elastin and Hypoxia

Obstructive sleep apnea (OSA) is highly prevalent in the USA and is recognized as an independent risk factor for atherosclerotic cardiovascular disease. Identification of atherosclerosis risk factor attributable to OSA may provide opportunity to develop preventive measures for cardiovascular risk reduction. Chronic intermittent hypoxia (CIH) is a prominent feature of OSA pathophysiology and may be a major mechanism linking OSA to arteriosclerosis. Animal studies demonstrated that CIH exposure facilitated high-cholesterol diet (HCD)-induced atherosclerosis, accelerated the progression of existing atherosclerosis, and induced atherosclerotic lesions in the absence of other atherosclerosis risk factors, demonstrating that CIH is an independent causal factor of atherosclerosis. Comparative studies revealed major differences between CIH-induced and the classic HCD-induced atherosclerosis. Systemically, CIH was a much weaker inducer of atherosclerosis. CIH and HCD differentially activated inflammatory pathways. Histologically, CIH-induced atherosclerotic plaques had no clear necrotic core, contained a large number of CD31+ endothelial cells, and had mainly elastin deposition, whereas HCD-induced plaques had typical necrotic cores and fibrous caps, contained few endothelial cells, and had mainly collagen deposition. Metabolically, CIH caused mild, but HCD caused more severe dyslipidemia. Mechanistically, CIH did not, but HCD did, cause macrophage foam cell formation. NF-κB p50 gene deletion augmented CIH-induced, but not HCD-induced atherosclerosis. These differences reflect the intrinsic differences between the two types of atherosclerosis in terms of pathological nature and underlying mechanisms and support the notion that CIH-induced atherosclerosis is a new paradigm that differs from the classic HCD-induced atherosclerosis.

Topics: Animals; Atherosclerosis; Cholesterol, Dietary; Collagen; Diet, Atherogenic; Disease Models, Animal; Elastin; Foam Cells; Gene Knockout Techniques; Humans; Hypoxia; NF-kappa B; Risk Factors; Signal Transduction; Sleep Apnea, Obstructive

The microangiopathy under hyperglycemia and diabetes develops only in the microcirculation component of circulatory system. In this area considerable amount of pericytes is concentrated. These cells contain myofibrils and in circulatory mode envelop capillaries being situated on the outside of basilemma. It is possible that in a phylogenetic sense this is the earliest functional unity of endothelium monolayer as a pacemaker and pericytes as contractile elements which are the earliest "propeller" because of implementing the function of advancement of lymph, hemolymph and blood in capillaries. Probably, endothelium and pericytes formed the first variation of peristaltic "pump" for the purpose of blood advancement longwise of capillaries. Most probably, the state of distal part of arterial race (muscular type arterioles) impact the parameters of proximal part of arterial race (elastic type arterioles) and myocardium itself in the same extent as the state of "pump" in capillaries, endothelium and pericytes function impact the function of local peristaltic pumps (muscular type arterioles) in paracrine cenosis. It is supposed that the pericytes are the regulators of physical, hydraulic factor of activation of biologic reaction of transcitosis--excretion of nutrients and humoral mediators from capillaries to the pool of intercellular medium to perform the biologic function of homeostasis. Hyperglycemia, glycotoxins formation, bivalent substances (glyoxal, methilglyoxal, malonic dialdehyde) reacting simultaneously by both ends of molecule result in formation within collagen of areolar tissue of short transversal cross-links (glycosylation end product) which significantly increase rigidity (hardness) of capillary wall. In these conditions, myofibrils of pericytes no longer form directed deformation of capillary wall to effect peristalsis and advancement of hemolymph (blood later on) along capillaries according the synthesis of monolayer endothelium NO as a dilatation factor. This is the cause of blood circulation disturbance on the level of exchange capillaries and formation of chronic hypoxemia resulting in the only increase of rate of glycosylation chemical reaction. The microangiopathy is formed in the cells and tissues in an integrated pool of intercellular medium and never occurs in the cerebrospinal fluid pool where no hyperglycemia develops.

Topics: Arterioles; Capillaries; Cell Communication; Collagen; Diabetes Mellitus; Diabetic Angiopathies; Elastin; Extracellular Matrix; Glycation End Products, Advanced; Glycosylation; Humans; Hyperglycemia; Hypoxia; Pericytes

During the past few years, proteomics has been extensively applied to various fields of medicine including nephrology. Current applications of renal and urinary proteomics are to better understand renal physiology, to explore the complexity of disease mechanisms, and to identify novel biomarkers and new therapeutic targets. This review provides some examples and perspectives of how proteomics can be applied to nephrology and how experimental data can be linked to physiology, functional significance and clinical applications. In some instances, proteomic analysis can be utilized to generate a new hypothesis from a set of candidates that are obtained from expression studies. The new hypothesis can then be addressed rapidly by conventional molecular biology methods, as demonstrated by identification of an altered renal elastin-elastase system in diabetic nephropathy and alterations in the renal kallikrein-kallistatin pathway in hypoxia-induced hypertension. The strengths and limitations of proteomics in renal research are summarized. Optimization of analytical protocols is required to overcome current limitations. Applications of proteomics to nephrology will then be more fruitful and successful.

Topics: Animals; Biomarkers; Carrier Proteins; Diabetic Nephropathies; Elastin; Gene Expression Regulation; Graft Rejection; Humans; Hypertension; Hypoxia; Kallikreins; Kidney; Kidney Transplantation; Models, Biological; Pancreatic Elastase; Proteomics; Serpins; Urine

Bronchopulmonary dysplasia (BPD) is a chronic lung disease that occurs in very premature infants and is characterized by impaired alveologenesis. This ultimate phase of lung development is mostly postnatal and allows growth of gas-exchange surface area to meet the needs of the organism. Alveologenesis is a highly integrated process that implies cooperative interactions between interstitial, epithelial, and vascular compartments of the lung. Understanding of its underlying mechanisms has considerably progressed recently with identification of structural, signaling, or remodeling molecules that are crucial in the process. Thus, the pivotal role of elastin deposition in lung walls has been demonstrated, and many key control-molecules have been identified, including various transcription factors, growth factors such as platelet-derived growth factor, fibroblast growth factors, and vascular endothelial growth factor, matrix-remodeling enzymes, and retinoids. BPD-associated changes in lung expression/content have been evidenced for most of these molecules, especially for signaling pathways, through both clinical investigations in premature infants and the use of animal models, including the premature baboon or lamb, neonatal exposure to hyperoxia in rodents, and maternal-fetal infection. These findings open therapeutic perspectives to correct imbalanced signaling. Unraveling the intimate molecular mechanisms of alveolar building appears as a prerequisite to define new strategies for the prevention and care of BPD.

Topics: Animals; Bronchopulmonary Dysplasia; Cell Differentiation; Cell Proliferation; Elastin; Fibroblast Growth Factors; Humans; Hypoxia; Infant, Newborn; Lung; Models, Biological; Platelet-Derived Growth Factor; Premature Birth; Pulmonary Alveoli; Pulmonary Gas Exchange; Retinoids; Signal Transduction; Vascular Endothelial Growth Factor A

Topics: Animals; Biological Factors; Bronchi; Child; Disease Models, Animal; Elastin; Extracellular Matrix; Humans; Hypoxia; Infant; Infant, Newborn; Infant, Premature, Diseases; Lung; Lung Diseases; Lung Transplantation; Mechanotransduction, Cellular; Oxygen; Pluripotent Stem Cells; Pulmonary Alveoli; Pulmonary Artery; Respiration; Species Specificity

Topics: Animals; Elastic Tissue; Elastin; Fibrillin-2; Fibroblasts; Gene Silencing; Humans; Hypoxia; Mice; Protein-Lysine 6-Oxidase; Ulcer

Chronic intermittent hypoxia (CIH) is a major determinant of the cardiovascular morbidity associated with obstructive sleep apnoea (OSA), and the magnitude of CIH impact may be influenced by ageing. Here, we assessed the role of ageing in the early cardiovascular structural remodelling induced by severe CIH in a murine model of OSA.. Early vascular remodelling was observed in young mice exposed to CIH as illustrated by intima-media thickening (mean change: 4.6 ± 2.6 μm; P = 0.02), elastin fibre disorganization (mean change: 9.2 ± 4.5%; P = 0.02) and fragmentation (mean change: 2.5 ± 0.8%; P = 0.03), and collagen (mean change: 3.2 ± 0.6%; P = 0.001) and mucopolysaccharide accumulation (mean change: 2.4 ± 0.8%; P = 0.01). In contrast, vascular remodelling was not apparent in aged mice exposed to CIH. Furthermore, left ventricular perivascular fibrosis (mean change: 0.71 ± 0.1; P < 0.001) and hypertrophy (mean change: 0.17 ± 0.1; P = 0.038) were increased by CIH exposure in young mice, but not in aged mice. Principal component analysis identified similar cardiovascular alterations among the young mice exposed to CIH and both older mouse groups, suggesting that CIH induces premature cardiovascular senescence.. Cardiovascular remodelling induced by severe CIH is affected by the age at which CIH onset occurs, suggesting that the deleterious cardiovascular effects associated with CIH may be more pronounced in younger populations, and such changes resemble chronological age-related declines in cardiovascular structural integrity.

Topics: Age Factors; Aging; Animals; Chronic Disease; Collagen; Disease Models, Animal; Elastin; Female; Glycosaminoglycans; Hypoxia; Mice; Mice, Inbred C57BL; Sleep Apnea, Obstructive; Tunica Intima; Vascular Remodeling

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

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

Preterm infants are at high risk for long-term abnormalities in cardiopulmonary function. Our objectives were to determine the long-term effects of hypoxia or hyperoxia on cardiopulmonary development and function in an immature animal model. Newborn C57BL/6 mice were exposed to air, hypoxia (12% oxygen), or hyperoxia (85% oxygen) from Postnatal Day 2-14, and then returned to air for 10 weeks (n = 2 litters per condition; > 10/group). Echocardiography, blood pressure, lung function, and lung development were evaluated at 12-14 weeks of age. Lungs from hyperoxia- or hypoxia-exposed mice were larger and more compliant (compliance: air, 0.034 ± 0.001 ml/cm H2O; hypoxia, 0.049 ± 0.002 ml/cm H2O; hyperoxia, 0.053 ± 0.002 ml/cm H2O; P < 0.001 air versus others). Increased airway reactivity, reduced bronchial M2 receptor staining, and increased bronchial α-smooth muscle actin content were noted in hyperoxia-exposed mice (maximal total lung resistance with methacholine: air, 1.89 ± 0.17 cm H2O ⋅ s/ml; hypoxia, 1.52 ± 0.34 cm H2O ⋅ s/ml; hyperoxia, 4.19 ± 0.77 cm H2O ⋅ s/ml; P < 0.004 air versus hyperoxia). Hyperoxia- or hypoxia-exposed mice had larger and fewer alveoli (mean linear intercept: air, 40.2 ± 0. 0.8 μm; hypoxia, 76.4 ± 2.4 μm; hyperoxia, 95.6 ± 4.6 μm; P < 0.001 air versus others; radial alveolar count [n]: air, 11.1 ± 0.4; hypoxia, 5.7 ± 0.3; hyperoxia, 5.6 ± 0.3; P < 0.001 air versus others). Hyperoxia-exposed adult mice had left ventricular dysfunction without systemic hypertension. In conclusion, exposure of newborn mice to hyperoxia or hypoxia leads to cardiopulmonary abnormalities in adult life, similar to that described in ex-preterm infants. This animal model may help to identify underlying mechanisms and to develop therapeutic strategies for pulmonary morbidity in former preterm infants.

Topics: Actins; Age Factors; Animals; Animals, Newborn; Blood Pressure; Bronchial Hyperreactivity; Bronchoconstriction; Cardiovascular System; Collagen; Disease Models, Animal; Elastin; Hyperoxia; Hypoxia; Lung; Lung Compliance; Mice, Inbred C57BL; Receptor, Muscarinic M2; Time Factors; Ventricular Dysfunction, Left; Ventricular Function, Left

The ductus arteriosus (DA), a fetal arterial connection between the main pulmonary artery and the descending aorta, normally closes immediately after birth. The oxygen concentration in the blood rises after birth, and in the DA this increase in oxygen concentration causes functional closure, which is induced by smooth muscle contraction. Previous studies have demonstrated that hypoxia and/or oxygenation affect vascular remodeling of various vessels. Therefore, we hypothesized that the rise in oxygen concentration would affect the vascular structure of the DA due to production of proteins secreted from DA smooth muscle cells (SMC).. Liquid chromatography-tandem mass spectrometry was used to comprehensively investigate the secreted proteins in the supernatant of rat DA SMC harvested under hypoxic conditions (1% oxygen) or under normoxic conditions (21% oxygen). We found that the rise in oxygen concentration reduced the secretion of elastin from DA SMC. On reverse transcription-polymerase chain reaction, the expression of elastin mRNA was not significantly changed in DA SMC from hypoxic to normoxic conditions.. Given that elastin forms internal elastic lamina and elastic fibers in the vascular muscle layers, and that a rise in oxygen concentration reduced the secretion of elastin, this suggests that the rise in blood oxygen concentration after birth reduces the secretion of elastin, and therefore may play a role in DA structural remodeling after birth.

Topics: Animals; Blotting, Western; Cells, Cultured; Disease Models, Animal; Ductus Arteriosus; Elastin; Female; Gas Chromatography-Mass Spectrometry; Gene Expression Regulation, Developmental; Hypoxia; Muscle, Smooth, Vascular; Oxygen; Oxygen Consumption; Pregnancy; Pregnancy, Animal; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA; Vascular Remodeling

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

This study sought to determine the effect of combined treatment of hypoxia plus indomethacin on pulmonary vascular remodeling in fetal rats.. Hypoxia and indomethacin were used to treat pregnant rats during 19-21 days of gestation. The adventitia, media, and intima of pulmonary arteries from fetal rats were assessed. Western blots were used for determining the abundance of smooth muscle specific alpha-actin protein (α-SMA), elastin, and endothelial nitric oxide synthase (eNOS) in lung tissues. Plasma brain-type natriuretic peptide (BNP) levels, reflecting the increased right ventricular load or pulmonary arterial pressure, were detected.. The ratio of left ventricular free wall plus septum to right ventricular weight significantly increased in hypoxia plus indomethacin-treated group. The medial thickness percentage of pulmonary arteries of < 100 μm and ≥100 μm in diameter from hypoxia plus indomethacin-treated group was higher than that from control or single treatment group. Vascular elastin area percentage and immunostaining density of eNOS from the combined-treated group were higher than other groups. The relative abundance of α-SMA, elastin, and eNOS and plasma BNP levels in hypoxia plus indomethacin-treated group also significantly increased compared with other groups.. Hypoxia and indomethacin had synergistic effect on fetal pulmonary vascular remodeling. This rat model induced by combined treatments can mimic human persistent pulmonary hypertension of the newborn.

Topics: Actins; Animals; Animals, Newborn; Blotting, Western; Cardiovascular Agents; Disease Models, Animal; Elastin; Female; Humans; Hypoxia; Indomethacin; Infant, Newborn; Lung; Myocardium; Natriuretic Peptide, Brain; Nitric Oxide Synthase Type III; Persistent Fetal Circulation Syndrome; Pulmonary Artery; Rats; Rats, Sprague-Dawley

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

Hypoxia impairs normal neonatal pulmonary artery remodeling and alveolar development. Matrix metalloproteinase-2 (MMP-2), which regulates collagen breakdown, is important during development. Our objective was to test the hypothesis that hypoxia attenuates the normal postnatal increase in MMP-2 and evaluate alveolar development and pulmonary arterial remodeling in Mmp2 mice. C57BL/6 wild-type (WT), Mmp2, Mmp2, and MMP-inhibited (with doxycycline) mice were exposed to hypoxia (12% O2) or air from birth to 2 wk of age. Pulmonary arterial remodeling, alveolar development, and vascular collagen and elastin were evaluated. MMP-2 was estimated by quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, immunohistochemistry, and zymography. We observed that 1) in WT mice, hypoxia led to thicker-walled pulmonary arteries and impaired alveolarization, accompanied by decreased MMP-2 and increased tissue inhibitor of metalloproteinases-2 (TIMP-2); 2) Mmp2 mice in air had thicker-walled arteries, impaired alveolarization, and increased perivascular collagen and elastin compared with WT; 3) hypoxia further inhibited alveolarization but did not alter arterial thickening in Mmp2 mice. Mmp2 and MMP-inhibited mice also had thicker-walled arteries than WT in air, but alveolarization was not different. We conclude that hypoxia reduces the postnatal MMP-2 increase in the lung, which may contribute to abnormal pulmonary arterial remodeling and impaired alveolarization.

Topics: Animals; Animals, Newborn; Collagen; Disease Models, Animal; Down-Regulation; Doxycycline; Elastin; Gene Expression Regulation, Enzymologic; Hypoxia; Lung; Matrix Metalloproteinase 2; Matrix Metalloproteinase Inhibitors; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Protease Inhibitors; Pulmonary Alveoli; Pulmonary Artery; RNA, Messenger; Time Factors; Tissue Inhibitor of Metalloproteinase-2

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

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

Endothelin-1 (ET-1) mediates hypoxia-mediated pulmonary vascular remodeling (HPVR), and endothelin-A receptor (ET-AR) blockade prevents HPVR in newborn mice. Our objective was to determine postnatal effects of chronic hypoxia and/or ET-AR blockade on lung ET-1, ET-AR, ET-BR, and vascular collagen and elastin. Newborn C57BL/6 mice (n = 6-8/gp) given either BQ610 (ET-AR blocker) or vehicle were exposed to air or hypoxia (12% O2) from birth for 1, 3, or 14 d. Lung ET-1 was assessed by ELISA, and ET-AR and ET-BR by immunohistochemistry. Vascular collagen and elastin were assessed by quantitative image analysis. ET-1, ET-AR, ET-BR, collagen I and III, and tropoelastin mRNA levels were assessed by real-time quantitative RT-PCR. We observed that: 1) hypoxia attenuated the normal postnatal decrease in ET-1 and collagen content; 2) ET-AR blockade reduced collagen independent of O2; 3) hypoxia increased elastin mRNA expression and attenuated the normal postnatal decrease in elastin content; and 4) BQ610 reduced elastin mRNA but not elastin content. We conclude that, in neonatal mice, hypoxia attenuates normal postnatal decreases in ET-1, vascular collagen, and elastin. ET-AR blockade reduces collagen fiber area but not mRNA, and does not decrease elastin despite reducing its expression.

Topics: Animals; Animals, Newborn; Blood Vessels; Collagen; Elastin; Endothelin A Receptor Antagonists; Endothelin B Receptor Antagonists; Endothelin-1; Female; Hypoxia; Lung; Mice; Mice, Inbred C57BL; Oligopeptides; Oxygen; Pregnancy; Receptor, Endothelin A; Receptor, Endothelin B

Primary pulmonary hypertension is a rare but deadly disease. Lungs extracted from PPH patients are deficient in endothelial nitric oxide synthase (eNOS), making the eNOS-null mouse a potentially useful model of the disease. To better understand the progression of pulmonary vascular remodeling in the congenital absence of eNOS, we induced pulmonary hypertension in eNOS-null mice using hypobaric hypoxia, and then quantified large artery structure and function in contralateral vessels. In particular, to assess structure we quantified diameter, wall thickness, and collagen, elastin and smooth muscle cell content; to quantify function we performed pressure-diameter tests. After remodeling, the pulmonary arteries had increased wall, collagen and elastin thicknesses compared to controls (P<0.05). The remodeled pulmonary arteries also had increased elastic moduli at low and high strains compared to controls (P<0.05). The increases in moduli at low and high strain correlated with increases in elastin and collagen thickness, respectively (P<0.05). These results provide insight into the mechanobiology of pulmonary vascular remodeling in the congenital absence of eNOS, and the coupled nature of these changes.

Topics: Animals; Biomechanical Phenomena; Collagen; Elastin; Female; Frozen Sections; Histocytochemistry; Hypoxia; Lung; Male; Mice; Mice, Knockout; Muscle, Smooth, Vascular; Nitric Oxide Synthase Type III; Pulmonary Artery; Random Allocation; Time Factors; Tomography, X-Ray Computed; Tunica Intima

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

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

Adaptive changes in cellular and connective tissue components of the lung after chronic exposure to reduced ambient oxygen and/or pressure were studied. Four-week-old male Sprague-Dawley rats were randomly divided into five groups (n = 12 each): 1) general control, room air (GC); 2) hypobaric normoxic; 3) normobaric hypoxic; 4) hypobaric hypoxic; and 5) weight-matched control, restricted food intake (WMC; weight matched to hypobaric hypoxic animals). Lung growth (lung weight and DNA, RNA, protein, hydroxyproline, and desmosine contents) diminished in WMC compared with GC. Somatic growth decreased in hypobaric and normobaric hypoxic rats compared with GC. Lung weight; DNA, RNA, protein, hydroxyproline, and desmosine contents; and RNA/DNA, protein/DNA, and desmosine/DNA ratios increased in both hypobaric and normobaric hypoxic rats compared with WMC. Hydroxyproline and desmosine contents and the hydroxyproline/DNA ratio were significantly higher in hypobaric than normobaric hypoxic rats. Hypobaric normoxia caused a slight somatic growth reduction, but biochemical parameters of lung growth remained unaffected. In conclusion, in growing animals, despite inhibition of lung growth due to reduced food consumption, accelerated lung growth in hypobaric or normobaric hypoxia occurs by hyperplastic and hypertrophic changes. Hypobaric normoxia does not affect lung growth, but connective tissue proteins accumulate slightly more in hypobaric hypoxia than in hypoxia alone.

Topics: Animals; Atmospheric Pressure; Body Weight; Collagen; DNA; Eating; Elastin; Hematocrit; Hypoxia; Lung; Male; Proteins; Rats; Rats, Sprague-Dawley; RNA

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

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

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

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: 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