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[Figure: see text].
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Proteínas de Ligação a DNA/metabolismo , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Artéria Femoral/metabolismo , Membro Posterior/irrigação sanguínea , Isquemia/metabolismo , Neovascularização Fisiológica , Fatores de Transcrição/metabolismo , Animais , Aorta/metabolismo , Aorta/fisiopatologia , Cálcio/metabolismo , Sinalização do Cálcio , Células Cultivadas , Circulação Colateral , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Endotélio Vascular/fisiopatologia , Artéria Femoral/fisiopatologia , Isquemia/genética , Isquemia/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiopatologia , Miócitos de Músculo Liso/metabolismo , Fluxo Sanguíneo Regional , Fatores de Transcrição/genéticaRESUMO
Alzheimer's disease (AD) has long been considered a brain-specific dementia syndrome. However, in recent decades, the occurrence of cardiovascular (CV) disease in the progression of AD has been confirmed by increasing epidemiological evidence. In this study, we conducted an in-depth cardiovascular characterization of a humanized amyloid precursor protein (APP) overexpressing mouse model (hAPP23+/-), which overexpresses the Swedish mutation (KM670/671NL). At the age of 6 mo, hAPP23+/- mice had a lower survival, lower body weight, and increased corticosterone and VMA levels compared with C57BL/6 littermates. Systolic blood pressure was increased in hAPP23+/- animals compared with C57BL/6 littermates, but diastolic blood pressure was not statistically different. Pulse pressure remained unchanged but abdominal and carotid pulse-wave velocity (aPWV and cPWV) were increased in hAPP23+/- compared with C57BL/6 mice. Echocardiography showed no differences in systolic or diastolic cardiac function. Ex vivo evaluation of vascular function showed decreased adreno receptor dependent vasoconstriction of hAPP23+/- aortic segments, although the isobaric biomechanics of the aortic wall were similar to C57BL/6 aortic segments. In conclusion, hAPP23+/- mice exhibited high serum corticosterone levels, elevated systolic blood pressure, and increased arterial stiffness in vivo. However, ex vivo aortic stiffness of hAPP23+/- aortic segments was not changed and vascular reactivity to α1-adrenoceptor stimulation was attenuated. These findings highlight the need for more frequent assessment of circulating stress hormone levels and PWV measurements in daily clinical practice for people at risk of AD.NEW & NOTEWORTHY We showed that male amyloid precursor protein (APP) transgenic mice have higher circulating stress hormone levels. As a result, higher systolic blood pressure and pulse-wave velocity were measured in vivo in addition to a smaller α-adrenergic receptor-dependent contraction upon ex vivo stimulation with phenylephrine. Our findings highlight the need for more frequent assessment of circulating stress hormone levels and PWV measurements in daily clinical practice for people at risk of Alzheimer's disease.
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Doença de Alzheimer/sangue , Precursor de Proteína beta-Amiloide/metabolismo , Aorta Torácica/metabolismo , Pressão Arterial , Corticosterona/sangue , Rigidez Vascular , Vasoconstrição , Agonistas de Receptores Adrenérgicos alfa 1/farmacologia , Doença de Alzheimer/genética , Doença de Alzheimer/fisiopatologia , Precursor de Proteína beta-Amiloide/genética , Animais , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/fisiopatologia , Modelos Animais de Doenças , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação , Receptores Adrenérgicos alfa 1/metabolismo , Regulação para Cima , Vasoconstrição/efeitos dos fármacosRESUMO
Endothelial cells (ECs) secrete different paracrine signals that modulate the function of adjacent cells; two examples of these paracrine signals are nitric oxide (NO) and neuregulin-1 (NRG1), a cardioprotective growth factor. Currently, it is undetermined whether one paracrine factor can compensate for the loss of another. Herein, we hypothesized that NRG1 can compensate for endothelial NO synthase (eNOS) deficiency. We characterized eNOS null and wild-type (WT) mice by cardiac ultrasound and histology and we determined circulating NRG1 levels. In a separate experiment, eight groups of mice were divided into four groups of eNOS null mice and WT mice; half of the mice received angiotensin II (ANG II) to induce a more severe phenotype. Mice were randomized to daily injections with NRG1 or vehicle for 28 days. eNOS deficiency increased NRG1 plasma levels, indicating that ECs increase their NRG1 expression when NO production is deleted. eNOS deficiency also increased blood pressure, lowered heart rate, induced cardiac fibrosis, and affected diastolic function. In eNOS null mice, ANG II administration not only increased cardiac fibrosis but also induced cardiac hypertrophy and renal fibrosis. NRG1 administration prevented cardiac and renal hypertrophy and fibrosis caused by ANG II infusion and eNOS deficiency. Moreover, Nrg1 expression in the myocardium is shown to be regulated by miR-134. This study indicates that administration of endothelium-derived NRG1 can compensate for eNOS deficiency in the heart and kidneys.NEW & NOTEWORTHY ECs compensate for eNOS deficiency by increasing the secretion of NRG1. NRG1 administration prevents cardiac and renal hypertrophy and fibrosis caused by ANG II infusion and eNOS deficiency. NRG1 expression is regulated by miR-134.
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Células Endoteliais/metabolismo , Frequência Cardíaca/genética , Coração/efeitos dos fármacos , MicroRNAs/metabolismo , Miocárdio/patologia , Neuregulina-1/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico/metabolismo , Angiotensina II/farmacologia , Animais , Cardiomegalia/genética , Cardiomegalia/patologia , Diástole/efeitos dos fármacos , Fibrose/genética , Fibrose/patologia , Regulação da Expressão Gênica , Frequência Cardíaca/efeitos dos fármacos , Rim/patologia , Camundongos , Camundongos Knockout , Neuregulina-1/farmacologia , Óxido Nítrico Sintase Tipo III/metabolismo , Distribuição Aleatória , Vasoconstritores/farmacologiaRESUMO
Increasing epidemiological evidence highlights the association between systemic insulin resistance and Alzheimer's disease (AD). As insulin resistance can be caused by high-stress hormone levels and since hypercortisolism appears to be an important risk factor of AD, we aimed to investigate the systemic insulin functionality and circulating stress hormone levels in a mutant humanized amyloid precursor protein (APP) overexpressing (hAPP23+/-) AD mouse model. Memory and spatial learning of male hAPP23+/- and C57BL/6 (wild type, WT) mice were assessed by a Morris Water Maze (MWM) test at the age of 4 and 12 months. The systemic metabolism was examined by intraperitoneal glucose and insulin tolerance tests (GTT, ITT). Insulin and corticosterone levels were determined in serum. In the hippocampus, parietal and occipital cortex of hAPP23+/- brains, amyloid-beta (Aß) deposits were present at 12 months of age. MWM demonstrated a cognitive decline in hAPP23+/- mice at 12 but not at 4 months, evidenced by increasing total path lengths and deteriorating probe trials compared to WT mice. hAPP23+/- animals presented increased serum corticosterone levels compared to WT mice at both 4 and 12 months. hAPP23+/- mice exhibited peripheral insulin resistance compared to WT mice at 4 months, which stabilized at 12 months of age. Serum insulin levels were similar between genotypes at 4 months of age but were significantly higher in hAPP23+/- mice at 12 months of age. Peripheral glucose homeostasis remained unchanged. These results indicate that peripheral insulin resistance combined with elevated circulating stress hormone levels could be potential biomarkers of the pre-symptomatic phase of AD.
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Doença de Alzheimer/sangue , Biomarcadores/sangue , Disfunção Cognitiva , Corticosterona/sangue , Resistência à Insulina , Doença de Alzheimer/diagnóstico , Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Precursor de Proteína beta-Amiloide/genética , Animais , Síndrome de Cushing/complicações , Modelos Animais de Doenças , Insulina/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos TransgênicosRESUMO
Arterial media calcification (AMC) is predominantly regulated by vascular smooth muscle cells (VSMCs), which transdifferentiate into pro-calcifying cells. In contrast, there is little evidence for endothelial cells playing a role in the disease. The current study investigates cellular functioning and molecular pathways underlying AMC, respectively by, an ex vivo isometric organ bath set-up to explore the interaction between VSMCs and ECs and quantitative proteomics followed by functional pathway interpretation. AMC development, which was induced in mice by dietary warfarin administration, was proved by positive Von Kossa staining and a significantly increased calcium content in the aorta compared to that of control mice. The ex vivo organ bath set-up showed calcified aortic segments to be significantly more sensitive to phenylephrine induced contraction, compared to control segments. This, together with the fact that calcified segments as compared to control segments, showed a significantly smaller contraction in the absence of extracellular calcium, argues for a reduced basal NO production in the calcified segments. Moreover, proteomic data revealed a reduced eNOS activation to be part of the vascular calcification process. In summary, this study identifies a poor endothelial function, next to classic pro-calcifying stimuli, as a possible initiator of arterial calcification.
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Células Endoteliais/patologia , Túnica Média/efeitos dos fármacos , Calcificação Vascular/induzido quimicamente , Calcificação Vascular/patologia , Varfarina/farmacologia , Animais , Aorta/efeitos dos fármacos , Aorta/metabolismo , Calcificação Fisiológica/efeitos dos fármacos , Cálcio/metabolismo , Transdiferenciação Celular/efeitos dos fármacos , Células Endoteliais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos DBA , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Osteogênese/efeitos dos fármacos , Túnica Média/metabolismo , Túnica Média/patologia , Calcificação Vascular/metabolismoRESUMO
An important physiological role of the aorta is to convert the pulsatile blood flow that originates in the heart to a nearly continuous flow in the peripheral vessels. Previously, we demonstrated that basal, unstimulated nitric oxide (NO) production is more abundant in large as compared with muscular arteries and that it is an important regulator of arterial (aortic) stiffness. Hence, endothelial function and NO bioavailability are important determinants of aortic biomechanics, and mouse models with altered NO signaling might be of interest to investigate the (patho)physiological role of the NO signaling as a dynamic regulator of arterial stiffness. We aimed to characterize the ex vivo biomechanical properties of aortic segments from mice with no (eNOS-/-), normal [wild type (WT)], or high (eNOS-tg) endothelial NO synthase (eNOS) expression. Isobaric aortic diameter and compliance were lower in eNOS-/- mice and increased in eNOS-tg mice as compared with WT mice. Interestingly, these differences remained when NO levels were pharmacologically restored ex vivo, suggesting that they were not merely the result of a lack or excess of the vasodilator effects of NO. Analysis of basal vascular smooth muscle cell tone and the phasic as well as the tonic contraction in response to α1-adrenergic stimulation with phenylephrine revealed that the chronic lack of eNOS expression affected aortic reactivity similarly but with different magnitude as compared with acute eNOS blockade using Nω-nitro-l-arginine methyl ester in WT and eNOS-tg mice, suggesting that chronical distortion of NO signaling triggered several compensatory mechanisms that reflect the organism's attempt to restore the contractile imbalance and maintain optimal central hemodynamics.NEW & NOTEWORTHY Endothelial function and NO bioavailability are important determinants of aortic biomechanics and function. With a new technique we investigated the ex vivo aortic segment biomechanics of different mouse models with altered NO signaling. Our experiments clearly show that chronic distortion of NO signaling triggered several compensatory mechanisms that reflect the organism's attempt to maintain optimal central hemodynamics.
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Aorta/fisiologia , Óxido Nítrico Sintase Tipo III/metabolismo , Rigidez Vascular , Animais , Aorta/metabolismo , Fenômenos Biomecânicos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tono Muscular , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiologia , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Transdução de Sinais , VasoconstriçãoRESUMO
KEY POINTS: Cyclic stretch is known to alter intracellular pathways involved in vessel tone regulation. We developed a novel set-up that allows straightforward characterization of the biomechanical properties of the mouse aorta while stretched at a physiological heart rate (600 beats min-1 ). Active vessel tone was shown to have surprisingly large effects on isobaric stiffness. The effect of structural vessel wall alterations was confirmed using a genetic mouse model. This set-up will contribute to a better understanding of how active vessel wall components and mechanical stimuli such as stretch frequency and amplitude regulate aortic mechanics. ABSTRACT: Cyclic stretch is a major contributor to vascular function. However, isolated mouse aortas are frequently studied at low stretch frequency or even in isometric conditions. Pacing experiments in rodents and humans show that arterial compliance is stretch frequency dependent. The Rodent Oscillatory Tension Set-up to study Arterial Compliance is an in-house developed organ bath set-up that clamps aortic segments to imposed preloads at physiological rates up to 600 beats min-1 . The technique enables us to derive pressure-diameter loops and assess biomechanical properties of the segment. To validate the applicability of this set-up we aimed to confirm the effects of distension pressure and vascular smooth muscle tone on arterial stiffness. At physiological stretch frequency (10 Hz), the Peterson modulus (EP ; 293 (10) mmHg) for wild-type mouse aorta increased 22% upon a rise in pressure from 80-120 mmHg to 100-140 mmHg, while, at normal pressure, EP increased 80% upon maximal contraction of the vascular smooth muscle cells. We further validated the method using a mouse model with a mutation in the fibrillin-1 gene and an endothelial nitric oxide synthase knock-out model. Both models are known to have increased arterial stiffness, and this was confirmed using the set-up. To our knowledge, this is the first set-up that facilitates the study of biomechanical properties of mouse aortic segments at physiological stretch frequency and pressure. We believe that this set-up can contribute to a better understanding of how cyclic stretch frequency, amplitude and active vessel wall components influence arterial stiffening.
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Aorta/fisiologia , Contração Muscular , Técnicas de Cultura de Órgãos/métodos , Amplificadores Eletrônicos , Animais , Fenômenos Biomecânicos , Camundongos , Camundongos Endogâmicos C57BL , Miografia/instrumentação , Miografia/métodos , Técnicas de Cultura de Órgãos/instrumentaçãoRESUMO
The mechanisms by which physical activity affects cardiovascular function and physiology are complex and multifactorial. In the present study, cardiac output during rest or acute physical activity was simulated in isolated aortic segments of healthy C57BL/6J wild-type mice. This was performed using the Rodent Oscillatory Tension Set-up to study Arterial Compliance (ROTSAC) by applying cyclic stretch of different amplitude, duration and frequency in well-controlled and manageable experimental conditions. Our data show that vascular smooth muscle cells (VSMCs) of the aorta have the intrinsic ability to "de-stiffen" or "relax" after periods of high cyclic stretch and to "re-stiffen" slowly thereafter upon return to normal distension pressures. Thereby, certain conditions have to be fulfilled: 1) VSMC contraction and repetitive stretching (loading/unloading cycles) are a prerequisite to induce post-exercise de-stiffening; 2) one bout of high cyclic stretch is enough to induce de- and re-stiffening. Aortic de-stiffening was highly dependent on cyclic stretch amplitude and on the manner and timing of contraction with probable involvement of focal adhesion phosphorylation/activation. Results of this study may have implications for the therapeutic potential of regular and acute physical activity and its role in the prevention and/or treatment of cardiovascular disease.
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Aorta , Miócitos de Músculo Liso , Camundongos , Animais , Pressão Sanguínea , Camundongos Endogâmicos C57BL , PressãoRESUMO
In vivo angiotensin II (AngII)-treatment is a widely used experimental model to induce cardiovascular disease and results in a high likelihood of abdominal aorta aneurysm (AAA) formation. This involves progressive and irreversible focal dilation of the abdominal aorta and induces adverse aortic connective tissue remodeling contributing to aortic wall stiffening through inflammation, elastin degradation, and collagen restructuring. Hence, the present study aimed to investigate how AAA formation in AngII-treated mice affects aortic function and biomechanics. To this end, C57Bl/6J mice were treated with AngII (1000 ng/[kg.min]) or PBS infusion for 28 days. Peripheral blood pressure, echocardiography, and aortic pulse wave velocity were measured in vivo. Thoracic aorta rings were studied ex vivo in organ chambers, while aortic vascular smooth muscle cell (VSMC) phenotype was investigated histologically. We confirmed peripheral hypertension, cardiac hypertrophy, aortic stiffening, and increased VSMC proliferation and migration after AngII-treatment. Abdominal aorta aneurysm formation was observed in 8/13 AngII-treated mice. Ex vivo thoracic aortic rings of both aneurysmal and non-aneurysmal AngII-treated mice showed high isobaric aortic stiffness, endothelial dysfunction, heightened α1 -adrenergic contractility, and altered VSMC contractile calcium signaling. However, aortic biomechanics were differently affected, with heightened α1 -adrenoreceptor mediated aortic stiffening in non-aneurysmal mice, whereas contraction-dependent stiffening was impaired in aneurysmal mice. In conclusion, although aneurysmal and non-aneurysmal 4-week AngII-treated mice displayed similar changes in aortic physiology, aortic biomechanics were dissimilarly affected.
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Angiotensina II , Aneurisma da Aorta Abdominal , Adrenérgicos/metabolismo , Angiotensina II/metabolismo , Animais , Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/metabolismo , Colágeno/metabolismo , Elastina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Análise de Onda de PulsoRESUMO
Introduction and Aims: Endothelial nitric oxide synthase (eNOS) knockout mice develop pronounced cardiovascular disease. In the present study, we describe the alterations in aortic physiology and biomechanics of eNOS knockout and C57Bl/6 control mice at 2-12 months of age, including a thorough physiological investigation of age and cyclic stretch-dependent VSMC contractility and aortic stiffness. Methods and Results: Peripheral blood pressure and aortic pulse wave velocity were measured in vivo, and aortic biomechanical studies and isometric contractions were investigated ex vivo. Age-dependent progression of aortic stiffness, peripheral hypertension, and aortic contractility in eNOS knockout mice was absent, attenuated, or similar to C57Bl/6 control mice. Voltage-gated calcium channel (VGCC)-dependent calcium influx inversely affected isometric contraction and aortic stiffening by α1-adrenergic stimulation in eNOS knockout mice. Baseline aortic stiffness was selectively reduced in eNOS knockout mice after ex vivo cyclic stretch exposure in an amplitude-dependent manner, which prompted us to investigate cyclic stretch dependent regulation of aortic contractility and stiffness. Aortic stiffness, both in baseline conditions and after activation of vascular smooth muscle cell (VSMC) contraction, was reduced with increasing cyclic stretch amplitude. This cyclic stretch dependency was attenuated with age, although aged eNOS knockout mice displayed better preservation of cyclic stretch-dependency compared to C57Bl/6 control mice. Store operated calcium entry-medicated aortic stiffening as induced by inhibiting sarcoplasmic reticulum calcium ATPase pumps with 10 µM CPA was most pronounced in the aorta of aged mice and at low cyclic stretch amplitude, but independent of eNOS. Basal aortic tonus and VSMC depolarization were highly dependent on eNOS, and were most pronounced at low cyclic stretch, with attenuation at increasing cyclic stretch amplitude. Conclusion: eNOS knockout mice display attenuated progression of arterial disease as compared to C57Bl/6 control mice. Basal VSMC tone in eNOS knockout mice could be reduced by ex vivo exposure to cyclic stretch through stretch-dependent regulation of cytosolic calcium. Both baseline and active aortic stiffness were highly dependent on cyclic stretch regulation, which was more pronounced in young versus aged mice. Other mediators of VSMC contraction and calcium handling were dependent on cyclic stretch mechanotransduction, but independent of eNOS.
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Introduction and Aims: Endothelial dysfunction is recognized as a cardiovascular aging hallmark. Administration of nitric oxide synthase blocker N-Ω-Nitro-L-arginine methyl ester hydrochloride (L-NAME) constitutes a well-known small animal model of cardiovascular aging. Despite extensive phenotypic characterization, the exact aortic function changes in L-NAME treated mice are largely unknown. Therefore, this study presents a longitudinal characterization of the aortic reactivity and biomechanical alterations in L-NAME treated C57Bl/6 mice. Methods and Results: Male C57Bl/6 mice were treated with L-NAME (0.5 mg/ml drinking water) for 1, 2, 4, 8, or 16 weeks. Peripheral blood pressure measurement (tail-cuff) and transthoracic echocardiograms were recorded, showing progressive hypertension after 4 weeks of treatment and progressive cardiac hypertrophy after 8-16 weeks of treatment. Aortic stiffness was measured in vivo as aortic pulse wave velocity (aPWV, ultrasound) and ex vivo as Peterson modulus (Ep). Aortic reactivity and biomechanics were investigated ex vivo in thoracic aortic rings, mounted isometrically or dynamically-stretched in organ bath set-ups. Aortic stiffening was heightened in L-NAME treated mice after all treatment durations, thereby preceding the development of hypertension and cardiac aging. L-NAME treatment doubled the rate of arterial stiffening compared to control mice, and displayed an attenuation of the elevated aortic stiffness at high distending pressure, possibly due to late-term reduction of medial collagen types I, III, and IV content. Remarkably, endothelial dysfunction, measured by acetylcholine concentration-response stimulation in precontracted aortic rings, was only observed after short-term (1-4 weeks) treatment, followed by restoration of endothelial function which coincided with increased phosphorylation of endothelial nitric oxide synthase (S1177). In the late-disease phase (8-16 weeks), vascular smooth muscle cell (VSMC) dysfunction developed, including increased contribution of voltage-dependent calcium channels (assessed by inhibition with diltiazem), basal VSMC cytoplasmic calcium loading (assessed by removal of extracellular calcium), and heightened intracellular contractile calcium handling (assessed by measurement of sarcoplasmic reticulum-mediated transient contractions). Conclusion: Arterial stiffness precedes peripheral hypertension and cardiac hypertrophy in chronic L-NAME treated male C57Bl/6 mice. The underlying aortic disease mechanisms underwent a distinct shift from early endothelial dysfunction to late-term VSMC dysfunction, with continued disease progression.
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During the last decades, most of the preclinical neurodegenerative research was performed in mouse models of amyloidosis, tauopathies or α-synucleinopathies preferentially maintained on a C57BL/6J background. However, comprehensive neurobehavioural data from C57BL/6J mice outlining the critical point of spontaneous cognitive decline are incomplete. In this study, we aimed for the neurobehavioural phenotyping of hippocampus-dependent spatial learning and memory of aging C57BL/6J mice. Neurobehavioural phenotyping was performed by means of a Morris Water Maze (MWM) and a Novel Object Recognition (NOR) test. MWM measurements revealed signs of age-related memory loss in C57BL/6J animals from the age of 6 months onward. The NOR assessment strengthened latter finding by decreasing discrimination indexes (DI) and recognition indexes (RI) starting from the age of 6 months. Taken together, these findings contribute to the current knowledge of spontaneous cognitive behaviours of this perhaps most widely used mouse strain and serve as a benchmark for dementia mouse models to distinguish spontaneous from pathological neurodegenerative behaviour.
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Envelhecimento/fisiologia , Disfunção Cognitiva/fisiopatologia , Memória/fisiologia , Camundongos Endogâmicos C57BL , Aprendizagem Espacial/fisiologia , Animais , Modelos Animais de Doenças , Hipocampo , Masculino , Camundongos , Teste do Labirinto Aquático de Morris , Teste de Campo AbertoRESUMO
Aortic stiffness is a hallmark of cardiovascular disease, but its pathophysiology remains incompletely understood. This study presents an in-dept characterization of aortic aging in male C57Bl/6 mice (2-24 months). Cardiovascular measurements include echocardiography, blood pressure measurement, and ex vivo organ chamber experiments. In vivo and ex vivo aortic stiffness increases with age, and precede the development of cardiac hypertrophy and peripheral blood pressure alterations. Contraction-independent stiffening (due to extracellular matrix changes) is pressure-dependent. Contraction-dependent aortic stiffening develops through heightened α1-adrenergic contractility, aberrant voltage-gated calcium channel function, and altered vascular smooth muscle cell calcium handling. Endothelial dysfunction is limited to a modest decrease in sensitivity to acetylcholine-induced relaxation with age. Our findings demonstrate that progressive arterial stiffening in C57Bl/6 mice precedes associated cardiovascular disease. Aortic aging is due to changes in extracellular matrix and vascular smooth muscle cell signalling, and not to altered endothelial function.
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Doenças Cardiovasculares , Rigidez Vascular , Envelhecimento , Animais , Matriz Extracelular , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular , Rigidez Vascular/fisiologiaRESUMO
The physiology of vascular smooth muscle (VSMC) cells is affected by autophagy, a catabolic cellular mechanism responsible for nutrient recycling. Autophagy-inducing compounds may reverse arterial stiffening, whereas congenital VSMC-specific autophagy deficiency promotes arterial stiffening. The elevated aortic stiffness in 3.5-month-old C57Bl/6 mice, in which the essential autophagy-related gene Atg7 was specifically deleted in the VSMCs (Atg7F/F SM22α-Cre+ mice) was mainly due to passive aortic wall remodeling. The present study investigated whether aortic stiffness was also modulated by a shorter duration of autophagy deficiency. Therefore, aortic segments of 2-month-old Atg7F/F SM22α-Cre+ mice were studied. Similarly to the older mice, autophagy deficiency in VSMCs promoted aortic stiffening by elastin degradation and elastin breaks, and increased the expression of the calcium binding protein S100A4 (+ 157%), the aortic wall thickness (+ 27%), the sensitivity of the VSMCs to depolarization and the contribution of VGCC mediated Ca2+ influx to α1 adrenergic contractions. Hence, all these phenomena occurred before the age of 2 months. When compared to autophagy deficiency in VSMCs at 3.5 months, shorter term autophagy deficiency led to higher segment diameter at 80 mmHg (+ 7% versus - 2%), normal baseline tonus (versus increased), unchanged IP3-mediated phasic contractions (versus enhanced), and enhanced endothelial cell function (versus normal). Overall, and because in vivo cardiac parameters or aortic pulse wave velocity were not affected, these observations indicate that congenital autophagy deficiency in VSMCs of Atg7F/F SM22α-Cre+ mice initiates compensatory mechanisms to maintain circulatory homeostasis.
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Músculo Liso Vascular , Análise de Onda de Pulso , Animais , Autofagia/fisiologia , Fenômenos Biomecânicos , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/metabolismoRESUMO
Aim: Cyclic stretch of vascular tissue at any given pressure reveals greater dimensions during unloading than during loading, which determines the cardiac beat-by-beat hysteresis loop on the pressure-diameter/volume relationship. The present study did not focus on hysteresis during a single stretch cycle but investigated whether aortic stiffness determined during continuous stretch at different pressures also displayed hysteresis phenomena. Methods: Aortic segments from C57Bl6 mice were mounted in the Rodent Oscillatory Set-up for Arterial Compliance (ROTSAC), where they were subjected to high frequency (10 Hz) cyclic stretch at alternating loads equivalent to a constant theoretical pulse pressure of 40 mm Hg. Diastolic and systolic diameter, compliance, and the Peterson elastic modulus (Ep), as a measure of aortic stiffness, was determined starting at cyclic stretch between alternating loads corresponding to 40 and 80 mm Hg, at each gradual load increase equivalent to 20 mm Hg, up to loads equivalent to pressures of 220 and 260 mm Hg (loading direction) and then repeated in the downward direction (unloading direction). This was performed in baseline conditions and following contraction by α1 adrenergic stimulation with phenylephrine or by depolarization with high extracellular K+ in aortas of young (5 months), aged (26 months) mice, and in segments treated with elastase. Results: In baseline conditions, diastolic/systolic diameters and compliance for a pulse pressure of 40 mm Hg were larger at any given pressure upon unloading (decreasing pressure) than loading (increasing pressure) of the aortic segments. The pressure-aortic stiffness (Ep) relationship was similar in the loading and unloading directions, and aortic hysteresis was absent. On the other hand, hysteresis was evident after activation of the VSMCs with the α1 adrenergic agonist phenylephrine and with depolarization by high extracellular K+, especially after inhibition of basal NO release with L-NAME. Aortic stiffness was significantly smaller in the unloading than in the loading direction. In comparison with young mice, old-mouse aortic segments also displayed contraction-dependent aortic hysteresis, but hysteresis was shifted to a lower pressure range. Elastase-treated segments showed higher stiffness upon unloading over nearly the whole pressure range. Conclusions: Mouse aortic segments display pressure- and contraction-dependent diameter, compliance, and stiffness hysteresis phenomena, which are modulated by age and VSMC-extracellular matrix interactions. This may have implications for aortic biomechanics in pathophysiological conditions and aging.
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GSK-7975A is described to inhibit stromal interaction molecule 1(STIM1)-mediated Ca2+ release-activated Ca2+ channels ORAI 1, ORAI 2 and ORAI 3 in different cell types. The present study investigated whether isometric contractions of mouse aortic segments were affected by this selective store-operated calcium channel inhibitor. Depending on the way by which Ca2+ influx pathways were activated during contraction, GSK-7975A inhibited contractility of mouse aortic segments with different affinity. When contractile effects were induced by depolarization as with elevated extracellular K+ and opening of voltage-gated calcium channels, the affinity was approximately 10 times lower than when contraction was elicited with Ca2+ influx via non-selective cation channels. GSK-7975A may repolarize the aortic smooth muscle cells by inhibiting non-selective cation channels, has no effect on IP3-mediated phenylephrine-induced phasic contractions or on refilling of the contractile sarcoplasmic reticulum Ca2+ store, but has significant effects on non-contractile store-operated Ca2+ influx.
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Aorta/efeitos dos fármacos , Benzamidas/farmacologia , Canais de Cálcio Ativados pela Liberação de Cálcio/antagonistas & inibidores , Contração Isométrica/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Pirazóis/farmacologia , Animais , Aorta/fisiologia , Cálcio/metabolismo , Camundongos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Retículo Sarcoplasmático/efeitos dos fármacos , Retículo Sarcoplasmático/metabolismoRESUMO
Increasing epidemiological and experimental evidence points to a link between arterial stiffness and rapid cognitive decline. However, the underlying mechanism linking the two diseases is still unknown. The importance of nitric oxide synthases in both diseases is well-defined. In this study, we introduced arterial stiffness in both genetic (eNOS-/-, endothelial nitric oxide synthase knockout) and pharmacological (N(G)-nitro-L-arginine methyl ester (L-NAME) treatment) NO dysfunction models to study their association with cognitive decline. Our findings demonstrate that the non-selective inhibition of NOS activity with L-NAME induces cardiac dysfunction, arterial stiffness, and a decline in hippocampal-dependent learning and memory. This outcome demonstrates the importance of neuronal NOS (nNOS) in both cardiovascular and neurological pathophysiology and its potential contribution in the convergence between arterial stiffness and cognitive decline.
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Autophagy is an important cellular survival process that enables degradation and recycling of defective organelles and proteins to maintain cellular homeostasis. Hence, defective autophagy plays a role in many age-associated diseases, such as atherosclerosis, arterial stiffening and hypertension. Recently, we showed in mice that autophagy in vascular smooth muscle cells (VSMCs) of large elastic arteries such as the aorta is important for Ca2+ mobilization and vascular reactivity. Whether autophagy plays a role in the smaller muscular arteries, such as the femoral artery, and thereby contributes to for example, blood pressure regulation is currently unknown. Therefore, we determined vascular reactivity of femoral artery segments of mice containing a VSMC specific deletion of the essential autophagy gene Atg7 (Atg7F/F SM22α-Cre+) and compared them to femoral artery segments of corresponding control mice (Atg7+/+ SM22α-Cre+). Our results indicate that similar to the aorta, femoral artery segments showed enhanced contractility. Specifically, femoral artery segments of Atg7F/F SM22α-Cre+ mice showed an increase in phasic phenylephrine (PE) induced contractions, together with an enhanced sensitivity to depolarization induced contractions. In addition, and importantly, VSMC sensitivity to exogenous nitric oxide (NO) was significantly increased in femoral artery segments of Atg7F/F SM22α-Cre+ mice. Notwithstanding the fact that small artery contractility is a significant pathophysiological determinant for the development of hypertension, 7 days of treatment with angiotensin II (AngII), which increased systolic blood pressure in control mice, was ineffective in Atg7F/F SM22α-Cre+ mice. It is likely that this was due to the increased sensitivity of VSMCs to NO in the femoral artery, although changes in the heart upon AngII treatment were also present, which could also be (partially) accountable for the lack of an AngII-induced rise in blood pressure in Atg7F/F SM22α-Cre+ mice. Overall, our study indicates that apart from previously shown effects on large elastic arteries, VSMC autophagy also plays a pivotal role in the regulation of the contractile and relaxing properties of the smaller muscular arteries. This may suggest a role for autophagy in vascular pathologies, such as hypertension and arterial stiffness.
RESUMO
Over the past few decades, isometric contraction studies of isolated thoracic aorta segments have significantly contributed to our overall understanding of the active, contractile properties of aortic vascular smooth muscle cells (VSMCs) and their cross-talk with endothelial cells. However, the physiological role of VSMC contraction or relaxation in the healthy aorta and its contribution to the pulse-smoothening capacity of the aorta is currently unclear. Therefore, we investigated the acute effects of VSMC contraction and relaxation on the isobaric biomechanical properties of healthy mouse aorta. An in-house developed set-up was used to measure isobaric stiffness parameters of periodically stretched (10 Hz) aortic segments at an extended pressure range, while pharmacologically modulating VSMC tone and endothelial cell function. We found that the effects of α1-adrenergic stimulation with phenylephrine on the pressure-stiffness relationship varied in sensitivity, magnitude and direction, with the basal, unstimulated NO production by the endothelium playing a pivotal role. We also investigated how arterial disease affected this system by using the angiotensin-II-treated mouse. Our results show that isobaric stiffness was increased and that the aortic segments demonstrated a reduced capacity for modulating the pressure-stiffness relationship. This suggests that not only increased isobaric stiffness at normal pressure, but also a reduced capacity of the VSMCs to limit the pressure-associated increase in aortic stiffness, may contribute to the pathogenesis of this mouse model. Overall, this study provides more insight in how aortic VSMC tone affects the pressure-dependency of aortic biomechanics at different physiological and pathological conditions.
Assuntos
Aorta/fisiologia , Relaxamento Muscular , Músculo Liso Vascular/fisiologia , Rigidez Vascular , Vasoconstrição , Angiotensina II/farmacologia , Animais , Aorta/citologia , Aorta/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tono Muscular , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/fisiologiaRESUMO
Induction of hypertension by angiotensin II (AngII) is a widely used experimental stimulus to study vascular aging in mice. It is associated with large artery stiffness, a hallmark of arterial aging and a root cause of increased cardiovascular risk. We reported earlier that long term (4 week) AngII treatment in mice altered the active, contractile properties of the arteries in a vascular bed-specific manner and that, in healthy mice aorta, active contractile properties of the aortic wall determine isobaric aortic stiffness. Given the huge physiological relevance of large artery stiffening, we aimed to characterize the early (1 week) changes in the active properties of the aorta of AngII-treated mice. We were not able to detect a significant effect of AngII treatment on anesthetized blood pressure or abdominal aorta pulse wave velocity. Ex vivo biomechanical and functional studies of the aorta revealed increased arterial stiffness and altered vascular smooth muscle cell (VSMC) and endothelial cell reactivity. Interestingly, the AngII-associated changes in the aorta could be largely attributed to alterations in basal VSMC tone and basal nitric oxide efficacy, indicating that, besides structural remodeling of the arterial wall, dysfunctional active components of the aorta play a crucial role in the pathophysiological mechanisms by which AngII treatment induces arterial stiffness.