elastin and Aortic-Valve-Stenosis

Calcific aortic stenosis is a progressive disease characterized by massive fibrosis andmineralization of the valve leaflets. The aim of this study was to determine whether the onset of native calcific aortic stenosis is associated primarily with matrix remodelling events, and particularly with elastin degradation.. The immunohistochemical expression profile of matrix degradating enzymes and tenascin-C was investigated in both healthy and native calcified aortic valves. Collagen and elastic tissue were studied by light microscopy and electron microscopy. Immunophenotypic analysis of inflammatory cells was carried out by using monoclonal antibodies to macrophages, T and B lymphocytes. Immunoreactivity for tenascin-C and matrix metalloproteinase-12 (MMP-12) was associated with areas of dense mineralization, which were characterized by fibrosis, fragmentation and calcification of elastic fibres a positive reaction was also found around small islands of calcification. MMP-11 was not detected in the diseased valves. Osteopontin and osteonectin were also found at sites of mineralization. All calcified valves examined showed inflammatory cell infiltration.. Our results demonstrate the direct involvement of MMP-12 in native aortic valve stenosis. MMP-mediated degradation of elastic fibres might contribute actively to valve mineralization by inducing calcium deposition onto fragmented elastin.

Topics: Aged; Aged, 80 and over; Aortic Valve; Aortic Valve Stenosis; Calcinosis; Elastin; Female; Humans; Immunohistochemistry; Inflammation; Male; Matrix Metalloproteinase 12; Microscopy, Electron, Transmission; Middle Aged; Osteonectin; Osteopontin; Tenascin

Elastin, the protein responsible for the elastic properties of vertebrate tissues, has been thought to be solely restricted to that role. As a consequence, elastin was conventionally described as an amorphous polymer. Recent results in the biomedical, biochemical and biophysical fields have lead to the conclusion that the presence of elastin in the extracellular space has very complex implications involving many other molecules. The present review describes the current state of knowledge concerning elastin as an elastic macromolecule. First, the genetic, biological, biochemical and biophysical processes leading to a functional polymer are described. Second, the elastic function of elastin is discussed. The controversy on elastin structure and elasticity is discussed and a novel dynamic mechanism of elasticity proposed. Finally, pathologies where the elastin molecule is involved are considered. This updated description of functional elastin provides the required background for the understanding of its pathologies and defines clearly the properties a substance should possess to be qualified as a good elastic biomaterial.

Topics: Animals; Aortic Valve Stenosis; Cutis Laxa; Elastic Tissue; Elastin; Humans; Skin Diseases; Structure-Activity Relationship; Williams Syndrome

Supravalvular aortic stenosis (SVAS) occurs as an autosomal dominant trait or as part of the phenotype of the usually sporadic condition Williams syndrome. SVAS is the result of mutation or deletion of the elastin gene (ELN), located at chromosome 7q11.23. Thus, SVAS may be more appropriately termed an elastin arteriopathy. Studies have demonstrated various point mutations and intragenic deletions of ELN resulting in nonsyndromic SVAS. Individuals with Williams syndrome are hemizygous for the elastin gene, owing to a 1 to 2 megabase deletion of a portion of the long arm of chromosome 7 that encompasses ELN. This submicroscopic deletion is readily detected by fluorescent in-situ hybridization, useful in the diagnosis of Williams syndrome. The severity of SVAS is quite variable, both in series of Williams syndrome patients and within SVAS kindreds, suggesting that other genetic factors are involved in expression of the phenotype. Experiments with elastin knockout mice will likely yield clues regarding the role of elastin in arterial morphogenesis and the pathogenesis of obstructive vascular disease.

Topics: Animals; Aortic Valve Stenosis; Chromosome Aberrations; Chromosome Deletion; Chromosome Disorders; Chromosomes, Human, Pair 7; DNA Mutational Analysis; Elastin; Gene Expression; Genes, Dominant; Humans; Mice; Phenotype; Williams Syndrome

Williams syndrome is a complex developmental disorder. The major cardiovascular component of Williams syndrome is supravalvular aortic stenosis, a progressive disease that may need surgical repair. Williams syndrome is associated with heterozygous microdeletion in the chromosomal region 7q11.23 encompassing the elastin gene. We have identified a new, highly informative tetranucleotide repeat polymorphism within the human elastin gene. This marker together with other, previously described elastin gene markers was used to show deletion of the elastin gene in nine sporadic Williams syndrome patients from Hungary. Application of polymorphisms within and flanking the elastin gene on chromosome 7 provides a fast, polymerase chain reaction based method for mutational analysis of Williams syndrome patients.

Topics: Aortic Valve Stenosis; Child; Child, Preschool; Chromosomes, Human, Pair 7; Elastin; Female; Genetic Markers; Heterozygote; Humans; Hungary; Male; Pedigree; Polymerase Chain Reaction; Polymorphism, Genetic; Williams Syndrome

Familial supravalvular aortic stenosis is a rare autosomal dominant condition. It may be distinguished from the Williams-Beuren syndrome by the absence of the characteristic dysmorphic appearances and of mental retardation. The case of a 9-year-old girl with a severe surgical stenosis led to the diagnosis of the same malformation in the mother and two brothers. This family adds to the 121 cases reported in the literature describing the main features of SVAS. Molecular biological advances have shown that familial SVAS and the Williams syndrome are due to mutation of the elastin gene located at 7q11-23. In the Williams syndrome the allele of this gene is completely absent and there is also probably deletion of contiguous genes, which explains involvement of cognitive function. In SVAS, the genetic lesion, mutation or microdeletion is more limited, explaining the usually isolated aortic malformation. Other studies are necessary to confirm these results.

Topics: Adult; Angiocardiography; Aortic Valve Stenosis; Child; Chromosomes, Human, Pair 7; Diagnosis, Differential; Elastin; Female; Genetic Techniques; Genotype; Humans; Male; Mutation; Pedigree; Sequence Deletion; Williams Syndrome

We have used single-strand conformation and heteroduplex analyses of genomic amplimers to identify point mutations within the elastin gene (ELN) in patients with non-syndromic supravalvular aortic stenosis (SVAS) from a total of eight unrelated families. Six novel point mutations were identified. We have collected detailed clinical information on mutation carriers and demonstrated significant non-penetrance in some of the families. Together with the new mutations described here, 14 point mutations have been reported in SVAS patients, and 10 of these result in premature stop codons (PTCs). We have analyzed the expression of ELN alleles in skin fibroblasts from one SVAS patient and shown that PTC mutations indeed result in selective elimination of mutant transcripts. Inhibition of the nonsense-mediated decay mechanism by cycloheximide resulted in the stabilization of mutant elastin mRNA. Allelic inactivation by the ELN mutation in this patient led to an overall decrease of the steady state levels of elastin mRNA. Finally, we have demonstrated reduced synthesis and secretion of tropoelastin by skin fibroblasts from the same SVAS patient. We conclude that PTC mutations in ELN result in nonsense-mediated decay of mutant mRNA in this patient. Given the predominance of PTC mutations in SVAS, we suggest that functional haploinsufficiency may be a pathomechanism underlying most cases of non-syndromic SVAS.

Topics: Adolescent; Adult; Aged; Alleles; Aortic Valve Stenosis; Cells, Cultured; Child; Child, Preschool; Cycloheximide; Elastin; Female; Fibroblasts; Gene Frequency; Gene Silencing; Genetic Carrier Screening; Genetic Testing; Humans; Infant; Male; Middle Aged; Penetrance; Point Mutation; Polymorphism, Genetic; RNA Processing, Post-Transcriptional; RNA, Messenger

Calcified aortic valve disease in its final stage leads to aortic valve stenosis, limiting cardiac function. To date, surgical intervention is the only option for treating calcific aortic valve stenosis. This study combined controlled drug delivery by nanoparticles (NPs) and active targeting by antibody conjugation. The chelating agent diethylenetriaminepentaacetic acid (DTPA) was covalently bound to human serum albumin (HSA)-based NP, and the NP surface was modified using conjugating antibodies (anti-elastin or isotype IgG control). Calcification was induced ex vivo in porcine aortic valves by preincubation in an osteogenic medium containing 2.5 mM sodium phosphate for five days. Valve calcifications mainly consisted of basic calcium phosphate crystals. Calcifications were effectively resolved by adding 1-5 mg DTPA/mL medium. Incubation with pure DTPA, however, was associated with a loss of cellular viability. Reversal of calcifications was also achieved with DTPA-coupled anti-elastin-targeted NPs containing 1 mg DTPA equivalent. The addition of these NPs to the conditioned media resulted in significant regression of the valve calcifications compared to that in the IgG-NP control without affecting cellular viability. These results represent a step further toward the development of targeted nanoparticular formulations to dissolve aortic valve calcifications.

Topics: Animals; Aortic Valve; Aortic Valve Stenosis; Elastin; Humans; Immunoglobulin G; Nanoparticles; Pentetic Acid; Swine

Clinical studies have revealed a greater risk of pulmonary autograft dilation after the Ross procedure in patients with preoperative aortic insufficiency (AI). The present study examined whether the morphologic, biomechanical, and cellular properties of the pulmonary artery (PA) from patients with AI were phenotypically different compared with patients diagnosed with aortic stenosis (AS).. PA segments were harvested from patients undergoing the Ross procedure for AS (n = 16) and AI (n = 6). Preoperative aortic annulus was significantly larger (P < 0.05) in patients with AI (28.5 ± 1.8 mm) vs AS (22.8 ± 1.2 mm). Morphologic, biomechanical, and cellular phenotypes of the PA were analyzed.. Collagen and elastin content in the media of the PA wall were similar in patients with AS and AI. Elastic modulus and energy loss of the PA were not significantly different between the groups. In the media of the PA, expression of a panel of vascular smooth muscle cell-specific proteins were similar in patients with AS and AI. In contrast, nonmuscle myosin IIB protein levels in the PA of AS patients were significantly higher compared with AI patients, and immunofluorescence identified staining in α-smooth muscle actin-positive vascular smooth muscle cells.. Despite similar morphological and biomechanical properties, the disparate expression of nonmuscle myosin IIB protein distinguishes the PA of patients with AI from patients with AS. The biological role in vascular smooth muscle cells and the potential contribution of nonmuscle myosin IIB to pulmonary autograft dilation in a subset of AI patients after the Ross procedure remain to be determined.

Topics: Actins; Aorta; Aortic Valve Insufficiency; Aortic Valve Stenosis; Autografts; Biomechanical Phenomena; Collagen; Echocardiography, Doppler; Elastic Modulus; Elastin; Female; Humans; Male; Middle Aged; Myocytes, Smooth Muscle; Nonmuscle Myosin Type IIB; Pulmonary Artery; Pulmonary Valve; Tunica Media

AR has a negative effect on the nondilated ascending aortic wall. Accordingly, our results support the need for more detailed studies of the aortic wall in relation to aortic valve disease and may ultimately lead to more aggressive clinical monitoring and/or surgical criteria for patients with relevant AR. Graphic Abstract: A graphic abstract is available for this article.

Topics: Adult; Aged; Aorta; Aortic Valve Insufficiency; Aortic Valve Stenosis; Apoptosis; Case-Control Studies; Collagen; Dilatation, Pathologic; Elastin; Endothelial Cells; Female; Fibrillins; Humans; Immunohistochemistry; Male; Middle Aged; Nitric Oxide Synthase Type III; Vascular Remodeling; Young Adult

Pore size is generally small in nanofibrous scaffolds prepared by electrospinning polymeric solutions. Increase of scaffold thickness leads to decrease in pore size, causing impediment to cell infiltration into the scaffolds during tissue engineering. In contrast, comparatively larger pore size can be realized in microfibrous scaffolds prepared from polymeric solutions at higher concentrations. Further, microfibrous scaffolds are conducive to infiltration of reparative M2 phenotype macrophages during in vivo/in situ tissue engineering. However, rise of mechanical properties of a fibrous scaffold with the increase of polymer concentration may limit the functionality of a scaffold-based, tissue-engineered heart valve. In this study, we developed microfibrous scaffolds from 14%, 16% and 18% (wt/v) polycaprolactone (PCL) polymer solutions prepared with chloroform solvent. Porcine valvular interstitial cells were cultured in the scaffolds for 14 d to investigate the effect of microfibers prepared with different PCL concentrations on the seeded cells. Further, fresh microfibrous scaffolds were implanted subcutaneously in a rat model for two months to investigate the effect of microfibers on infiltrated cells. Cell proliferation, and its morphologies, gene expression and deposition of different extracellular matrix proteins in the in vitro study were characterized. During the in vivo study, we characterized cell infiltration, and myofibroblast and M1/M2 phenotypes expression of the infiltrated cells. Among different PCL concentrations, microfibrous scaffolds from 14% solution were suitable for heart valve tissue engineering for their sufficient pore size and low but adequate tensile properties, which promoted cell adhesion to and proliferation in the scaffolds, and effective gene expression and extracellular matrix deposition by the cells in vitro. They also encouraged the cells in vivo for their infiltration and effective gene expression, including M2 phenotype expression.

Topics: Animals; Aortic Valve; Aortic Valve Stenosis; Calcinosis; Cell Adhesion; Cell Proliferation; Cells, Cultured; Collagen; Elastin; Extracellular Matrix; Glycosaminoglycans; Mesenchymal Stem Cells; Nanofibers; Polyesters; Polymers; Rats; Rats, Sprague-Dawley; Stress, Mechanical; Swine; Tensile Strength; Tissue Engineering; Tissue Scaffolds; Vimentin

Degenerative heart valve disease is a life-threatening disease affecting about 3% of the population over 65 years. Up to date, cardiac surgery with heart valve replacement is the only available therapy. The disease is characterized by degenerative disorganization of the heart valve structure and alterations in the residing cell populations. Causes and mechanisms of disease genesis are still not fully understood and until now pharmacological therapies are not available. Thus there is enormous interest in new technologies that enable a better characterization of structure and composition of diseased valves. Currently most research techniques demand for extensive processing of extracted valve material. We present a novel approach combining coherent anti-Stokes Raman scattering, endogenous two-photon excited fluorescence and second harmonic generation. Cusp constituents can be examined simultaneously, three-dimensionally and without extensive manipulation of the sample enabling impressive insights into a complex disease.

Topics: Adipocytes; Aged; Aged, 80 and over; Aorta; Aortic Valve; Aortic Valve Insufficiency; Aortic Valve Stenosis; Collagen; Compressive Strength; DNA; Elastin; Humans; Image Processing, Computer-Assisted; Lithostathine; Male; Microscopy, Fluorescence; Optics and Photonics; Photons; Spectrum Analysis, Raman

Currently, percutaneous aortic valve (PAV) replacement devices are being investigated to treat aortic stenosis in patients deemed to be of too high a risk for conventional open-chest surgery. Successful PAV deployment and function are heavily reliant on the tissue-stent interaction. Many PAV feasibility trials have been conducted with porcine models under the assumption that these tissues are similar to human; however, this assumption may not be valid. The goal of this study was to characterize and compare the biomechanical properties of aged human and porcine aortic tissues.. The biaxial mechanical properties of the left coronary sinus, right coronary sinus, non-coronary sinus, and ascending aorta of eight aged human (90.1 ± 6.8 years) and 10 porcine (6-9 months) hearts were quantified. Tissue structure was analyzed via histological techniques.. Aged human aortic tissues were significantly stiffer than the corresponding porcine tissues in both the circumferential and longitudinal directions (p < 0.001). In addition, the nearly linear stress-strain behavior of the porcine tissues, compared with the highly nonlinear response of the human tissues at a low strain range, suggested structural differences between the aortic tissues from these two species. Histological analysis revealed that porcine samples were composed of more elastin and less collagen fibers than the respective human samples.. Significant material and structural differences were observed between the human and porcine tissues, which raise questions on the validity of using porcine models to investigate the biomechanics involved in PAV intervention.

Topics: Aged; Aged, 80 and over; Aging; Animals; Aorta; Aortic Valve Stenosis; Collagen; Disease Models, Animal; Elasticity; Elastin; Female; Heart Valve Prosthesis Implantation; Humans; Male; Models, Cardiovascular; Sinus of Valsalva; Species Specificity; Stress, Mechanical; Sus scrofa

Clinical studies have demonstrated that 50% of individuals with chronic renal disease (CRD) die of cardiovascular causes, including advanced calcific arterial and valvular disease; however, the mechanisms of accelerated calcification in CRD remain obscure, and no therapies can prevent disease progression. We recently demonstrated in vivo that inflammation triggers cardiovascular calcification. In vitro evidence also indicates that elastin degradation products may promote osteogenesis. Here, we used genetically modified mice and molecular imaging to test the hypothesis in vivo that cathepsin S (catS), a potent elastolytic proteinase, accelerates calcification in atherosclerotic mice with CRD induced by 5/6 nephrectomy.. Apolipoprotein-deficient (apoE(-/-))/catS(+/+) (n=24) and apoE(-/-)/catS(-/-) (n=24) mice were assigned to CRD and control groups. CRD mice had significantly higher serum phosphate, creatinine, and cystatin C levels than those without CRD. To visualize catS activity and osteogenesis in vivo, we coadministered catS-activatable and calcification-targeted molecular imaging agents 10 weeks after nephrectomy. Imaging coregistered increased catS and osteogenic activities in the CRD apoE(-/-)/catS(+/+) cohort, whereas CRD apoE(-/-)/catS(-/-) mice exhibited less calcification. Quantitative histology demonstrated greater catS-associated elastin fragmentation and calcification in CRD apoE(-/-)/catS(+/+) than CRD apoE(-/-)/catS(-/-) aortas and aortic valves. Notably, catS deletion did not cause compensatory increases in RNA levels of other elastolytic cathepsins or matrix metalloproteinases. Elastin peptide and recombinant catS significantly increased calcification in smooth muscle cells in vitro, a process further amplified in phosphate-enriched culture medium.. The present study provides direct in vivo evidence that catS-induced elastolysis accelerates arterial and aortic valve calcification in CRD, providing new insight into the pathophysiology of cardiovascular calcification.

Topics: Animals; Aorta; Aortic Valve; Aortic Valve Stenosis; Apolipoproteins E; Calcinosis; Carotid Artery Diseases; Cathepsins; Cells, Cultured; Creatinine; Cystatin C; Elastin; Humans; Kidney Failure, Chronic; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Muscle, Smooth, Vascular; Nephrectomy; Osteogenesis; Phosphates; Tunica Intima; Tunica Media

Despite extensive strides in understanding pressure overload induced heart failure, there is very little known about oxidative stress induced matrix metalloproteinase (MMP) activation, collagen degradation and remodeling in pressure overload heart failure. We hypothesize that pressure overload leads to redox imbalance causing increased expression/activity of MMP-2/9 producing collagen degradation and heart failure. To test this hypothesis, we created pressure overload heart failure by abdominal aortic stenosis (AS) in wild-type C57BL/6J and collagen mutant (Col1a1 with 129 s background) mice. At 4 weeks, post surgery, functional parameters were measured. Left ventricle (LV) tissue sections were analyzed by histology, Western Blot and PCR. The results suggest an increase in iNOS with a decrease in eNOS, an increase in nitrated protein modification and depletion of antioxidants thioredoxin and SOD in pressure overload. MMP-2/9 expression/activity and collagen degradation were increased in the AS animals. To determine whether a mutation in the collagen gene at the site of MMP cleavage mitigates cardiac hypertrophy, we used Col1a1 mice. In these mice, the AS induced LV hypertrophy (LVH) was ameliorated. In conclusion, our results suggest that AS leads to increased oxidative stress, expression/activity of MMP-2/9 and a decrease in antioxidant expression producing collagen degradation and heart failure.

Topics: Analysis of Variance; Animals; Aortic Valve Stenosis; Blood Pressure; Blotting, Western; Chronic Disease; Collagen; Disease Models, Animal; Echocardiography; Elastin; Electrocardiography; Enzyme Activation; Heart Failure; Hypertrophy, Left Ventricular; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Oxidative Stress; Research Design; Reverse Transcriptase Polymerase Chain Reaction; Stroke Volume; Ventricular Dysfunction, Left

Topics: Animals; Aortic Valve Stenosis; Cardiac Output; Cell Proliferation; Elastin; Gene Expression Regulation; Genetic Therapy; Humans; Hypertension; Mice; Mice, Transgenic; Muscle, Smooth, Vascular; RNA, Messenger

Aortic stenosis (AS) is characterized by extensive remodelling of the valves, including infiltration of inflammatory cells, extracellular matrix degradation, and fibrosis. The molecular mechanisms behind this adverse remodelling have remained obscure. In this article, we study whether cathepsin G, an angiotensin II (Ang II)-forming elastolytic enzyme, contributes to progression of AS.. Stenotic aortic valves (n = 86) and control valves (n = 17) were analysed for cathepsin G, transforming growth factor-beta1 (TGF-beta1), and collagens I and III with RT-PCR and immunohistochemistry. Valvular collagen/elastin ratio was quantified by histochemistry. In stenotic valves, cathepsin G was present in mast cells and showed increased expression (P < 0.001), which correlated positively (P < 0.001) with the expression levels of TGF-beta1 and collagens I and III. TGF-beta1 was also present in mast cell-rich areas and cathepsin G induced losartan-sensitive TGF-beta1 expression in cultured fibroblasts. Collagen/elastin ratio was increased in stenotic valves (P < 0.001) and correlated positively with smoking (P = 0.02). Nicotine in cigarette smoke activated mast cells and induced TGF-beta1 expression in cultured fibroblasts. Fragmented elastin was observed in stenotic valves containing activated cathepsin G-secreting mast cells and in normal valves treated with cathepsin G.. In stenotic aortic valves, mast cell-derived cathepsin G may cause adverse valve remodelling and AS progression.

Topics: Adult; Aged; Aged, 80 and over; Aortic Valve Stenosis; Cathepsin G; Cathepsins; Collagen; Elastin; Female; Humans; Immunohistochemistry; Male; Mast Cells; Middle Aged; Nicotiana; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Serine Endopeptidases; Smoke; Transforming Growth Factor beta1

To investigate the possible role of elastolytic cathepsins S, K, and V and their endogenous inhibitor cystatin C in adverse extracellular matrix remodeling of stenotic aortic valves.. Stenotic aortic valves were collected at valve replacement surgery and control valves at cardiac transplantations. The expression of cathepsins S, K, and V and cystatin C was studied by conventional and real-time polymerase chain reaction and by immunohistochemistry. Total cathepsin activity in the aortic valves was quantified by a fluorometric microassay. When compared with control valves, stenotic valves showed increased mRNA expression of cathepsins S, K, and V (P<0.05 for each) and a higher total cathepsin activity (P<0.001). In stenotic valves, cystatin C mRNA was increased (P<0.05), and cystatin C protein was found particularly in areas with infiltrates of inflammatory cells. Both cathepsin S and cystatin C were present in bony areas of the valves, whereas cathepsin V localized to endothelial cells in areas rich of neovascularization. Incubation of thin sections of aortic valves with cathepsins S, K, and V resulted in severe disruption of elastin fibers, and this cathepsin effect could be blocked by adding cystatin C to the incubation system.. Stenotic aortic valves show increased expression and activity of elastolytic cathepsins S, K, and V. These cathepsins may accelerate the destruction of aortic valvular extracellular matrix, so promoting the progression of aortic stenosis.

Topics: Aortic Valve; Aortic Valve Stenosis; Case-Control Studies; Cathepsin K; Cathepsins; Cystatin C; Cystatins; Cysteine Endopeptidases; Elastin; Fluorometry; Humans; Immunohistochemistry; In Vitro Techniques; RNA, Messenger

Williams-Beuren syndrome is a rare and usually sporadic genetic anomaly with an estimated frequency of 1:25,000, that also has cardiac defects due to the effect on the elastin locus of a deletion on the 7th chromosome. Identical twin boys presented with exercise-induced syncope. Echocardiographic examination revealed severe calcification at the aortic valves, mitral anterior leaflets, and mitral annuli in both cases. A basal interventricular septum was also involved in one case. Doppler evaluation demonstrated severe aortic stenosis with a peak gradient of 112 and 118 mmHg in both cases. Moderate mitral stenosis was also detected in one twin. We performed aortic mechanical valve replacement and dilated the aortic annulus with Nick's procedure and evaluated the diagnosis and therapy methods in light of the literature. As a result, we determined that these boys are the first monozygotic twins who were diagnosed during childhood.

Topics: Aortic Valve; Aortic Valve Stenosis; Calcinosis; Diseases in Twins; Elastin; Gene Deletion; Heart Valve Prosthesis Implantation; Humans; Male; Mitral Valve Stenosis; Twins, Monozygotic; Williams Syndrome

We have screened the elastin gene for mutations responsible for supravalvular aortic stenosis (SVAS) in two large, independently collected families with isolated (nonsyndromic) SVAS. By single-strand conformation polymorphism and heteroduplex analysis, we have identified a C to G transversion within the acceptor splice site of exon 16 in SVAS patients from both families. This mutation segregates in both families with high penetrance of SVAS, and all affected individuals carry the mutation. Haplotype analysis by using closely linked polymorphisms, including a previously unreported BfaI restriction fragment length polymorphism within the 3'-UTR of the elastin gene, indicates that the mutations found in the two apparently non-overlapping kindreds are identical by descent. To study the effect of the mutation on the expression of the mutant allele, we have established a primary skin fibroblast culture from one of the affected individuals. Reverse transcription/polymerase chain reaction analysis of elastin mRNA species indicates that the mutation results in two abnormal elastin mRNA species. One mutant elastin mRNA is generated by the activation of a cryptic splice site that lies within intron 15 and that adds 44 bp of intronic sequence to the sequence encoded by exon 16. This insertion creates a frame shift that results in a 59-amino-acid-long abnormal protein sequence and leads to a termination codon in the mRNA sequence encoded by exon 17. The smaller abnormal mRNA species arises as a consequence of the skipping of exon 16. This study demonstrates, for the first time, the expression of mutant alleles of the elastin gene in patients with isolated SVAS.

Topics: Alternative Splicing; Aortic Valve Stenosis; Binding Sites; Elastin; Female; Gene Expression Regulation; Humans; Male; Mutation; Pedigree; RNA Splicing

A large supravalvular aortic stenosis kindred, with a point mutation in exon 18 and a stop codon in exon 22 of the elastin gene, is described. Clinically, the disease severity appeared to increase in successive generations in this family.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Aortic Valve Stenosis; Child; Child, Preschool; Codon, Terminator; Elastin; Exons; Female; Frameshift Mutation; Genotype; Haplotypes; Humans; Infant; Male; Middle Aged; Pedigree; Point Mutation

The elastin gene is consistently deleted in Williams syndrome and as this protein represents the major component of the elastic fibers of the dermis, we sought to investigate skin elastic fibers in Williams syndrome as a key to unraveling extracellular matrix disorganization in this condition. Both morphometric parameters analyzed by using automated image analysis and immunofluorescence labeling with monoclonal antibodies against elastin and fibrillin 1 showed a disorganized pre-elastic (oxytalan and elaunin) and mature elastic fibers in the dermis of 10 Williams syndrome patients compared with five healthy children and one patient with isolated supravalvular aortic stenosis. Skin biopsies in Williams syndrome patients provide a simple mean to elucidate extracellular matrix anomalies. Hopefully, this method could give clues to the understanding of the elastic network anomalies in this condition and even to the consequences of these latter on elasticity and resilience of other tissues such as the arterial tree.

Topics: Adolescent; Aortic Valve Stenosis; Child; Child, Preschool; Chromosomes, Human, Pair 7; Elastic Tissue; Elastin; Extracellular Matrix; Fibrillin-1; Fibrillins; Fluorescent Antibody Technique, Indirect; Humans; Microfilament Proteins; Skin Abnormalities; Williams Syndrome

Topics: Aortic Valve Stenosis; Elastin; Female; Gene Frequency; Genetic Markers; Homozygote; Humans; Male; Microsatellite Repeats; Pedigree; Polymerase Chain Reaction; Polymorphism, Genetic; Williams Syndrome

Both Williams syndrome and isolated supravalvular aortic stenosis (SVAS) are caused by mutations at the elastin locus. Deletion demonstrable by FISH is the hallmark of Williams syndrome, whereas the mutations reported so far in SVAS have been more subtle. FISH positive elastin hemizygosity has not been reported in isolated SVAS. This report records our experience of FISH for elastin deletion in isolated SVAS and specifically reports a patient with non-Williams related SVAS, positive for the elastin deletion by FISH.

Topics: Adolescent; Alleles; Aortic Valve Stenosis; Elastin; Humans; In Situ Hybridization, Fluorescence; Male; Williams Syndrome

Supravalvular aortic stenosis (SVAS) is an inherited obstructive vascular disease that affects the aorta, carotid, coronary and pulmonary arteries. Previous molecular genetic data have led to the hypothesis that SVAS results from mutations in the elastin gene, ELN. In these studies, the disease phenotype was linked to gross DNA rearrangements (35 and 85 kb deletions and a translocation) in three SVAS families. However, gross rearrangements of ELN have not been identified in most cases of autosomal dominant SVAS. To define the spectrum of ELN mutations responsible for this disorder, we refined the genomic structure of human ELN and used this information in mutational analyses. ELN point mutations co-segregate with the disease in four familial cases and are associated with SVAS in three sporadic cases. Two of the mutations are nonsense, one is a single base pair deletion and four are splice site mutations. In one sporadic case, the mutation arose de novo. These data demonstrate that point mutations of ELN cause autosomal dominant SVAS.

Topics: Aortic Valve Stenosis; Cloning, Molecular; DNA Mutational Analysis; DNA Primers; Elastin; Female; Frameshift Mutation; Humans; Male; Molecular Sequence Data; Mutation; Pedigree; Polymorphism, Single-Stranded Conformational; RNA Splicing

We describe the complete exon-intron structure of the human elastin (ELN) gene located at chromosome 7q11.23. There are 34 exons occupying approximately 47 kb of genomic DNA. All exons are in-frame, allowing exon skipping without disrupting the reading frame. Microsatellites are located in introns 17 and 18. Deletions of all or large parts of the ELN gene have been previously reported in two patients with supravalvular aortic stenosis (SVAS), and SVAS is also a frequent feature of Williams syndrome, where patients are hemizygous for ELN. We list primer pairs for amplifying each exon, with flanking intron, from genomic DNA to allow detection of point mutations in the ELN gene. We show that some patients with isolated SVAS have point mutations that are predicted to lead to premature chain termination. Knowledge of the genomic structure will allow more extensive mutation screening in genomic DNA of patients with SVAS and other conditions.

Topics: Amino Acid Sequence; Aortic Valve Stenosis; Base Sequence; Child, Preschool; Elastin; Exons; Genes, Dominant; Humans; Infant; Infant, Newborn; Introns; Male; Microsatellite Repeats; Molecular Sequence Data; Point Mutation; Repetitive Sequences, Nucleic Acid

To investigate the frequency of deletions of the elastin gene in patients with Williams syndrome (WS), we screened 44 patients by both FISH and PCR amplification of a dinucleotide repeat polymorphism. FISH was performed using cosmids containing either the 5' or the 3' end of the elastin gene. PCR analysis was performed on the patients and their parents with a (CA)n repeat polymorphism found in intron 17 of the elastin locus. Of the 44 patients screened, 91% were shown to be deleted by FISH. Using the DNA polymorphism, both maternally (39%) and paternally (61%) derived deletions were found. Four patients were not deleted for elastin but have clinical features of WS. Since deletions of elastin cannot account for several features found in WS, these patients will be valuable in further delineation of the critical region responsible for the WS phenotype. Although PCR can be useful for determining the parental origin of the deletion, our results demonstrate that FISH analysis of the elastin locus provides a more rapid and informative test to confirm a clinical diagnosis of WS. The presence of two copies of the elastin locus in a patient does not, however, rule out WS as a diagnosis.

Topics: Abnormalities, Multiple; Aortic Valve Stenosis; Chromosomes, Human, Pair 7; Elastin; Face; Female; Gene Deletion; Growth Disorders; Humans; In Situ Hybridization, Fluorescence; Intellectual Disability; Karyotyping; Male; Polymerase Chain Reaction

Although family history can be an important risk factor for cardiovascular disease, relatively little is known about the nature of specific genetic risk factors. One approach to this problem is to identify and characterize genes responsible for inherited disorders in the hope that this information will also provide mechanistic insight into common forms of cardiovascular disease.. Over the last decade, it has become possible to identify genes that cause human disease by use of the techniques of molecular genetics, specifically genetic linkage analysis, positional cloning, and mutational analyses. We have used these techniques to study three inherited cardiovascular disorders: supravalvular aortic stenosis, Williams syndrome, and long-QT syndrome. We have discovered that the vascular pathology of supravalvular aortic stenosis and Williams syndrome results from mutations involving the elastin gene on chromosome 7q11.23. These mutations include intragenic deletions, translocations, and complete deletion of the elastin gene, suggesting that a quantitative reduction in elastin during vascular development is pathogenically important. To date, only the elastin gene has proved important for supravalvular aortic stenosis. By contrast, genetic linkage analyses in families with long-QT syndrome indicate that at least four distinct genes can cause this disorder. We have identified three LQT loci: LQT1 on chromosome 11p15.5, LQT2 on 7q35-36, and LQT3 on 3p21-24. Recently, we demonstrated that mutations in a putative cardiac potassium channel gene, HERG, are responsible for the chromosome 7-linked form of long-QT syndrome, whereas mutations in the cardiac sodium channel gene SCN5A cause the chromosome 3-linked form of this disorder. HERG mutations and potassium channel biophysics suggest a dominant-negative molecular mechanism and reduced repolarization currents. By contrast, SCN5A mutations probably cause subtle alterations of cardiac sodium channel function and prolonged depolarizing currents.. Molecular genetic analyses of long-QT syndrome, supravalvular aortic stenosis, and Williams syndrome have begun to unravel the mechanisms underlying these inherited disorders. Rapid genetic testing for Williams syndrome is now available using a simple cytogenetic test, fluorescence in situ hybridization, but additional work will be required for long-QT syndrome and autosomal-dominant supravalvular aortic stenosis. Improved diagnosis and mechanistic understanding of these disorders should lead to rational treatment and prevention.

Topics: Aortic Valve Stenosis; Chromosome Mapping; Elastin; Electrocardiography; Humans; Long QT Syndrome; Mutation; Sodium Channels; Syndrome

Topics: Aortic Valve Stenosis; Base Sequence; DNA, Complementary; Elastin; Genotype; Humans; Molecular Sequence Data; Phenotype; Sequence Deletion

Since structure-function correlations have not been determined for cryopreserved allograft heart valves, we studied 20 explanted valves in place several hours to nine years, as either orthotopic aortic valves/root replacements or right ventricle to pulmonary artery conduits. They were explanted primarily due to growth-related conduit or valve stenosis, valve regurgitation, or infection. Controls included seven unused cryopreserved allograft valves and 16 aortic valves removed from transplanted allograft hearts obtained at either autopsy (n = 15) or retransplantation (n = 1), two days to four years postoperatively, following myocardial rejection (n = 4), graft coronary arteriosclerosis (n = 4), and other (n = 8). Analysis included gross inspection, radiography, light microscopy, electron microscopy, and immunohistochemical studies (to allow identification/localization of endothelial cells, mononuclear inflammatory cells, and T and B lymphocyte subsets). Cryopreserved allograft valves implanted more than one day had progressively severe loss of normal structure and were devoid of surface endothelium and stainable deep connective tissue cells. Inflammatory cellularity varied from none (most valves) to prominent (primarily T lymphocytes in one valve). Transmission electron microscopy of three long term valvular allografts revealed no viable cells, remarkable preservation of the collagenous valve matrix and focal cell-oriented calcification. In contrast, aortic valves from transplanted hearts showed remarkable structural preservation, including layered architecture, endothelium and deep connective tissue cells; inflammatory infiltrates were generally sparse, even in cases with fatal myocardial rejection. We conclude that cryopreserved allograft valves are morphologically non-viable valves, whose structural basis for function seems primarily related to the largely preserved collagen.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aortic Valve; Aortic Valve Insufficiency; Aortic Valve Stenosis; B-Lymphocyte Subsets; Calcinosis; Cell Survival; Collagen; Connective Tissue; Coronary Artery Disease; Cryopreservation; Elastin; Endocarditis, Bacterial; Endothelium; Follow-Up Studies; Graft Rejection; Humans; Immunohistochemistry; Leukocytes, Mononuclear; Microscopy, Electron; Pulmonary Valve; Pulmonary Valve Insufficiency; Pulmonary Valve Stenosis; Radiography; T-Lymphocyte Subsets; Tissue Preservation; Transplantation, Homologous

Topics: Aortic Valve Stenosis; Chromosomes, Human, Pair 7; Elastin; Female; Genetic Linkage; Humans; Male; Pedigree

Williams syndrome (WS) is a developmental disorder affecting connective tissue and the central nervous system. A common feature of WS, supravalvular aortic stenosis, is also a distinct autosomal dominant disorder caused by mutations in the elastin gene. In this study, we identified hemizygosity at the elastin locus using genetic analyses in four familial and five sporadic cases of WS. Fluorescent in situ hybridization and quantitative Southern analyses confirmed these findings, demonstrating inherited and de novo deletions of the elastin gene. These data indicate that deletions involving one elastin allele cause WS and implicate elastin hemizygosity in the pathogenesis of the disease.

Topics: Adult; Alleles; Aortic Valve Stenosis; Arteries; Blotting, Southern; Child; Child, Preschool; Chromosomes, Human, Pair 7; Connective Tissue Diseases; Developmental Disabilities; Elastin; Genes; Genotype; Humans; In Situ Hybridization, Fluorescence; Intellectual Disability; Pedigree; Sequence Deletion; Syndrome

To identify genes involved in vascular disease, we investigated patients with supravalvular aortic stenosis (SVAS), an inherited vascular disorder that causes hemodynamically significant narrowing of large elastic arteries. Pulsed-field gel and Southern analyses showed that a translocation near the elastin gene cosegregated with SVAS in one family. DNA sequence analyses demonstrated that the translocation disrupted the elastin gene and localized the breakpoint to exon 28. Taken together with our previous study linking SVAS to the elastin gene in two additional families and existing knowledge of vascular biology, these data suggest that mutations in the elastin gene can cause SVAS.

Topics: Amino Acid Sequence; Aortic Valve Stenosis; Base Sequence; Chromosome Mapping; Chromosomes, Human, Pair 6; Chromosomes, Human, Pair 7; Elastin; Female; Genetic Linkage; Humans; Hybrid Cells; Male; Molecular Sequence Data; Pedigree; Polymorphism, Restriction Fragment Length; Syndrome; Translocation, Genetic

A rare case of Williams syndrome diagnosed at the age of 41 is documented. The first subjective symptom was chest pain and the patient displayed many other features in addition to severe supravalvular aortic stenosis with a systolic gradient of 60 mmHg. The stenotic lesion had an area of 0.5 cm2, and was associated with dilated and tortuous coronary arteries. Extended aortoplasty was successfully performed and the postoperative course has been excellent without any cardiac symptoms. In spite of the severe cardiac lesions, this case had been largely asymptomatic and presented unusual features related to the diagnosis and management of this syndrome in an adult. The pattern of abnormalities found in this case suggested problems in relation to the calcitonin/calcitonin gene related peptide (CGRP) and the elastin gene occurring in embryonic organogenesis.

Topics: Abnormalities, Multiple; Adult; Aortic Valve Stenosis; Calcitonin Gene-Related Peptide; Elastin; Face; Female; Humans; Intellectual Disability; Syndrome; Time Factors

Supravalvular aortic stenosis (SVAS) is an autosomal dominant disorder characterized by abnormalities of development of the great vessels. SVAS is also commonly part of Williams syndrome. Linkage to the elastin gene on chromosome 7q11 has recently been reported in two kindreds with SVAS. Previous reports of patients with 7q11 deletions have noted great vessel abnormalities in some. We report on a family in which SVAS is cosegregating with a balanced reciprocal translocation, t(6:7) (p21.1;q11.23), providing further evidence that SVAS is the result of a mutation of elastin at 7q11.23 region. The propositus of the translocation family has some minor anomalies which occur in Williams syndrome, suggesting that elastin abnormalities may cause some of the abnormalities found in Williams syndrome.

Topics: Adult; Aged; Aged, 80 and over; Aortic Valve Stenosis; Child; Child, Preschool; Chromosomes, Human, Pair 6; Chromosomes, Human, Pair 7; Elastin; Female; Humans; Infant, Newborn; Karyotyping; Male; Pedigree; Syndrome; Translocation, Genetic; Ultrasonography

Supravalvular aortic stenosis (SVAS) is a localized or diffuse congenital narrowing of the ascending aorta which may occur sporadically, as a familial defect, or in association with Williams syndrome. Familial cases suggest an autosomal dominant gene defect but the underlying molecular basis of SVAS is unknown. In this study, we sought to localize the genetic defect in familial SVAS by linkage analysis in a large three generation family. A total of 44 polymorphic markers were examined for linkage, including 17 Southern blot-based RFLPs, 2 PCR-based RFLPs, and 25 microsatellites, primarily of the (CA)n repeat type. We report linkage of the disease phenotype to a highly informative (CA)n repeat marker, Mfd 50, at locus D7S440 which has been localized to chromosome arm 7q. Using a 100% penetrance model, which was more conservative than lower values of penetrance, a peak LOD score of 4.66 at a recombination frequency of 0.043 was found. A number of candidate genes have been localized to this region, including collagen 1A2, laminin B1, and elastin. Based on our preliminary linkage data, the abnormal microscopic appearance of aortic elastic fibers in SVAS, and analogous animal and human diseases associated with elastic fiber and vascular abnormalities, there is indirect evidence suggesting elastin as a possible candidate gene for this disorder.

Topics: Aortic Valve Stenosis; Chromosome Mapping; Chromosomes, Human, Pair 7; Elastin; Female; Genes, Dominant; Genetic Linkage; Genetic Markers; Humans; Male; Oligodeoxyribonucleotides; Pedigree; Polymorphism, Genetic; Repetitive Sequences, Nucleic Acid

The pathogenesis of vascular disease is unclear, but genetic factors play an important role. In this study we performed linkage analyses in two families with supravalvular aortic stenosis, an inherited vascular disorder that causes narrowing of major arteries and may lead to cardiac overload and failure. DNA markers on the long arm of chromosome 7 (D7S371, D7S395, D7S448, and ELN) were linked to supravalvular aortic stenosis in both families with a combined logarithm of likelihood for linkage (lod score) of 5.9 at the ELN locus. These findings indicate that a gene for supravalvular aortic stenosis is located in the same chromosomal subunit as elastin, which becomes a candidate for the disease gene.

Topics: Aortic Valve Stenosis; Base Sequence; Chromosome Mapping; Chromosomes, Human, Pair 7; DNA; Elastin; Female; Genetic Linkage; Genetic Markers; Humans; Lod Score; Male; Molecular Sequence Data; Oligodeoxyribonucleotides; Pedigree; Phenotype; Polymerase Chain Reaction; Recombination, Genetic

Histological sections through the walls of abdominal aortic aneurysms showed scarce and disrupted elastic tissue. The elastin content of the aneurysmal aortic media was only 8.1 +/- 3.2% dry defatted weight (n = 11). The elastin content of grossly normal age and anatomically matched aortic media was 35.0 +/- 3.2% dry weight (n = 4) and the elastin content of severely atherosclerotic, stenosed infrarenal aortic media was 22.0 +/- 7.2% dry weight (n = 6). There was an inverse correlation of elastin content with the elastinolytic activity of aortic media homogenates, r = -0.78. Elastase activity, measured by the hydrolysis of [3H]elastin, was highest in aneurysmal aortic homogenates, 92.1 +/- 43.7 U/mg protein (n = 18), falling to 46.9 +/- 13.3 U/mg protein (n = 13) in severely stenosed atherosclerotic aortic homogenates and 35.5 +/- 11.9 U/mg (n = 6) in grossly normal aortic homogenates. The elastinolytic activity of stenotic aorta contained leukocyte elastase as an important component. In aneurysmal homogenates leukocyte elastase was also found but the increased elastase activity resulted from a protease(s) (Mr 95,000) extracted in 2 M urea, having minimal specificity for alanyl bonds and no immunological cross-reactivity with leukocyte elastase.

Topics: Aged; Amino Acids; Aorta, Abdominal; Aortic Aneurysm; Aortic Valve Stenosis; Elastin; Female; Histocytochemistry; Humans; Male; Middle Aged; Molecular Weight; Pancreatic Elastase; Tissue Extracts