elastin and Hypertension--Pulmonary

Increased elastin production and accumulation is a rapid and sensitive response to elevated vascular wall stress in both systemic and pulmonary hypertension. While initially protecting the vessel wall, these structural changes may in the longer term result in reinforcement of the hypertensive state and contribute to the persistence of the pathology of hypertension. Rapid responses apparently uncorrelated with increased elastin mRNA, at least in the case of systemic vessels, suggest novel mechanisms perhaps including increased efficiency of message translation or matrix accumulation of the protein. Investigations using in vitro organ and cell culture models have indicated a role for phospholipases and protein kinases, including protein kinase C, in stretch-induced elastin synthesis. In addition, tyrosine phosphorylation of membrane/sub-membrane/cytoskeletal sensors, including focal adhesion kinase and members of the lipocortin family, have been shown to be important in this transduction mechanism. Because its turnover is normally very slow, additional vascular elastin accumulated during hypertensive episodes, together with its consequences for the physical properties of the vessel wall, may persist long after blood pressure is restored to normal levels. Thus, recent interest has been drawn to the possibility of achieving regression of accumulated matrix elastin by promoting turnover of this protein through activation of endogenous vascular elastase and collagenase activities.

Topics: Animals; Blood Pressure; Blood Vessels; Connective Tissue; Elastin; Humans; Hypertension; Hypertension, Pulmonary

Pulmonary hypertension worsens outcome in left heart disease. Stiffening of the pulmonary artery may drive this pathology by increasing right ventricular dysfunction and lung vascular remodeling. Here we show increased stiffness of pulmonary arteries from patients with left heart disease that correlates with impaired pulmonary hemodynamics. Extracellular matrix remodeling in the pulmonary arterial wall, manifested by dysregulated genes implicated in elastin degradation, precedes the onset of pulmonary hypertension. The resulting degradation of elastic fibers is paralleled by an accumulation of fibrillar collagens. Pentagalloyl glucose preserves arterial elastic fibers from elastolysis, reduces inflammation and collagen accumulation, improves pulmonary artery biomechanics, and normalizes right ventricular and pulmonary hemodynamics in a rat model of pulmonary hypertension due to left heart disease. Thus, targeting extracellular matrix remodeling may present a therapeutic approach for pulmonary hypertension due to left heart disease.

Topics: Animals; Biomechanical Phenomena; Elastin; Heart Diseases; Humans; Hypertension, Pulmonary; Pulmonary Artery; Rats

Pulmonary hypertension (PH) associated with congenital heart disease is a progressive hemodynamic disease that can lead to increased pulmonary vascular resistance, vascular remodeling, and even right heart failure and death. LF3 is a novel inhibitor of the reporter gene activity of β-catenin/TCF4 interaction in the Wnt/β-catenin signal pathway. However, whether this action of LF3 can prevent PH development remains unclear. In this study, we investigated the therapeutic effect of LF3 in rat primary pulmonary artery smooth muscle cells (PASMCs) of the PH model. We found that LF3 inhibited the decrease in pulmonary artery acceleration time and ejection time by ultra-high-resolution ultrasound imaging and blocked the increase of pulmonary artery systolic pressure by using the BL420 biological function experimental system and right ventricular hypertrophy index by the electronic scales. Simultaneously, it prevented the increase of α-smooth muscle actin and fibronectin and the decrease of elastin in pulmonary arteries of rats in the PH group, as revealed by an immunohistochemical analysis. Moreover, cell proliferation and migration assays showed that LF3 significantly reduced the proliferation and migration of PASMCs. Western blotting and quantitative real-time polymerase chain reaction analyses revealed that LF3 suppressed the expression of proliferating cell nuclear antigens and Bcl-2 and increased the expression of Bax but did not alter the expressions of β-catenin and TCF4. Taken together, LF3 can reduce the migration and proliferation of PASMCs and induce their apoptosis to prevent the development of PH. It would be worthwhile to explore the potential use of LF3 in the treatment of PH.

Topics: Actins; Animals; Apoptosis; Benzenesulfonamides; beta Catenin; Cell Movement; Cell Proliferation; Cells, Cultured; Disease Models, Animal; Elastin; Fibronectins; Hemodynamics; Hypertension, Pulmonary; Male; Myocytes, Smooth Muscle; Proliferating Cell Nuclear Antigen; Pulmonary Artery; Rats, Sprague-Dawley; Sulfonamides; Transcription Factor 4; Vascular Remodeling; Wnt Signaling Pathway

A critical event in the adaptation to extrauterine life is relaxation of the pulmonary vasculature at birth, allowing for a rapid increase in pulmonary blood flow that is essential for efficient gas exchange. Failure of this transition leads to pulmonary hypertension (PH), a major cause of newborn mortality associated with preterm birth, infection, hypoxia, and malformations including congenital diaphragmatic hernia (CDH). While individual vasoconstrictor and dilator genes have been identified, the coordination of their expression is not well understood. Here, we found that lung mesenchyme-specific deletion of CDH-implicated genes encoding pre-B cell leukemia transcription factors (Pbx) led to lethal PH in mice shortly after birth. Loss of Pbx genes resulted in the misexpression of both vasoconstrictors and vasodilators in multiple pathways that converge to increase phosphorylation of myosin in vascular smooth muscle (VSM) cells, causing persistent constriction. While targeting endothelin and angiotensin, which are upstream regulators that promote VSM contraction, was not effective, treatment with the Rho-kinase inhibitor Y-27632 reduced vessel constriction and PH in Pbx-mutant mice. These results demonstrate a lung-intrinsic, herniation-independent cause of PH in CDH. More broadly, our findings indicate that neonatal PH can result from perturbation of multiple pathways and suggest that targeting the downstream common effectors may be a more effective treatment for neonatal PH.

Topics: Alleles; Animals; Apoptosis; Cell Proliferation; Disease Models, Animal; Echocardiography; Elastin; Female; Gene Deletion; Hernias, Diaphragmatic, Congenital; Homeodomain Proteins; Hypertension, Pulmonary; Lung; Mice; Mice, Knockout; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Myosins; Parturition; Phosphorylation; Pre-B-Cell Leukemia Transcription Factor 1; Proto-Oncogene Proteins; Pulmonary Artery; Respiration; Vasoconstriction

Arterial stiffness and wave reflection are important components of the ventricular afterload. Therefore, we aimed to assess the arterial wave characteristics and mechanical properties of the proximal pulmonary arteries (PAs) in the hypoxic pulmonary hypertensive rat model. After 21 days in normoxic or hypoxic chambers (24 animals/group), animals underwent transthoracic echocardiography and PA catheterization with a dual-tipped pressure and Doppler flow sensor wire. Wave intensity analysis was performed. Artery rings obtained from the pulmonary trunk, right and left PAs, and aorta were subjected to a tensile test to rupture. Collagen and elastin content were determined. In hypoxic rats, proximal PA wall thickness, collagen content, tensile strength per unit collagen, maximal elastic modulus, and wall viscosity increased, whereas the elastin-to-collagen ratio and arterial distensibility decreased. Arterial pulse wave velocity was also increased, and the increase was more prominent in vivo than ex vivo. Wave intensity was similar in hypoxic and normoxic animals with negligible wave reflection. In contrast, the aortic maximal elastic modulus remained unchanged, whereas wall viscosity decreased. In conclusion, there was no evidence of altered arterial wave propagation in proximal PAs of hypoxic rats while the extracellular matrix protein composition was altered and collagen tensile strength increased. This was accompanied by altered mechanical properties in vivo and ex vivo. NEW & NOTEWORTHY In rats exposed to chronic hypoxia, we have shown that pulse wave velocity in the proximal pulmonary arteries increased and pressure dependence of the pulse wave velocity was steeper in vivo than ex vivo leading to a more prominent increase in vivo.

Topics: Animals; Aorta; Arterial Pressure; Biomechanical Phenomena; Chronic Disease; Collagen; Disease Models, Animal; Elastic Modulus; Elastin; Hypertension, Pulmonary; Hypoxia; Male; Models, Cardiovascular; Pulmonary Artery; Pulse Wave Analysis; Rats, Sprague-Dawley; Tensile Strength; Time Factors; Vascular Remodeling; Vascular Stiffness; Viscosity

Development of right ventricular (RV) hypertension eventually contributes to RV and left ventricular (LV) myocardial fibrosis and dysfunction. The molecular mechanisms are not fully elucidated.. Pulmonary artery banding was used to induce RV hypertension in rats in vivo. Then, we evaluated cardiac function and regional remodeling 6 weeks after pulmonary artery banding. To further elucidate mechanisms responsible for regional cardiac remodeling, we also mimicked RV hypertensive stress by cyclic mechanical stretching applied to confluent cultures of cardiac fibroblasts, isolated from the RV free wall, septal hinge points, and LV free wall. Echocardiography and catheter evaluation demonstrated that rats in the pulmonary artery banding group developed RV hypertension with leftward septal displacement, LV compression, and increased LV end-diastolic pressures. Picrosirius red staining indicated that pulmonary artery banding induced marked RV fibrosis and dysfunction, with prominent fibrosis and elastin deposition at the septal hinge points but less LV fibrosis. These changes were associated with proportionally increased expressions of integrin-β1 and profibrotic signaling proteins, including phosphorylated Smad2/3 and transforming growth factor-β1. Moreover, mechanically stretched fibroblasts also expressed significantly increased levels of α-smooth muscle actin, integrin-β1, transforming growth factor-β1, collagen I deposition, and wrinkle formation on gel assays, consistent with myofibroblast transformation. These changes were not observed in parallel cultures of mechanically stretched fibroblasts, preincubated with the integrin inhibitor (BTT-3033).. Experimentally induced RV hypertension triggers regional RV, hinge-point, and LV integrin β1-dependent mechanotransduction signaling pathways that eventually trigger myocardial fibrosis via transforming growth factor-β1 signaling. Reduced LV fibrosis and preserved global function, despite geometrical and pressure aberrations, suggest a possible elastin-mediated protective mechanism at the septal hinge points.

Topics: Animals; Arterial Pressure; Cells, Cultured; Collagen Type I; Disease Models, Animal; Elastin; Fibrosis; Heart Ventricles; Hypertension, Pulmonary; Hypertrophy, Left Ventricular; Hypertrophy, Right Ventricular; Integrin beta1; Male; Mechanotransduction, Cellular; Pulmonary Artery; Rats, Sprague-Dawley; Transforming Growth Factor beta1; Ventricular Function, Left; Ventricular Function, Right; Ventricular Remodeling

We determined in patients with pulmonary arterial (PA) hypertension (PAH) whether in addition to increased production of elastase by PA smooth muscle cells previously reported, PA elastic fibers are susceptible to degradation because of their abnormal assembly.. Fibrillin-1 and elastin are the major components of elastic fibers, and fibrillin-1 binds bone morphogenetic proteins (BMPs) and the large latent complex of transforming growth factor-β1 (TGFβ1). Thus, we considered whether BMPs like TGFβ1 contribute to elastic fiber assembly and whether this process is perturbed in PAH particularly when the BMP receptor, BMPR2, is mutant. We also assessed whether in mice with. Disrupting BMPR2 impairs TGFβ1- and BMP4-mediated elastic fiber assembly and is of pathophysiologic significance in PAH.

Topics: Animals; Bone Morphogenetic Protein 4; Bone Morphogenetic Protein Receptors, Type I; Bone Morphogenetic Protein Receptors, Type II; Case-Control Studies; Cells, Cultured; Disease Models, Animal; Elastic Tissue; Elastin; Familial Primary Pulmonary Hypertension; Fibrillin-1; Fibroblasts; Genetic Predisposition to Disease; Humans; Hypertension, Pulmonary; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; Mutation; Myocytes, Smooth Muscle; Phenotype; Pulmonary Artery; RNA Interference; Transfection; Transforming Growth Factor beta; Vascular Remodeling

During the progression of pulmonary hypertension (PH), proximal pulmonary arteries (PAs) increase in both thickness and stiffness. Collagen, a component of the extracellular matrix, is mainly responsible for these changes via increased collagen fiber amount (or content) and crosslinking. We sought to differentiate the effects of collagen content and cross-linking on mouse PA mechanical changes using a constitutive model with parameters derived from experiments in which collagen content and cross-linking were decoupled during hypoxic pulmonary hypertension (HPH). We employed an eight-chain orthotropic element model to characterize collagen's mechanical behavior and an isotropic neo-Hookean form to represent elastin. Our results showed a strong correlation between the material parameter related to collagen content and measured collagen content (R(2)=0.82, P<0.0001) and a moderate correlation between the material parameter related to collagen crosslinking and measured crosslinking (R(2)=0.24, P=0.06). There was no significant change in either the material parameter related to elastin or the measured elastin content from histology. The model-predicted pressure at which collagen begins to engage was ∼25mmHg, which is consistent with experimental observations. We conclude that this model may allow us to predict changes in the arterial extracellular matrix from measured mechanical behavior in PH patients, which may provide insight into prognoses and the effects of therapy.. The literature has proposed several constitutive models to describe the mechanical effects of arterial collagen but none separates collagen content from crosslinking. Given that both are critical to arterial mechanics, the novel model described here does so. Furthermore, our novel model is well tested by experimental data; model parameters were reasonably correlated with measured collagen content and crosslinking and the model-predicted collagen transition stretch was consistent with that obtained experimentally. Given that arterial collagen structural changes and collagen engagement are critical to arterial stiffening in several disease states, this model, by linking mechanical and biological properties, may allow us to predict important biological changes during disease progression from measured mechanical behavior.

Topics: Aminopropionitrile; Animals; Collagen; Collagen Type I; Collagen Type I, alpha 1 Chain; Elasticity; Elastin; Extracellular Matrix; Hydroxyproline; Hypertension, Pulmonary; Hypoxia; Materials Testing; Mice; Models, Anatomic; Mutation; Pressure; Pulmonary Artery; Stress, Mechanical

Exposure of neonatal mice to hyperoxia results in pulmonary vascular remodeling and aberrant phosphodiesterase type 5 (PDE5) signaling. Although glucocorticoids are frequently utilized in the NICU, little is known about their effects on the developing pulmonary vasculature and on PDE5. We sought to determine the effects of hydrocortisone (HC) on pulmonary vascular development and on PDE5 in a neonatal mouse model of hyperoxic lung injury.. C57BL/6 mice were placed in 21% O2 or 75% O2 within 24 h of birth and received HC (1, 5, or 10 mg/kg subcutaneously every other day) or vehicle. At 14 d, right ventricular hypertrophy (RVH), medial wall thickness (MWT), lung morphometry, and pulmonary artery (PA) PDE5 activity were assessed. PDE5 activity was measured in isolated pulmonary artery smooth muscle cells exposed to 21 or 95% O2 ± 100 nmol/l HC for 24 h.. Hyperoxia resulted in alveolar simplification, RVH, increased MWT, and increased PA PDE5 activity. HC decreased hyperoxia-induced RVH and attenuated MWT. HC had dose-dependent effects on alveolar simplification. HC decreased hyperoxia-induced PDE5 activity both in vivo and in vitro.. HC decreases hyperoxia-induced pulmonary vascular remodeling and attenuates PDE5 activity. These findings suggest that HC may protect against hyperoxic injury in the developing pulmonary vasculature.

Topics: Animals; Cyclic Nucleotide Phosphodiesterases, Type 5; Dose-Response Relationship, Drug; Elastin; Glucocorticoids; Humans; Hydrocortisone; Hyperoxia; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Lung; Lung Injury; Mice; Mice, Inbred C57BL; Pulmonary Alveoli; Pulmonary Artery; Signal Transduction

Optimal histochemical staining is critical to ensure excellent quality stained sections to enable light microscopic and histomorphometric image analysis.  Verhoeff-van Gieson is the most widely used histochemical stain for the visualization of vascular elastic fibers. However, it is notoriously difficult to differentiate fine elastic fibers of small vasculature to enable histomorphometric image analysis of vasculature size characterization especially in organs such as the lung. A tissue fixation regime of 10% neutral buffered formalin with subsequent fixation in 70% ethanol further compounds the problem of small vessel staining and identification.  Therefore, a modified Verhoeff's elastin stain was developed as a reliable method to optimally highlight the internal and external elastic lamina of small arteries (50-100 µm) and intra-acinar vessels (10-50 µm) in 3 µm thick lung tissue sections from models of pulmonary arterial hypertension.  This modified Verhoeff's elastin stain demonstrated optimal staining of fine elastic fibers of pulmonary blood vessels. As a result, high-quality histomorphometric image analysis evaluation of vessel wall thickness in small arteries and intra-acinar vessels was successfully accomplished.  In conclusion, modification of the standard Verhoeff-van Gieson histochemical stain is needed to visualize small caliber vessels' elastic fibers especially in tissues not fixed in 10% neutral buffered formalin only.

Topics: Animals; Elastin; Ferric Compounds; Hematoxylin; Hypertension, Pulmonary; Iodides; Pulmonary Artery; Rats; Rats, Sprague-Dawley; Staining and Labeling

High pulmonary vascular resistance (PVR), proximal pulmonary artery (PA) impedance, and right ventricular (RV) afterload due to remodeling contribute to the pathogenesis and severity of pulmonary hypertension (PH). Intra-amniotic exposure to endotoxin (ETX) causes sustained PH and high mortality in rat pups at birth, which are associated with impaired vascular growth and RV hypertrophy in survivors. Treatment of ETX-exposed pups with antenatal vitamin D (vit D) improves survival and lung growth, but the effects of ETX exposure on RV-PA coupling in the neonatal lung are unknown. We hypothesized that intrauterine ETX impairs RV-PA coupling through sustained abnormalities of PA stiffening and RV performance that are attenuated with vit D therapy. Fetal rats were exposed to intra-amniotic injections of ETX, ETX+vit D, or saline at 20 days gestation (term = 22 days). At postnatal day 14, pups had pressure-volume measurements of the RV and isolated proximal PA, respectively. Lung homogenates were assayed for extracellular matrix (ECM) composition by Western blot. We found that ETX lungs contain decreased α-elastin, lysyl oxidase, collagen I, and collagen III proteins (P < 0.05) compared control and ETX+vit D lungs. ETX-exposed animals have increased RV mechanical stroke work (P < 0.05 vs. control and ETX+vit D) and elastic potential energy (P < 0.05 vs. control and ETX+vit D). Mechanical stiffness and ECM remodeling are increased in the PA (P < 0.05 vs. control and ETX+vit D). We conclude that intrauterine exposure of fetal rats to ETX during late gestation causes persistent impairment of RV-PA coupling throughout infancy that can be prevented with early vit D treatment.

Topics: Animals; Animals, Newborn; Elastin; Endotoxins; Female; Heart Ventricles; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Lung; Pregnancy; Pulmonary Artery; Rats; Rats, Sprague-Dawley; Respiratory Physiological Phenomena; Vascular Resistance; Vitamin D

Pulmonary vascular remodeling, the pathological hallmark of pulmonary arterial hypertension, is attributed to proliferation, apoptosis resistance, and migration of vascular cells. A role of dysregulated matrix cross-linking and stability as a pathogenic mechanism has received little attention. We aimed to assess whether matrix cross-linking enzymes played a causal role in experimental pulmonary hypertension (PH).. All 5 lysyl oxidases were detected in concentric and plexiform vascular lesions of patients with idiopathic pulmonary arterial hypertension. Lox, LoxL1, LoxL2, and LoxL4 expression was elevated in lungs of patients with idiopathic pulmonary arterial hypertension, whereas LoxL2 and LoxL3 expression was elevated in laser-capture microdissected vascular lesions. Lox expression was hypoxia-responsive in pulmonary artery smooth muscle cells and adventitial fibroblasts, whereas LoxL1 and LoxL2 expression was hypoxia-responsive in adventitial fibroblasts. Lox expression was increased in lungs from hypoxia-exposed mice and in lungs and pulmonary artery smooth muscle cells of monocrotaline-treated rats, which developed PH. Pulmonary hypertensive mice exhibited increased muscularization and perturbed matrix structures in vessel walls of small pulmonary arteries. Hypoxia exposure led to increased collagen cross-linking, by dihydroxylysinonorleucine and hydroxylysinonorleucine cross-links. Administration of the lysyl oxidase inhibitor β-aminopropionitrile attenuated the effect of hypoxia, limiting perturbations to right ventricular systolic pressure, right ventricular hypertrophy, and vessel muscularization and normalizing collagen cross-linking and vessel matrix architecture.. Lysyl oxidases are dysregulated in clinical and experimental PH. Lysyl oxidases play a causal role in experimental PH and represent a candidate therapeutic target. Our proof-of-principle study demonstrated that modulation of lung matrix cross-linking can affect pulmonary vascular remodeling associated with PH.

Topics: Adult; Aged, 80 and over; Animals; Antihypertensive Agents; Case-Control Studies; Cell Hypoxia; Cells, Cultured; Collagen; Disease Models, Animal; Elastin; Enzyme Inhibitors; Familial Primary Pulmonary Hypertension; Female; Fibroblasts; Gene Expression Regulation, Enzymologic; Humans; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Hypoxia; Isoenzymes; Male; Mice; Middle Aged; Monocrotaline; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Protein-Lysine 6-Oxidase; Pulmonary Artery; Rats; RNA, Messenger; Ventricular Dysfunction, Right; Young Adult

Hypoxic pulmonary hypertension (HPH) causes extralobar pulmonary artery (PA) stiffening, which potentially impairs right ventricular systolic function. Changes in the extracellular matrix proteins collagen and elastin have been suggested to contribute to this arterial stiffening. We hypothesized that vascular collagen accumulation is a major cause of extralobar PA stiffening in HPH and tested our hypothesis with transgenic mice that synthesize collagen type I resistant to collagenase degradation (Col1a1(R/R)). These mice and littermate controls that have normal collagen degradation (Col1a1(+/+)) were exposed to hypoxia for 10 days; some were allowed to recover for 32 days. In vivo PA pressure and isolated PA mechanical properties and collagen and elastin content were measured for all groups. Vasoactive studies were also performed with U-46619, Y-27632, or calcium- and magnesium-free medium. Pulmonary hypertension occurred in both mouse strains due to chronic hypoxia and resolved with recovery. HPH caused significant PA mechanical changes in both mouse strains: circumferential stretch decreased, and mid-to-high-strain circumferential elastic modulus increased (P < 0.05 for both). Impaired collagen type I degradation prevented a return to baseline mechanical properties with recovery and, in fact, led to an increase in the low and mid-to-high-strain moduli compared with hypoxia (P < 0.05 for both). Significant changes in collagen content were found, which tended to follow changes in mid-to-high-strain elastic modulus. No significant changes in elastin content or vasoactivity were observed. Our results demonstrate that collagen content is important to extralobar PA stiffening caused by chronic hypoxia.

Topics: Animals; Biomechanical Phenomena; Blood Pressure; Chronic Disease; Collagen Type I; Collagen Type I, alpha 1 Chain; Disease Models, Animal; Elastic Modulus; Elastin; Hydroxyproline; Hypertension, Pulmonary; Hypoxia; Mechanotransduction, Cellular; Mice; Mice, Transgenic; Mutation; Pulmonary Artery; Recovery of Function; Time Factors; Vasoconstriction; Vasoconstrictor Agents

Elastin is a major structural component of large elastic arteries and a principal determinant of arterial biomechanical properties. Elastin loss-of-function mutations in humans have been linked to the autosomal-dominant disease supravalvular aortic stenosis, which is characterized by stenotic lesions in both the systemic and pulmonary circulations. To better understand how elastin insufficiency influences the pulmonary circulation, we evaluated pulmonary cardiovascular physiology in a unique set of transgenic and knockout mice with graded vascular elastin dosage (range 45-120% of wild type). The central pulmonary arteries of elastin-insufficient mice had smaller internal diameters (P < 0.0001), thinner walls (P = 0.002), and increased opening angles (P = 0.002) compared with wild-type controls. Pulmonary circulatory pressures, measured by right ventricular catheterization, were significantly elevated in elastin-insufficient mice (P < 0.0001) and showed an inverse correlation with elastin level. Although elastin-insufficient animals exhibited mild to moderate right ventricular hypertrophy (P = 0.0001) and intrapulmonary vascular remodeling, the changes were less than expected, given the high right ventricular pressures, and were attenuated compared with those seen in hypoxia-induced models of pulmonary arterial hypertension. The absence of extensive pathological cardiac remodeling at the high pressures in these animals suggests a developmental adaptation designed to maintain right-sided cardiac output in a vascular system with altered elastin content.

Topics: Adaptation, Physiological; Animals; Blood Pressure; Elastin; Female; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Pulmonary Artery; Pulmonary Circulation; Stress, Mechanical; Ventricular Function, Right; Ventricular Pressure

Vascular expression of bone morphogenetic type IA receptor (Bmpr1a) is reduced in lungs of patients with pulmonary arterial hypertension, but the significance of this observation is poorly understood. To elucidate the role of Bmpr1a in the vascular pathology of pulmonary arterial hypertension and associated right ventricular (RV) dysfunction, we deleted Bmpr1a in vascular smooth muscle cells and in cardiac myocytes in mice using the SM22alpha;TRE-Cre/LoxP;R26R system. The LacZ distribution reflected patchy deletion of Bmpr1a in the lung vessels, aorta, and heart of SM22alpha;TRE-Cre;R26R;Bmpr1a(flox/+) and flox/flox mutants. This reduction in BMPR-IA expression was confirmed by Western immunoblot and immunohistochemistry in the flox/flox group. This did not affect pulmonary vasoreactivity to acute hypoxia (10% O2) or the increase in RV systolic pressure and RV hypertrophy following 3 weeks in chronic hypoxia. However, both SM22alpha;TRE-Cre;R26R;Bmpr1a(flox/+) and flox/flox mutant mice had fewer muscularized distal pulmonary arteries and attenuated loss of peripheral pulmonary arteries compared with age-matched control littermates in hypoxia. When Bmpr1a expression was reduced by short interference RNA in cultured pulmonary arterial smooth muscle cells, serum-induced proliferation was attenuated explaining decreased hypoxia-mediated muscularization of distal vessels. When Bmpr1a was reduced in cultured microvascular pericytes by short interference RNA, resistance to apoptosis was observed and this could account for protection against hypoxia-mediated vessel loss. The similar elevation in RV systolic pressure and RV hypertrophy, despite the attenuated remodeling with chronic hypoxia in the flox/flox mutants versus controls, was not a function of elevated left ventricular end diastolic pressure but was associated with increased periadventitial deposition of elastin and collagen, potentially influencing vascular stiffness.

Topics: Animals; Aorta; Arteries; Bone Morphogenetic Protein Receptors, Type I; Cells, Cultured; Collagen; Coronary Circulation; Elastin; Humans; Hypertension, Pulmonary; Hypoxia; Lung; Mice; Mice, Knockout; Microfilament Proteins; Muscle Proteins; Myocardial Contraction; Myocardium; Myocytes, Smooth Muscle; Neovascularization, Pathologic

S100A4/Mts-overexpressing mice have thick elastic laminae and mild pulmonary arterial hypertension (PAH), and the occasional older mouse develops occlusive neointimal lesions and perivascular inflammation. We hypothesized that a vasculotropic virus could induce neointimal lesions in the S100A4/Mts1 mouse by facilitating breakdown of elastin and migration and proliferation of smooth muscle cells. To test this hypothesis, we infected S100A4/Mts1 mice with gammaherpesvirus 68 (gammaHV68). We observed, 6 mo after gammaHV68 [4 x 10(3) plaque-forming units (PFU)], perivascular inflammation in 10/15 S100A4/Mts1 mice and occlusive neointimal formation in 3/10 mice, accompanied by striking degradation of elastin. We then compared the early response after high-dose gammaHV68 (4 x 10(6) PFU) in C57Bl/6 and S100A4/Mts1 mice. In S100A4/Mts1 mice only, significant PAH, muscularization of distal vessels, and elastase activity were observed 6 wk after gammaHV68. These features resolved by 3 mo without neointimal formation. We therefore infected mice with the M1-gammaHV68 strain that reactivates from latency with higher efficiency and observed neointimal lesions at 3 mo in 2/5 C57Bl/6 (5-9% of vessels) and in 5/5 S100A4/Mts1 mice (13-40% of vessels) accompanied by mild PAH, heightened lung elastase activity, and intravascular viral expression. This suggested that enhanced generation of elastin peptides in S100A4/Mts1 mice may promote increased viral entry in the vessel wall. Using S100A4/Mts1 PA organ culture, we showed, in response to elastase activity, heightened production of elastin peptides associated with invasion of inflammatory cells and intravascular viral antigen. We therefore propose that early viral access to the vessel wall may be a critical determinant of the extent of vascular pathology following reactivation.

Topics: Animals; Antigens, Viral; Blood Pressure; Elastin; Gammaherpesvirinae; Herpesviridae Infections; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Mice; Mice, Inbred C57BL; Models, Biological; Mutation; Peptides; Protein Processing, Post-Translational; Pulmonary Artery; S100 Calcium-Binding Protein A4; S100 Proteins; Viral Load; Virus Activation; Virus Internalization

Thiazolidinediones (TZDs) are insulin-sensitizing agents that also decrease systemic blood pressure, attenuate the formation of atherosclerotic lesions, and block remodeling of injured arterial walls. Recently, TZDs were shown to prevent pulmonary arterial (PA) remodeling in rats treated with monocrotaline. Presently we report studies testing the ability of the TZD rosiglitazone (ROSI) to attenuate pathological arterial remodeling in the lung and prevent the development of pulmonary hypertension (PH) in rats subjected to chronic hypoxia. PA remodeling was reduced in ROSI-treated animals exposed to hypoxia compared with animals exposed to hypoxia alone. ROSI treatment blocked muscularization of distal pulmonary arterioles and reversed remodeling and neomuscularization in lungs of animals previously exposed to chronic hypoxia. Decreased PA remodeling in ROSI-treated animals was associated with decreased smooth muscle cell proliferation, decreased collagen and elastin deposition, and increased matrix metalloproteinase-2 activity in the PA wall. Cells expressing the c-Kit cell surface marker were observed in the PA adventitia of untreated animals exposed to hypoxia but not in ROSI-treated hypoxic rats. Right ventricular hypertrophy and cardiomyocyte hypertrophy were also blunted in ROSI-treated hypoxic animals. Interestingly, mean PA pressures were elevated equally in the untreated and ROSI-treated groups, indicating that ROSI had no effect on the development of PH. However, mean PA pressure was normalized acutely in both groups of hypoxia-exposed animals by Fasudil, an agent that inhibits RhoA/Rho kinase-mediated vasoconstriction. We conclude that ROSI can attenuate and reverse PA remodeling and neomuscularization associated with hypoxic PH. However, this agent fails to block the development of PH, apparently because of its inability to repress sustained Rho kinase-mediated arterial vasoconstriction.

Topics: Animals; Blood Pressure; Collagen; Elastin; Extracellular Matrix; Female; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Hypoxia; Male; PPAR gamma; Proto-Oncogene Proteins c-kit; Pulmonary Artery; Rats; Rats, Inbred WKY; Rosiglitazone; Thiazolidinediones

Right heart failure due to pulmonary hypertension causes significant morbidity and mortality. To study the linked vascular mechanical and biological changes that are induced by pulmonary hypertension, we mechanically tested isolated left main pulmonary arteries from mice exposed to chronic hypobaric hypoxia and performed histological assays on contralateral vessels. In isolated vessel tests, hypoxic vessels stretched less in response to pressure than controls at all pressure levels. Given the short length and large diameter of the pulmonary artery, the tangent Young's modulus could not be measured; instead, an effective elastic modulus was calculated that increased significantly with hypoxia [(280 kPa (SD 53) and 296 kPa (SD 50) for 10 and 15 days, respectively, vs. 222 kPa (SD 35) for control; P < 0.02)]. Hypoxic vessels also had higher damping coefficients [(0.063 (SD 0.017) and 0.054 (SD 0.014) for 10 and 15 days, respectively, vs. 0.033 (SD 0.016) for control; P < 0.002)], indicating increased energy dissipation. The increased stiffness with hypoxia correlated with an increase in collagen thickness (percent collagen multiplied by wall thickness) as well as the sum of elastin and collagen thicknesses measured histologically in the artery wall. These results highlight the mechanobiological changes in the pulmonary vasculature that occur in response to hypoxia-induced pulmonary hypertension. Furthermore, they demonstrate significant vascular mechanical and biological changes that would increase pulmonary vascular impedance, leading to right heart failure.

Topics: Animals; Collagen; Elasticity; Elastin; Hypertension, Pulmonary; Hypoxia; Male; Mice; Mice, Inbred C57BL; Pulmonary Artery; Stress, Mechanical

Transgenic mice overexpressing the calcium binding protein, S100A4/Mts1, occasionally develop severe pulmonary vascular obstructive disease. To understand what underlies this propensity, we compared the pulmonary vascular hemodynamic and structural features of S100A4/Mts1 with control C57Bl/6 mice at baseline, following a 2-week exposure to chronic hypoxia, and after 1 and 3 months "recovery" in room air. S100A4/Mts1 mice had greater right ventricular systolic pressure and right ventricular hypertrophy at baseline, which increased further with chronic hypoxia and was sustained after 3 months "recovery" in room air. These findings correlated with a heightened response to acute hypoxia and failure to vasodilate with nitric oxide or oxygen. S100A4/Mts1 mice, when compared with C57Bl/6 mice, also had impaired cardiac function judged by reduced ventricular elastance and decreased cardiac output. Despite higher right ventricular systolic pressures with chronic hypoxia, S100A4/Mts1 mice did not develop more severe PVD, but in contrast to C57Bl/6 mice, these features did not regress on return to room air. Microarray analysis of lung tissue identified a number of genes differentially upregulated in S100A4/Mts1 versus control mice. One of these, fibulin-5, is a matrix component necessary for normal elastin fiber assembly. Fibulin-5 was localized to pulmonary arteries and associated with thickened elastic laminae. This feature could underlie attenuation of pulmonary vascular changes in response to elevated pressure, as well as impaired reversibility.

Topics: Animals; Elastin; Extracellular Matrix Proteins; Female; Hypertension, Pulmonary; Hypoxia; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Oligonucleotide Array Sequence Analysis; Pancreatic Elastase; Recombinant Proteins; RNA, Messenger; S100 Calcium-Binding Protein A4; S100 Proteins; Systole

Increased serine elastase activity has been implicated in the vascular remodeling associated with chronic hypoxia-related pulmonary hypertension in rats.. In this study we determined the time course of hypoxia-induced serine elastase activity in the murine lung and related this to initiation of a proteolytic cascade characterized by an increase in matrix metalloproteinases (MMPs). We then used transgenic mice in which overexpression of the selective serine elastase inhibitor elafin was targeted to the cardiovascular system to determine whether upregulation of a naturally occurring serine elastase inhibitor suppresses MMPs and the hemodynamic and structural response to chronic hypoxia (air at 380 mm Hg). In nontransgenic but not in elafin-transgenic mice, we documented a transient increase in serine elastase activity after 12 hours of hypoxic exposure attributed to a 30-kDa protein as determined by elastin zymography and fluorophosphonate/fluorophosphate-biotin labeling. Two days after hypoxia, the pro-forms of MMP-2 and MMP-9 were induced in the nontransgenic mice, but MMP-9 was suppressed in elafin-transgenic mice. Acute hypoxic vasoconstriction was similar in nontransgenic and elafin-transgenic littermates. Chronic hypoxia for 26 days resulted in >1-fold increase in right ventricular pressure (P<0.004) in nontransgenic compared with control or elafin-transgenic littermates. In the latter mice, normalization of the right ventricular pressure was associated with reduced muscularization and preservation of the number of distal vessels (P<0.04 for both comparisons).. Modulation of the severity of chronic hypoxia-induced pulmonary vascular disease could be a function of endogenously expressed serine elastase inhibitors.

Topics: Actins; Animals; Elastin; Endothelin-1; Hematocrit; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Hypoxia; Kinetics; Matrix Metalloproteinases; Mice; Mice, Transgenic; Pressure; Proteinase Inhibitory Proteins, Secretory; Proteins; Pulmonary Artery; RNA, Messenger; Serine Proteinase Inhibitors; Up-Regulation; Ventricular Pressure

Sustained fetal tracheal occlusion (TO) results in accelerated lung growth but causes severe type II cell depletion. Temporary TO fails to cause lung growth in a congenital diaphragmatic hernia (CDH) model but preserves type II cells and corrects pulmonary hypertension. Herein, we study the pulmonary vascular changes caused by temporary TO.. CDH was created in 12 fetal lambs (65-70 d; term, 145 days). In 6 lambs, the trachea was occluded for 2 weeks (CDH + TO; 108-122 d). Animals were killed at 136 days. The lungs were processed with elastin stains and anti-alpha-smooth muscle actin antibody. Partial or circumferential presence of inner and outer elastic lamina was used to determine muscularization of pulmonary arterioles. The percent of medial wall thickness was plotted against vessel diameter for each group.. Lung weight/body weight was smaller in lambs with CDH (1. 35% +/- 0.56%) and CDH + TO (1.70% +/- 0.34%) than in control lambs (3.55% +/- 0.56%; P <.05, single-factor analysis of variance). The smallest muscularized vessel was 113 +/- 50 microm, and the largest nonmuscularized vessel was 138 +/- 49 microm in lambs with CDH, significantly different from control lambs (185 +/- 69 microm and 350 +/- 116 microm, respectively) and lambs with CDH + TO (185 +/- 97 microm and 245 +/- 100 microm, respectively; P <.05). In lambs with CDH, only 25% of vessels of less than 60 microm were nonmuscularized, compared with 81% in control lambs (P <.05) and 74% in lambs with CDH + TO.Conclusions. Temporary tracheal occlusion, from 108 to 122 days, corrects the abnormal muscularization of pulmonary arterioles seen in CDH. These morphometric findings parallel physiologic results at birth and further suggest that short-term occlusion, which preserves surfactant-producing type II pneumocytes without lung growth, may be sufficient to improve neonatal outcome of diaphragmatic hernia.

Topics: Actins; Animals; Arterioles; Body Weight; Elastin; Female; Hernia, Diaphragmatic; Hypertension, Pulmonary; Lung; Organ Size; Pregnancy; Pulmonary Circulation; Sheep; Trachea

Topics: Animals; Cell Differentiation; Cell Division; Cell Lineage; Collagen; Elastin; Hypertension, Pulmonary; Hypoxia; Muscle, Smooth, Vascular

Continuous air embolization (CAE) into the pulmonary arterial circulation of sheep results in functional and structural changes of chronic pulmonary hypertension. Release of elastin peptides into lung lymph during CAE and attenuation of CAE-induced pulmonary hypertension by neutrophil depletion suggest that neutrophil elastase may contribute to these changes. To investigate this notion, we treated awake sheep with a potent neutrophil elastase inhibitor, recombinant secretory leukoprotease inhibitor (SLPI) (100 mg/day by aerosol), during 12 days of CAE (CAE+SLPI; n = 7). Controls included sheep receiving CAE + vehicle (VEH) (n = 6), VEH alone (n = 3), and SLPI alone (n = 3). SLPI significantly attenuated the CAE-induced increases in lung lymph flow (day 8; 2.3 +/- 0.5 vs. 5.6 +/- 1.7 ml/15 min), protein clearance (day 8; 1.36 +/- 0.32 vs. 3.08 +/- 0.84 ml/15 min), and elastin peptide concentration (day 8; 243 +/- 41 vs. 398 +/- 44 ng/ml). SLPI delayed the onset of sustained pulmonary hypertension from day 8 to day 12. Both CAE groups showed similar structural changes in the pulmonary arteries. SLPI was well tolerated in control sheep and did not affect hemodynamics or structure. We conclude that serine proteases may contribute to the early initiation of chronic pulmonary hypertension but do not play a striking role in its eventual development.

Topics: Aerosols; Animals; Capillary Permeability; Elastin; Embolism, Air; Hemodynamics; Humans; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Lung; Lung Injury; Lymph; Proteinase Inhibitory Proteins, Secretory; Proteins; Pulmonary Circulation; Pulmonary Gas Exchange; Recombinant Proteins; Secretory Leukocyte Peptidase Inhibitor; Serine Proteinase Inhibitors; Sheep; Vascular Resistance

Vascular remodeling is commonly associated with pulmonary hypertension (PH) postnatally, but little is known about its presence in fetuses. In sixteen fetal sheep (126 wk gestation), the ductus arteriosus (DA) was ligated, and the animals were studied at 4, 8, and 14 days after surgery. The uninstrumented twins served as controls. Four days post-DA ligation PH resulted in an increase in the right and left ventricular free wall weight ratio (1.2 +/- 0.1 vs. 1.0 +/- 0.1 in the controls; P < 0.01), with a further progressive increase at 8 (1.4 +/- 0.1 vs. 1.0 +/- 0.1; P < 0.01) and 14 days (1.5 +/- 0.2 vs. 1.0 +/- 0.1; P < 0.01). An increase in vascular percent medial thickness was observed after 4 days of DA ligation and was restricted to small vessels. The large arteries collagen and elastin contents were 28.9 +/- 3.4 and 27.1 +/- 3.4 micrograms/micrograms of DNA, respectively, and were not significantly different from control values even after 14 days DA ligation. We further compared elastin synthesis in fetal and neonatal arteries in vitro. Synthesis in the fetus was greater than the newborn (10.6 +/- 1.4 vs. 4.6 +/- 0.7 cpm.mg wet wt-1.h-1; P < 0.01). Vessel endothelium denudation reduced synthesis to 60 +/- 8% of controls in the fetus, whereas no change was seen in the newborn. After an increase in wall stress, synthesis increased in the fetus (194 +/- 28% of control P < 0.01) and newborn (173 +/- 25%; P < 0.01). Removal of the endothelium abolished the response.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Animals, Newborn; Blood Vessels; Ductus Arteriosus; Elastin; Endothelium, Vascular; Fetus; Hypertension, Pulmonary; Ligation; Sheep

Topics: Animals; Cystic Fibrosis; Elastin; Endothelium, Vascular; Humans; Hypertension, Pulmonary; Hypoxia; Lung Diseases, Obstructive; Muscle, Smooth, Vascular; Pancreatic Elastase

In a rat model of pulmonary hypertension induced by monocrotaline, medial hypertrophy of the pulmonary arteries is associated with enhanced production (synthesis) of insoluble elastin relative to accumulation and an increased number of elastin fragments, features suggestive of an elastolytic process. In the present study, we measured and characterized pulmonary artery (PA) elastolytic activity at time points before as well as coincident with the progression of medial hypertrophy in monocrotaline-injected adult male Sprague-Dawley rats. We also determined whether medial hypertrophy is preceded by ultrastructural changes in elastin. Since medial hypertrophy develops but fails to progress in rats injected with monocrotaline at 8 days of age, we assessed whether, compared with adult rats, there were also structural and biochemical differences in elastin and elastolytic activity. A twofold increase in elastolytic activity per milligram tissue was observed 2 days after monocrotaline injection in adult rats (p less than 0.01), and there was an increased number of breaks in the internal elastic lamina (IEL) at 4 days (p less than 0.05) (i.e., before the development of medial hypertrophy). Associated with the progression of medial hypertrophy between 16 and 28 days after monocrotaline injection, there was a further threefold increase in elastolytic activity per milligram tissue by 28 days (p less than 0.01). Susceptibility of the elastolytic activity to specific inhibitors suggested that one or more serine elastases is involved. In infant rats in which medial and right ventricular hypertrophy fail to progress in severity between 16 and 28 days after monocrotaline injection, we did not measure an increase in elastolytic activity, nor was there evidence of an increase in the number of breaks in the IEL at 4 days, suggesting a lack of increased elastolytic activity at an earlier time point. The total content of PA elastin in infant rats, although increased compared with control rats (p less than 0.01), was not associated with heightened production and appeared ultrastructurally as thicker laminae (p less than 0.05) rather than as fragments previously reported in adult rats.

Topics: Age Factors; Animals; Collagen; Elastin; Hypertension, Pulmonary; Hypertrophy; Male; Monocrotaline; Pancreatic Elastase; Pulmonary Artery; Rats; Rats, Inbred Strains

The progression of structural changes in the pulmonary arterial bed were followed in a model of chronic pulmonary hypertension. Chronically instrumented awake sheep received continuous air embolization for 0 (controls), 1, 4, 8, or 12 days (n = 5-6/group). After the period of embolization, the lungs were removed, the pulmonary arteries were distended with barium-gelatin, and the lungs were fixed via the airways with formal-saline. Quantitative techniques were applied to sections from random blocks from the lungs of each animal. One day of embolization resulted in granulocyte sequestration in the lung interstitium and in small vessels; additionally, intraalveolar and perivascular edema was present. By 4 days, increased medial thickness, appearance of muscle in smaller arteries than normal (e.g., muscular arteries at alveolar duct level: control = 1.2 +/- 1.2%; day 4 = 22.7 +/- 7.7) and reduction in number of barium-filled intraacinar arteries was found. The arterial changes progressed in severity to day 8 and were similar at day 12. Since arterial remodelling involves increased elastin deposition, the concentration of elastin peptides was measured in lung lymph. Increased flux of elastin peptides was apparent from day 2 of embolization and continued to increase to a level 20 x baseline by day 12 (baseline 351 +/- 86 micrograms/15 min; day 12 = 6338 +/- 2999). Comparison of the onset of the structural changes with previous findings shows that the arterial remodelling parallels the onset of sustained pulmonary hypertension. The increase in lung-lymph elastin peptides by day 2 provides evidence that vascular remodelling is initiated before day 4 of embolization. The early sequestration of granulocytes and appearance of edema suggest that these may be part of the trigger to the development of the structural changes.

Topics: Angiography; Animals; Biopsy; Blood Vessels; Cardiomegaly; Chronic Disease; Elastin; Embolism, Air; Hypertension, Pulmonary; Lung; Osmolar Concentration; Peptides; Pulmonary Circulation; Sheep

Previously in rats injected with the toxin monocrotaline and administered SC-39026, a serine elastase inhibitor, pulmonary hypertension was decreased in association with reduced muscularization of peripheral pulmonary arteries. To determine whether inhibition of elastolytic activity might prevent this vascular change in other conditions producing pulmonary hypertension, we administered SC-39026 to rats during a 10-day exposure to chronic hypobaric hypoxia. We also measured elastolytic activity in the central pulmonary arteries of rats using [3H]elastin substrate and determined whether there was an increase in activity either as early as 2 days or at completion of the hypoxic exposure, which could be inhibited by SC-39026. to further determine whether the mechanism of muscularization of peripheral arteries is modulated by degradation of elastin or other elastase-susceptible extracellular matrix proteins, we assessed desmosine excretion and ultrastructural alterations in elastin as well as in type IV collagen, fibronectin, and laminin. SC-39026 reduced the number of muscularized arteries and the level of pulmonary arterial pressure during exposure to chronic hypoxia. Elastolytic activity was fourfold higher in central pulmonary arteries 2 days after hypoxia when compared with values in control vessels, and the activity was inhibited by SC-39026. In small peripheral pulmonary arteries there were no significant changes with hypoxia reflected in desmosines or in the immunocytochemistry of elastase-susceptible glycoproteins, with the exception of decreased laminin. This feature was not inhibited by SC-39026. To further assess whether the protective effect of SC-39026 was related to its inhibition of elastase, an extended study was carried out using a different elastase inhibitor, alpha 1-proteinase inhibitor. An even greater reduction in hypoxia-induced pulmonary hypertension and vascular changes was observed with this elastase inhibitor and the latter included medial hypertrophy.

Topics: alpha 1-Antitrypsin; Animals; Cardiomegaly; Chlorobenzoates; Collagen; Elastic Tissue; Elastin; Fibronectins; Hemodynamics; Hypertension, Pulmonary; Hypoxia; Immunohistochemistry; Laminin; Male; Microscopy, Electron; Pancreatic Elastase; Pulmonary Artery; Rats; Rats, Inbred Strains

Topics: Animals; Biomechanical Phenomena; Collagen; Elastin; Hypertension, Pulmonary; Immunohistochemistry; Male; Microbial Collagenase; Pulmonary Artery; Rats; Rats, Inbred Strains

Pulmonary hypertension is associated with abnormal connective tissue deposition in the media of pulmonary arteries. Lobar arteries from calves maintained for up to 15 days at simulated high altitude showed a 35% increase in collagen and a greater than 40% increase in crosslinked elastin per microgram protein. Labeling of artery tissue with [14C]proline revealed a nearly twofold increase in relative collagen synthesis. There was increased incorporation into Types I, III, IV, and V collagen with an increase in the proportion of newly synthesized Type IV collagen. Quantitation of collagen mRNA by slot-blot assay demonstrated increased levels of Types I and IV collagen message. In addition, medial smooth muscle cells isolated from the hypertensive calves demonstrated a nearly twofold increase in relative collagen synthesis, a twofold increase in the accumulation of newly synthesized collagen per microgram DNA, and increased levels of Types I and IV collagen mRNA. Exposure of pulmonary artery smooth muscle cells, adventitial cells, and fetal calf ligament fibroblasts to conditioned calf serum harvested from cultures of medial cells from hypertensive animals increased their levels of collagen as well as elastin mRNA. These studies suggest that the increased production of collagen in hypertensive arteries is mediated at a pre-translational level by soluble factor(s) generated by medial smooth muscle cells.

Topics: Animals; Animals, Newborn; Cattle; Cells, Cultured; Collagen; Elastin; Hypertension, Pulmonary; Hypoxia; Muscle, Smooth, Vascular; Organ Culture Techniques; Pulmonary Artery; RNA; RNA, Messenger

In situ hybridization was used to determine the morphologic distribution of tropoelastin and alpha 1(I) procollagen mRNA expression in elastic intralobar arteries from neonatal calves with hypoxic pulmonary hypertension induced by a 15-day exposure to a simulated altitude of 1500 m. In vessels from normotensive control animals, low levels of hybridizable tropoelastin mRNA were detected in smooth muscle cells (SMC) of the inner media associated with large elastic lamellae. Compared to control arteries, vessels from hypertensive animals demonstrated a markedly different pattern of hybridization. In these arteries, strong hybridization signals for tropoelastin mRNA were seen in SMC lying between the elastic lamellae of the outer media, and the density of labeling associated with these medial cells decreased progressively toward the lumen. Endothelial and adventitial cells in both control and hypertensive arteries were negative for tropoelastin mRNA. Type I procollagen mRNA was dispersed through the media of control arteries, and in hypertensive calves, the hybridization signal was more intense and was unevenly distributed through the media similarly to that for tropoelastin mRNA. Adventitial cells were strongly positive for procollagen mRNA, and the signal was equally intense for both control and hypertensive arteries. Cells that had no detectable tropoelastin mRNA were noted in the outer media of both control and hypertensive vessels. These cells occurred as broad circumferential bands in the normotensive artery and as nodular foci in the hypertensive artery. Immunocytochemical studies with antibodies to smooth muscle specific actin, desmin, and vimentin demonstrated that cells within these foci, as well as tropoelastin mRNA-positive cells, were SMC. These studies demonstrate that expression of tropoelastin and procollagen mRNA was differentially stimulated by pulmonary hypertension within specific regions and SMC populations of the vascular wall.

Topics: Animals; Animals, Newborn; Arteries; Cattle; Collagen; Elastin; Gene Expression; Hypertension, Pulmonary; Immunohistochemistry; Nucleic Acid Hybridization; Oxygen; Procollagen; Pulmonary Circulation; RNA Probes; RNA, Messenger; Tropoelastin

Changes in elastin and collagen synthesis in the pulmonary artery wall, assessed both biochemically and ultrastructurally, were related to the development of progressive pulmonary hypertension induced by the toxin monocrotaline. Male Sprague-Dawley rats (200 to 225 gm) were injected subcutaneously in the hind flank with either monocrotaline (60 mg/kg) or an equivalent volume of saline and studied 8, 16 and 28 days later. At each time point, the right ventricle and left ventricle with septum were separated and weighed to follow the development of right ventricular hypertrophy. The hilar pulmonary artery was assessed by light microscopy for medial hypertrophy and by electron microscopy for changes in the endothelium, subendothelium and media. The mainstem pulmonary artery was used to determine synthesis of elastin and collagen by in vitro incorporation of [14C]proline into nonelastin, [14C]hydroxyproline into collagen, and [3H]valine into cyanogen bromide-insoluble elastin. In addition, total content of insoluble elastin was determined by weight of the residue after cyanogen bromide digestion and of collagen by total hydroxyproline content in sodium dodecyl sulfate and cyanogen bromide extracts. Eight days after monocrotaline injection, there was pulmonary artery endothelial swelling and a significant decrease in the number of myoendothelial junctions (p less than 0.05) associated with a decreased proportion of amorphous elastin in the media (p less than 0.01). Sixteen days after monocrotaline injection, a decrease in the proportion of elastin in the media was still evident (p less than 0.01) despite an apparent increase in insoluble elastin synthesis. Moreover, the amorphous elastin was distributed preferentially in small islands rather than in laminae (p less than 0.05). Twenty-eight days after monocrotaline injection, medial and right ventricular hypertrophy had developed (p less than 0.05 and p less than 0.01, respectively). At the same time, there was a striking increase in both insoluble elastin synthesis and total insoluble elastin content (p less than 0.01 for both) and an increase in collagen synthesis and total collagen content (p less than 0.05 for both). In addition, the ratio of insoluble elastin synthesis to collagen synthesis was greater than in controls (p less than 0.01), whereas the ratio of total insoluble elastin to total collagen did not change. On ultrastructural analysis, the proportion of amorphous elastin in the vessel wall relative

Topics: Animals; Aorta; Collagen; Elastin; Endothelium, Vascular; Hypertension, Pulmonary; Male; Microscopy, Electron; Monocrotaline; Plants, Toxic; Pulmonary Artery; Pyrrolizidine Alkaloids; Rats; Senecio

We suggest that hypoxia-induced pulmonary hypertension in the newborn calf is an attractive model for studying the mechanisms underlying alterations in extracellular matrix accumulation which occur in pulmonary vascular disease. Our data support a model (Fig 7) in which the SMC, perhaps as a result of hypoxic and/or pressure-induced vessel wall injury, becomes phenotypically altered. This phenotypically altered SMC generates a factor, termed smooth muscle derived extracellular matrix factor (SMEF), and possibly other factors. SMEF, in turn, stimulates or induces elastin and collagen synthesis in fibroblasts and endothelial cells. SMEF, or an associated activity derived from phenotypically altered smooth muscle cells, also induces elastin receptor expression on the cell surface and affects the chemotactic responsiveness of vascular cells. Thus, the SMC may be able to affect both the secretory and responsive properties of cell types in the vascular wall. The SMC may be critical in the vascular remodeling in pulmonary hypertension. The possible autocrine or paracrine alteration of cellular phenotypes by smooth muscle-derived mediators provides an important new direction for future research into molecular and cellular mechanisms of connective tissue regulation in diseased vessels.

Topics: Altitude; Animals; Animals, Newborn; Cattle; Cattle Diseases; Collagen; Disease Models, Animal; Elastin; Hypertension, Pulmonary; Hypoxia; Muscle, Smooth, Vascular; Phenotype; Pulmonary Artery

Abnormal accumulation of connective tissue in blood vessels contributes to alterations in vascular physiology associated with disease states such as hypertension and atherosclerosis. Elastin synthesis was studied in blood vessels from newborn calves with severe pulmonary hypertension induced by alveolar hypoxia in order to investigate the cellular stimuli that elicit changes in pulmonary arterial connective tissue production. A two- to fourfold increase in elastin production was observed in pulmonary artery tissue and medial smooth muscle cells from hypertensive calves. This stimulation of elastin production was accompanied by a corresponding increase in elastin messenger RNA consistent with regulation at the transcriptional level. Conditioned serum harvested from cultures of pulmonary artery smooth muscle cells isolated from hypertensive animals contained one or more low molecular weight elastogenic factors that stimulated the production of elastin in both fibroblasts and smooth muscle cells and altered the chemotactic responsiveness of fibroblasts to elastin peptides. These results suggest that connective tissue changes in the pulmonary vasculature in response to pulmonary hypertension are orchestrated by the medial smooth muscle cell through the generation of specific differentiation factors that alter both the secretory phenotype and responsive properties of surrounding cells.

Topics: Animals; Cattle; Connective Tissue; Disease Models, Animal; Elastin; Humans; Hypertension, Pulmonary; Hypoxia; Muscle, Smooth, Vascular; RNA, Messenger; Transcription, Genetic

Topics: Animals; Aorta; Collagen; Elastin; Gene Expression Regulation; Hypertension, Pulmonary; Hypoxia; Pulmonary Artery; Rats; Rats, Inbred Strains; RNA, Messenger

Topics: Aging; Animals; Arterioles; Blood Flow Velocity; Capillaries; Cats; Collagen; Connective Tissue; Cytoplasm; Elasticity; Elastin; Erythrocytes; Fixatives; Freezing; Humans; Hypertension, Pulmonary; Lung; Lung Compliance; Microcirculation; Rats; Venules

Internal diameter, thickness of media, and surface density of elastin in media of both pulmonary trunk and aorta were determined in a random group of 100 adults. Because of contrasting results of histologic vs chemical studies of the amount of elastin in the pulmonary trunk, we have used a television image analyzer for objective determination of surface density of elastin in histologic slides, applying the same method to the aorta for comparison. Both caliber and medial thickness of the pulmonary trunk increase somewhat with age. The surface density of elastin in the pulmonary trunk is not influenced by age and averages approximately 26%, with considerable individual variation. In patients with pulmonary hypertension, this percentage is consistently higher. We conclude that the decrease in extensibility of the aging pulmonary trunk is caused by changes in physical properties of the constituents of its wall, rather than by a decreasing elastin content.

Topics: Adolescent; Adult; Aged; Aging; Aorta, Thoracic; Arteriosclerosis; Densitometry; Elastin; Female; Humans; Hypertension, Pulmonary; Male; Middle Aged; Pulmonary Artery; Surface Properties

Topics: Altitude; Animals; Atmosphere Exposure Chambers; Carotid Body; Elastic Tissue; Elastin; Heart Ventricles; Hypertension, Pulmonary; Hypertrophy; Hypoxia; Male; Muscle, Smooth; Organ Size; Pulmonary Artery; Pulmonary Heart Disease; Rats

Topics: Adolescent; Adult; Age Factors; Aged; Aorta; Child; Child, Preschool; Elastin; Female; Fluorescence; Formates; Heart Defects, Congenital; Humans; Hypertension, Pulmonary; Infant; Infant, Newborn; Male; Middle Aged; Pulmonary Artery; Pulmonary Valve Stenosis; Tyrosine