Progressive aortic stiffness in aging C57Bl/6 mice displays altered contractile behaviour and extracellular matrix changes.
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ABSTRACT:
SUBMITTER: De Moudt S
PROVIDER: S-EPMC9203497 | biostudies-literature | 2022 Jun
REPOSITORIES: biostudies-literature
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Progressive aortic stiffness in aging C57Bl/6 mice displays altered contractile behaviour and extracellular matrix changes.
De Moudt Sofie S Hendrickx Jhana O JO Neutel Cédric C De Munck Dorien D Leloup Arthur A De Meyer Guido R Y GRY Martinet Wim W Fransen Paul P
Communications biology 20220617 1
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-i ...[more]
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Project description:BackgroundArrhythmogenic cardiomyopathy (ACM) is an inherited genetic disorder of desmosomal dysfunction, and PKP2 (plakophilin-2) has been reported to be the most common disease-causing gene when mutation-positive. In the early concealed phase, the ACM heart is at high risk of sudden cardiac death before cardiac remodeling occurs because of mistargeted ion channels and altered Ca2+ handling. However, the results of pathogenic PKP2 variants on myocyte contraction in ACM pathogenesis remain unknown.MethodsWe studied the outcomes of a human truncating variant of PKP2 on myocyte contraction using a novel knock-in mouse model with insertion of thymidine in exon 5 of Pkp2, which mimics a familial case of ACM (PKP2-L404fsX5). We used serial echocardiography, electrocardiography, blood pressure measurements, histology, cardiomyocyte contraction, intracellular calcium measurements, and gene and protein expression studies.ResultsSerial echocardiography of Pkp2 heterozygous (Pkp2-Het) mice revealed progressive failure of the right ventricle (RV) in animals older than 3 months. By contrast, left ventricular function remained normal. ECGs of 6-month-old anesthetized Pkp2-Het mice showed normal baseline heart rates and QRS complexes. Cardiac responses to β-adrenergic agonist isoproterenol (2 mg/kg) plus caffeine (120 mg/kg) were also normal. However, adrenergic stimulation enhanced the susceptibility of Pkp2-Het hearts to tachyarrhythmia and sudden cardiac death. Histological staining showed no significant fibrosis or adipocyte infiltration in the RVs and left ventricles of 6- and 12-month-old Pkp2-Het hearts. Contractility assessment of isolated myocytes demonstrated progressively reduced Pkp2-Het RV cardiomyocyte function consistent with RV failure measured by echocardiography. However, aging Pkp2-Het and control RV myocytes loaded with intracellular Ca2+ indicator Fura-2 showed comparable Ca2+ transients. Western blotting of Pkp2-RV homogenates revealed a 40% decrease in actin, whereas actin immunoprecipitation followed by a 2,4-dinitrophenylhydrazine staining showed doubled oxidation level. This correlated with a 39% increase in troponin-I phosphorylation. In contrast, Pkp2-Het left ventricular myocytes had normal contraction, actin expression and oxidation, and troponin-I phosphorylation. Last, Western blotting of cardiac biopsies revealed that actin expression was 40% decreased in RVs of patients with end-stage ACM.ConclusionsDuring the early concealed phase of ACM, reduced actin expression drives loss of RV myocyte contraction, contributing to progressive RV dysfunction.
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Project description: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.
| S-EPMC9254682 | biostudies-literature
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| S-EPMC4808390 | biostudies-literature
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| S-EPMC11892926 | biostudies-literature
Project description:This study utilized a standard rat model of aging and global gene expression analyses to attempt to identify the most appropriate time points to study vascular aging and to identify molecules associated with the development of pathology. Recent studies have established that age is the major risk factor for vascular disease. Numerous changes occur in vascular structure and function during aging, and animal models, including rodents, are the primary means to determine the underlying mechanisms of age-mediated vascular pathology. This study utilized a standard rat model of aging and both morphometric and global gene expression analyses to attempt to identify the most appropriate time points to study vascular aging and to identify molecules associated with the development of pathology. Morphometric analysis of Fischer 344/Brown Norway F1 hybrid (F344xBN) rat thoracic aorta indicated, in contrast to some previous studies, that progressive increases in intimal and medial thickness, as well as smooth muscle cell-containing intimal protrusions, occurred with age. This structural vascular pathology was associated with a progressive increase in global differential gene expression. Specific molecules with altered mRNA and protein expression included the adhesion molecules ICAM-1 and VCAM-1, and the bone morphogenic proteins osteopontin and bone sialoprotein-1. Intimal-associated macrophages were detected with immunohistochemistry in paraffin sections of thoracic aorta and were found to increase significantly in number with age. Both systemic and tissue markers of oxidant stress, serum 8-isoprostane and 3-nitrotyrosine, respectively, were found to increase during aging. The results demonstrate that increased inflammation and oxidant stress occur progressively during vascular aging, and suggest therapeutic targets to limit or reverse aging-associated vascular pathology and dysfunction. Experiment Overall Design: Samples were taken from rats at 4 ages, 3mo, 6mo, 15mo and 28 mo, with 4 replicates in each age group.
2008-06-15 | E-GEOD-7281 | biostudies-arrayexpress
Project description:BACKGROUND: Previous genomic studies with human tissues have compared differential gene expression between 2 conditions (ie, normal versus diseased) to identify altered gene expression in a binary manner; however, a potentially more informative approach is to correlate the levels of gene expression with quantitative physiological parameters. METHODS AND RESULTS: In this study, we have used this approach to examine genes whose expression correlates with arterial stiffness in human aortic specimens. Our data identify 2 distinct groups of genes, those associated with cell signaling and those associated with the mechanical regulation of vascular structure (cytoskeletal-cell membrane-extracellular matrix). Although previous studies have concentrated on the contribution of the latter group toward arterial stiffness, our data suggest that changes in expression of signaling molecules play an equally important role. Alterations in the profiles of signaling molecules could be involved in the regulation of cell cytoskeletal organization, cell-matrix interactions, or the contractile state of the cell. CONCLUSIONS: Although the influence of smooth muscle contraction/relaxation on arterial stiffness could be controversial, our provocative data would suggest that further studies on this subject are indicated. Keywords: other
2003-08-28 | GSE420 | GEO
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Project description:BackgroundVascular calcification and increased extracellular matrix (ECM) stiffness are hallmarks of vascular aging. Sox9 (SRY-box transcription factor 9) has been implicated in vascular smooth muscle cell (VSMC) osteo/chondrogenic conversion; however, its relationship with aging and calcification has not been studied.MethodsImmunohistochemistry was performed on human aortic samples from young and aged patients. Young and senescent primary human VSMCs were induced to produce ECM, and Sox9 expression was manipulated using adenoviral overexpression and depletion. ECM properties were characterized using atomic force microscopy and proteomics, and VSMC phenotype on hydrogels and the ECM were examined using confocal microscopy.ResultsIn vivo, Sox9 was not spatially associated with vascular calcification but correlated with the senescence marker p16 (cyclin-dependent kinase inhibitor 2A). In vitro Sox9 showed mechanosensitive responses with increased expression and nuclear translocation in senescent cells and on stiff matrices. Sox9 was found to regulate ECM stiffness and organization by orchestrating changes in collagen (Col) expression and reducing VSMC contractility, leading to the formation of an ECM that mirrored that of senescent cells. These ECM changes promoted phenotypic modulation of VSMCs, whereby senescent cells plated on ECM synthesized from cells depleted of Sox9 returned to a proliferative state, while proliferating cells on a matrix produced by Sox9 expressing cells showed reduced proliferation and increased DNA damage, reiterating features of senescent cells. LH3 (procollagen-lysine, 2-oxoglutarate 5-dioxygenase 3) was identified as an Sox9 target and key regulator of ECM stiffness. LH3 is packaged into extracellular vesicles and Sox9 promotes extracellular vesicle secretion, leading to increased LH3 deposition within the ECM.ConclusionsThese findings highlight the crucial role of ECM structure and composition in regulating VSMC phenotype. We identify a positive feedback cycle, whereby cellular senescence and increased ECM stiffening promote Sox9 expression, which, in turn, drives further ECM modifications to further accelerate stiffening and senescence.
| S-EPMC10826924 | biostudies-literature
Project description: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.
| S-EPMC8507434 | biostudies-literature
Project description:This study utilized a standard rat model of aging and global gene expression analyses to attempt to identify the most appropriate time points to study vascular aging and to identify molecules associated with the development of pathology. Recent studies have established that age is the major risk factor for vascular disease. Numerous changes occur in vascular structure and function during aging, and animal models, including rodents, are the primary means to determine the underlying mechanisms of age-mediated vascular pathology. This study utilized a standard rat model of aging and both morphometric and global gene expression analyses to attempt to identify the most appropriate time points to study vascular aging and to identify molecules associated with the development of pathology. Morphometric analysis of Fischer 344/Brown Norway F1 hybrid (F344xBN) rat thoracic aorta indicated, in contrast to some previous studies, that progressive increases in intimal and medial thickness, as well as smooth muscle cell-containing intimal protrusions, occurred with age. This structural vascular pathology was associated with a progressive increase in global differential gene expression. Specific molecules with altered mRNA and protein expression included the adhesion molecules ICAM-1 and VCAM-1, and the bone morphogenic proteins osteopontin and bone sialoprotein-1. Intimal-associated macrophages were detected with immunohistochemistry in paraffin sections of thoracic aorta and were found to increase significantly in number with age. Both systemic and tissue markers of oxidant stress, serum 8-isoprostane and 3-nitrotyrosine, respectively, were found to increase during aging. The results demonstrate that increased inflammation and oxidant stress occur progressively during vascular aging, and suggest therapeutic targets to limit or reverse aging-associated vascular pathology and dysfunction. Keywords: time course
2007-10-29 | GSE7281 | GEO