1 de Angiotensina II/metabolismo , Proteínas de la Matriz Extracelular , Insuficiencia Cardíaca/fisiopatología , Receptor de Angiotensina Tipo 1/metabolismo , Renina/metabolismo , Animales , Proteínas de la Matriz Extracelular/genética , Proteínas de la Matriz Extracelular/metabolismo , Ratones , Ratones Transgénicos , Renina/antagonistas & inhibidoresRESUMEN
The renin-angiotensin system (RAS) plays a critical role in the pathogenesis of many types of cardiovascular diseases including cardiomyopathy, valvular heart disease, aneurysms, stroke, coronary artery disease and vascular injury. Besides the classical regulatory effects on blood pressure and sodium homoeostasis, the RAS is involved in the regulation of contractility and remodelling of the vessel wall. Numerous studies have shown beneficial effect of inhibition of this system in the pathogenesis of cardiovascular diseases. However, dysregulation and overexpression of the RAS, through different molecular mechanisms, also induces, the initiation of vascular damage. The key effector peptide of the RAS, angiotensin II (Ang II) promotes cell proliferation, apoptosis, fibrosis, oxidative stress and inflammation, processes known to contribute to remodelling of the vasculature. In this review, we focus on the components that are under the influence of the RAS and contribute to the development and progression of vascular disease; extracellular matrix defects, atherosclerosis and ageing. Furthermore, the beneficial therapeutic effects of inhibition of the RAS on the vasculature are discussed, as well as the need for additive effects on top of RAS inhibition.
Asunto(s)
Sistema Renina-Angiotensina , Enfermedades Vasculares/metabolismo , Animales , Progresión de la Enfermedad , Humanos , Terapia Molecular Dirigida , Sistema Renina-Angiotensina/efectos de los fármacos , Enfermedades Vasculares/tratamiento farmacológico , Enfermedades Vasculares/patologíaRESUMEN
BACKGROUND: In this study we set out to investigate the clinically observed relationship between chronic obstructive pulmonary disease (COPD) and aortic aneurysms. We tested the hypothesis that an inherited deficiency of connective tissue might play a role in the combined development of pulmonary emphysema and vascular disease. METHODS: We first determined the prevalence of chronic obstructive pulmonary disease in a clinical cohort of aortic aneurysms patients and arterial occlusive disease patients. Subsequently, we used a combined approach comprising pathological, functional, molecular imaging, immunological and gene expression analysis to reveal the sequence of events that culminates in pulmonary emphysema in aneurysmal Fibulin-4 deficient (Fibulin-4(R)) mice. RESULTS: Here we show that COPD is significantly more prevalent in aneurysm patients compared to arterial occlusive disease patients, independent of smoking, other clinical risk factors and inflammation. In addition, we demonstrate that aneurysmal Fibulin-4(R/R) mice display severe developmental lung emphysema, whereas Fibulin-4(+/R) mice acquire alveolar breakdown with age and upon infectious stress. This vicious circle is further exacerbated by the diminished antiprotease capacity of the lungs and ultimately results in the development of pulmonary emphysema. CONCLUSIONS: Our experimental data identify genetic susceptibility to extracellular matrix degradation and secondary inflammation as the common mechanisms in both COPD and aneurysm formation.