transforming-growth-factor-beta and Heart-Valve-Diseases

Dilatation of the aorta is a constantly evolving condition that can lead to the ultimate life-threatening event, acute aortic dissection. Recent research has tried to identify quantifiable biomarkers, with both diagnostic and prognostic roles in different aortopathies. Most studies have focused on the bicuspid aortic valve, the most frequent congenital heart disease (CHD), and majorly evolved around matrix metalloproteinases (MMPs). Other candidate biomarkers, such as asymmetric dimethylarginine, soluble receptor for advanced glycation end-products or transforming growth factor beta have also gained a lot of attention recently. Most of the aortic anomalies and dilatation-related studies have reported expression variation of tissular biomarkers. The ultimate goal remains, though, the identification of biomarkers among the serum plasma, with the upregulation of circulating MMP-1, MMP-2, MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), asymmetric dimethylarginine (ADMA), soluble receptor for advanced glycation end-products (sRAGE) and transforming growth factor beta (TGF-β) being reported in association to several aortopathies and related complications in recent research. These molecules are apparently quantifiable from the early ages and have been linked to several CHDs and hereditary aortopathies. Pediatric data on the matter is still limited, and further studies are warranted to elucidate the role of plasmatic biomarkers in the long term follow-up of potentially evolving congenital aortopathies.

Topics: Aorta; Aortic Valve; Biomarkers; Child; Dilatation, Pathologic; Heart Defects, Congenital; Heart Valve Diseases; Humans; Matrix Metalloproteinases; Receptor for Advanced Glycation End Products; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta

Marfan Syndrome (MFS) and Loeys-Dietz Syndrome (LDS) represent heritable connective tissue disorders that cosegregate with a similar pattern of cardiovascular defects (thoracic aortic aneurysm, mitral valve prolapse/regurgitation, and aortic root dilatation with regurgitation). This pattern of cardiovascular defects appears to be expressed along a spectrum of severity in many heritable connective tissue disorders and raises suspicion of a relationship between the normal development of connective tissues and the cardiovascular system. Given the evidence of increased transforming growth factor-beta (TGF-β) signaling in MFS and LDS, this signaling pathway may represent the common link in this relationship. To further explore this hypothetical link, this chapter will review the TGF-β signaling pathway, heritable connective tissue syndromes related to TGF-β receptor (TGFBR) mutations, and discuss the pathogenic contribution of TGF-β to these syndromes with a primary focus on the cardiovascular system.

Topics: Adrenergic beta-Antagonists; Angiotensin II Type 1 Receptor Blockers; Antibodies, Neutralizing; Aortic Aneurysm, Thoracic; Aortic Valve; Bicuspid Aortic Valve Disease; Gene Expression Regulation; Heart Defects, Congenital; Heart Valve Diseases; Humans; Loeys-Dietz Syndrome; Marfan Syndrome; Mutation; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad Proteins; Transforming Growth Factor beta

Topics: Animals; Aortic Valve; Bicuspid Aortic Valve Disease; Fibrillins; Heart Valve Diseases; Heart Valves; Humans; Matrix Metalloproteinase 2; Microfilament Proteins; Mitral Valve Prolapse; Receptor, Notch1; Signal Transduction; Transforming Growth Factor beta

Recent studies have suggested an important role for periostin and transforming growth factor beta (TGF beta) and bone morphogenetic protein (BMP) ligands in heart valve formation and valvular heart diseases. The function of these molecules in cardiovascular development has previously been individually reviewed, but their association has not been thoroughly examined. Here, we summarize the current understanding of the association between periostin and TGF beta and BMP ligands, and discuss the implications of this association in the context of the role of these molecules in heart valve development and valvular homeostasis. Information about hierarchal connections between periostin and TGF beta and BMP ligands in valvulogenesis will increase our understanding of the pathogenesis, progression, and medical treatment of human valve diseases.

Topics: Bone Morphogenetic Proteins; Cell Adhesion Molecules; Cell Differentiation; Heart Valve Diseases; Heart Valves; Humans; Models, Biological; Organogenesis; Signal Transduction; Transforming Growth Factor beta

Marfan syndrome (MFS) is a systemic disorder of the connective tissue with pleiotropic manifestations due to heterozygous FBN1 mutations and consequent upregulation of TGFβ signaling in affected tissues. Myxomatous thickening and elongation of the mitral valve (MV) leaflets commonly occur in this condition. Investigation of murine models of this disease has led to improved understanding of the mechanisms that underlie many of the phenotypic features of MFS, including MV disease. Loeys-Dietz syndrome (LDS) is a related disorder due to heterozygous mutations in the genes encoding subunits of the TGFβ receptor, and it may also involve the MV leaflets with similar elongation and thickening of the MV leaflets. Although the genetic basis and pathogenesis of nonsyndromic MV prolapse has been elusive to date, insights derived from monogenic disorders like MFS and LDS can be informative with regard to novel gene discovery and investigation into the pathogenesis of MV disease. This manuscript will review the prevalence of MV disease in MFS, its pathogenic basis as determined in mice with Fbn1 mutations, and ongoing studies that seek to better understand MV disease in the context of fibrillin-1 deficiency or excessive TGFβ signaling.

Topics: Animals; Disease Models, Animal; Fibrillin-1; Fibrillins; Genetic Predisposition to Disease; Heart Valve Diseases; Humans; Loeys-Dietz Syndrome; Marfan Syndrome; Mice; Mice, Transgenic; Microfilament Proteins; Mitral Valve; Phenotype; Receptor, Angiotensin, Type 2; Receptors, Transforming Growth Factor beta; Signal Transduction; Transforming Growth Factor beta

The study of the cellular and molecular pathogenesis of heart valve disease is an emerging area of research made possible by the availability of cultures of valve interstitial cells (VICs) and valve endothelial cells (VECs) and by the design and use of in vitro and in vivo experimental systems that model elements of valve biological and pathobiological activity. VICs are the most common cells in the valve and are distinct from other mesenchymal cell types in other organs. We present a conceptual approach to the investigation of VICs by focusing on VIC phenotype-function relationships. Our review suggests that there are five identifiable phenotypes of VICs that define the current understanding of their cellular and molecular functions. These include embryonic progenitor endothelial/mesenchymal cells, quiescent VICs (qVICs), activated VICs (aVICs), progenitor VICs (pVICs), and osteoblastic VICs (obVICs). Although these may exhibit plasticity and may convert from one form to another, compartmentalizing VIC function into distinct phenotypes is useful in bringing clarity to our understanding of VIC pathobiology. We present a conceptual model that is useful in the design and interpretation of studies on the function of an important phenotype in disease, the activated VIC. We hope this review will inspire members of the investigative pathology community to consider valve pathobiology as an exciting new frontier exploring pathogenesis and discovering new therapeutic targets in cardiovascular diseases.

Topics: Animals; Cell Lineage; Embryonic Stem Cells; Endothelial Cells; Heart Valve Diseases; Heart Valves; Humans; Inflammation; Mesenchymal Stem Cells; Neovascularization, Pathologic; Neovascularization, Physiologic; Osteoblasts; Transforming Growth Factor beta

The importance of fibrosis in organ pathology and dysfunction appears to be increasingly relevant to a variety of distinct diseases. In particular, a number of different cardiac pathologies seem to be caused by a common fibrotic process. Within the heart, this fibrosis is thought to be partially mediated by transforming growth factor-beta1 (TGF-beta1), a potent stimulator of collagen-producing cardiac fibroblasts. Previously, TGF-beta1 had been implicated solely as a modulator of the myocardial remodelling seen after infarction. However, recent studies indicate that dilated, ischaemic and hypertrophic cardiomyopathies are all associated with raised levels of TGF-beta1. In fact, the pathogenic effects of TGF-beta1 have now been suggested to play a major role in valvular disease and arrhythmia, particularly atrial fibrillation. Thus far, medical therapy targeting TGF-beta1 has shown promise in a multitude of heart diseases. These therapies provide great hope, not only for treatment of symptoms but also for prevention of cardiac pathology as well. As is stated in the introduction, most reviews have focused on the effects of cytokines in remodelling after myocardial infarction. This article attempts to underline the significance of TGF-beta1 not only in the post-ischaemic setting, but also in dilated and hypertrophic cardiomyopathies, valvular diseases and arrhythmias (focusing on atrial fibrillation). It also aims to show that TGF-beta1 is an appropriate target for therapy in a variety of cardiovascular diseases.

Topics: Arrhythmias, Cardiac; Cardiomyopathies; Fibrosis; Heart Diseases; Heart Valve Diseases; Humans; Matrix Metalloproteinases; Myocardium; Transforming Growth Factor beta; Transforming Growth Factor beta1

We sought to determine whether there are differences in transforming growth factor-beta (TGFß) signaling in aneurysms associated with bicuspid (BAV) and unicuspid (UAV) aortic valves versus normal aortic valves. Ascending aortic aneurysms are frequently associated with BAV and UAV. The mechanisms are not yet clearly defined, but similarities to transforming growth factor-beta TGFß vasculopathies (i.e. Marfan, Loeys-Dietz syndromes) are reported. Non-dilated (ND) and aneurysmal (D) ascending aortic tissue was collected intra-operatively from individuals with a TAV (N = 10ND, 10D), BAV (N = 7ND, 8D) or UAV (N = 7ND, 8D). TGFß signaling and aortic remodeling were assessed through immuno-assays and histological analyses. TGFß1 was increased in BAV/UAV-ND aortas versus TAV (P = 0.02 and 0.04, respectively). Interestingly, TGFß1 increased with dilatation in TAV (P = 0.03) and decreased in BAV/UAV (P = 0.001). In TAV, SMAD2 and SMAD3 phosphorylation (pSMAD2, pSMAD3) increased with dilatation (all P = 0.04) and with TGFß1 concentration (P = 0.04 and 0.03). No relationship between TGFß1 and pSMAD2 or pSMAD3 was observed for BAV/UAV (all P > 0.05). pSMAD3 increased with dilatation in BAV/UAV aortas (P = 0.01), whereas no relationship with pSMAD2 was observed (P = 0.56). Elastin breaks increased with dilatation in all groups (all P < 0.05). In TAV, elastin degradation correlated with TGFß1, pSMAD2 and pSMAD3 (all P < 0.05), whereas in BAV and UAV aortas, elastin degradation correlated only with pSMAD3 (P = 0.0007). TGFß signaling through SMAD2/SMAD3 contributes to aortic remodeling in TAV, whereas TGFß-independent activation of SMAD3 may underlie aneurysm formation in BAV/UAV aortas. Therefore, SMAD3 should be further investigated as a therapeutic target against ascending aortic dilatation in general, and particularly in BAV/UAV patients.

Topics: Aortic Diseases; Dilatation; Dilatation, Pathologic; Elastin; Heart Valve Diseases; Humans; Smad3 Protein; Transforming Growth Factor beta; Transforming Growth Factors

The pathobiology of ascending aorta aneurysms (AAA) onset and progression is not well understood and only partially characterized. AAA are also complicated in case of bicuspid aorta valve (BAV) anatomy. There is emerging evidence about the crucial role of endothelium-related pathways, which show in AAA an altered expression and function. Here, we examined the involvement of ERG-related pathways in the differential progression of disease in aortic tissues from patients having a BAV or tricuspid aorta valve (TAV) with or without AAA. Our findings identified ERG as a novel endothelial-specific regulator of TGF-β-SMAD, Notch, and NO pathways, by modulating a differential fibrotic or calcified AAA progression in BAV and TAV aortas. We provided evidence that calcification is correlated to different ERG expression (as gene and protein), which appears to be under control of Notch signaling. The latter, when increased, associated with an early calcification in aortas with BAV valve and aneurysmatic, was demonstrated to favor the progression versus severe complications, i.e., dissection or rupture. In TAV aneurysmatic aortas, ERG appeared to modulate fibrosis. Therefore, we proposed that ERG may represent a sensitive tissue biomarker to monitor AAA progression and a target to develop therapeutic strategies and influence surgical procedures.

Topics: Aorta; Aortic Valve; Bicuspid Aortic Valve Disease; Biomarkers; Endothelium; Heart Valve Diseases; Humans; Transcription Factors; Transcriptional Regulator ERG; Transforming Growth Factor beta

Several clinical reports indicate that the use of amphetaminic anorectic drugs or ergot derivatives could cause valvular heart disease (VHD). We sought to investigate whether valvular lesions develop in response to long-term oral administration of these drugs and to identify drug-targeted biological processes that may lead to VHD. Treatment of New Zealand White rabbits with pergolide, dexfenfluramine, or high-dose serotonin for 16 weeks induced valvular alterations characterized by extracellular matrix remodeling. Transcriptome profiling of tricuspid valves using RNA sequencing revealed distinct patterns of differentially expressed genes (DEGs) that clustered according to the different treatments. Genes that were affected by the three treatments were functionally enriched for reduced cell metabolism processes. The two drugs yielded more changes in gene expression than serotonin and shared most of the DEGs. These DEGs were mostly enriched for decreased biosynthetic processes, increased cell-matrix interaction, and cell response to growth factors, including TGF-β, which was associated with p38 MAPK activation. Treatment with pergolide specifically affected genes involved in homeostasis, which was corroborated by the activation of the master regulator of cell energy homeostasis, AMPK-α, as well as decreased levels of metabolism-related miR-107. Thus, both pergolide and dexfenfluramine may cause VHD through valve metabolic reprogramming and matrix remodeling.

Topics: Administration, Oral; AMP-Activated Protein Kinases; Animals; Cell Proliferation; Cluster Analysis; Dexfenfluramine; Enzyme Activation; Extracellular Matrix; Female; Gene Expression Regulation; Heart Valve Diseases; Homeostasis; MicroRNAs; p38 Mitogen-Activated Protein Kinases; Pergolide; Rabbits; Sequence Analysis, RNA; Serotonin; Transcriptome; Transforming Growth Factor beta; Tricuspid Valve

Atrial fibrillation (AF) is the most frequently diagnosed cardiac arrhythmia worldwide. Patients with permanent atrial fibrillation are at an increased risk of developing valvular heart disease. Atrial fibrosis occurs in this pathophysiological setting. LIM kinase 1 (LIMK1) is a serine/threonine kinase that regulates microtubule stability and actin polymerization in fibroblasts. LIMK1 has been implicated in the pathogenesis of atrial fibrillation. Clinical data and biopsies of the right atrial appendage were collected from 50 patients with valvular heart disease who underwent heart valve replacement surgery. Data from patients with permanent atrial fibrillation (AF) and patients with sinus rhythm (SR) were compared. We found that AF patients had upregulated expression of LIMK1 as well as higher fibrosis. Transforming growth factor-β (TGF-β) stimulation induced the differentiation of cardiac fibroblasts into myofibroblasts as well as upregulated expression of LIMK1. Downregulation of LIMK1 by siRNA inhibited TGF-β induced fibroblast-myofibroblast transition, as evidenced by the downregulation of the expression of several differentiation markers, namely alpha-smooth muscle actin and type I and III collagen. Our findings revealed that increased LIMK1 protein levels may contribute to atrial fibrosis, and suggested that LIMK1 might be involved in AF development by promoting fibrogenesis associated with TGF-β.

Topics: Actins; Animals; Atrial Fibrillation; Biopsy; Collagen Type I; Female; Fibroblasts; Fibrosis; Gene Expression Regulation; Heart Atria; Heart Valve Diseases; Humans; Lim Kinases; Male; Mice; Middle Aged; Myofibroblasts; RNA, Small Interfering; Transforming Growth Factor beta

Ascending thoracic aortic aneurysm (ATAA) is a major cause of morbidity and mortality worldwide. The pathogenesis of medial degeneration of the aorta remains undefined. High-throughput secretome analysis by mass spectrometry may be useful to elucidate the molecular mechanisms involved in aneurysm formation as well as to identify biomarkers for early diagnosis or targets of therapy. The purpose of the present study was to analyze the secreted/released proteins from ATAA specimens of both tricuspid aortic valve (TAV) and bicuspid aortic valve (BAV) patients.. Aortic specimens were collected from patients undergoing elective surgery and requiring graft replacement of the ascending aorta. Each sample of the ascending aortic aneurysm, 4 BAV (3 males; aged 53.5±11.4 years) and 4 TAV (1 male; 78±7.5 years), was incubated for 24h in serum-free medium. Released proteins were digested with trypsin. Peptide mixtures were fractioned by nano-high performance liquid chromatography and analyzed by mass spectrometry. Following identification of differentially expressed proteins, quantitative real time polymerase chain reaction (qRT-PCR) analysis was performed.. The comparison between the proteins released from BAV and TAV aneurysmatic tissues showed significantly diverging expression fingerprints in the two groups of patients. Bioinformatics analysis revealed 38 differentially released proteins; in particular 7 proteins were down-regulated while 31 were up-regulated in BAV with respect to TAV. Most of the proteins that were up-released in BAV were related to the activation of transforming growth factor (TGF)-β signaling. Latent TGF-β binding protein 4 (LTBP4) exhibited one of the highest significant under-expressions (10-fold change) in BAV secretomes with respect to TAV. qRT-PCR analysis validated this significant difference at LTBP4 gene level (BAV: 1.03±0.9 vs TAV: 3.6±3.2; p<0.05).. Hypothesis-free secretome profiling clearly showed diverging expression fingerprints in the ATAA of TAV and BAV patients, confirming the crucial role of TGF-β signaling in modulating ATAA development in bicuspid patients.

Topics: Aged; Aorta; Aortic Aneurysm; Aortic Aneurysm, Thoracic; Aortic Valve; Bicuspid Aortic Valve Disease; Female; Heart Valve Diseases; Humans; Latent TGF-beta Binding Proteins; Male; Middle Aged; Signal Transduction; Transforming Growth Factor beta; Tricuspid Valve

Individuals with bicuspid aortic valves (BAV) are at a higher risk of developing thoracic aortic aneurysms (TAA) than patients with trileaflet aortic valves (TAV). The aneurysms associated with BAV most commonly involve the ascending aorta and spare the descending aorta. Smooth muscle cells (SMCs) in the ascending and descending aorta arise from neural crest (NC) and paraxial mesoderm (PM), respectively. We hypothesized defective differentiation of the neural crest stem cells (NCSCs)-derived SMCs but not paraxial mesoderm cells (PMCs)-derived SMCs contributes to the aortopathy associated with BAV. When induced pluripotent stem cells (iPSCs) from BAV/TAA patients were differentiated into NCSC-derived SMCs, these cells demonstrated significantly decreased expression of marker of SMC differentiation (MYH11) and impaired contraction compared to normal control. In contrast, the PMC-derived SMCs were similar to control cells in these aspects. The NCSC-SMCs from the BAV/TAA also showed decreased TGF-β signaling based on phosphorylation of SMAD2, and increased mTOR signaling. Inhibition of mTOR pathway using rapamycin rescued the aberrant differentiation. Our data demonstrates that decreased differentiation and contraction of patient's NCSC-derived SMCs may contribute to that aortopathy associated with BAV.

Topics: Aortic Diseases; Aortic Valve; Bicuspid Aortic Valve Disease; Biomarkers; Cell Culture Techniques; Cell Differentiation; Heart Valve Diseases; Humans; Immunophenotyping; Induced Pluripotent Stem Cells; Muscle Contraction; Myocytes, Smooth Muscle; Myosin Heavy Chains; Neural Crest; Phenotype; Signal Transduction; TOR Serine-Threonine Kinases; Transforming Growth Factor beta

Patients with bicuspid aortic valve (BAV) have an increased risk of developing ascending aortic aneurysms. Transforming growth factor-β (TGFβ) is a crucial factor of vascular remodeling, the impaired signaling of which can alter the structure and composition of the extracellular matrix. In this study, we analyzed the activity of TGFβ in aneurysmal and nonaneurysmal ascending aorta from BAV patients, using tricuspid aortic valve (TAV) patients as a reference group.. The response to exogenous TGFβ was analyzed with regard to gene expression in primary aortic smooth muscle cells that were isolated from 7 BAV and 5 TAV patients and in valve fibroblasts from 7 BAV and 8 TAV patients. The set of genes that were significantly changed by TGFβ (217 genes) was compared with gene expression profiles of the ascending aorta from BAV and TAV patients (139 arrays). By principle component analysis, based on the 217 genes, gene expression differed significantly in the intima/media region between aneurysmal BAV and TAV aortas, driven by the response in TAV patients. During aneurysm development the levels of phosphorylated SMADs and the availability of free TGFβ were lower in BAV patients compared with TAV. Confocal microscopy analysis showed a higher colocalization of latency associated peptide and latent TGFβ binding protein 3 in BAV aortas.. Our findings suggest that TGFβ activation during aneurysm formation is muted in patients with BAV, possibly as a result of an increased TGFβ sequestration in the extracellular space.

Topics: Adult; Aged; Aged, 80 and over; Aortic Aneurysm; Aortic Valve; Bicuspid Aortic Valve Disease; Cells, Cultured; Extracellular Matrix; Female; Gene Expression Profiling; Gene Expression Regulation; Heart Valve Diseases; Humans; Latent TGF-beta Binding Proteins; Male; Middle Aged; Phosphorylation; Principal Component Analysis; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad Proteins; Transforming Growth Factor beta

We studied the mechanistic links between fibrocalcific changes in the aortic valve and aortic valve function in mice homozygous for a hypomorphic epidermal growth factor receptor mutation (Wave mice). We also studied myocardial responses to aortic valve dysfunction in Wave mice.. At 1.5 months of age, before development of valve fibrosis and calcification, aortic regurgitation, but not aortic stenosis, was common in Wave mice. Aortic valve fibrosis, profibrotic signaling, calcification, osteogenic markers, lipid deposition, and apoptosis increased dramatically by 6 and 12 months of age in Wave mice. Aortic regurgitation remained prevalent, however, and aortic stenosis was rare, at all ages. Proteoglycan content was abnormally increased in aortic valves of Wave mice at all ages. Treatment with pioglitazone prevented abnormal valve calcification, but did not protect valve function. There was significant left ventricular volume overload, hypertrophy, and fetal gene expression, at all ages in Wave mice with aortic regurgitation. Left ventricular systolic function was normal until 6 months of age in Wave mice, but became impaired by 12 months of age. Myocardial transverse tubules were normal in the presence of left ventricular hypertrophy at 1.5 and 3 months of age, but became disrupted by 12 months of age.. We present the first comprehensive phenotypic and molecular characterization of spontaneous aortic regurgitation and volume-overload cardiomyopathy in an experimental model. In Wave mice, fibrocalcific changes are not linked to valve dysfunction and are epiphenomena arising from structurally incompetent myxomatous valves.

Topics: Actins; Animals; Aortic Valve; Aortic Valve Insufficiency; Calcinosis; Cell Death; Disease Progression; Fibrosis; Gene Expression; Heart Valve Diseases; Lipid Metabolism; Mice; Mice, Mutant Strains; Osteocalcin; Pioglitazone; Proteoglycans; Sp7 Transcription Factor; Systole; Thiazolidinediones; Transcription Factors; Transforming Growth Factor beta

To establish human heart valve interstitial cells calcification culture model in vitro, and observe the effect of bone morphogenetic protein-2 (BMP-2) on calcification of human heart valve interstitial cells.. Human heart valve interstitial cells were cultured in vitro, and divided into control group: cells were cultured in conventional media plus recombinant human BMP-2 treatment and experimental group: besides above treaments, calcification inducers ( recombinant human BMP-2, β-glycerophosphate, L-ascorbic acid, dexamethasone) were added to the culture media. The two group of cells were cultured for 14 days and were stained by Von Kossa, then the cell calcification was observed in this valvular interstitial cells calcification culture model in vitro. Protein expression of intercellular adhesion molecule 1 (ICAM-1), interleukin 8, BMP-2 and BMP-4 was determined by Western blot and BMP-2 secretion was measured by ELISA.. In the control group, the structure of human heart valve interstitial cells was clear, and the spindle and radial growth shaped cellular morphology was visible, and Von Kossa staining was negative. In the experimental group, the nuclei become darker in color, and granular sediment distribution was seen surrounding cells, and Von Kossa staining was positive, the cells were forming nodules of calcification. The protein expression of ICAM-1, interleukin 8, BMP-2 and BMP-4 in the experimental was significantly higher than that of the control group (all P < 0.05). The expression of BMP-2 in the experimental group was also significantly higher than that in control group ((92.5 ± 4.9) pg/ml vs. (22.2 ± 1.9) pg/ml, P < 0.05).. Human BMP-2, β-glycerophosphate, L-ascorbic acid, and dexamethasone can induce human heart valve interstitial cells calcification and enhance inflammation in vitro by stimulating the secretion of BMP-2.

Topics: Ascorbic Acid; Bone Morphogenetic Protein 2; Calcinosis; Cells, Cultured; Glycerophosphates; Heart Valve Diseases; Humans; Recombinant Proteins; Transforming Growth Factor beta

Aortic valve disease (AVD) is characterized by elastic fiber fragmentation (EFF), fibrosis and aberrant angiogenesis. Emilin1 is an elastin-binding glycoprotein that regulates elastogenesis and inhibits TGF-β signaling, but the role of Emilin1 in valve tissue is unknown. We tested the hypothesis that Emilin1 deficiency results in AVD, mediated by non-canonical (MAPK/phosphorylated Erk1 and Erk2) TGF-β dysregulation. Using histology, immunohistochemistry, electron microscopy, quantitative gene expression analysis, immunoblotting and echocardiography, we examined the effects of Emilin1 deficiency (Emilin1-/-) in mouse aortic valve tissue. Emilin1 deficiency results in early postnatal cell-matrix defects in aortic valve tissue, including EFF, that progress to latent AVD and premature death. The Emilin1-/- aortic valve displays early aberrant provisional angiogenesis and late neovascularization. In addition, Emilin1-/- aortic valves are characterized by early valve interstitial cell activation and proliferation and late myofibroblast-like cell activation and fibrosis. Interestingly, canonical TGF-β signaling (phosphorylated Smad2 and Smad3) is upregulated constitutively from birth to senescence, whereas non-canonical TGF-β signaling (phosphorylated Erk1 and Erk2) progressively increases over time. Emilin1 deficiency recapitulates human fibrotic AVD, and advanced disease is mediated by non-canonical (MAPK/phosphorylated Erk1 and Erk2) TGF-β activation. The early manifestation of EFF and aberrant angiogenesis suggests that these processes are crucial intermediate factors involved in disease progression and therefore might provide new therapeutic targets for human AVD.

Topics: Animals; Aortic Valve; Bicuspid Aortic Valve Disease; Calcinosis; Cell Proliferation; Cutis Laxa; Disease Models, Animal; Disease Progression; Elastic Tissue; Fibrosis; Heart Defects, Congenital; Heart Valve Diseases; Inflammation; Membrane Glycoproteins; Mice; Models, Biological; Myofibroblasts; Neovascularization, Pathologic; Signal Transduction; Transforming Growth Factor beta; Ultrasonography

The clinical course of many patients with a bicuspid aortic valve (BAV) is complicated by ascending aortic dilatation. Currently, the indication for aortic surgery is solely based on the aortic diameter and subsequently only a small proportion of BAV patients undergoing valve surgery require concomitant ascending aortic replacement based on these recommendations. Unfortunately, a substantial number of BAV patients still develop aortic dilatation in the future and would potentially benefit from a more aggressive approach towards ascending aortic replacement. We, therefore, designed this study to identify molecular biological markers in the aortic wall predictive of aortopathy in BAV.. Ascending aortic wall specimen of BAV (n = 36) and tricuspid aortic valve (TAV) (n = 23), both without and with (>44 mm) dilatation were investigated histologically and immunohistochemically for the expression of markers for vascular remodelling [transforming growth factor (TGF)-β, phosphorylated Smad2, matrix metalloproteinase 9 (MMP9)], cellular differentiation [c-Kit, phosphorylated-c-Kit, hypoxia-inducable factor-1 alpha (HIF1α)] and haemodynamic influences on the aortic wall [endothelial nitric oxide (eNOS)].. All BAV patients showed significantly less inflammation (P < 0.001) and an altered intima/media ratio when compared with TAV patients. The expression of markers of a signalling pathway characteristic for cellular dedifferentiation, as exemplified by the marked expression of c-Kit, phosphorylated c-Kit and HIF1α; in the dilated BAV group was however completely comparable with only a subgroup of the non-dilated BAV (BAb), whereas the remainder of the non-dilated BAV group (BAa) was significantly distinct. This difference between the dilated BAV and BAa was further confirmed in the expression of TGF-β, phosphorylated Smad2, MMP9 and eNOS. Besides the expression pattern, similarity in the dilated BAV and BAb was also noted clinically in the most common variant of commissure position and conjoined raphe of the BAV. Based on these observations, we consider the BAb group a likely candidate for future dilatation as opposed to the BAa group.. Using a panel of molecular tissue markers, the non-dilated BAV patients can be divided into groups susceptible and non-susceptible to aortopathy.

Topics: Adult; Aged; Aorta; Aortic Diseases; Aortic Valve; Bicuspid Aortic Valve Disease; Biomarkers; Cohort Studies; Female; Heart Valve Diseases; Humans; Immunohistochemistry; Male; Middle Aged; Muscle, Smooth, Vascular; Nitric Oxide; Phosphorylation; Prognosis; Proto-Oncogene Proteins c-kit; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta

Previous studies on BAV (bicuspid aortic valve)-related aortopathy, whose aetiology is still debated, have focused mainly on severe dilatations. In the present study, we aimed to detect earlier signs of aortopathy. Specimens were collected from the 'concavity' (lesser curvature) and the 'convexity' (greater curvature) of mildly dilated AAs (ascending aortas; diameter ≤4 cm) with stenotic TAV (tricuspid aortic valve) or BAV and from donor normal aortas. Specimens were submitted to morphometry, immunohistochemistry and differential gene-expression analysis, focusing on SMC (smooth muscle cell) phenotype, remodelling, MF (myofibroblast) differentiation and TGFβ (transforming growth factor β) pathway. Smoothelin and myocardin mRNAs decreased in all the samples from patients, with the exception of those from BAV convexity, where a change in orientation of smoothelin-positive SMCs and an increase of α-SMA (α-smooth muscle actin) mRNA occurred. Dilated aortas from BAV and TAV patients showed both shared and distinct alterations concerning the TGFβ pathway, including an increased TGFβ and TGFβR2 (TGFβ receptor 2) expression in both groups and a decreased TGFβR1 expression in BAV samples only. Despite a decrease of the mRNA coding for the ED-A (extra domain-A) isoform of FN (fibronectin) in the BAV convexity, the onset of the expression of the corresponding protein in the media was observed in dilated aortas, whereas the normal media from donors was negative for this isoform. This discrepancy could be related to modifications in the intima, normally expressing ED-A FN and showing an altered structure in mild aortic dilatations in comparison with donor aorta. Our results suggest that changes in SMC phenotype and, likely, MF differentiation, occur early in the aortopathy associated with valve stenosis. The defective expression of TGFβR1 in BAV might be a constitutive feature, while other changes we reported could be influenced by haemodynamics.

Topics: Adult; Aged; Aged, 80 and over; Analysis of Variance; Antibodies, Monoclonal; Aortic Valve; Aortic Valve Stenosis; Bicuspid Aortic Valve Disease; Body Weights and Measures; Cell Differentiation; Cytoskeletal Proteins; DNA Primers; Female; Fibronectins; Heart Valve Diseases; Humans; Immunohistochemistry; Male; Middle Aged; Muscle Proteins; Myocytes, Smooth Muscle; Myofibroblasts; Nuclear Proteins; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Statistics, Nonparametric; Trans-Activators; Transforming Growth Factor beta

A number of drugs and drug candidates, including fenfluramine and ergot derivatives, are associated with valvulopathy in humans; however, these responses are poorly predicted from animal studies. In vitro and in vivo evidence suggests that these compounds exert their pathological effect through activation of serotonin 2B receptor (5HT2BR) signaling. However, the variable effect of fenfluramine and other 5HT2BR agonists in rodents has cast doubt on the relevance of animal findings to predicting human risk. Herein, a candidate compound, RO3013, induced subendocardial cell proliferation in the mitral and tricuspid valves in rats after only 3 days of daily dosing. Additionally, there was a treatment-related increase in immunostaining of the proliferation marker Ki67, and phosphorylated Smad3 in the heart indicative of TGFβ signaling co-localized with 5HT2BR expression. To substantiate the hypothesis that RO3013-induced valvular proliferation is secondary to 5HT2BR activation, the compound was evaluated in vitro and found to bind to the human 5HT2BR with a K(i) of 3.8μM; however, it was virtually devoid of agonist activity in a functional assay in human cells. By contrast, RO3013 bound to the rat 5HT2BR with a K(i) of 1.2μM and activated the receptor with an EC50 of 0.5μM. This agonist potency estimate is in good agreement with the free plasma concentrations of RO3013 at which valvular proliferation was observed. These results suggest that the rat may be susceptible to 5HT2BR-mediated valvular proliferation similar to humans; yet, the significant differences between binding and functional activities can be a possible explanation for the observed species-selective receptor responses.

Topics: Animals; Cell Proliferation; Heart Valve Diseases; Humans; Ki-67 Antigen; Male; Myocardium; Rats; Rats, Wistar; Receptor, Serotonin, 5-HT2B; Serotonin 5-HT2 Receptor Agonists; Smad3 Protein; Transforming Growth Factor beta

The secreted periostin protein, which marks mesenchymal cells in endocardial cushions following epithelial-mesenchymal transformation and in mature valves following remodeling, is a putative valvulogenesis target molecule. Indeed, periostin is expressed throughout cardiovascular morphogenesis and in all 4 adult mice valves (annulus and leaflets). Additionally, periostin is expressed throughout the fibrous cardiac skeleton and endocardial cushions in the developing heart but is absent from both normal and/or pathological mouse cardiomyocytes. Periostin (peri(lacZ)) knockout mice exhibit viable valve disease, with neonatal lethality in a minority and latent disease with leaflet abnormalities in the viable majority. Surviving peri(lacZ)-null leaflets are truncated, contain ectopic cardiomyocytes and smooth muscle, misexpress the cartilage proteoglycan aggrecan, demonstrate disorganized matrix stratification, and exhibit reduced transforming growth factor-beta signaling. Neonatal peri(lacZ) nulls that die (14%) display additional defects, including leaflet discontinuities, delamination defects, and deposition of acellular extracellular matrix. Assessment of collagen production, 3D lattice formation ability, and transforming growth factor-beta responsiveness indicate periostin-deficient fibroblasts are unable to support normal valvular remodeling and establishment of a mature cardiac skeleton. Furthermore, pediatric stenotic bicuspid aortic valves that have lost normal extracellular matrix trilaminar stratification have greatly reduced periostin. This suggests that loss of periostin results in inappropriate differentiation of mesenchymal cushion cells and valvular abnormalities via a transforming growth factor-beta-dependent pathway during establishment of the mature heart. Thus, peri(lacZ) knockouts provide a new model of viable latent valve disease.

Topics: Animals; Cell Adhesion Molecules; Cell Differentiation; Cell Proliferation; Cytoskeleton; Disease Models, Animal; Endocardium; Extracellular Matrix; Fibroblasts; Heart Valve Diseases; Mesoderm; Mice; Mice, Knockout; Myocardium; Myocytes, Cardiac; Transforming Growth Factor beta

The Fibulins are a 6-member protein family hypothesized to function as intermolecular bridges that stabilize the organization of extracellular matrix structures. Here, we show that reduced expression of Fibulin-4 leads to aneurysm formation, dissection of the aortic wall and cardiac abnormalities. Fibulin-4 knockdown mice with a hypomorphic expression allele arose from targeted disruption of the adjacent Mus81 endonuclease gene. Mice homozygous for the Fibulin-4 reduced expression allele (Fibulin-4(R/R)) show dilatation of the ascending aorta and a tortuous and stiffened aorta, resulting from disorganized elastic fiber networks. They display thickened aortic valvular leaflets that are associated with aortic valve stenosis and insufficiency. Strikingly, already a modest reduction in expression of Fibulin-4 in the heterozygous Fibulin-4(+/R) mice occasionally resulted in small aneurysm formation. To get insight into the underlying molecular pathways involved in aneurysm formation and response to aortic failure, we determined the aorta transcriptome of Fibulin-4(+/R) and Fibulin-4(R/R) animals and identified distinct and overlapping biological processes that were significantly overrepresented including cytoskeleton organization, cell adhesion, apoptosis and several novel gene targets. Transcriptome and protein expression analysis implicated perturbation of TGF-beta signaling in the pathogenesis of aneurysm in fibulin-4 deficient mice. Our results show that the dosage of a single gene can determine the severity of aneurysm formation and imply that disturbed TGF-beta signaling underlies multiple aneurysm phenotypes.

Topics: Animals; Aorta, Thoracic; Aortic Aneurysm; Aortic Valve; Extracellular Matrix Proteins; Heart Valve Diseases; Homeostasis; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Mice, Transgenic; Signal Transduction; Transcription, Genetic; Transforming Growth Factor beta

Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia in humans. The pathophysiology of AF involves electrical, structural and contractile remodeling, which is associated with changes in cardiac gene expression. Previous studies of gene-expression changes in clinical AF have mostly been limited to a small number of candidate genes and have not all been well controlled for underlying heart disease. The present study assessed AF-related gene-expression changes in valve-disease patients with microarrays representing the cardiac transcriptome. Right atrial appendages from 11 patients with chronic AF and underlying valvular heart disease (AF-VHD) and seven patients in sinus rhythm with VHD (SR-VHD) were individually compared to an age-matched sinus-rhythm control group (SR-CTRL, 11 patients) using cardiac-specific microarray analysis. One-class statistical analysis was used to identify genes differentially expressed between SR-VHD and SR-CTRL patients. Two-class statistical analysis was used to identify genes differentially expressed between AF-VHD and SR-VHD patients. Out of 3863 analyzed genes, 832 genes were differentially expressed between SR-VHD and SR-CTRL patients, and 169 genes were differentially expressed between AF-VHD and SR-VHD patients. Striking AF-related changes included altered expression of nine genes pointing towards the development of fibrosis (e.g. upregulation of transforming growth factor beta1), and changes in eight genes potentially related to an increased risk of thromboembolic events (e.g. upregulation of alpha2 macroglobulin). Microarray results were confirmed by quantitative PCR. Our results suggest that AF produces a characteristic profile of gene-expression changes that may be related to the pathophysiology of the arrhythmia.

Topics: Aged; alpha-Macroglobulins; Atrial Fibrillation; Female; Gene Expression Profiling; Heart Valve Diseases; Humans; Male; Oligonucleotide Array Sequence Analysis; Polymerase Chain Reaction; Transforming Growth Factor beta; Transforming Growth Factor beta1

Scarring and collagen deposition in the valves and destruction of myocytes may result from the combined effects of a smoldering rheumatic process and a constant trauma to the mitral valve or aortic valve by the turbulent flow in rheumatic heart disease (RHD). Transforming growth factor-beta1 (TGF-beta1) may be responsible for the increased valvular fibrosis and calcification in the pathogenesis of RHD. However, the role of TGF-beta1 genetic variant in RHD has not been studied. This case-controlled study was carried out to investigate the possible relationship between the TGF-beta1 gene C-509T and T869C polymorphisms and RHD among the Chinese population in Taiwan.. A group of 115 patients with RHD documented by using echocardiography and 100 age- and sex-matched healthy control patients were studied. TGF-beta1 gene C-509T and T869C polymorphisms were identified with polymerase chain reaction-based restriction analysis.. A significant difference was seen in the distribution of genotypes between patients with RHD and control patients for either TGF-beta1 C-509T polymorphism (P <.0001) or T869C polymorphism (P <.0001). The frequency of TGF-beta1 C-509T CC genotype was lower in the RHD group than in the control group (chi2 = 19.05, P <.0001), which suggests that this genotype may confer protective effects against RHD. A significant difference was seen in the distribution of allelic frequency between patients with RHD and control patients for TGF-beta1 T869C polymorphism (P =.04). The odds ratio (OR) for risk of RHD associated with TGF-beta1 T869C T allele was 1.49 (95% CI, 1.02-2.19). Further categorization of patients with RHD into mitral valve disease and combined valve disease subgroups revealed no statistical difference in these gene polymorphisms when compared with the 2 subgroups.. Patients with RHD have a lower frequency of TGF-beta1 C-509T CC genotype and a higher frequency of T869C T allele, which supports a role for the TGF-beta1 gene C-509T and T869C polymorphisms in determining the risk/protection of RHD in Taiwan Chinese patients.

Topics: Adult; Aged; Case-Control Studies; China; Female; Genetics, Population; Genotype; Heart Valve Diseases; Humans; Male; Middle Aged; Mitral Valve Insufficiency; Mitral Valve Stenosis; Polymorphism, Genetic; Rheumatic Heart Disease; Severity of Illness Index; Taiwan; Transforming Growth Factor beta; Transforming Growth Factor beta1; Ultrasonography

Clinical disorders associated with increased serotonin [5-hydroxytryptamine (5-HT)] levels, such as carcinoid syndrome, and the use of serotonin agonists, such as fenfluoramine have been associated with a valvulopathy characterized by hyperplastic valvular and endocardial lesions with increased extracellular matrix. Furthermore, 5-HT has been demonstrated to up-regulate transforming growth factor (TGF)-beta in mesangial cells via G-protein signal transduction. We investigated the hypothesis that increased exposure of heart valve interstitial cells to 5-HT may result in increased TGF-beta1 expression and activity because of serotonin receptor-mediated signal transduction with activation of Galphaq, and subsequently up-regulation of phospholipase C. Thus, in the present study we performed a clinical-pathological investigation of retrieved carcinoid and normal valve cusps using immunohistochemical techniques to detect the presence of TGF-beta1 and other proteins associated with TGF-beta expression, including TGF-beta receptors I and II, latent TGF-beta-associated peptide (LAP), and alpha-smooth muscle actin. Carcinoid valve cusps demonstrated the unusual finding of widespread smooth muscle actin involving the interstitial cells in the periphery of carcinoid nodules; these same cells were also positive for LAP. Normal valve cusps were only focally positive for smooth muscle actin and LAP. In sheep aortic valve interstitial cell cultures 5-HT induced TGF-beta1 mRNA production and increased TGF-beta1 activity. 5-HT also increased collagen biosynthesis at the dosages studied. Furthermore, TGF-beta1 added to SAVIC cultures increased the production of sulfated glycan and hyaluronic acid. In addition, overexpression of Galphaq using an adenoviral expression vector for a constitutively active Galphaq mutant (Q209L-Galphaq) resulted in increased phospholipase C activity as well as up-regulation of TGF-beta expression and activity. These results strongly support the view that G-protein-related signal transduction is involved in 5-HT up-regulation of TGF-beta1. In conclusion, 5-HT-associated valve disease may be, in part, because of TGF-beta1 mechanisms.

Topics: Actins; Adenoviridae; Aged; Aged, 80 and over; Animals; Aortic Valve; Cells, Cultured; Female; GTP-Binding Protein alpha Subunits, Gq-G11; Heart Valve Diseases; Heterotrimeric GTP-Binding Proteins; Humans; Hyaluronic Acid; Male; Second Messenger Systems; Serotonin; Sheep; Transforming Growth Factor beta; Transforming Growth Factor beta1; Type C Phospholipases; Up-Regulation