RESUMO
Objective: A cardinal feature of Marfan syndrome is thoracic aortic aneurysm. The contribution of the renin-angiotensin system via AT1aR (Ang II [angiotensin II] receptor type 1a) to thoracic aortic aneurysm progression remains controversial because the beneficial effects of angiotensin receptor blockers have been ascribed to off-target effects. This study used genetic and pharmacological modes of attenuating angiotensin receptor and ligand, respectively, to determine their roles on thoracic aortic aneurysm in mice with fibrillin-1 haploinsufficiency (Fbn1C1041G/+). Approach and Results: Thoracic aortic aneurysm in Fbn1C1041G/+ mice was found to be strikingly sexual dimorphic. Males displayed aortic dilation over 12 months while aortic dilation in Fbn1C1041G/+ females did not differ significantly from wild-type mice. To determine the role of AT1aR, Fbn1C1041G/+ mice that were either +/+ or -/- for AT1aR were generated. AT1aR deletion reduced expansion of ascending aorta and aortic root diameter from 1 to 12 months of age in males. Medial thickening and elastin fragmentation were attenuated. An antisense oligonucleotide against angiotensinogen was administered to male Fbn1C1041G/+ mice to determine the effects of Ang II depletion. Antisense oligonucleotide against angiotensinogen administration attenuated dilation of the ascending aorta and aortic root and reduced extracellular remodeling. Aortic transcriptome analyses identified potential targets by which inhibition of the renin-angiotensin system reduced aortic dilation in Fbn1C1041G/+ mice. Conclusions: Deletion of AT1aR or inhibition of Ang II production exerted similar effects in attenuating pathologies in the proximal thoracic aorta of male Fbn1C1041G/+ mice. Inhibition of the renin-angiotensin system attenuated dysregulation of genes within the aorta related to pathology of Fbn1C1041G/+ mice.
Assuntos
Angiotensinogênio/metabolismo , Aorta Torácica/metabolismo , Aneurisma da Aorta Torácica/prevenção & controle , Fibrilina-1/genética , Deleção de Genes , Síndrome de Marfan/genética , Receptor Tipo 1 de Angiotensina/genética , Sistema Renina-Angiotensina , Angiotensinogênio/genética , Animais , Aorta Torácica/patologia , Aneurisma da Aorta Torácica/genética , Aneurisma da Aorta Torácica/metabolismo , Aneurisma da Aorta Torácica/patologia , Modelos Animais de Doenças , Feminino , Fibrilina-1/metabolismo , Predisposição Genética para Doença , Haploinsuficiência , Masculino , Síndrome de Marfan/metabolismo , Síndrome de Marfan/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Fenótipo , Receptor Tipo 1 de Angiotensina/deficiência , Sistema Renina-Angiotensina/genética , Caracteres Sexuais , Fatores Sexuais , TranscriptomaRESUMO
The renin-angiotensin system (RAS) is a highly complex hormonal cascade that spans multiple organs and cell types to regulate solute and fluid balance along with cardiovascular function. Much of our current understanding of the functions of the RAS has emerged from a series of key studies in genetically-modified animals. Here, we review key findings from ground-breaking transgenic models, spanning decades of research into the RAS, with a focus on their use in studying blood pressure. We review the physiological importance of this regulatory system as evident through the examination of mouse models for several major RAS components: angiotensinogen, renin, ACE, ACE2, and the type 1 A angiotensin receptor. Both whole-animal and cell-specific knockout models have permitted critical RAS functions to be defined and demonstrate how redundancy and multiplicity within the RAS allow for compensatory adjustments to maintain homeostasis. Moreover, these models present exciting opportunities for continued discovery surrounding the role of the RAS in disease pathogenesis and treatment for cardiovascular disease and beyond.
Assuntos
Angiotensinogênio/genética , Doenças Cardiovasculares/genética , Modelos Animais de Doenças , Sistema Renina-Angiotensina/genética , Renina/genética , Equilíbrio Hidroeletrolítico/genética , Enzima de Conversão de Angiotensina 2/deficiência , Enzima de Conversão de Angiotensina 2/genética , Angiotensinogênio/deficiência , Animais , Pressão Sanguínea/genética , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/patologia , Regulação da Expressão Gênica , Humanos , Rim/citologia , Rim/metabolismo , Camundongos , Camundongos Knockout , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 2 de Angiotensina/deficiência , Receptor Tipo 2 de Angiotensina/genética , Renina/deficiência , Transdução de SinaisRESUMO
The present study tested the hypotheses that overexpression of an intracellular Ang II (angiotensin II) fusion protein, mito-ECFP/Ang II, selectively in the mitochondria of mouse proximal tubule cells induces mitochondrial oxidative and glycolytic responses and elevates blood pressure via the Ang II/AT1a receptor/superoxide/NHE3 (the Na+/H+ exchanger 3)-dependent mechanisms. A PT-selective, mitochondria-targeting adenoviral construct encoding Ad-sglt2-mito-ECFP/Ang II was used to test the hypotheses. The expression of mito-ECFP/Ang II was colocalized primarily with Mito-Tracker Red FM in mouse PT cells or with TMRM in kidney PTs. Mito-ECFP/Ang II markedly increased oxygen consumption rate as an index of mitochondrial oxidative response (69.5%; P<0.01) and extracellular acidification rate as an index of mitochondrial glycolytic response (34%; P<0.01). The mito-ECFP/Ang II-induced oxygen consumption rate and extracellular acidification rate responses were blocked by AT1 blocker losartan (P<0.01) and a mitochondria-targeting superoxide scavenger mito-TEMPO (P<0.01). By contrast, the nonselective NO inhibitor L-NAME alone increased, whereas the mitochondria-targeting expression of AT2 receptors (mito-AT2/GFP) attenuated the effects of mito-ECFP/Ang II (P<0.01). In the kidney, overexpression of mito-ECFP/Ang II in the mitochondria of the PTs increased systolic blood pressure 12±3 mm Hg (P<0.01), and the response was attenuated in PT-specific PT-Agtr1a-/- and PT-Nhe3-/- mice (P<0.01). Conversely, overexpression of AT2 receptors selectively in the mitochondria of the PTs induced natriuretic responses in PT-Agtr1a-/- and PT-Nhe3-/- mice (P<0.01). Taken together, these results provide new evidence for a physiological role of PT mitochondrial Ang II/AT1a/superoxide/NHE3 and Ang II/AT2/NO/NHE3 signaling pathways in maintaining blood pressure homeostasis.
Assuntos
Angiotensina II/fisiologia , Túbulos Renais Proximais/fisiologia , Mitocôndrias/fisiologia , Receptor Tipo 1 de Angiotensina/fisiologia , Receptor Tipo 2 de Angiotensina/fisiologia , Transdução de Sinais/fisiologia , Bloqueadores do Receptor Tipo 2 de Angiotensina II/farmacologia , Animais , Células Cultivadas , Glicólise , Hipertensão/fisiopatologia , Imidazóis/farmacologia , Córtex Renal/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NG-Nitroarginina Metil Éster/farmacologia , Compostos Organofosforados/farmacologia , Piperidinas/farmacologia , Piridinas/farmacologia , Receptor Tipo 1 de Angiotensina/deficiência , Sódio/metabolismo , Simportadores de Sódio-Bicarbonato/metabolismo , Trocador 1 de Sódio-Hidrogênio/deficiência , Trocador 1 de Sódio-Hidrogênio/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismoRESUMO
BACKGROUND: Inhibition of the renin-angiotensin system remains a cornerstone in reducing proteinuria and progression of kidney failure, effects believed to be the result of reduction in BP and glomerular hyperfiltration. However, studies have yielded conflicting results on whether podocyte-specific angiotensin II (AngII) signaling directly induces podocyte injury. Previous research has found that after AngII stimulation, ß-arrestin-bound angiotensin II receptor type 1 (AT1R) is internalized in a clathrin- and dynamin-dependent manner, and that Dynamin1 and Dynamin2 double-knockout mice exhibit impaired clathrin-mediated endocytosis. METHODS: We used podocyte-specific Dyn double-knockout mice to examine AngII-stimulated AT1R internalization and signaling in primary podocytes and controls. We also examined the in vivo effect of AngII in these double-knockout mice through renin-angiotensin system blockers and through deletion of Agtr1a (which encodes the predominant AT1R isoform expressed in kidney, AT1aR). We tested calcium influx, Rac1 activation, and lamellipodial extension in control and primary podocytes of Dnm double-knockout mice treated with AngII. RESULTS: We confirmed augmented AngII-stimulated AT1R signaling in primary Dnm double-knockout podocytes resulting from arrest of clathrin-coated pit turnover. Genetic ablation of podocyte Agtr1a in Dnm double-knockout mice demonstrated improved albuminuria and kidney function compared with the double-knockout mice. Isolation of podocytes from Dnm double-knockout mice revealed abnormal membrane dynamics, with increased Rac1 activation and lamellipodial extension, which was attenuated in Dnm double-knockout podocytes lacking AT1aR. CONCLUSIONS: Our results indicate that inhibiting aberrant podocyte-associated AT1aR signaling pathways has a protective effect in maintaining the integrity of the glomerular filtration barrier.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Vesículas Revestidas por Clatrina/fisiologia , Podócitos/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/efeitos dos fármacos , Albuminúria/fisiopatologia , Angiotensina II/farmacologia , Animais , Sinalização do Cálcio , Células Cultivadas , Creatinina/sangue , Creatinina/urina , Dinamina I/deficiência , Dinamina I/fisiologia , Dinamina II/deficiência , Dinamina II/fisiologia , Endocitose , Glomerulonefrite/genética , Glomerulonefrite/fisiopatologia , Hemodinâmica , Glomérulos Renais/patologia , Masculino , Camundongos , Camundongos Knockout , Neuropeptídeos/fisiologia , Podócitos/efeitos dos fármacos , Podócitos/ultraestrutura , Pseudópodes/fisiologia , Receptor Tipo 1 de Angiotensina/deficiência , Proteínas rac1 de Ligação ao GTP/fisiologiaRESUMO
An intact renin-angiotensin system involving ANG II type 1 (AT1) receptors is crucial for normal kidney development. It is still unclear in which cell types AT1 receptor signaling is required for normal kidney development, maturation, and function. Because all kidney cells deriving from stroma progenitor cells express AT1 receptors and because stromal cells fundamentally influence nephrogenesis and tubular maturation, we investigated the relevance of AT1 receptors in stromal progenitors and their descendants for renal development and function. For this aim, we generated and analyzed mice with conditional deletion of AT1A receptor in the FoxD1 cell lineage in combination with global disruption of the AT1B receptor gene. These FoxD1-AT1ko mice developed normally. Their kidneys showed neither structural nor functional abnormalities compared with wild-type mice, whereas in isolated perfused FoxD1-AT1ko kidneys, the vasoconstrictor and renin inhibitory effects of ANG II were absent. In vivo, however, plasma renin concentration and renal renin expression were normal in FoxD1-AT1ko mice, as were blood pressure and glomerular filtration rate. These findings suggest that a strong reduction of AT1 receptors in renal stromal progenitors and their descendants does not disturb normal kidney development.
Assuntos
Linhagem da Célula , Fatores de Transcrição Forkhead/metabolismo , Rim/metabolismo , Receptor Tipo 1 de Angiotensina/deficiência , Sistema Renina-Angiotensina , Células-Tronco/metabolismo , Células Estromais/metabolismo , Animais , Pressão Sanguínea , Feminino , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Taxa de Filtração Glomerular , Rim/citologia , Masculino , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Organogênese , Fenótipo , Receptor Tipo 1 de Angiotensina/genética , Renina/sangue , Sistema Renina-Angiotensina/genética , Transdução de SinaisRESUMO
BACKGROUND: Previously, we demonstrated that obese mice have marked elevations in systemic concentrations of angiotensin II (AngII). Drugs that inhibit the renin-angiotensin system (RAS), including angiotensin type 1 receptor (AT1R) antagonists, have been reported to delay the onset of type 2 diabetes (T2D), suggesting improvements in insulin sensitivity or regulation of pancreatic insulin secretion. Pancreatic islets possess components of the RAS, including AT1R, but it is unclear if AngII acts at islets to regulate insulin secretion during the development of T2D. METHODS: We deleted AT1aR from pancreatic islets and examined effects on insulin secretion in mice fed a low-fat (LF) or high-fat (HF) diet. In separate studies, to exacerbate the system, we infused HF-fed mice of each genotype with AngII. RESULTS: Pancreatic AT1aR deficiency impaired glucose tolerance and elevated plasma glucose concentrations in HF, but not LF-fed mice. In HF-fed mice, high glucose increased insulin secretion from islets of AT1aRfl/fl, but not AT1aRpdx mice. In AngII-infused mice, following glucose challenge, plasma glucose or insulin concentrations were not significantly different between genotypes. Moreover, high glucose stimulated insulin secretion from islets of AT1aRfl/fl and AT1aRpdx mice, presumably related to weight loss, and improved insulin sensitivity in both groups of AngII-infused HF-fed mice. CONCLUSIONS: Our results suggest that during the adaptive response to insulin resistance from HF feeding, AngII promotes insulin secretion from islets through an AT1aR mechanism. These results suggest the timing of initiation of AT1R blockade may be important in the progression from prediabetes to T2D with ß-cell failure.
Assuntos
Dieta Hiperlipídica , Resistência à Insulina , Insulina/sangue , Ilhotas Pancreáticas/metabolismo , Obesidade/sangue , Receptor Tipo 1 de Angiotensina/deficiência , Sistema Renina-Angiotensina , Adaptação Fisiológica , Angiotensina II/administração & dosagem , Animais , Biomarcadores/sangue , Glicemia/metabolismo , Dieta com Restrição de Gorduras , Modelos Animais de Doenças , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Obesidade/genética , Obesidade/fisiopatologia , Receptor Tipo 1 de Angiotensina/genética , Sistema Renina-Angiotensina/efeitos dos fármacos , Via SecretóriaRESUMO
ENaC-mediated sodium reabsorption in the collecting duct (CD) is a critical determinant of urinary sodium excretion. Existing evidence suggest direct stimulatory actions of Angiotensin II (Ang II) on ENaC in the CD, independently of the aldosterone-mineralocorticoid receptor (MR) signaling. Deletion of the major renal AT1 receptor isoform, AT1a R, decreases blood pressure and reduces ENaC abundance despite elevated aldosterone levels. The mechanism of this insufficient compensation is not known. Here, we used patch clamp electrophysiology in freshly isolated split-opened CDs to investigate how AT1a R dysfunction compromises functional ENaC activity and its regulation by dietary salt intake. Ang II had no effect on ENaC activity in CDs from AT1a R -/- mice suggesting no complementary contribution of AT2 receptors. We next found that AT1a R deficient mice had lower ENaC activity when fed with low (<0.01% Na+ ) and regular (0.32% Na+ ) but not with high (â¼2% Na+ ) salt diet, when compared to the respective values obtained in Wild type (WT) animals. Inhibition of AT1 R with losartan in wild-type animals reproduces the effects of genetic ablation of AT1a R on ENaC activity arguing against contribution of developmental factors. Interestingly, manipulation with aldosterone-MR signaling via deoxycosterone acetate (DOCA) and spironolactone had much reduced influence on ENaC activity upon AT1a R deletion. Consistently, AT1a R -/- mice have a markedly diminished MR abundance in cytosol. Overall, we conclude that AT1a R deficiency elicits a complex inhibitory effect on ENaC activity by attenuating ENaC Po and precluding adequate compensation via aldosterone cascade due to decreased MR availability.
Assuntos
Canais Epiteliais de Sódio/metabolismo , Túbulos Renais Coletores/metabolismo , Receptor Tipo 1 de Angiotensina/deficiência , Aldosterona/farmacologia , Angiotensina II/farmacologia , Animais , Losartan/farmacologia , Masculino , Camundongos Endogâmicos C57BL , Receptor Tipo 1 de Angiotensina/metabolismo , Receptores de Mineralocorticoides/metabolismo , Transdução de Sinais/efeitos dos fármacos , Cloreto de Sódio na Dieta/farmacologiaRESUMO
OBJECTIVE: Two were the aims of this study: first, to translate whole-genome expression profiles into computational predictions of functional associations between signaling pathways that regulate aorta homeostasis and the activity of angiotensin II type 1a receptor (At1ar) in either vascular endothelial or smooth muscle cells; and second, to characterize the impact of endothelial cell- or smooth muscle cell-specific At1ar disruption on the development of thoracic aortic aneurysm in fibrillin-1 hypomorphic (Fbn1mgR/mgR ) mice, a validated animal model of early onset progressively severe Marfan syndrome. APPROACH AND RESULTS: Cdh5-Cre and Sm22-Cre transgenic mice were used to inactivate the At1ar-coding gene (Agt1ar) in either intimal or medial cells of both wild type and Marfan syndrome mice, respectively. Computational analyses of differentially expressed genes predicted dysregulated signaling pathways of cell survival and matrix remodeling in Agt1arCdh5-/- aortas and of cell adhesion and contractility in Agt1arSm22-/- aortas. Characterization of Fbn1mgR/mgR;Agt1arCdh5-/- mice revealed increased median survival associated with mitigated aneurysm growth and media degeneration, as well as reduced levels of phosphorylated (p-) Erk1/2 but not p-Smad2. By contrast, levels of both p-Erk1/2 and p-Smad2 proteins were normalized in Fbn1mgR/mgR;Agt1arSm22-/- aortas in spite of them showing no appreciable changes in thoracic aortic aneurysm pathology. CONCLUSIONS: Physiological At1ar signaling in the intimal and medial layers is associated with distinct regulatory processes of aorta homeostasis and function; improper At1ar activity in the vascular endothelium is a significant determinant of thoracic aortic aneurysm development in Marfan syndrome mice.
Assuntos
Aorta Torácica/metabolismo , Aneurisma da Aorta Torácica/metabolismo , Células Endoteliais/metabolismo , Miócitos de Músculo Liso/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismo , Animais , Aorta Torácica/patologia , Aorta Torácica/fisiopatologia , Aneurisma da Aorta Torácica/genética , Aneurisma da Aorta Torácica/patologia , Aneurisma da Aorta Torácica/fisiopatologia , Biologia Computacional , Dilatação Patológica , Modelos Animais de Doenças , Células Endoteliais/patologia , Fibrilina-1/genética , Fibrilina-1/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Homeostase , Masculino , Síndrome de Marfan/genética , Síndrome de Marfan/metabolismo , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miócitos de Músculo Liso/patologia , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Transdução de SinaisRESUMO
OBJECTIVE: Angiotensin (Ang) II type 1 receptor (AT1) activation is essential for the development of exogenous Ang II-induced abdominal aortic aneurysms (AAAs) in hyperlipidemic animals. Experimental data derived from this modeling system, however, provide limited insight into the role of endogenous Ang II in aneurysm pathogenesis. Consequently, the potential translational value of AT1 inhibition in clinical AAA disease management remains incompletely understood on the basis of the existing literature. METHODS: AAAs were created in wild-type (WT) and AT1a knockout (KO) mice by intra-aortic infusion of porcine pancreatic elastase (PPE). WT mice were treated with the AT1 receptor antagonist telmisartan, 10 mg/kg/d in chow, or the peroxisome proliferator-activated receptor γ (PPARγ) antagonist GW9662, 3 mg/kg/d through oral gavage, beginning 1 week before or 3 days after PPE infusion. Influences on aneurysm progression as well as mechanistic insights into AT1-mediated pathogenic processes were determined using noninvasive ultrasound imaging, histopathology, aortic gene expression profiling, and flow cytometric analysis. RESULTS: After PPE infusion, aortic enlargement was almost completely abrogated in AT1a KO mice compared with WT mice. As defined by a ≥50% increase in aortic diameter, no PPE-infused, AT1a KO mouse actually developed an AAA. On histologic evaluation, medial smooth muscle cellularity and elastic lamellae were preserved in AT1a KO mice compared with WT mice, with marked attenuation of mural angiogenesis and leukocyte infiltration. In WT mice, telmisartan administration effectively suppressed aneurysm pathogenesis after PPE infusion as well, regardless of whether treatment was initiated before or after aneurysm creation or continued for a limited or extended time. Telmisartan treatment was associated with reduced messenger RNA levels for CCL5 and matrix metalloproteinases 2 and 9 in aneurysmal aortae, with no apparent effect on PPARγ-regulated gene expression. Administration of the PPARγ antagonist GW9662 failed to "rescue" the aneurysm phenotype in telmisartan-treated, PPE-infused WT mice. Neither effector T-cell differentiation nor regulatory T-cell cellularity was affected by telmisartan treatment status. CONCLUSIONS: Telmisartan effectively suppresses the progression of elastase-induced AAAs without apparent effect on PPARγ activation or T-cell differentiation. These findings reinforce the critical importance of endogenous AT1 activation in experimental AAA pathogenesis and reinforce the translational potential of AT1 inhibition in medical aneurysm disease management.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Aorta Abdominal/efeitos dos fármacos , Aneurisma da Aorta Abdominal/prevenção & controle , Benzimidazóis/farmacologia , Benzoatos/farmacologia , Elastase Pancreática , Receptor Tipo 1 de Angiotensina/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/deficiência , Animais , Aorta Abdominal/diagnóstico por imagem , Aorta Abdominal/metabolismo , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/genética , Aneurisma da Aorta Abdominal/metabolismo , Dilatação Patológica , Modelos Animais de Doenças , Regulação da Expressão Gênica , Predisposição Genética para Doença , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Receptor Tipo 1 de Angiotensina/genética , Transdução de Sinais/efeitos dos fármacos , Telmisartan , Fatores de Tempo , TranscriptomaRESUMO
Hypertensive patients have an increased risk of developing kidney cancer. We have shown in vivo that besides elevating blood pressure, angiotensin II causes DNA damage dose dependently. Here, the role of blood pressure in the formation of DNA damage is studied. Mice lacking one of the two murine angiotensin II type 1 receptor (AT1R) subtypes, AT1aR, were equipped with osmotic minipumps, delivering angiotensin II during 28 days. Parameters of oxidative stress and DNA damage of kidneys and hearts of AT1aR-knockout mice were compared with wild-type (C57BL/6) mice receiving angiotensin II, and additionally, with wild-type mice treated with candesartan, an antagonist of both AT1R subtypes. In wild-type mice, angiotensin II induced hypertension, reduced kidney function, and led to a significant formation of reactive oxygen species (ROS). Furthermore, genomic damage was markedly increased in this group. All these responses to angiotensin II could be attenuated by concurrent administration of candesartan. In AT1aR-deficient mice treated with angiotensin II, systolic pressure was not increased, and renal function was not affected. However, angiotensin II still led to an increase of ROS in kidneys and hearts of these animals. Additionally, genomic damage in the form of double-strand breaks was significantly induced in kidneys of AT1aR-deficient mice. Our results show that angiotensin II induced ROS production and DNA damage even without the presence of AT1aR and independently of blood pressure changes.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/deficiência , Angiotensina II/farmacologia , Animais , Hipertensão/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Espécies Reativas de Oxigênio/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismoRESUMO
The aim of this study was to confirm the renoprotective effect of xanthine oxidoreductase (XOR) inhibitor, topiroxostat, compared with another XOR inhibitor, febuxostat, under decreased angiotensin II type 1a (AT1a) receptor expression in the model of renal injury caused by adenine. To evaluate the degree of tubular damage using urinary liver-type fatty acid-binding protein (L-FABP) under decreased AT1a expression, we used AT1a receptor knockdown hetero and human L-FABP chromosomal transgenic (Tg) mice (AT1a+/-L-FABP+/-). Male AT1a+/-L-FABP+/- mice were divided into two groups: the adenine diet group (n = 40) was given a diet containing only 0.2% w/w adenine, and the normal diet group (n = 5) was given a normal diet. When renal dysfunction was confirmed in the adenine diet group 4 weeks after starting the diet, the adenine diet group was further divided into five groups. The adenine diet group (n = 8) was continuously given only the adenine diet. Each group receiving high-dose (3mg/kg) or low-dose (1mg/kg) topiroxostat (Topiroxostat-H group, n = 8, Topiroxostat-L group, n = 8) or febuxostat (Febuxostat-H group, n = 8, Febuxostat-L group, n = 8) was given the adenine diet including the drug for another 4 weeks. The levels of renal XOR, renal dysfunction, urinary L-FABP, tubulointerstitial damage, hypoxia, and oxidative stress were decreased or attenuated after treatment with topiroxostat or febuxostat compared with the adenine diet group. Furthermore, antioxidant capacity was maintained owing to these treatments. In conclusion, topiroxostat and febuxostat attenuated renal damage under decreased AT1a expression in the adenine-induced renal injury model.
Assuntos
Citoproteção/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Rim/efeitos dos fármacos , Nitrilas/farmacologia , Piridinas/farmacologia , Receptor Tipo 1 de Angiotensina/metabolismo , Xantina Desidrogenase/antagonistas & inibidores , Angiotensina II/urina , Animais , Pressão Sanguínea/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Hipóxia Celular/efeitos dos fármacos , Colágeno Tipo III/metabolismo , Proteínas de Ligação a Ácido Graxo/genética , Técnicas de Silenciamento de Genes , Humanos , Rim/citologia , Rim/metabolismo , Masculino , Camundongos , Estresse Oxidativo/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genéticaRESUMO
Angiotensin II (AngII) promotes hypertension, atherogenesis, vascular aneurysm and impairs post-ischemic cardiac remodeling through concerted roles on vascular cells, monocytes and T lymphocytes. However, the role of AngII in B lymphocyte responses is largely unexplored. Here, we show that chronic B cell depletion (Baffr deficiency) significantly reduces atherosclerosis in Apoe -/- mice infused with AngII. While adoptive transfer of B cells in Apoe -/- /Baffr -/- mice reversed atheroprotection in the absence of AngII, infusion of AngII in B cell replenished Apoe -/- /Baffr -/- mice unexpectedly prevented the progression of atherosclerosis. Atheroprotection observed in these mice was associated with a significant increase in regulatory CD1dhiCD5+ B cells, which produced high levels of interleukin (IL)-10 (B10 cells). Replenishment of Apoe -/- /Baffr -/- mice with Il10 -/- B cells reversed AngII-induced B cell-dependent atheroprotection, thus highlighting a protective role of IL-10+ regulatory B cells in this setting. Transfer of AngII type 1A receptor deficient (Agtr1a -/-) B cells into Apoe -/- /Baffr -/- mice substantially reduced the production of IL-10 by B cells and prevented the AngII-dependent atheroprotective B cell phenotype. Consistent with the in vivo data, AngII synergized with BAFF to induce IL-10 production by B cells in vitro via AngII type 1A receptor. Our data demonstrate a previously unknown synergy between AngII and BAFF in inducing IL-10 production by B cells, resulting in atheroprotection.
Assuntos
Angiotensina II/metabolismo , Aterosclerose/etiologia , Aterosclerose/metabolismo , Fator Ativador de Células B/genética , Linfócitos B Reguladores/metabolismo , Animais , Aterosclerose/patologia , Linfócitos B Reguladores/imunologia , Biomarcadores , Células Cultivadas , Modelos Animais de Doenças , Imuno-Histoquímica , Interleucina-10/biossíntese , Camundongos , Camundongos Knockout , Fenótipo , Receptor Tipo 1 de Angiotensina/deficiênciaRESUMO
The Frank-Starling law of the heart is a physiological phenomenon that describes an intrinsic property of heart muscle in which increased cardiac filling leads to enhanced cardiac contractility. Identified more than a century ago, the Frank-Starling relationship is currently known to involve length-dependent enhancement of cardiac myofilament Ca2+ sensitivity. However, the upstream molecular events that link cellular stretch to the length-dependent myofilament Ca2+ sensitivity are poorly understood. Because the angiotensin II type 1 receptor (AT1R) and the multifunctional transducer protein ß-arrestin have been shown to mediate mechanosensitive cellular signaling, we tested the hypothesis that these two proteins are involved in the Frank-Starling mechanism of the heart. Using invasive hemodynamics, we found that mice lacking ß-arrestin 1, ß-arrestin 2, or AT1R were unable to generate a Frank-Starling force in response to changes in cardiac volume. Although wild-type mice pretreated with the conventional AT1R blocker losartan were unable to enhance cardiac contractility with volume loading, treatment with a ß-arrestin-biased AT1R ligand to selectively activate ß-arrestin signaling preserved the Frank-Starling relationship. Importantly, in skinned muscle fiber preparations, we found markedly impaired length-dependent myofilament Ca2+ sensitivity in ß-arrestin 1, ß-arrestin 2, and AT1R knockout mice. Our data reveal ß-arrestin 1, ß-arrestin 2, and AT1R as key regulatory molecules in the Frank-Starling mechanism, which potentially can be targeted therapeutically with ß-arrestin-biased AT1R ligands.
Assuntos
Modelos Cardiovasculares , Contração Miocárdica/fisiologia , beta-Arrestina 1/fisiologia , beta-Arrestina 2/fisiologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Sinalização do Cálcio/fisiologia , Técnicas In Vitro , Losartan/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Miocárdica/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/fisiologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , beta-Arrestina 1/deficiência , beta-Arrestina 1/genética , beta-Arrestina 2/deficiência , beta-Arrestina 2/genéticaRESUMO
Angiotensin II (Ang II) and its receptor AT1 (AT1R), an important effector axis of renin-angiotensin system (RAS), have been demonstrated to regulate T-cell responses. However, these studies characterized Ang II and AT1R effects using pharmacological tools, which do not target only Ang II/AT1R axis. The specific role of AT1R expressed by antigen-specific CD8+ T cells is unknown. Then we immunized transgenic mice expressing a T-cell receptor specific for SIINFEKL epitope (OT-I mice) with sporozoites of the rodent malaria parasite Plasmodium berghei expressing the cytotoxic epitope SIINFEKL. Early priming events after immunization were not affected but the expansion and contraction of AT1R-deficient (AT1R-/-) OT-I cells was decreased. Moreover, they seemed more activated, express higher levels of CTLA-4, PD-1, LAG-3, and have decreased functional capacity during the effector phase. Memory AT1R-/- OT-I cells exhibited higher IL-7Rα expression, activation, and exhaustion phenotypes but less cytotoxic capacity. Importantly, AT1R-/- OT-I cells show better control of blood parasitemia burden and ameliorate mice survival during lethal disease induced by blood-stage malaria. Our study reveals that AT1R in antigen-specific CD8+ T cells regulates expansion, differentiation, and function during effector and memory phases of the response against Plasmodium, which could apply to different infectious agents.
Assuntos
Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Receptor Tipo 1 de Angiotensina/imunologia , Animais , Linfócitos T CD8-Positivos/metabolismo , Diferenciação Celular , Citotoxicidade Imunológica , Imunização , Epitopos Imunodominantes/genética , Memória Imunológica , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Ovalbumina/imunologia , Fragmentos de Peptídeos/imunologia , Plasmodium berghei/imunologia , Plasmodium berghei/patogenicidade , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Esporozoítos/imunologia , Esporozoítos/patogenicidadeRESUMO
The action of angiotensin II (AngII) on the Ca(2+) signals driving pyeloureteric peristalsis was investigated using both conventional and angiotensin receptor (ATr) ATr1A and ATr2 knockout ((-/-)) mice. Contractility in the renal pelvis of adult ATr1A(-/-) and ATr2(-/-) mice was compared to their respective wildtype (ATr1A(+/+) and ATr2(+/+)) controls of the same genetic background (FVB/N and C57Bl/6 respectively) using video microscopy. The effects of AngII on the Ca(2+) signals in typical and atypical smooth muscle cells (TSMCs and ASMCs, respectively) within the pelvic wall of conventional mice were recorded using Fluo-4 Ca(2+) imaging. Compared to ATr1A(+/+) , ATr2(+/+) and ATr2(-/-) mice, kidneys of the ATr1A(-/-) mouse were mildly-to-severely hydronephrotic, associated with an enlarged calyx, an atrophic papilla and a hypoplastic renal pelvis. Contraction frequencies in the renal pelvis of moderately hydronephrotic ATr1A(-/-) and unaffected ATr2(-/-) mice were not significantly different from their ATr1A(+/+), ATr2(+/+) controls. No contractions were observed in severely-hydronephrotic ATr1A(-/-) kidneys. AngII increased the spontaneous contraction frequency of the renal pelvis in ATr1A(+/+), ATr2(+/+) and ATr2(-/-) mice, but had little effect on the contractions in the mildly-hydronephrotic ATr1A(-/-) renal pelvis. The ATr1 blocker, candesartan prevented the positive chronotropic effects of AngII. AngII increased the frequency and synchronicity of Ca(2+) transients in both TSMCs and ASMCs. It was concluded that the hydronephrosis observed in ATr1A(-/-) mouse kidneys does not arise from a failure in the development of the essential pacemaker and contractile machinery driving pyeloureteric peristalsis.
Assuntos
Técnicas de Inativação de Genes , Hidronefrose/genética , Hidronefrose/fisiopatologia , Pelve Renal/fisiopatologia , Peristaltismo/genética , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Angiotensina II/farmacologia , Animais , Relógios Biológicos/efeitos dos fármacos , Relógios Biológicos/genética , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/genética , Feminino , Hidronefrose/patologia , Pelve Renal/efeitos dos fármacos , Pelve Renal/patologia , Masculino , Camundongos , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/patologia , Peristaltismo/efeitos dos fármacos , Receptor Tipo 2 de Angiotensina/deficiência , Receptor Tipo 2 de Angiotensina/genéticaRESUMO
Alzheimer's disease is characterized by neuronal loss and cerebral accumulation of amyloid-ß protein (Aß) and lowering the generation of Aß is a pivotal approach in the strategy of Alzheimer's disease treatment. Midlife hypertension is a major risk factor for the future onset of sporadic Alzheimer's disease and the use of some antihypertensive drugs may decrease the incidence of Alzheimer's disease. However, it is largely unknown how the blood pressure regulation system is associated with the pathogenesis of Alzheimer's disease. Here we found that the deficiency of angiotensin type 1a receptor (AT1a), a key receptor for regulating blood pressure, significantly decreased Aß generation and amyloid plaque formation in a mouse model of Alzheimer's disease. The lack of AT1a inhibited the endocleavage of presenilin-1 (PS1), which is essential for γ-secretase complex formation and Aß generation. Notably, the ligand of AT1a, angiotensin II, enhanced Aß generation, PS1 endocleavage and γ-secretase complex formation. Our results suggest that AT1a activation is closely associated with Aß generation and brain amyloid accumulation by regulating γ-secretase complex formation. Thus, removal of life style factors or stresses that stimulate AT1a to elevate blood pressure may decrease Aß generation and brain amyloid accumulation, thereby preventing the pathogenesis of Alzheimer's disease.
Assuntos
Peptídeos beta-Amiloides/metabolismo , Amiloide/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Receptor Tipo 1 de Angiotensina/genética , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/metabolismo , Angiotensina II/metabolismo , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Knockout , Fosfatidilinositol 3-Quinases/metabolismo , Placa Amiloide/genética , Placa Amiloide/patologia , Agregação Patológica de Proteínas/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor Tipo 1 de Angiotensina/deficiência , Transdução de SinaisRESUMO
BACKGROUND: Whether biomechanical force on the heart can induce exosome secretion to modulate cardiovascular function is not known. We investigated the secretion and activity of exosomes containing a key receptor in cardiovascular function, the angiotensin II type I receptor (AT1R). METHODS AND RESULTS: Exosomes containing AT1Rs were isolated from the media overlying AT1R-overexpressing cells exposed to osmotic stretch and from sera of mice undergoing cardiac pressure overload. The presence of AT1Rs in exosomes was confirmed by both electron microscopy and radioligand receptor binding assays and shown to require ß-arrestin2, a multifunctional adaptor protein essential for receptor trafficking. We show that functional AT1Rs are transferred via exosomes in an in vitro model of cellular stretch. Using mice with global and cardiomyocyte conditional deletion of ß-arrestin2, we show that under conditions of in vivo pressure overload the cellular source of the exocytosis of exosomes containing AT1R is the cardiomyocyte. Exogenously administered AT1R-enriched exosomes target cardiomyocytes, skeletal myocytes, and mesenteric resistance vessels and are sufficient to confer blood pressure responsiveness to angiotensin II infusion in AT1R knockout mice. CONCLUSIONS: AT1R-enriched exosomes are released from the heart under conditions of in vivo cellular stress to likely modulate vascular responses to neurohormonal stimulation. In the context of the whole organism, the concept of G protein-coupled receptor trafficking should consider circulating exosomes as part of the reservoir of functional AT1Rs.
Assuntos
Exossomos/química , Miócitos Cardíacos/química , Receptor Tipo 1 de Angiotensina/sangue , Estresse Mecânico , Animais , Arrestinas/deficiência , Arrestinas/genética , Arrestinas/fisiologia , Pressão Sanguínea , Constrição , Exossomos/fisiologia , Feminino , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Eletrônica , Células Musculares/metabolismo , Miócitos Cardíacos/ultraestrutura , Pressão Osmótica , Transporte Proteico , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Ensaio Radioligante , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Resistência Vascular , beta-ArrestinasRESUMO
BACKGROUND: Perivascular adipose tissue is implicated in vasoreactivity; however, its effect on atherosclerosis remains undefined. METHODS AND RESULTS: We examined the effect of a high-cholesterol diet (HCD) on phenotypic alterations of the thoracic periaortic adipose tissue (tPAT) in apoE-deficient (apoE(-/-)) mice. Gene expression of the components of the renin angiotensin system and that of macrophage markers were significantly higher in apoE(-/-) mice fed an HCD than in those fed a chow diet (CD). These changes were absent both in angiotensin II (AngII) receptor blocker (ARB)-treated apoE(-/-) mice and in Ang II type 1 (AT1) receptor-deficient apoE(-/-) (Agtr1(-/-)/apoE(-/-)) mice. To evaluate their effect on atherosclerosis, we transplanted tPAT into apoE(-/-) mice alongside the distal abdominal aorta. Transplanted tPAT was harvested from apoE(-/-) and Agtr1(-/-)/apoE(-/-) mice fed a CD (tPAT-CD/apoE(-/-), tPAT-CD/Agtr1(-/-)/apoE(-/-)), HCD (tPAT-HCD/apoE(-/-), tPAT-HCD/Agtr1(-/-)/apoE(-/-)), or HCD in combination with ARB treatment (tPAT-HCD/ARB/apoE(-/-)). Four weeks after transplantation, a significantly increased oil red O-positive area was observed in the aorta of tPAT-HCD/apoE(-/-) mice than in tPAT-CD/apoE(-/-) mice. Such a change was absent in tPAT-HCD/ARB/apoE(-/-) and tPAT-HCD/Agtr1(-/-)/apoE(-/-) mice. CONCLUSIONS: Our findings demonstrated that AT1 receptor plays a crucial role in HCD-induced phenotypic alterations of tPAT, modulation of which could exert beneficial effects on atherosclerosis.
Assuntos
Tecido Adiposo/transplante , Bloqueadores do Receptor Tipo 2 de Angiotensina II/farmacologia , Apolipoproteínas E/genética , Aterosclerose/genética , Aterosclerose/prevenção & controle , Adipocinas/metabolismo , Animais , Aorta/metabolismo , Apolipoproteínas E/deficiência , Aterosclerose/patologia , Hemodinâmica , Inflamação/prevenção & controle , Lipídeos/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genéticaRESUMO
OBJECTIVE: Myogenic vasoconstriction is mediated by vascular smooth muscle cells of resistance arteries sensing mechanical stretch. Angiotensin II AT1 receptors and in particular AT1BRs in murine vascular smooth muscle cells have been characterized as mechanosensors that cannot fully account for myogenic vasoconstriction observed. Therefore, we aimed at uncovering novel vascular mechanosensors by expression profiling and functional characterization of candidate proteins. APPROACH AND RESULTS: Analyzing myogenic tone of isolated murine mesenteric arteries of AT1A and AT1B receptor double gene-deficient (AT1A/1B (-/-)) mice ex vivo, we observed a decreased myogenic tone at high intraluminal pressures and an unexpected hyper-reactivity at low intraluminal pressures because of upregulation of cysteinyl leukotriene 1 receptors (CysLT1Rs). Pharmacological blockade of CysLT1Rs with pranlukast significantly reduced myogenic tone not only in AT1A/1B (-/-) but also in wild-type arteries. In wild-type arteries, additional blockade of angiotensin II AT1 receptors with candesartan resulted in an additive reduction of myogenic tone. Furthermore, calcium imaging experiments were performed with fura-2-loaded human embryonic kidney 293 cells overexpressing CysLT1Rs and with isolated mesenteric vascular smooth muscle cells. Hypo-osmotically induced membrane stretch provoked calcium transients that were significantly reduced by pranlukast. Incubations of isolated mesenteric vascular smooth muscle cells with the 5-lipoxygenase inhibitor zileuton had no effect. Furthermore, the Gq/11-protein inhibitor YM 254890 profoundly reduced myogenic tone to the same extent as induced by the application of pranlukast plus candesartan. CONCLUSIONS: Here, we identify a novel, hitherto unappreciated role of CysLT1Rs in vascular regulation. We identified CysLT1Rs as novel mechanosensors in the vasculature involved in myogenic vasoconstriction. Moreover, our findings suggest that myogenic tone is determined by AT1 and CysLT1 receptors acting together as mechanosensors via Gq/11-protein activation.
Assuntos
Mecanorreceptores/metabolismo , Mecanotransdução Celular , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismo , Receptores de Leucotrienos/metabolismo , Vasoconstrição , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Pressão Arterial , Sinalização do Cálcio , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/metabolismo , Células HEK293 , Humanos , Antagonistas de Leucotrienos/farmacologia , Masculino , Mecanorreceptores/efeitos dos fármacos , Mecanotransdução Celular/efeitos dos fármacos , Artérias Mesentéricas/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/deficiência , Receptor Tipo 1 de Angiotensina/genética , Receptores de Leucotrienos/efeitos dos fármacos , Receptores de Leucotrienos/genética , Transfecção , Vasoconstrição/efeitos dos fármacosRESUMO
Enhanced central sympathetic outflow worsens left ventricular (LV) remodeling and prognosis in heart failure after myocardial infarction (MI). Previous studies suggested that activation of brain angiotensin II type 1 receptors (AT1R) in the brain stem leads to sympathoexcitation due to neuronal AT1R upregulation. Recent studies, however, revealed the importance of astrocytes for modulating neuronal activity, but whether changes in astrocytes influence central sympathetic outflow in heart failure is unknown. In the normal state, AT1R are only weakly expressed in astrocytes. We hypothesized that AT1R in astrocytes are upregulated in heart failure and modulate the activity of adjacent neurons, leading to enhanced sympathetic outflow. In the present study, by targeting deletion of astrocyte-specific AT1R, we investigated whether AT1R in astrocytes have a key role in enhancing central sympathetic outflow, and thereby influencing LV remodeling process and the prognosis of MI-induced heart failure. Using the Cre-LoxP system, we generated glial fibrillary acidic protein (GFAP)-specific AT1R knockout (GFAP/AT1RKO) mice. Urinary norepinephrine excretion for 24 h, as an indicator of sympathoexcitation, was significantly lower in GFAP/AT1RKO-MI mice than in control-MI mice. LV size and heart weight after MI were significantly smaller in GFAP/AT1RKO mice than in control mice. Prognosis was significantly improved in GFAP/AT1RKO-MI mice compared with control-MI mice. Our findings indicated that AT1R expression was upregulated in brain stem astrocytes in MI-induced heart failure, which worsened LV remodeling and prognosis via sympathoexcitation. Thus, in addition to neuronal AT1R, AT1R in astrocytes appear to have a key role in enhancing central sympathetic outflow in heart failure.