RESUMEN

Antimicrobial peptides (AMP), part of the innate immune system, are well studied for their ability to kill pathogenic microorganisms. However, many also possess important immunomodulatory effects, and this area has potential for the development of novel therapies to supplement traditional methods such as the use of antibiotics. Here, we characterise the microbicidal and immunomodulatory potential of the proline-rich bovine AMP, Bactenecin 5 (Bac5). We demonstrate broad antimicrobial activity, including against some mycobacterial species, which are important pathogens of fish, cattle and humans. Bac5 is able to activate macrophage-like THP-1 cells and can synergistically trigger the upregulation of tnf-α when co-stimulated with M. marinum. Furthermore, Bac5 sensitises A549 epithelial cells to stimulation with TNF-α. For the first time, we characterise the activity of Bac5 in vivo, and show it to be a potent chemokine for macrophages in the zebrafish (Danio rerio) embryo model of infection. Bac5 also supports the early recruitment of neutrophils in the presence of M. marinum. In the absence of host adaptive immunity, exogenous injected Bac5 is able to slow, although not prevent, infection of zebrafish with M. marinum.

Asunto(s)

Péptidos Catiónicos Antimicrobianos/farmacología , Péptidos Cíclicos/farmacología , Células A549 , Secuencia de Aminoácidos , Animales , Péptidos Catiónicos Antimicrobianos/química , Bacillus/efectos de los fármacos , Bovinos , Quimiocinas/metabolismo , Humanos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Mycobacterium marinum/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Células THP-1 , Transcripción Genética/efectos de los fármacos , Pez Cebra/embriología

RESUMEN

It is of the utmost importance to increase the activity of bone cells on the surface of materials used in the design of orthopaedic implants. Increased activity of such cells can promote either integration of these materials into surrounding bone or complete replacement with naturally produced bone if biodegradable materials are used. Osteoblasts are bone-producing cells and, for that reason, are the cells of interest in initial studies of new orthopaedic implants. If these cells are functioning normally, they lay down bone matrix onto both existing bone and prosthetic materials implanted into the body. It is generally accepted that a successful material should enhance osteoblast function, leading to more bone deposition and, consequently, increased strength of the interface between the material and juxtaposed bone. The present study provided the first evidence of greater osteoblast function on carbon and alumina formulations that mimic the nano-dimensional crystal geometry of hydroxyapatite found in bone.

Asunto(s)

Óxido de Aluminio , Carbono , Osteoblastos/fisiología , Prótesis e Implantes , Materiales Biocompatibles , Fibra de Carbono , Adhesión Celular , Humanos , Nanotecnología/métodos

RESUMEN

In this study, we investigated cardiomyocyte cytoarchitecture in a mouse model for dilated cardiomyopathy (DCM), the muscle LIM protein (MLP) knockout mouse and substantiated several observations in a second DCM model, the tropomodulin-overexpressing transgenic (TOT) mouse. Freshly isolated cardiomyocytes from both strains are characterized by a more irregular shape compared with wild-type cells. Alterations are observed at the intercalated disks, the specialized areas of mechanical coupling between cardiomyocytes, whereas the subcellular organization of contractile proteins in the sarcomeres of MLP knockout mice appears unchanged. Distinct parts of the intercalated disks are affected differently. Components from the adherens junctions are upregulated, desmosomal proteins are unchanged, and gap junction proteins are downregulated. In addition, the expression of N-RAP, a LIM domain- containing protein located at the intercalated disks, is upregulated in MLP knockout as well as in TOT mice. Detailed analysis of intercalated disk composition during postnatal development reveals that an upregulation of N-RAP expression might serve as an early marker for the development of DCM. Altered expression levels of cytoskeletal proteins (either the lack of MLP or an increased expression of tropomodulin) apparently lead to impaired function of the myofibrillar apparatus and to physiological stress that ultimately results in DCM and is accompanied by an altered appearance and composition of the intercalated disks.

Asunto(s)

Proteínas de Microfilamentos , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/ultraestructura , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Animales , Cardiomiopatía Dilatada/metabolismo , Cardiomiopatía Dilatada/patología , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Uniones Comunicantes/metabolismo , Expresión Génica/fisiología , Proteínas con Dominio LIM , Ratones , Ratones Noqueados , Microscopía Electrónica , Fibras Musculares Esqueléticas/patología , Sarcómeros/metabolismo , Tropomodulina

RESUMEN

In the heart, the intercellular geometry of myocyte coupling by Connexin43-gap junctions (Cx43-gjs) is a determinant of normal and abnormal patterns of propagation of electrical excitation. ZO-1 has been suggested to play a role in determining the pattern of intercellular coupling between myocytes. We therefore investigated the co-distribution of Cx43 with ZO-1 in ventricular myocytes of the adult rat using quantitative immunoconfocal microscopy. Our data indicates that low-moderate levels of co-immunolocalization occur between Cx43 and ZO-1 in normal ventricular myocardium. However, rapid and significant increases in relative co-localization occur between Cx43 and ZO-1 following dissociation of myocytes from ventricular myocardium--a treatment inducing internalization of Cx43-gjs. This increased relative co-localization may represent an increase in Cx43-ZO-1 interaction, suggesting a role for ZO-1 in the remodeling of myocardial Cx43-gjs. A more comprehensive study, including immunoprecipitation and immunoelectron microscopy analyses has been carried out (Barker et al. Circ. Res., in press, 2002 and as presented to the 2001 International GJ Conference). This study further assesses the biological relevance of the increased association between ZO-1 and Cx43 accompanying internalization of Cx43-gjs.

Asunto(s)

Conexina 43/metabolismo , Proteínas de la Membrana/metabolismo , Miocitos Cardíacos/metabolismo , Fosfoproteínas/metabolismo , Animales , Comunicación Celular/fisiología , Ventrículos Cardíacos/citología , Ventrículos Cardíacos/metabolismo , Inmunohistoquímica , Microscopía Confocal , Miocitos Cardíacos/citología , Ratas , Ratas Sprague-Dawley , Proteína de la Zonula Occludens-1

RESUMEN

BACKGROUND: To determine potential mechanisms of the transition from hypertrophy to very early failure, we examined apoptosis in a model of ascending aortic stenosis (AS) in male FVB/n mice. METHODS AND RESULTS: Compared with age-matched controls, 4-week and 7-week AS animals (n=12 to 16 per group) had increased ratios of left ventricular weight to body weight (4.7+/-0.7 versus 3.1+/-0.2 and 5. 7+/-0.4 versus 2.7+/-0.1 mg/g, respectively, P<0.05) with similar body weights. Myocyte width was also increased in 4-week and 7-week AS mice compared with controls (19.0+/-0.8 and 25.2+/-1.8 versus 14. 1+/-0.5 microm, respectively, P<0.01). By 7 weeks, AS myocytes displayed branching with distinct differences in intercalated disk size and staining for beta(1)-integrin on both cell surface and adjacent extracellular matrix. In vivo left ventricular systolic developed pressure per gram as well as endocardial fractional shortening were similar in 4-week AS and controls but depressed in 7-week AS mice. Myocyte apoptosis estimated by in situ nick end-labeling (TUNEL) was extremely rare in 4-week AS and control mice; however, a low prevalence of TUNEL-positive myocytes and DNA laddering were detected in 7-week AS mice. The specificity of TUNEL labeling was confirmed by in situ ligation of hairpin oligonucleotides. CONCLUSIONS: Our findings indicate that myocyte apoptosis develops during the transition from hypertrophy to early failure in mice with chronic biomechanical stress and support the hypothesis that the disruption of normal myocyte anchorage to adjacent extracellular matrix and cells, a process called anoikis, may signal apoptosis.

Asunto(s)

Estenosis de la Válvula Aórtica/complicaciones , Animales , Apoptosis/fisiología , Gasto Cardíaco Bajo/etiología , Comunicación Celular/fisiología , Progresión de la Enfermedad , Ecocardiografía , Hemodinámica , Hipertrofia Ventricular Izquierda/etiología , Hipertrofia Ventricular Izquierda/patología , Hipertrofia Ventricular Izquierda/fisiopatología , Integrina beta1/metabolismo , Masculino , Ratones , Ratones Endogámicos , Microscopía Confocal , Distribución Tisular

RESUMEN

The ras family of small GTP-binding proteins exerts powerful effects upon cell structure and function. One member of this family, rac, induces actin cytoskeletal reorganization in nonmuscle cells and hypertrophic changes in cultured cardiomyocytes. To examine the effect of rac1 activation upon cardiac structure and function, transgenic mice were created that express constitutively activated rac1 specifically in the myocardium. Transgenic rac1 protein was expressed at levels comparable to endogenous rac levels, with activation of the rac1 signaling pathway resulting in two distinct cardiomyopathic phenotypes: a lethal dilated phenotype associated with neonatal activation of the transgene and a transient cardiac hypertrophy seen among juvenile mice that resolved with age. Neither phenotype showed myofibril disarray and hypertrophic hearts were hypercontractilein working heart analyses. The rac1 target p21-activated kinase translocated from a cytosolic to a cytoskeletal distribution, suggesting that rac1 activation was inducing focal adhesion reorganization. Corroborating results showed altered localizations of src in dilated cardiomyopathy and paxillin in both cardiomyopathic phenotypes. This study, the first examination of rac1-mediated cardiac effects in vivo, demonstrates that dilation and hypertrophy can share a common molecular origin and presents evidence that both timing and concurrent signaling from multiple pathways can influence cardiac remodeling.

Asunto(s)

Cardiomiopatía Dilatada/metabolismo , Adhesión Celular , Proteína de Unión al GTP rac1/metabolismo , Animales , Cardiomiopatía Dilatada/mortalidad , Cardiomiopatía Dilatada/patología , Células Cultivadas , Expresión Génica , Corazón , Ratones , Ratones Transgénicos , Miocardio/citología , Miocardio/patología , Fenotipo , Transducción de Señal , Proteína de Unión al GTP rac1/genética

RESUMEN

Dilated cardiomyopathy is characterized by decreased contractile function and loss of myofibril organization. Previously unexplored structural and molecular events that precede and initiate dilation can now be studied in tropomodulin-overexpressing transgenic (TOT) mice exhibiting progressive dilated cardiomyopathy. Onset of dilation did not correspond to a change in transgene expression levels, which were more than threefold above normal at birth and remained elevated throughout postnatal life. Similarly, mitogen-activated protein kinase activation (p38, ERK1/ERK2, JNK1/JNK2) was not associated with dilation. In contrast, calcineurin was activated before dilation, presumably due to doubling of intracellular diastolic calcium levels in TOT cardiomyocytes. Amplitude of systolic calcium transients was greatly increased as well, demonstrating the novel and unique calcium handling profile of TOT cardiomyocytes. Loss of myofibril organization was not apparent by confocal microscopy until over 1 week after birth, although neonatal sarcomeric abnormalities were revealed by ultrastructural analysis. Rapid postnatal increases in heart:body weight ratio at 1.5 weeks were followed by two waves of mortality between 2 and 3 weeks after birth coincident with maturational stress. Ultimately, TOT pathogenesis is a compensatory response to altered sarcomeric structure driven by calcineurin activation within days after birth, making TOTs an excellent paradigm for studying the role of calcium overload in dilated cardiomyopathy.

Asunto(s)

Cardiomiopatía Dilatada/etiología , Cardiomiopatía Dilatada/patología , Proteínas Portadoras/genética , Animales , Calcio/metabolismo , Cardiomiopatía Dilatada/metabolismo , Proteínas Portadoras/biosíntesis , Immunoblotting , Ratones , Ratones Transgénicos , Proteínas de Microfilamentos/biosíntesis , Transducción de Señal , Tropomodulina

RESUMEN

Cardiac hypertrophy is the fundamental adaptation of the adult heart to mechanical load. Recent work has shown that inhibition of calcineurin activity with cyclosporine suppresses the development of hypertrophy in calcineurin transgenic mice and in in vitro systems of neonatal rat cardiocytes stimulated with peptide growth factors. To test the hypothesis that the calcineurin signaling pathway is critical for load-induced hypertrophy in vivo, we examined the effects of cyclosporine treatment on left ventricular hypertrophy induced by experimental ascending aortic stenosis for 4 weeks in mice. Left ventricular systolic pressure was elevated to a similar level in aortic stenosis mice that were treated with cyclosporine versus no drug. Left ventricular mass and myocyte size were similar in treated and untreated aortic stenosis animals and significantly greater than control animals, showing that cyclosporine treatment does not suppress hypertrophic growth. Both treated and untreated animals showed increased left ventricular expression of the load-sensitive gene atrial natriuretic factor. Calcineurin activity was measured in the left ventricle and the spleen from control mice and aortic stenosis mice treated with cyclosporine versus no drug. Levels of calcineurin activity were similar in the spleens of control and untreated aortic stenosis mice. However, calcineurin activity was severely depressed in left ventricular tissue of untreated aortic stenosis mice compared with control mice and was further reduced by cyclosporine treatment. Thus, pathological hypertrophy and cardiac-restricted gene expression induced by pressure overload in vivo are not suppressed by treatment with cyclosporine and do not appear to depend on the elevation of left ventricular calcineurin activity.

Asunto(s)

Inhibidores de la Calcineurina , Ciclosporina/farmacología , Hipertensión/complicaciones , Hipertrofia Ventricular Izquierda/enzimología , Hipertrofia Ventricular Izquierda/etiología , Animales , Aorta/cirugía , Calcineurina/fisiología , Hipertrofia Ventricular Izquierda/patología , Ligadura , Masculino , Ratones , Ratones Endogámicos , Miocardio/patología , Distribución Aleatoria

RESUMEN

This experiment was designed to evaluate the effects of pectin (PE), guar gum (GG) and psyllium (PSY) intake on VLDL and LDL metabolism in female guinea pigs fed high dietary cholesterol. Guinea pigs were fed a 15 g/100 g fat diet containing 0.25 g/100 g cholesterol with 12.5 g/100 g PE, 12.5 g/100 g GG, 7.5 g/100 g PSY or 12.5 g/100 g cellulose (control diet) for 4 wk. Plasma cholesterol concentrations were 29, 43 and 39% lower in guinea pigs fed PE, GG or PSY, respectively, compared with the control group (P < 0.0001). Plasma apolipoprotein (apo) B concentrations were 16-22% lower in the groups fed soluble fiber compared with the control group (P < 0.01). In contrast, hepatic cholesterol and triglyceride concentrations were not different among the PE, GG, PSY and control groups. No differences in triacylglycerol (TAG) or apo B secretion rates, measured by blocking VLDL catabolism by triton (WR 1339) injection, were observed, whereas plasma LDL apo B fractional catabolic rates (FCR), determined by injection of radiolabeled LDL, were higher in guinea pigs fed GG or PSY than in those from the control group. All sources of dietary soluble fiber reduced LDL apo B flux (P < 0.05). These results suggest that the mechanisms of plasma LDL cholesterol lowering by dietary soluble fiber are distinctive for each fiber source and result in specific alterations in lipoprotein metabolism in female guinea pigs. Differences between male and female guinea pigs in response to these diets are discussed.

Asunto(s)

LDL-Colesterol/análisis , Fibras de la Dieta/farmacología , Lipoproteínas/metabolismo , Animales , Apolipoproteínas B/análisis , HDL-Colesterol/análisis , LDL-Colesterol/sangre , Femenino , Galactanos/farmacología , Cobayas , Lipoproteínas/sangre , Lipoproteínas VLDL/análisis , Masculino , Mananos/farmacología , Microsomas Hepáticos/metabolismo , Pectinas/farmacología , Gomas de Plantas , Psyllium/farmacología , Factores Sexuales , Triglicéridos/análisis

RESUMEN

Two major myosin light chain 2 isoforms are coexpressed in the early stages of murine cardiogenesis, a cardiac ventricular isoform and a cardiac atrial isoform, each of which is tightly regulated in a muscle cell-type-specific manner during embryogenesis (Chien, K. R., Zhu, H., Knowlton, K. U., Miller-Hance, W., van Bilsen, M., O'Brien, T. X., and Evans, S. M. (1993) Annu. Rev. Physiol. 55, 77-95). We have disrupted myosin light chain 2v gene in mice and monitored in vivo cardiac function in living myosin light chain 2v -/- embryos. The mutant embryos die at approximately embryonic day 12.5. In mutant ventricles, the myosin light chain 2a protein level is increased and reaches levels comparable to the myosin light chain 2v in the ventricles of wild type littermates and is appropriately incorporated into the thick filaments of mutant embryonic hearts. However, despite the substitution of myosin light chain 2a, ultrastructural analysis revealed defects in sarcomeric assembly and an embryonic form of dilated cardiomyopathy characterized by a significantly reduced left ventricular ejection fraction in mutant embryos compared with wild type littermates. We conclude that myosin light chain 2v may have a unique function in the maintenance of cardiac contractility and ventricular chamber morphogenesis during mammalian cardiogenesis and that a chamber-specific combinatorial code for sarcomeric assembly may exist that ultimately requires myosin light chain 2v in ventricular muscle cells.

Asunto(s)

Miosinas Cardíacas , Corazón/embriología , Corazón/fisiología , Cadenas Ligeras de Miosina/metabolismo , Animales , Femenino , Masculino , Ratones , Microscopía Electrónica , Miocardio/ultraestructura , Cadenas Ligeras de Miosina/genética , Mutación Puntual , Recombinación Genética

RESUMEN

Tropomodulin is a tropomyosin-binding protein that terminates "pointed-end" actin filament polymerization. To test the hypothesis that regulation of tropomodulin:actin filament stoichiometry is critical for maintenance of actin filament length, tropomodulin levels were altered in cells by infection with recombinant adenoviral expression vectors, which produce either sense or antisense tropomodulin mRNA. Neonatal rat cardiomyocytes were infected, and sarcomeric actin filament organization was examined. Confocal microscopy indicated that overexpression of tropomodulin protein shortened actin filaments and caused myofibril degeneration. In contrast, decreased tropomodulin content resulted in the formation of abnormally long actin filament bundles. Despite changes in myofibril structure caused by altered tropomodulin expression, total protein turnover of the cardiomyocytes was unaffected. Biochemical analyses of infected cardiomyocytes indicated that changes in actin distribution, rather than altered actin content, accounted for myofibril reorganization. Ultrastructural analysis showed thin-filament disarray and revealed the presence of leptomeres after tropomodulin overexpression. Tropomodulin-mediated effects constitute a novel mechanism to control actin filaments, and our findings demonstrate that regulated tropomodulin expression is necessary to maintain stabilized actin filament structures in cardiac muscle cells.

Asunto(s)

Proteínas Portadoras/genética , Proteínas de Microfilamentos , Miocardio/metabolismo , Miofibrillas/metabolismo , Actinas/metabolismo , Adenoviridae/genética , Adenoviridae/fisiología , Animales , Proteínas Portadoras/fisiología , Células Cultivadas , ADN Recombinante , Expresión Génica/genética , Expresión Génica/fisiología , Vectores Genéticos/genética , Humanos , Ratones , Miocardio/citología , Miofibrillas/ultraestructura , Miofibrillas/virología , Proteínas/metabolismo , ARN Mensajero/metabolismo , ARN Viral/metabolismo , Ratas , Sarcómeros/química , Sarcómeros/ultraestructura , Sarcómeros/virología , Transfección/genética , Transfección/fisiología , Tropomodulina

Asunto(s)

Curación Mental , Humanos , Control Interno-Externo , Relaciones Profesional-Paciente

RESUMEN

To investigate the possible role of phosphorylation of protein tyrosine during myofibrillogenesis (6- to 13-somite stages) of the chicken embryonic heart tube, immunolocalization of phosphotyrosine (P-Tyr) and the relationship between P-Tyr and developing myofibrils were studied by means of confocal scanning laser microscopy and immuno-electron microscopy. The staining pattern of P-Tyr varied in different sites of myocytes at different stages of embryonic development: At the cell-cell boundaries, P-Tyr was localized at the adhesion belt of outer myocardial layer cells (6- to 13-somite stages), non-junctional cell-cell contacts (6- to 13-somite stages) and early intercalated disks of both the outer and inner myocardial layer cells (8- to 13-somite stages). At the cell-extracellular matrix boundaries of inner layer cells, the first stages of myofibril formation appeared as serially aligned areas of P-Tyr localization closely associated with circumferentially aligned thick actin bundles (8- to 9-somite stages). This P-Tyr immunostaining decreased when the thick actin bundles developed into mature striated myofibrils at the 10- to 13-somite stages. These findings suggest that the phosphorylation of protein tyrosine residues is primarily concentrated at the modulating cell-cell and cell-matrix adhesion sites of developing myocytes and myofibrils.

Asunto(s)

Corazón/embriología , Miocardio/metabolismo , Miofibrillas/metabolismo , Fosfotirosina/metabolismo , Actinas/análisis , Animales , Adhesión Celular , Embrión de Pollo , Inmunohistoquímica , Uniones Intercelulares/metabolismo , Microscopía Confocal , Microscopía Fluorescente , Microscopía Inmunoelectrónica , Miocardio/química , Miofibrillas/química , Fosfotirosina/fisiología , Factores de Tiempo , Distribución Tisular

RESUMEN

The long-range goal of this research is to establish an in vitro system that will permit pertubation of mammalian heart development and in situ examination of the cellular and molecular events underlying cardiac morphogenesis. Rat embryos at 9.5-11.5 days of gestation were placed in culture bottles containing rat serum and Tyrode's solution. Embryos cultured for 24 and 48 h were compared to age-matched in vivo controls for morphological score, morphometric analysis of heart development, and confocal and electron microscopic analysis of myofiber pattern formation. Morphological scores indicated that embryos cultured for 24 h from day 9.5 to 10.5 had essentially normal development when compared to age-matched embryos allowed to develop in vivo. Development of embryos maintained for 48 h in culture was slightly delayed at 66-68% of age matched in vivo embryos. Analysis of hearts from embryos allowed to develop 9.5-11.5 days in vivo plus 24 and 48 h in culture showed that the ventricular thickness and height, as well as the truncal, atrial and ventricular diameters were equivalent to those of hearts from age-matched in vivo controls. Hearts from embryos allowed to develop from 11.5-12.5 days in vitro and cultured for 24 and 48 h had smaller left ventricular and atrial dimensions than controls. Cardiac myofibrillogenesis and myofibrillar pattern formation in embryos cultured from 9.5 days of in vivo development for 48 h were also normal. These studies indicate that the rat whole embryo culture system is a useful model to study several critical periods in mammalian heart development.

Asunto(s)

Desarrollo Embrionario y Fetal , Corazón/embriología , Animales , Técnicas de Cultivo , Embrión de Mamíferos/ultraestructura , Edad Gestacional , Microscopía Confocal , Microscopía Electrónica , Miocardio/ultraestructura , Miofibrillas/ultraestructura , Ratas , Ratas Sprague-Dawley

RESUMEN

The role of angiotensin II (Ang II) in the early embryonic development of the heart has not been examined. We have used RT-PCR to identify mRNA for angiotensinogen, angiotensin-converting enzyme, and the Ang II AT1 and AT2 receptors in embryonic day 10.25 Sprague-Dawley rats, and have used confocal microscopy to localize the AT1 receptor to the greater curvature of the developing ventricle in these animals at embryonic days (ED) 9.25 and 10.25. The antibodies used in immunolocalization studies did not distinguish between the AT1a and AT1b receptor subtypes. In whole embryo culture, Ang II added to the culture media resulted in increased ventricular growth and myocyte hypertrophy when treated embryos were compared to cultured littermate controls. Use of Losartan and PD123,319 to block the Ang II AT1 and AT2 receptors resulted in reduced ventricular development and cardiac dilation when compared to control and Ang II-treated embryos. Addition of Ang II and PD123,319 to the culture media also resulted in cardiac loop inversions which may be associated with disruption of normal myofibrillar development. These results clearly indicate an important role for Ang II in the early embryonic development of the heart.

Asunto(s)

Angiotensina II/farmacología , Antagonistas de Receptores de Angiotensina , Corazón Fetal/efectos de los fármacos , Actinas/análisis , Angiotensina II/fisiología , Animales , Cardiomegalia/inducido químicamente , Corazón Fetal/anomalías , Corazón Fetal/ultraestructura , Fibroblastos/patología , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/embriología , Hipertrofia , Imidazoles/farmacología , Imidazoles/toxicidad , Losartán/farmacología , Losartán/toxicidad , Morfogénesis/efectos de los fármacos , Miocardio/patología , Cadenas Pesadas de Miosina/análisis , Técnicas de Cultivo de Órganos , Piridinas/farmacología , Piridinas/toxicidad , Ratas , Ratas Sprague-Dawley , Receptor de Angiotensina Tipo 1 , Receptor de Angiotensina Tipo 2 , Receptores de Angiotensina/fisiología

RESUMEN

BACKGROUND: In chickens, cytodifferentiation, right side dominance in myofibril development, and variations in myofibrillar patterns in different areas and layers of the myocardial wall exist which have been implicated in the process of heart looping. Little comparable information is available for developing myofibrillar patterns in the early development of mammalian hearts. METHODS: We have used transmission electron microscopy (TEM), confocal scanning laser microscopy (CSLM), and 3-D reconstruction techniques also present in the looping hearts of embryonic day (ED) 9.5 to 11.5 rat hearts. RESULTS: Local and regional variations and right side dominance in myofibrillar patterns were shown during looping in 9.5 through 11.5 days of development in embryonic rat heart. At 9.5 days of development, myofibrils near the lumen of the myocardial wall were primarily in circumferential bands while near the pericardial surface they were primarily in longitudinal bands. In older embryos, regional variations in myofibrillar organization was found in areas associated with the cardiac cushions, trabeculae, and myocardial wall of the developing heart chambers. Based on sarcomeric structure, myofibrils in the ventricle and outflow tract were more advanced than those found in the atrial wall. CONCLUSIONS: The local and regional patterns of myofibrils in looping rat hearts are similar to those which have been found in developing chicken hearts. This study and others indicate cytodifferentiation and development of the contractile apparatus has a crucial role in the process of heart looping.

Asunto(s)

Corazón/embriología , Miofibrillas/ultraestructura , Animales , Diferenciación Celular , Femenino , Atrios Cardíacos/citología , Atrios Cardíacos/embriología , Ventrículos Cardíacos/citología , Ventrículos Cardíacos/embriología , Procesamiento de Imagen Asistido por Computador , Microscopía Confocal , Microscopía Electrónica de Rastreo , Embarazo , Ratas , Ratas Sprague-Dawley , Tronco Arterial/citología , Tronco Arterial/embriología

RESUMEN

Mechanical forces play an essential role in regulating the synthesis and assembly of contractile proteins into the sarcomeres of cardiac myocytes. To examine if physical forces might also regulate the turnover of contractile proteins at a posttranslational site of control, beating and nonbeating neonatal cardiac myocytes (NCM) were subjected to a 5% static stretch. The L-type calcium channel blocker nifedipine (12 microM) was used to inhibit contraction. Pulse-chase biosynthetic labeling experiments demonstrated that contractile arrest accelerated the loss of isotopic tracer from the total myofibrillar protein fraction, myosin heavy chain (MHC), and actin, but not desmin. Myofibrillar abnormalities developed in parallel with these metabolic changes. A 5% static load appeared to partially stabilize myofibrillar structure in nonbeating NCM and suppressed the loss of isotopic tracer from the total myofibrillar protein fraction, MHC, and actin in beating and nonbeating NCM. Contractile activity and/or a static stretch promoted the accumulation of MHC, actin, and desmin. Applying a static load to myocytes that lacked preexisting myofibrils did not promote the assembly of sarcomeres or alter protein turnover. These data indicate that the turnover of MHC and actin is correlated with the organizational state of the myofibrillar apparatus.

Asunto(s)

Proteínas Contráctiles/metabolismo , Miocardio/metabolismo , Miofibrillas/ultraestructura , Animales , Animales Recién Nacidos , Células Cultivadas , Microscopía Confocal , Microscopía Electrónica , Contracción Miocárdica/efectos de los fármacos , Miocardio/citología , Miocardio/ultraestructura , Estimulación Física , Ratas

RESUMEN

Chronic hypertension, known to affect the collagen profile of the heart, and exercise result in impaired or improved heart function, respectively. Collagen types I [alpha 1(I)2 and alpha 2(I)] and III [alpha 1(III)3] are the predominant interstitial collagens thought to influence cardiac function, and the ratio of type III to I (collagen III/I) is thought to be a significant factor in the altered relaxation observed in hypertrophy. The present study tested the hypothesis that the myocardial structure and function are different in chronically exercise-trained vs. hypertensive rat hearts. Male rats were either chronically exercised (XTr) or submitted to experimental hypertension by coarctation of the abdominal aorta (Hyp) for 10 wks. Heart rate, blood pressure, and maximal rate of fall of the left ventricular pressure (-dp/dt) were recorded during isoproterenol stimulation. Results showed that both Hyp and XTr had higher heart weight and left ventricular weight-to-body weight ratios (P < 0.05). Mean arterial pressure (MAP) was higher in Hyp and lower in XTr (P < 0.05), whereas (-dP/dt)/MAP was diminished in Hyp but enhanced in XTr. Left ventricular collagen was higher in Hyp than XTr, whereas collagen III/I was reduced in Hyp compared with XTr (P < 0.05). Scanning and transmission electron microscopy also supported an accumulation of left ventricular collagen in Hyp compared with XTr. A negative correlation was observed between collagen III/I and (-dP/dt)/ MAP (r = -0.91; P < 0.05). These results suggest an important relationship between adaptations in left ventricular collagen and the changes in diastolic function observed in both chronic hypertension and exercise cardiac stress.

Asunto(s)

Colágeno/metabolismo , Hipertensión/fisiopatología , Miocardio/metabolismo , Esfuerzo Físico , Función Ventricular Izquierda , Animales , Peso Corporal , Diástole , Hipertensión/patología , Masculino , Microscopía Electrónica , Microscopía Electrónica de Rastreo , Miocardio/patología , Tamaño de los Órganos , Ratas , Ratas Sprague-Dawley

RESUMEN

Fumonisins, produced by Fusarium moniliforme, have been recognized as an important group of chemicals which cause health risks in domestic animals and humans. Decontamination procedures for fumonisin B1 (FB1) were evaluated to determine chemical modification and reduction in toxic/carcinogenic potentials. Ammoniation, a procedure used for decontamination of aflatoxins, yielded a 79% reduction in FB1 levels in naturally contaminated corn. Authentic FB1 and FB1-contaminated corn were exposed to alternative treatments containing various combinations of Ca(OH)2, NaHCO3, and H2O2 simulating a modified nixtamalization procedure. Treatments also included NH4Cl alone or in combination with H2O2 or horseradish peroxidase. The brine shrimp assay (Artemia spp.) was used to monitor toxicity of reaction products and the Salmonella/microsomal mutagenicity assay, using tester strains TA-100 and TA-102, was used to evaluate mutagenicity. Treatments of FB1-contaminated corn simulating modified nixtamalization (Ca(OH)2 alone or with Na-HCO3 + H2O2) gave 100% reduction of FB1 and reduced brine shrimp toxicity by ca. 40%. The positive mutagenic potential (without S-9) for extracts of corn naturally contaminated with FB1 was eliminated following exposure to modified nixtamalization. Reaction products formed when pure FB1 was treated with Ca(OH)2 and H2O2/NaHCO3 were inhibitory to Bacillus cereus, B. subtilis, and B. megaterium. No inhibitory potential was evident for contaminated corn extracts following the chemical treatments.

Asunto(s)

Carcinógenos Ambientales , Contaminación de Alimentos , Fumonisinas , Micotoxinas/química , Micotoxinas/toxicidad , Zea mays/química , Animales , Artemia , Hidróxido de Calcio/farmacología , Carbonatos/farmacología , Descontaminación/métodos , Caballos , Peróxido de Hidrógeno/farmacología , Pruebas de Mutagenicidad , Micotoxinas/análisis , Salmonella , Pruebas de Toxicidad

RESUMEN

The excitation-contraction coupling cycle (ECC) consists of a complex cascade of electrochemical and mechanical events; however, the relative contributions of these different processes in the regulation of cardiac myofibrillar structure are not well understood. There is extensive evidence to suggest that the mechanical aspects of the ECC play a crucial role in controlling the availability of contractile proteins for myofibrillar assembly. To examine if these physical forces might also serve to stabilize the structure of preexisting myofibrils, beating and nonbeating cultures of neonatal cardiac myocytes (NCM) were subjected to a 5% static stretch. Contractile arrest was achieved by treating NCM with 12 microM nifedipine, which resulted in immediate and sustained contractile arrest and initiated the evolution of marked myofibrillar abnormalities within 24 hours. As judged by scanning confocal and transmission electron microscopic examination, an external load appears to partially stabilize myofibrillar structure in nonbeating NCM. These results suggest that the maintenance of myofibrillar structure may be highly dependent upon the mechanical aspects of ECC.

Asunto(s)

Corazón/fisiología , Miocardio/citología , Miofibrillas/ultraestructura , Animales , Animales Recién Nacidos , Células Cultivadas , Técnicas de Cultivo/instrumentación , Técnicas de Cultivo/métodos , Corazón/efectos de los fármacos , Microscopía Confocal , Microscopía Electrónica , Miocardio/ultraestructura , Miofibrillas/efectos de los fármacos , Nifedipino/farmacología , Ratas , Estrés Mecánico