RESUMEN
The tricuspid valve ensures unidirectional blood flow from the right atrium to the right ventricle. The three tricuspid leaflets operate within a dynamic stress environment of shear, bending, tensile, and compressive forces, which is cyclically repeated nearly three billion times in a lifetime. Ostensibly, the microstructural and mechanical properties of the tricuspid leaflets have mechanobiologically evolved to optimally support their function under those forces. Yet, how the tricuspid leaflet microstructure determines its mechanical properties and whether this relationship differs between the three leaflets is unknown. Here we perform a microstructural and mechanical analysis in matched ovine tricuspid leaflet samples. We found that the microstructure and mechanical properties vary among the three tricuspid leaflets in sheep. Specifically, we found that tricuspid leaflet composition, collagen orientation, and valve cell nuclear morphology are spatially heterogeneous and vary across leaflet type. Furthermore, under biaxial tension, the leaflets' mechanical behaviors exhibited unequal degrees of mechanical anisotropy. Most importantly, we found that the septal leaflet was stiffer in the radial direction and not the circumferential direction as with the other two leaflets. The differences we observed in leaflet microstructure coincide with the varying biaxial mechanics among leaflets. Our results demonstrate the structure-function relationship for each leaflet in the tricuspid valve. We anticipate our results to be vital toward developing more accurate, leaflet-specific tricuspid valve computational models. Furthermore, our results may be clinically important, informing differential surgical treatments of the tricuspid valve leaflets. Finally, the identified structure-function relationships may provide insight into the homeostatic and remodeling potential of valvular cells in altered mechanical environments, such as in diseased or repaired tricuspid valves. STATEMENT OF SIGNIFICANCE: Our work is significant as we investigated the structure-function relationship of ovine tricuspid valve leaflets. This is important as tricuspid valves fail frequently and our current approach to repairing them is suboptimal. Specifically, we related the distribution of structural and cellular elements, such as collagen, glycosaminoglycans, and cell nuclei, to each leaflet's mechanical properties. We found that leaflets have different structures and that their mechanics differ. This may, in the future, inform leaflet-specific treatment strategies and help optimize surgical outcomes.
Asunto(s)
Válvula Tricúspide/fisiología , Animales , Anisotropía , Fenómenos Biomecánicos , Núcleo Celular/fisiología , Colágeno/fisiología , Colágeno/ultraestructura , Pruebas Mecánicas , Ovinos , Válvula Tricúspide/citología , Válvula Tricúspide/ultraestructuraRESUMEN
The atrioventricular heart valve (AHV) leaflets are critical to the facilitation of proper unidirectional blood flow through the heart. Previously, studies have been conducted to understand the tissue mechanics of healthy AHV leaflets to inform the development of valve-specific computational models and replacement materials for use in diagnosing and treating valvular heart disease. Generally, these studies involved biaxial mechanical testing of the AHV leaflet tissue specimens to extract relevant mechanical properties. Most of those studies considered freezing-based storage systems based on previous findings for other connective tissues such as aortic tissue or skin. However, there remains no study that specifically examines the effects of freezing storage on the characterized mechanical properties of the AHV leaflets. In this study, we aimed to address this gap in knowledge by performing biaxial mechanical characterizations of the tricuspid valve anterior leaflet (TVAL) tissue both before and after a 48-h freezing period. Primary findings of this study include: (i) a statistically insignificant change in the tissue extensibilities, with the frozen tissues being slightly stiffer and more anisotropic than the fresh tissues; and (ii) minimal variations in the stress relaxation behaviors between the fresh and frozen tissues, with the frozen tissues demonstrating slightly lessened relaxation. The findings from this study suggested that freezing-based storage does not significantly impact the observed mechanical properties of one of the five AHV leaflets-the TVAL. The results from this study are useful for reaffirming the experimental methodologies in the previous studies, as well as informing the tissue preservation methods of future investigations of AHV leaflet mechanics.
Asunto(s)
Criopreservación , Fenómenos Mecánicos , Válvula Tricúspide/citología , Animales , Fenómenos Biomecánicos , Porcinos , Válvula Tricúspide/fisiologíaRESUMEN
Atrioventricular heart valves (AHVs) are composed of structurally complex and morphologically heterogeneous leaflets. The coaptation of these leaflets during the cardiac cycle facilitates unidirectional blood flow. Valve regurgitation is treated preferably by surgical repair if possible or replacement based on the disease state of the valve tissue. A comprehensive understanding of valvular morphology and mechanical properties is crucial to refining computational models, serving as a patient-specific diagnostic and surgical tool for preoperative planning. Previous studies have modeled the stress distribution throughout the leaflet's thickness, but validations with layer-specific biaxial mechanical experiments are missing. In this study, we sought to fill this gap in literature by investigating the impact of microstructure constituents on mechanical behavior throughout the thickness of the AHVs' anterior leaflets. Porcine mitral valve anterior leaflets (MVAL) and tricuspid valve anterior leaflets (TVAL) were micro-dissected into three layers (atrialis/spongiosa, fibrosa, and ventricular) and two layers (atrialis/spongiosa and fibrosa/ventricularis), respectively, based on their relative distributions of extracellular matrix components as quantified by histological analyses: collagen, elastin, and glycosaminoglycans. Our results suggest that (i) for both valves, the atrialis/spongiosa layer is the most extensible and anisotropic layer, possibly due to its relatively low collagen content as compared to other layers, (ii) the intact TVAL response is stiffer than the atrialis/spongiosa layer but more compliant than the fibrosa/ventricularis layer, and (iii) the MVAL fibrosa and ventricularis layers behave nearly isotropic. These novel findings emphasize the biomechanical variances throughout the AHV leaflets, and our results could better inform future AHV computational model developments. STATEMENT OF SIGNIFICANCE: This study, which is the first of its kind for atrioventricular heart valve (AHV) leaflet tissue layers, rendered a mechanical characterization of the biaxial mechanical properties and distributions of extracellular matrix components (collagen, elastin, and glycosaminoglycans) of the mitral and tricuspid valve anterior leaflet layers. The novel findings from the present study emphasize the biomechanical variances throughout the thickness of AHV leaflets, and our results indicate that the previously-adopted homogenous leaflet in the AHV biomechanical modeling may be an oversimplification of the complex leaflet anatomy. Such improvement in the understanding of valvular morphology and tissue mechanics is crucial to future refinement of AHV computational models, serving as a patient-specific diagnostic and surgical tool, at the preoperative stage, for treating valvular heart diseases.
Asunto(s)
Válvula Mitral/fisiología , Válvula Tricúspide/fisiología , Animales , Anisotropía , Fenómenos Biomecánicos , Válvula Mitral/citología , Porcinos , Válvula Tricúspide/citologíaRESUMEN
AIMS: Calcific aortic valve stenosis (CAVS) is the most frequent manifestation of aortic valve disease and the third leading cause of cardiovascular disease in the Western countries associated with significant morbidity and mortality. An active biological progression involving inflammation and oxidation leading to valve endothelial damage is considered a hallmark of the early stages of valve degeneration. However, tricuspid (TAV) and bicuspid (BAV) aortic valve deterioration are considered to differ only by shear stress. We hypothesized that endothelial cells (EC) derived from BAV and TAV patients have different miRNA expression patterns and thus distinct pathways could lead to endothelial damage in BAV than TAV patients. METHODS AND RESULTS: We isolated ECs from patients with bicuspid or tricuspid aortic valve, which underwent surgery due to CAVS. MiRNA expression profile by PCR revealed eight upregulated miRNAs between BAV and TAV ECs. Functional analysis identified that BAV ECs presented altered cellular response to oxidative stress and DNA damage stimulus via p53 and alteration in the intrinsic apoptotic signaling pathway. GPX3 and SRXN1 mRNA were express at lower levels in BAV compared to TAV ECs, leading to an increment of DNA double-strand breaks. BAV ECs had a sustained apoptosis activation when compared to TAV ECs. This difference was exacerbated by oxidative stress stimulus leading to a reduced survival rate but completely reverted by miR-328-3p inhibition. CONCLUSION: The present data showed molecular differences in oxidative stress susceptibility, DNA damage magnitude, and apoptosis induction between ECs derived from BAV and TAV patients.
Asunto(s)
Válvula Aórtica/anomalías , MicroARNs/metabolismo , Válvula Tricúspide/citología , Anciano , Válvula Aórtica/citología , Enfermedad de la Válvula Aórtica Bicúspide , Western Blotting , Células Cultivadas , Células Endoteliales/citología , Células Endoteliales/metabolismo , Citometría de Flujo , Enfermedades de las Válvulas Cardíacas , Humanos , Persona de Mediana Edad , Estrés Oxidativo/genética , Estrés Oxidativo/fisiología , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
BACKGROUND: Aortic valve calcification is characterized as the active process of aortic valve interstitial cells (AVICs), and considered as an inflammatory disease. As an antioxidant, the anti-inflammatory activity of Lazaroid has been exhibited in various models. We hypothesized that Lazaroid U-74389G would inhibit the osteoblastic differentiation of AVICs induced by IL-1ß. METHODS: Normal tricuspid aortic valve leaflets were collected from patients with acute aortic dissection (Type A) undergoing the Bentall procedure. AVICs were isolated and stimulated with IL-1ß in presence or absence of U-74389G in culture. Cell lysates were analyzed for osteogenic markers and nuclear factor-κB using real-time PCR and Immunoblotting. Culture media was analyzed for IL-6 and IL-8 with enzyme-linked immunosorbent assay. Alizarin Red Staining was adopted to demonstrate the calcium deposition. RESULTS: The expression of alkaline phosphatase and bone morphogenetic protein, accompanied by the production of IL-6 and IL-8, was up-regulated in response to IL-1ß and was inhibited by the addition of U-74389G. The NF-κB pathway was activated by IL-1ß and involved in the suppression of U-74389G on osteoblastic differentiation in AVICs. The negative effects of U-74389G on ostengenic gene expression and mineralization of AVICs were blocked by glucocorticoid receptor antagonist mifepristone and the NF-κB inhibitor Bay 11-7082. CONCLUSIONS: U-74389G inhibits the pro-osteogenic response to IL-1ß stimulation in AVICs. The osteoblastic differentiation and mineralization of AVICs were inhabited by U-74389G though the modulation of NF-κB activation, and this pathway could be potential therapeutic targets for medical treatment of calcified aortic valve disease.
Asunto(s)
Interleucina-1beta/farmacología , Osteogénesis/efectos de los fármacos , Pregnatrienos/farmacología , Receptores de Glucocorticoides/metabolismo , Factor de Transcripción ReIA/antagonistas & inhibidores , Fosfatasa Alcalina/biosíntesis , Válvula Aórtica/citología , Válvula Aórtica/patología , Estenosis de la Válvula Aórtica/patología , Proteínas Morfogenéticas Óseas/biosíntesis , Calcinosis/patología , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Humanos , Mediadores de Inflamación/metabolismo , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Masculino , Mifepristona/farmacología , Miocitos del Músculo Liso/metabolismo , Nitrilos/farmacología , Osteoblastos/citología , Fosforilación/efectos de los fármacos , Receptores de Glucocorticoides/antagonistas & inhibidores , Sulfonas/farmacología , Válvula Tricúspide/citologíaRESUMEN
Previous research in our lab suggested that heart valve tissues cultured without mechanical stimulation do not retain their in vivo microstructure, i.e., cell density decreased within the deep tissue layers and increased at the periphery. In this study, a splashing rotating bioreactor was designed to apply mechanical stimulation to a mitral valve leaflet segment. Porcine valve segments (n = 9-10 per group) were cultured in the bioreactor for 2 weeks (dynamic culture), negative controls were cultured without mechanical stimulation (static culture), and baseline controls were fresh uncultured samples. Overall changes in cellularity and extracellular matrix (ECM) structure were assessed by H&E and Movat pentachrome stains. Tissues were also immunostained for multiple ECM components and turnover mediators. After 2 weeks of culture, proliferating cells were distributed throughout the tissue in segments cultured in the bioreactor, in contrast to segments cultured without mechanical stimulation. Most ECM components, especially collagen types I and III, better maintained normal expression patterns and magnitudes (as found in baseline controls) over 2 weeks of dynamic organ culture compared to static culture. Lack of mechanical stimulation changed several aspects of the tissue microstructure, including the cell distribution and ECM locations. In conclusion, mechanical stimulation by the bioreactor maintained tissue integrity, which will enable future in vitro investigation of mitral valve remodeling.
Asunto(s)
Reactores Biológicos , Proliferación Celular , Matriz Extracelular/metabolismo , Válvula Tricúspide/citología , Válvula Tricúspide/metabolismo , Animales , Técnicas de Cultivo de Órganos/métodos , PorcinosRESUMEN
A ecocardiografia representa hoje técnica não-invasiva de investigação diagnóstica amplamente empregada na avaliação de pacientes portadores de valvopatias, de cardiomiopatias não relacionadas a doença arterial coronária, assim como de indivíduos apresentando doenças do pericárdio. A ecocardiografia caracteriza-se por ser método de investigação cardíaca anatômica não-invasiva, não-radioativa, apresentando alta reprodutibilidade, fácil acesso, baixo custo e grande correlação com métodos invasivos hemodinâmicos de aferição de pressões cardíacas...
Asunto(s)
Humanos , Enfermedades de las Válvulas Cardíacas/complicaciones , Ecocardiografía/métodos , Estenosis de la Válvula Mitral , Cardiomiopatías/sangre , Endocarditis/historia , Válvula Tricúspide/citologíaRESUMEN
Pigmentation of murine cardiac tricuspid valve leaflet is associated with melanocyte concentration, which affects its stiffness. Owing to its biological and viscoelastic nature, estimation of the in situ stiffness measurement becomes a challenging task. Therefore, quasi-static and nanodynamic mechanical analysis of the leaflets of the mouse tricuspid valve is performed in the current work. The mechanical properties along the leaflet vary with the degree of pigmentation. Pigmented regions of the valve leaflet that contain melanocytes displayed higher storage modulus (7-10 GPa) than non-pigmented areas (2.5-4 GPa). These results suggest that the presence of melanocytes affects the viscoelastic properties of the mouse atrioventricular valves and are important for their proper functioning in the organism.
Asunto(s)
Elasticidad , Melanocitos/fisiología , Pigmentación/fisiología , Válvula Tricúspide/fisiología , Animales , Fenómenos Biomecánicos , Ratones , Válvula Tricúspide/citologíaRESUMEN
Regulation of normal cardiac development involves numerous transcription factors, cytoskeletal proteins, signaling molecules, and extracellular matrix proteins. These key molecular components act in concert to induce morphological changes essential for the proper development of a functional four-chambered heart. Growth factors such as BMPs and TGFbeta's play a role in migration, proliferation and differentiation during cardiac development and are important regulators of the extracellular matrix (ECM). Genes responsive to these morphogens are likely to play an equally significant role during cardiac development. Therefore, we sought to clone the chicken TGFbeta induced gene betaig-H3 and evaluate its spatio-temporal expression during heart morphogenesis. Our studies show by Northern analysis, whole mount and section in situ hybridization experiments that betaig-H3 is expressed primarily in the mesenchyme of the atrioventricular and outflow tract cushions and later in the right and left atrioventricular valve leaflets and supporting valve structures. The mRNA expression domains of betaig-H3 show a complementary pattern compared to that of its highly homologous relative, periostin.
Asunto(s)
Proteínas de la Matriz Extracelular/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Corazón/embriología , Organogénesis/genética , Factor de Crecimiento Transformador beta/metabolismo , Animales , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular/metabolismo , Embrión de Pollo , Secuencia Conservada , Proteínas de la Matriz Extracelular/genética , Regulación de la Expresión Génica/genética , Ventrículos Cardíacos/citología , Ventrículos Cardíacos/embriología , Ventrículos Cardíacos/metabolismo , Humanos , Mesodermo/citología , Mesodermo/metabolismo , Válvula Mitral/citología , Válvula Mitral/embriología , Válvula Mitral/metabolismo , ARN Mensajero/metabolismo , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie , Factor de Crecimiento Transformador beta/genética , Válvula Tricúspide/citología , Válvula Tricúspide/embriología , Válvula Tricúspide/metabolismoRESUMEN
We report the consistent distribution of a population of pigmented trp-1-positive cells in several important septal and valvular structures of the normal mouse (C57BL/6) heart. The pigmented cell population was first apparent by E16.5 p.c. in the right atrial wall and extended into the atrium along the interatrial septum. By E17.5, these cells were found along the apical membranous interventricular septum near or below the surface of the endocardium. The most striking distribution of dark pigmented cells was found in the tricuspid and mitral valvular leaflets and chordae tendineae. The normal distribution of pigmented cells in the valvuloseptal apparatus of C57BL/6 adult heart suggests that a premelanocytic lineage may participate in the earlier morphogenesis of the valve leaflets and chordae tendineae. The origin of the premelanocyte lineage is currently unknown. The most likely candidate populations include the neural crest and the epicardially derived cells. The only cell type in the heart previously shown to form melanocytes is the neural crest. The presence of neural crest cells, but not melanocytes, in some of the regions we describe has been reported by others. However, previous reports have not shown a contribution of melanocytes or neural crest derivatives to the atrioventricular valve leaflets or chordae tendineae in mouse hearts. If these cells are of neural crest origin, it would suggest a possibly greater contribution and persistence of neural crest cells to the valvuloseptal apparatus than has been previously understood.
Asunto(s)
Válvulas Cardíacas/citología , Corazón/anatomía & histología , Melanocitos/citología , Ratones Endogámicos C57BL/anatomía & histología , Animales , Técnicas de Cultivo de Célula , Linaje de la Célula , Cuerdas Tendinosas/química , Cuerdas Tendinosas/citología , Corazón Fetal/química , Corazón Fetal/citología , Corazón/embriología , Tabiques Cardíacos/química , Tabiques Cardíacos/citología , Válvulas Cardíacas/química , Válvulas Cardíacas/embriología , Inmunohistoquímica , Melanocitos/química , Ratones , Válvula Mitral/química , Válvula Mitral/citología , Oxidorreductasas/análisis , Válvula Tricúspide/química , Válvula Tricúspide/citologíaRESUMEN
The objective of this study was to evaluate the capacity of three clinically useful tissue sources: tricuspid valve leaflet (TVL), carotid artery (CA), and jugular vein (JV), to generate myofibroblasts for potential use in a tissue-engineered cardiac valve replacement. Tissue biopsies of clinically appropriate sizes obtained from juvenile sheep were used for this work. Cells obtained from all three tissue sources exhibited a myofibroblast phenotype in vitro, as demonstrated by their immunoreactivity with antibodies directed against vimentin, alpha-smooth muscle actin, fibronectin, and chondroitin sulfate. Protein synthesis characteristics were defined for the key extracellular matrix components: collagen, glycosaminoglycans, and elastin. Among the three sources, JV generated the highest numbers of cells, and JV cells produced the largest amount of collagen per cell. These data suggest that venous tissue, with its relative ease of accessibility, may generate myofibroblasts potentially useful for the interstitial cellular component of a tissue-engineered cardiac valve.
Asunto(s)
Prótesis Valvulares Cardíacas , Válvulas Cardíacas/citología , Músculo Liso Vascular/citología , Ingeniería de Tejidos/métodos , Actinas/metabolismo , Animales , Biopsia , Arterias Carótidas/citología , Técnicas de Cultivo de Célula/métodos , Proliferación Celular , Células Cultivadas , Sulfatos de Condroitina/metabolismo , Colágeno/biosíntesis , Matriz Extracelular/química , Matriz Extracelular/metabolismo , Fibronectinas/metabolismo , Válvulas Cardíacas/fisiología , Válvulas Cardíacas/trasplante , Venas Yugulares/citología , Modelos Animales , Músculo Liso Vascular/fisiología , Ovinos , Válvula Tricúspide/citología , Vimentina/metabolismoRESUMEN
We used a genetic lineage-labeling system to establish the material contributions of the progeny of 3 specific cell types to the cardiac valves. Thus, we labeled irreversibly the myocardial (alphaMHC-Cre+), endocardial (Tie2-Cre+), and neural crest (Wnt1-Cre+) cells during development and assessed their eventual contribution to the definitive valvar complexes. The leaflets and tendinous cords of the mitral and tricuspid valves, the atrioventricular fibrous continuity, and the leaflets of the outflow tract valves were all found to be generated from mesenchyme derived from the endocardium, with no substantial contribution from cells of the myocardial and neural crest lineages. Analysis of chicken-quail chimeras revealed absence of any substantial contribution from proepicardially derived cells. Molecular and morphogenetic analysis revealed several new aspects of atrioventricular valvar formation. Marked similarities are seen during the formation of the mural leaflets of the mitral and tricuspid valves. These leaflets form by protrusion and growth of a sheet of atrioventricular myocardium into the ventricular lumen, with subsequent formation of valvar mesenchyme on its surface rather than by delamination of lateral cushions from the ventricular myocardial wall. The myocardial layer is subsequently removed by the process of apoptosis. In contrast, the aortic leaflet of the mitral valve, the septal leaflet of the tricuspid valve, and the atrioventricular fibrous continuity between these valves develop from the mesenchyme of the inferior and superior atrioventricular cushions. The tricuspid septal leaflet then delaminates from the muscular ventricular septum late in development.
Asunto(s)
Endocardio/citología , Válvulas Cardíacas/embriología , Mesodermo/citología , Animales , Apoptosis , Linaje de la Célula , Movimiento Celular , Embrión de Pollo , Quimera/embriología , Cuerdas Tendinosas/citología , Cuerdas Tendinosas/embriología , Coturnix/embriología , Corazón Fetal/citología , Genes Reporteros , Edad Gestacional , Válvulas Cardíacas/citología , Imagenología Tridimensional , Integrasas/genética , Péptidos y Proteínas de Señalización Intercelular/genética , Ratones , Ratones Transgénicos , Válvula Mitral/citología , Válvula Mitral/embriología , Morfogénesis , Miocardio/citología , Cresta Neural/citología , Receptor TIE-2/genética , Eliminación de Secuencia , Válvula Tricúspide/citología , Válvula Tricúspide/embriología , Proteínas Virales/genética , Proteínas Wnt , Proteína Wnt1RESUMEN
BACKGROUND AND AIM OF THE STUDY: Organ cultures maintain cells within their native microstructural environment, and thus offer greater potential for studying tissue disease and remodeling than do monolayer cell cultures or pathological examinations of diseased tissue. To validate an in-vitro heart valve organ culture model, cell viability was examined within valve tissues over sustained culture periods. METHODS: Following culture of blocks of valve tissue for 1 to 49 days, cross-sections were cut with a vibratome, stained with a LIVE/DEAD kit, and imaged with confocal microscopy to quantify the number of live and dead cells present. RESULTS: In numerous organ cultures, valvular interstitial cells were found to be viable beyond 30 days. Live cells were abundant in the central region of the valve, but more sparse in the deepest central regions. Dead cells were found mainly on the surface of both fresh tissues and tissues after prolonged culture, with few dead cells occurring centrally. CONCLUSION: This is the first reported mapping of cell viability within heart valve organ cultures, and results suggest that extended organ culture of valve leaflets is indeed possible. The derived viability staining methods have wide applicability for organ cultures of other tissues as well as tissue-engineered matrices.
Asunto(s)
Válvula Aórtica/citología , Válvula Mitral/citología , Válvula Tricúspide/citología , Animales , Válvula Aórtica/metabolismo , Bovinos , Recuento de Células , Supervivencia Celular/fisiología , Células Cultivadas , Colorantes , Perros , Células Endoteliales/citología , Células Endoteliales/metabolismo , Eosina Amarillenta-(YS) , Hematoxilina , Humanos , Microscopía Electrónica de Rastreo , Válvula Mitral/metabolismo , Modelos Cardiovasculares , Técnicas de Cultivo de Órganos , Porcinos , Tiempo , Válvula Tricúspide/metabolismoRESUMEN
We determined how long cryopreserved aortic, pulmonic, mitral, and tricuspid valve homografts could be stored in a deep freezer (-80 degrees C) without compromising fibroblast viability. Valves harvested from 20 anesthetized mongrel dogs were grouped into nonfrozen control, frozen and stored in liquid nitrogen (-196 degrees C), and frozen and stored in a deep freezer (-80 degrees C). Frozen groups were divided into subgroups and stored for 2, 4, 8, or 12 weeks. A leaflet of each valve was divided into three fragments, and fibroblast viability was analyzed by flow cytometry. Cell viability was defined as staining by fluorescent diacetate but not by propidium iodide. The viability of untreated control valves from all four sites was about 70%, decreasing to about 50% when treated with low doses of antibiotics. The viability of frozen valves stored in liquid nitrogen was about 45% without a significant difference among storage periods. The viability of valves frozen and stored in a deep freezer was significantly lower than for the liquid nitrogen group at 2 weeks for the mitral valve and at 4 weeks for other valves. These results suggest that homografts can be stored in a deep freezer for up to 2 weeks without deterioration.
Asunto(s)
Frío , Criopreservación , Válvulas Cardíacas/fisiología , Animales , Válvula Aórtica/citología , Válvula Aórtica/fisiología , Supervivencia Celular , Perros , Válvulas Cardíacas/citología , Válvula Mitral/citología , Válvula Mitral/fisiología , Válvula Pulmonar/citología , Válvula Pulmonar/fisiología , Trasplante Homólogo , Válvula Tricúspide/citología , Válvula Tricúspide/fisiologíaRESUMEN
The distribution of vasoactive intestinal polypeptide-, neuropeptide Y-, and calcitonin gene-related peptide-immunoreactive nerve fibres was investigated in the atrioventricular valves of the rat. These nerve fibres were visualized by immunostaining of whole-mount preparations by the avidin-biotin-peroxidase complex method. Vasoactive intestinal polypeptide-immunoreactive nerve fibres were observed mainly in the anterior cusp of the mitral valve and, to a lesser extent, in the medial cusp of the tricuspid valve. Numerous neuropeptide Y-immunoreactive nerve fibres were found covering all of the cusps. Both types of peptidergic nerve fibre formed dense networks that consisted of interlacing and anastomosing nerve fibres. Calcitonin gene-related peptide-immunoreactive nerve fibres were seen in every cusp, but did not form a fine network. These results provide detailed anatomical information for evaluation of the possible roles of each type of peptide-containing nerve fibre in the function of atrioventricular valves.
Asunto(s)
Válvulas Cardíacas/inervación , Fibras Nerviosas/metabolismo , Neuropéptidos/metabolismo , Animales , Péptido Relacionado con Gen de Calcitonina/metabolismo , Femenino , Válvulas Cardíacas/citología , Inmunohistoquímica , Válvula Mitral/citología , Válvula Mitral/inervación , Neuropéptido Y/metabolismo , Ratas , Ratas Wistar , Sustancia P/metabolismo , Válvula Tricúspide/citología , Válvula Tricúspide/inervación , Péptido Intestinal Vasoactivo/metabolismoRESUMEN
Immunohistochemical research on atrioventricular valves in normotensive rats revealed that valvular myocardiocytes are the seat of synthesis of the atrial natriuretic factor (ANF). The endocardial cells that border the atrial and ventricular surfaces also had granules which were positive for ANF. The ANF which is also synthesized in the valvular myocardiocytes moved towards the cardiac cavity and crossed the endocardial cells in the more distal areas of the valvular edge. At the same time, the ANF was routed into the blood vessels in the areas close to implantation.
Asunto(s)
Factor Natriurético Atrial/biosíntesis , Endocardio/metabolismo , Válvula Mitral/química , Miocardio/metabolismo , Válvula Tricúspide/química , Animales , Factor Natriurético Atrial/análisis , Factor Natriurético Atrial/inmunología , Gránulos Citoplasmáticos , Inmunohistoquímica , Válvula Mitral/citología , Ratas , Ratas Wistar , Válvula Tricúspide/citologíaRESUMEN
Dense distribution of varicose fibres containing neuropeptide Y-like immunoreactivity (NPY-LI) was found in the atrioventricular valves of the Japanese monkey, and moderately in the rat. The immunoelectron microscopy using immunogolds resulted in the localization of NPY-LI within the dense-cored vesicles which existed with the small clear vesicles in the unmyelinated axons near the endocardium. These NPY-LI-containing fibres may participate in regulation of vasomotor role or other functions of the atrioventricular valves.
Asunto(s)
Válvula Mitral/inervación , Fibras Nerviosas/ultraestructura , Neuropéptido Y/análisis , Válvula Tricúspide/inervación , Animales , Nodo Atrioventricular/citología , Nodo Atrioventricular/ultraestructura , Axones/ultraestructura , Endotelio/inervación , Endotelio/ultraestructura , Inmunohistoquímica/métodos , Macaca , Microscopía Inmunoelectrónica/métodos , Válvula Mitral/citología , Válvula Mitral/ultraestructura , Ratas , Ratas Wistar , Válvula Tricúspide/citología , Válvula Tricúspide/ultraestructuraRESUMEN
More microappendages are present on both surfaces of the guinea pig tricuspid valve than on the mitral valve. The distal edge of all valve surfaces has more microappendages than the remainder of the valve surface. These results suggest that microappendage number may be the result of haemodynamic forces on the valve surface: the greater the force the smaller the number of microappendages. The function of microappendages on the surfaces of the heart valves is unknown, but two important roles have been suggested, namely to increase the surface area of the endothelial cells and to participate in the metabolism of these cells.
Asunto(s)
Válvulas Cardíacas/citología , Animales , Endotelio/citología , Femenino , Cobayas , Masculino , Válvula Mitral/citología , Válvula Tricúspide/citologíaRESUMEN
1. The electrophysiology of single myocytes isolated from the rabbit tricuspid valve was studied using the patch-clamp method (whole-cell configuration). Cell dispersion was achieved by collagenase treatment, using the Langendorff retrograde perfusion procedure. 2. After isolation, and while incubating in the recovery (Kraftbrühe) solution, cells had clear striations and were mostly spindle-shaped, or rod-like (less than 10%), with length varying from 35 microns to over 150 microns, and diameter from 3 to 10 microns. 3. Upon exposure to Tyrode solution, the calcium-tolerant cells were mostly rounded with smooth surfaces and well-defined borders. The mean diameter of these cells was 15 +/- 5 microns (S.D., n = 9). A smaller percentage (about 30%) retained the original elongated shape. 4. Patch pipette recordings showed the presence of spontaneous activity in about 30% of round cells, and less frequently in elongated cells. Maximum diastolic potentials (MDPs) in the round cells averaged -82 +/- 6 mV, with a take-off potential of -56 +/- 3 mV (n = 9), and an average maximum upstroke velocity (Vmax) value of 6.3 +/- 0.6 V/s (n = 4). In quiescent cells, the mean resting potential was 69 +/- 12 mV (n = 43). 5. Voltage clamp ramps revealed a steady-state I-V relation with a negative slope region. The mean input resistance value was 25 +/- 9 M omega (n = 16) for the elongated, and 883 +/- 481 M omega (n = 8) for the round cells. 6. Hyperpolarizing 5 s pulses (holding potential = -50 mV) occasionally revealed a slow, time-dependent inward current whose peak increased progressively as a function of clamp potential. The slowly activating current was sensitive to caesium 2 mM), indicating its similarity to the so-called 'pacemaker current' (iF). In alternate voltage- and current-clamp experiments, blocking of iF did not stop pacemaker activity, but there was up to a fourfold increase in pacemaker cycle length. 7. In some cells, 5 s hyperpolarizing steps from a holding potential of -40 or -50 mV produced large, inwardly directed and voltage-dependent current surges that decayed rapidly with time, similar to the inactivation described for the inward rectifier current, iK1. The current was very prominent at voltages more negative than -100 mV, and its decay process was best fitted by two time constants, one fast and one slow. For example, at -150 mV the time constants were 61 and 634 ms. The inward current was blocked by barium (1 mM).(ABSTRACT TRUNCATED AT 400 WORDS)
Asunto(s)
Válvula Tricúspide/fisiología , Potenciales de Acción , Animales , Calcio/metabolismo , Células Cultivadas , Cinética , Potenciales de la Membrana , Conejos , Tetrodotoxina/farmacología , Válvula Tricúspide/citologíaRESUMEN
The hypothesis that cocaine has Class I-type antiarrhythmic drug effects was tested in tissues isolated from rabbit heart with standard microelectrode methods. Propranolol (1 microM) was used to block beta-adrenergic effects. The actions of cocaine on cellular electrophysiology were concentration- and time-dependent and were reversible. In paced right atrial (RA) and right ventricular papillary (RVP) tissues, cocaine produced a profound prolongation of the effective refractory period (ERP) assessed by either premature stimulation or minimum pacing interval. ERP was increased up to eightfold in RA tissue and doubled in RVP tissue by 60 microM cocaine. This concentration of cocaine depressed action potential phase 0 depolarization 80% in RA tissue and 53% in RVP tissue but had no effect on resting membrane potentials. Automaticity was moderately depressed in sinus node (34% decrease in rate) but not in tricuspid valve cells. Phase 0 depolarization was not altered in these spontaneously active slow-response cells. Repolarization was depressed in RA, tricuspid valve, and sinus node cells leading to a twofold increase in action potential duration during exposure to cocaine. Evidence from the effects on cellular action potentials suggests that cocaine affects both fast Na+ channels and repolarizing K+ but not Ca2+ channels. We conclude that cocaine has Class I-type activity and the effects on ERP are extreme.