RESUMEN
It is widely believed that perinatal cardiomyocyte terminal differentiation blocks cytokinesis, thereby causing binucleation and limiting regenerative repair after injury. This suggests that heart growth should occur entirely by cardiomyocyte hypertrophy during preadolescence when, in mice, cardiac mass increases many-fold over a few weeks. Here, we show that a thyroid hormone surge activates the IGF-1/IGF-1-R/Akt pathway on postnatal day 15 and initiates a brief but intense proliferative burst of predominantly binuclear cardiomyocytes. This proliferation increases cardiomyocyte numbers by ~40%, causing a major disparity between heart and cardiomyocyte growth. Also, the response to cardiac injury at postnatal day 15 is intermediate between that observed at postnatal days 2 and 21, further suggesting persistence of cardiomyocyte proliferative capacity beyond the perinatal period. If replicated in humans, this may allow novel regenerative therapies for heart diseases.
Asunto(s)
Diferenciación Celular , Proliferación Celular , Corazón/crecimiento & desarrollo , Miocitos Cardíacos/citología , Animales , Separación Celular , Masculino , Ratones , Ratones Endogámicos C57BL , Miocitos Cardíacos/fisiología , Triyodotironina/metabolismoRESUMEN
The existence of naturally occurring ribosome heterogeneity is now a well-acknowledged phenomenon. However, whether this heterogeneity leads to functionally diverse 'specialized ribosomes' is still a controversial topic. Here, we explore the biological function of RPL3L (uL3L), a ribosomal protein (RP) paralogue of RPL3 (uL3) that is exclusively expressed in skeletal muscle and heart tissues, by generating a viable homozygous Rpl3l knockout mouse strain. We identify a rescue mechanism in which, upon RPL3L depletion, RPL3 becomes up-regulated, yielding RPL3-containing ribosomes instead of RPL3L-containing ribosomes that are typically found in cardiomyocytes. Using both ribosome profiling (Ribo-seq) and a novel orthogonal approach consisting of ribosome pulldown coupled to nanopore sequencing (Nano-TRAP), we find that RPL3L modulates neither translational efficiency nor ribosome affinity towards a specific subset of transcripts. In contrast, we show that depletion of RPL3L leads to increased ribosome-mitochondria interactions in cardiomyocytes, which is accompanied by a significant increase in ATP levels, potentially as a result of fine-tuning of mitochondrial activity. Our results demonstrate that the existence of tissue-specific RP paralogues does not necessarily lead to enhanced translation of specific transcripts or modulation of translational output. Instead, we reveal a complex cellular scenario in which RPL3L modulates the expression of RPL3, which in turn affects ribosomal subcellular localization and, ultimately, mitochondrial activity.
Ribosomes are macromolecular machines responsible for protein synthesis in all living beings. Recent studies have shown that ribosomes can be heterogeneous in their structure, possibly leading to a specialized function. Here, we focus on RPL3L, a ribosomal protein expressed exclusively in striated muscles. We find that the deletion of the Rpl3l gene in a mouse model triggers a compensation mechanism, in which the missing RPL3L protein is replaced by its paralogue, RPL3. Furthermore, we find that RPL3-containing ribosomes establish closer interactions with mitochondria, cellular organelles responsible for energy production, leading to higher energy production when compared with RPL3L-containing ribosomes. Finally, we show that the RPL3RPL3L compensation mechanism is also triggered in heart disease conditions, such as hypertrophy and myocardial infarction.
Asunto(s)
Corazón , Mitocondrias , Proteínas Ribosómicas , Ribosomas , Animales , Ratones , Mitocondrias/metabolismo , Músculo Esquelético/metabolismo , Biosíntesis de Proteínas , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/metabolismo , Ribosomas/genética , Ribosomas/metabolismoRESUMEN
Congenital heart disease (CHD) is the most common type of birth defect. In recent years, research has focussed on identifying the genetic causes of CHD. However, only a minority of CHD cases can be attributed to single gene mutations. In addition, studies have identified different environmental stressors that promote CHD, but the additive effect of genetic susceptibility and environmental factors is poorly understood. In this context, we have investigated the effects of short-term gestational hypoxia on mouse embryos genetically predisposed to heart defects. Exposure of mouse embryos heterozygous for Tbx1 or Fgfr1/Fgfr2 to hypoxia in utero increased the incidence and severity of heart defects while Nkx2-5+/- embryos died within 2 days of hypoxic exposure. We identified the molecular consequences of the interaction between Nkx2-5 and short-term gestational hypoxia, which suggest that reduced Nkx2-5 expression and a prolonged hypoxia-inducible factor 1α response together precipitate embryo death. Our study provides insight into the causes of embryo loss and variable penetrance of monogenic CHD, and raises the possibility that cases of foetal death and CHD in humans could be caused by similar gene-environment interactions.
Asunto(s)
Interacción Gen-Ambiente , Cardiopatías Congénitas/genética , Corazón/embriología , Proteína Homeótica Nkx-2.5/genética , Proteínas de Homeodominio/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Animales , Apoptosis , Proliferación Celular , Embrión de Mamíferos/metabolismo , Femenino , Predisposición Genética a la Enfermedad , Corazón/diagnóstico por imagen , Heterocigoto , Proteína Homeótica Nkx-2.5/fisiología , Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Oxígeno/metabolismo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Proteínas de Dominio T Box/genética , Factores de TiempoRESUMEN
The 'fight or flight' response to physiological stress involves sympathetic nervous system activation, catecholamine release and adrenergic receptor stimulation. In the heart, this induces positive inotropy, previously attributed to the ß1-adrenergic receptor subtype. However, the role of the α1A-adrenergic receptor, which has been suggested to be protective in cardiac pathology, has not been investigated in the setting of physiological stress. To explore this, we developed a tamoxifen-inducible, cardiomyocyte-specific α1A-adrenergic receptor knock-down mouse model, challenged mice to four weeks of endurance swim training and assessed cardiac outcomes. With 4-OH tamoxifen treatment, expression of the α1A-adrenergic receptor was knocked down by 80-89%, without any compensatory changes in the expression of other adrenergic receptors, or changes to baseline cardiac structure and function. Swim training caused eccentric hypertrophy, regardless of genotype, demonstrated by an increase in heart weight/tibia length ratio (30% and 22% in vehicle- and tamoxifen-treated animals, respectively) and an increase in left ventricular end diastolic volume (30% and 24% in vehicle- and tamoxifen-treated animals, respectively) without any change in the wall thickness/chamber radius ratio. Consistent with physiological hypertrophy, there was no increase in fetal gene program (Myh7, Nppa, Nppb or Acta1) expression. In response to exercise-induced volume overload, stroke volume (39% and 30% in vehicle- and tamoxifen-treated animals, respectively), cardiac output/tibia length ratio (41% in vehicle-treated animals) and stroke work (61% and 33% in vehicle- and tamoxifen-treated animals, respectively) increased, regardless of genotype. These findings demonstrate that cardiomyocyte α1A-adrenergic receptors are not necessary for cardiac adaptation to endurance exercise stress and their acute ablation is not deleterious.
Asunto(s)
Adaptación Fisiológica , Modelos Biológicos , Miocitos Cardíacos/metabolismo , Condicionamiento Físico Animal , Receptores Adrenérgicos alfa 1/metabolismo , Estrés Fisiológico , Animales , Biomarcadores , Gasto Cardíaco , Cardiomegalia/diagnóstico , Cardiomegalia/etiología , Cardiomegalia/metabolismo , Ecocardiografía de Estrés , Genotipo , Hemodinámica , Ratones , Ratones Noqueados , Ratones Transgénicos , Modelos Animales , Contracción Miocárdica , Receptores Adrenérgicos alfa 1/genéticaRESUMEN
Echocardiography is an invaluable tool for characterizing cardiac structure and function in vivo. Technological advances in high-frequency ultrasound over the past 3 decades have increased spatial and temporal resolution, and facilitated many important clinical and basic science discoveries. Successful reverse translation of established echocardiographic techniques, including M-mode, B-mode, color Doppler, pulsed-wave Doppler, tissue Doppler and, most recently, myocardial deformation imaging, from clinical cardiology into the basic science laboratory has enabled researchers to achieve a deeper understanding of myocardial phenotypes in health and disease. With high-frequency echocardiography, detailed evaluation of ventricular systolic function in a range of small animal models is now possible. Furthermore, improvements in frame rate and the advent of diastolic strain rate imaging, when coupled with the use of select pulsed-wave Doppler parameters, such as isovolumic relaxation time and E wave deceleration, have enabled nuanced interpretation of ventricular diastolic function. Comparing pulsed-wave Doppler indices of atrioventricular inflow during early and late diastole with parameters that describe the simultaneous myocardial deformation (e.g., tissue Doppler é and á, global longitudinal strain rate and global longitudinal velocity) may yield additional insights related to myocardial compliance. This review will provide a historical perspective of the development of high-frequency echocardiography and consider how ongoing innovation will help future-proof this important imaging modality for 21st century translational research.
Asunto(s)
Ecocardiografía/tendencias , Animales , Diagnóstico por Imagen/métodos , Ecocardiografía/instrumentación , Ecocardiografía/métodos , Ecocardiografía Doppler de Pulso , Corazón/diagnóstico por imagen , Humanos , Ratones , Investigación/instrumentación , Investigación/tendencias , Pez CebraRESUMEN
RATIONALE: Mutations in the LMNA gene, which encodes the nuclear lamina proteins lamin A and lamin C, are the most common cause of familial dilated cardiomyopathy (DCM). Mechanical stress-induced apoptosis has been proposed as the mechanism underpinning DCM in lamin A/C-deficient hearts, but supporting in vivo evidence has been lacking. OBJECTIVE: Our aim was to study interventions to modify mechanical stress in heterozygous Lmna knockout (Lmna(+/-)) mice. METHODS AND RESULTS: Cardiac structure and function were evaluated before and after exercise training, thoracic aortic constriction, and carvedilol treatment. Lmna(+/-) mice develop adult-onset DCM with relatively more severe disease in males. Lmna(+/-) cardiomyocytes show altered nuclear morphology and perinuclear desmin organization, with enhanced responses to hypo-osmotic stress indicative of cytoskeletal instability. Despite these structural defects that provide a template for mechanical stress-induced damage, young Lmna(+/-) mice subjected to 6 weeks of moderate or strenuous exercise training did not show induction of apoptosis or accelerated DCM. In contrast, regular moderate exercise attenuated DCM development in male Lmna(+/-) mice. Sustained pressure overload generated by thoracic aortic constriction depressed ventricular contraction in young wild-type and Lmna(+/-) mice with no sex or genotype differences in the time-course or severity of response. Treatment of male Lmna(+/-) mice from 12 to 40 weeks with the beta-blocker, carvedilol, prevented the dilatation and contractile dysfunction that was observed in placebo-treated mice. CONCLUSIONS: These data suggest that factors other than mechanical stress-induced apoptosis contribute to DCM and provide the first demonstration that regular moderate exercise and carvedilol can modify disease progression in lamin A/C-deficient hearts.
Asunto(s)
Antagonistas Adrenérgicos beta/uso terapéutico , Carbazoles/uso terapéutico , Cardiomiopatía Dilatada/genética , Corazón/fisiopatología , Lamina Tipo A/deficiencia , Miocardio/patología , Propanolaminas/uso terapéutico , Estrés Mecánico , Animales , Aorta Torácica , Apoptosis , Cardiomiopatía Dilatada/tratamiento farmacológico , Cardiomiopatía Dilatada/patología , Cardiomiopatía Dilatada/fisiopatología , Carvedilol , Constricción , Desmina/análisis , Femenino , Genotipo , Lamina Tipo A/genética , Masculino , Ratones , Ratones Noqueados , Presión Osmótica , Condicionamiento Físico Animal , Disfunción Ventricular Izquierda/patología , Disfunción Ventricular Izquierda/fisiopatologíaRESUMEN
Lamin A/C mutations are the most common cause of familial dilated cardiomyopathy (DCM) but the pathogenetic mechanisms are incompletely understood. Nesprins are spectrin repeat-containing proteins that interact with lamin A/C and are components of the linker-of-nucleoskeleton-and-cytoskeleton (LINC) complex that connects the nuclear envelope to the actin cytoskeleton. Our aim was to determine whether changes in nesprin-1 and actin might contribute to DCM in homozygous Lmna knockout (Lmna(-/-)) mice. Here we find that Lmna(-/-) cardiomyocytes have altered nuclear envelope morphology, disorganization of nesprin-1 and heterogeneity in the distribution of nuclear and cytoskeletal actin. Functional interactions of nesprin-1 with nuclear G-actin and with the cytoskeletal γ-actin, α-cardiac actin and α-smooth muscle actin (α-SMA) isoforms were shown by immunoprecipitation and Western blotting. At 4-6 weeks of age, Lmna(-/-) mice had normal levels of γ-actin and α-cardiac actin, but α-SMA expression was increased by 50%. In contrast to the predominant vascular distribution of α-SMA in WT ventricular sections, α-SMA had a diffuse staining pattern in Lmna(-/-) sections. Osmotic swelling studies showed enhanced radial swelling in Lmna(-/-) cardiomyocytes indicative of cytoskeletal instability. The distensibility of Lmna(-/-) cardiomyocytes with osmotic stress was reduced by addition of α-SMA-specific fusion peptide. Our findings support a model in which uncoupling of the nucleus and cytoskeleton associated with disruption of the LINC complex promotes mechanical instability and defective force transmission in cardiomyocytes. Changes in the distribution and expression patterns of nuclear and cytoskeletal actin suggest that diverse transcriptional and structural defects may also contribute to DCM in Lmna(-/-) mice.
Asunto(s)
Actinas/metabolismo , Lamina Tipo A/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares/metabolismo , Actinas/genética , Animales , Cardiomiopatía Dilatada/genética , Cardiomiopatía Dilatada/metabolismo , Cardiomiopatía Dilatada/patología , Núcleo Celular/genética , Núcleo Celular/metabolismo , Núcleo Celular/patología , Proteínas del Citoesqueleto , Citoesqueleto/genética , Citoesqueleto/metabolismo , Citoesqueleto/patología , Homocigoto , Lamina Tipo A/deficiencia , Lamina Tipo A/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Membrana Nuclear/genética , Membrana Nuclear/metabolismo , Membrana Nuclear/patologíaRESUMEN
RATIONALE: Gq-coupled receptors are thought to play a critical role in the induction of left ventricular hypertrophy (LVH) secondary to pressure overload, although mechano-sensitive channel activation by a variety of mechanisms has also been proposed, and the relative importance of calcineurin- and calmodulin kinase II (CaMKII)-dependent hypertrophic pathways remains controversial. OBJECTIVE: To determine the mechanisms regulating the induction of LVH in response to mechanical pressure overload. METHODS AND RESULTS: Transgenic mice with cardiac-targeted inhibition of Gq-coupled receptors (GqI mice) and their non-transgenic littermates (NTL) were subjected to neurohumoral stimulation (continuous, subcutaneous angiotensin II (AngII) infusion for 14 days) or mechanical pressure overload (transverse aortic arch constriction (TAC) for 21 days) to induce LVH. Candidate signaling pathway activation was examined. As expected, LVH observed in NTL mice with AngII infusion was attenuated in heterozygous (GqI+/-) mice and absent in homozygous (GqI-/-) mice. In contrast, LVH due to TAC was unaltered by either heterozygous or homozygous Gq inhibition. Gene expression of atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP) and α-skeletal actin (α-SA) was increased 48 h after AngII infusion or TAC in NTL mice; in GqI mice, the increases in ANP, BNP and α-SA in response to AngII were completely absent, as expected, but all three increased after TAC. Increased nuclear translocation of nuclear factor of activated T-cells c4 (NFATc4), indicating calcineurin pathway activation, occurred in NTL mice with AngII infusion but not TAC, and was prevented in GqI mice infused with AngII. Nuclear and cytoplasmic CaMKIIδ levels increased in both NTL and GqI mice after TAC but not AngII infusion, with increased cytoplasmic phospho- and total histone deacetylase 4 (HDAC4) and increased nuclear myocyte enhancer factor 2 (MEF2) levels. CONCLUSION: Cardiac Gq receptors and calcineurin activation are required for neurohumorally mediated LVH but not for LVH induced by mechanical pressure overload (TAC). Rather, TAC-induced LVH is associated with activation of the CaMKII-HDAC4-MEF2 pathway.
RESUMEN
Pathological left ventricular hypertrophy (LVH) occurs in response to pressure overload and remains the single most important clinical predictor of cardiac mortality. The molecular pathways in the induction of pressure overload LVH are potential targets for therapeutic intervention. Current treatments aim to remove the pressure overload stimulus for LVH, but do not completely reverse adverse cardiac remodelling. Although numerous molecular signalling steps in the induction of LVH have been identified, the initial step by which mechanical stretch associated with cardiac pressure overload is converted into a chemical signal that initiates hypertrophic signalling remains unresolved. In this study, we show that selective deletion of transient receptor potential melastatin 4 (TRPM4) channels in mouse cardiomyocytes results in an approximately 50% reduction in the LVH induced by transverse aortic constriction. Our results suggest that TRPM4 channel is an important component of the mechanosensory signalling pathway that induces LVH in response to pressure overload and represents a potential novel therapeutic target for the prevention of pathological LVH.
Asunto(s)
Eliminación de Gen , Hipertrofia Ventricular Izquierda/genética , Miocitos Cardíacos/metabolismo , Canales Catiónicos TRPM/genética , Animales , Hipertrofia Ventricular Izquierda/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Transducción de Señal , Canales Catiónicos TRPM/efectos adversos , Canales Catiónicos TRPM/metabolismoRESUMEN
Animal models of pressure overload are valuable for understanding hypertensive heart disease. We characterised a surgical model of pressure overload-induced hypertrophy in C57BL/6J mice produced by suprarenal aortic constriction (SAC). Compared to sham controls, at one week post-SAC systolic blood pressure was significantly elevated and left ventricular (LV) hypertrophy was evident by a 50% increase in the LV weight-to-tibia length ratio due to cardiomyocyte hypertrophy. As a result, LV end-diastolic wall thickness-to-chamber radius (h/R) ratio increased, consistent with the development of concentric hypertrophy. LV wall thickening was not sufficient to normalise LV wall stress, which also increased, resulting in LV systolic dysfunction with reductions in ejection fraction and fractional shortening, but no evidence of heart failure. Pathological LV remodelling was evident by the re-expression of fetal genes and coronary artery perivascular fibrosis, with ischaemia indicated by enhanced cardiomyocyte Hif1a expression. The expression of stem cell factor receptor, c-Kit, was low basally in cardiomyocytes and did not change following the development of robust hypertrophy, suggesting there is no role for cardiomyocyte c-Kit signalling in pathological LV remodelling following pressure overload.
Asunto(s)
Hipertrofia Ventricular Izquierda/patología , Miocitos Cardíacos/patología , Proteínas Proto-Oncogénicas c-kit/metabolismo , Animales , Aorta/fisiopatología , Constricción Patológica , Regulación de la Expresión Génica , Hipertensión/etiología , Hipertensión/fisiopatología , Hipertrofia Ventricular Izquierda/genética , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Presión , Proteínas Proto-Oncogénicas c-kit/genética , Circulación Renal , Renina/genética , Disfunción Ventricular Izquierda/genética , Disfunción Ventricular Izquierda/fisiopatología , Remodelación Ventricular/genética , Remodelación Ventricular/fisiologíaRESUMEN
Therapies that target scar formation after myocardial infarction (MI) could prevent ensuing heart failure or death from ventricular arrhythmias. We have previously shown that recombinant human platelet-derived growth factor-AB (rhPDGF-AB) improves cardiac function in a rodent model of MI. To progress clinical translation, we evaluated rhPDGF-AB treatment in a clinically relevant porcine model of myocardial ischemia-reperfusion. Thirty-six pigs were randomized to sham procedure or balloon occlusion of the proximal left anterior descending coronary artery with 7-day intravenous infusion of rhPDGF-AB or vehicle. One month after MI, rhPDGF-AB improved survival by 40% compared with vehicle, and cardiac magnetic resonance imaging showed left ventricular (LV) ejection fraction improved by 11.5%, driven by reduced LV end-systolic volumes. Pressure volume loop analyses revealed improved myocardial contractility and energetics after rhPDGF-AB treatment with minimal effect on ventricular compliance. rhPDGF-AB enhanced angiogenesis and increased scar anisotropy (high fiber alignment) without affecting overall scar size or stiffness. rhPDGF-AB reduced inducible ventricular tachycardia by decreasing heterogeneity of the ventricular scar that provides a substrate for reentrant circuits. In summary, we demonstrated that rhPDGF-AB promotes post-MI cardiac wound repair by altering the mechanics of the infarct scar, resulting in robust cardiac functional improvement, decreased ventricular arrhythmias, and improved survival. Our findings suggest a strong translational potential for rhPDGF-AB as an adjunct to current MI treatment and possibly to modulate scar in other organs.
Asunto(s)
Cicatriz/patología , Infarto del Miocardio/patología , Factor de Crecimiento Derivado de Plaquetas/farmacología , Animales , Arritmias Cardíacas/complicaciones , Arritmias Cardíacas/patología , Arritmias Cardíacas/fisiopatología , Arteriolas/efectos de los fármacos , Arteriolas/patología , Arteriolas/fisiopatología , Cicatriz/complicaciones , Cicatriz/tratamiento farmacológico , Cicatriz/fisiopatología , Colágeno/metabolismo , Fibrosis , Pruebas de Función Cardíaca/efectos de los fármacos , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/patología , Ventrículos Cardíacos/fisiopatología , Humanos , Contracción Miocárdica/efectos de los fármacos , Infarto del Miocardio/complicaciones , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/fisiopatología , Neovascularización Fisiológica/efectos de los fármacos , Factor de Crecimiento Derivado de Plaquetas/uso terapéutico , Proteínas Recombinantes/farmacología , Análisis de Supervivencia , Porcinos , Cicatrización de Heridas/efectos de los fármacosRESUMEN
We have previously demonstrated that adult transgenic C57BL/6J mice with CM-restricted overexpression of the dominant negative W v mutant protein (dn-c-kit-Tg) respond to pressure overload with robust cardiomyocyte (CM) cell cycle entry. Here, we tested if outcomes after myocardial infarction (MI) due to coronary artery ligation are improved in this transgenic model. Compared to non-transgenic littermates (NTLs), adult male dn-c-kit-Tg mice displayed CM hypertrophy and concentric left ventricular (LV) hypertrophy in the absence of an increase in workload. Stroke volume and cardiac output were preserved and LV wall stress was markedly lower than that in NTLs, leading to a more energy-efficient heart. In response to MI, infarct size in adult (16-week old) dn-c-kit-Tg hearts was similar to that of NTL after 24 h but was half that in NTL hearts 12 weeks post-MI. Cumulative CM cell cycle entry was only modestly increased in dn-c-kit-Tg hearts. However, dn-c-kit-Tg mice were more resistant to infarct expansion, adverse LV remodelling and contractile dysfunction, and suffered no early death from LV rupture, relative to NTL mice. Thus, pre-existing cardiac hypertrophy lowers wall stress in dn-c-kit-Tg hearts, limits infarct expansion and prevents death from myocardial rupture.
Asunto(s)
Cardiomegalia/patología , Infarto del Miocardio/patología , Animales , Cardiomegalia/genética , Modelos Animales de Enfermedad , Hipertrofia Ventricular Izquierda/genética , Hipertrofia Ventricular Izquierda/patología , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutación , Infarto del Miocardio/genética , Miocardio/patología , Proteínas Proto-Oncogénicas c-kit/genética , Disfunción Ventricular Izquierda/genética , Disfunción Ventricular Izquierda/patologíaRESUMEN
Laminopathies are a group of disorders caused by mutations in the LMNA gene that encodes the nuclear lamina proteins, lamin A and lamin C; their pathophysiological basis is unknown. We report that lamin A/C-deficient (Lmna(-/-)) mice develop rapidly progressive dilated cardiomyopathy (DCM) characterized by left ventricular (LV) dilation and reduced systolic contraction. Isolated Lmna(-/-) myocytes show reduced shortening with normal baseline and peak amplitude of Ca(2+) transients. Lmna(-/-) LV myocyte nuclei have marked alterations of shape and size with central displacement and fragmentation of heterochromatin; these changes are present but less severe in left atrial nuclei. Electron microscopy of Lmna(-/-) cardiomyocytes shows disorganization and detachment of desmin filaments from the nuclear surface with progressive disruption of the cytoskeletal desmin network. Alterations in nuclear architecture are associated with defective nuclear function evidenced by decreased SREBP1 import, reduced PPARgamma expression, and a lack of hypertrophic gene activation. These findings suggest a model in which the primary pathophysiological mechanism in Lmna(-/-) mice is defective force transmission resulting from disruption of lamin interactions with the muscle-specific desmin network and loss of cytoskeletal tension. Despite severe DCM, defects in nuclear function prevent Lmna(-/-) cardiomyocytes from developing compensatory hypertrophy and accelerate disease progression.
Asunto(s)
Cardiomiopatía Dilatada/genética , Lamina Tipo A/deficiencia , Animales , Apoptosis/genética , Apoptosis/fisiología , Cardiomiopatía Dilatada/patología , Cardiomiopatía Dilatada/fisiopatología , Núcleo Celular/patología , Ecocardiografía , Electrocardiografía , Lamina Tipo A/genética , Ratones , Miocardio/patología , Miocitos Cardíacos/patología , Factores de TiempoRESUMEN
The zebrafish (Danio rerio) is an increasingly popular model organism in cardiovascular research. Major insights into cardiac developmental processes have been gained by studies of embryonic zebrafish. However, the utility of zebrafish for modeling adult-onset heart disease has been limited by a lack of robust methods for in vivo evaluation of cardiac function. We established a physiological protocol for underwater zebrafish echocardiography using high frequency ultrasound, and evaluated its reliability in detecting altered cardiac function in two disease models. Serial assessment of cardiac function was performed in wild-type zebrafish aged 3 to 12â months and the effects of anesthetic agents, age, sex and background strain were evaluated. There was a varying extent of bradycardia and ventricular contractile impairment with different anesthetic drugs and doses, with tricaine 0.75â mmolâ l-1 having a relatively more favorable profile. When compared with males, female fish were larger and had more measurement variability. Although age-related increments in ventricular chamber size were greater in females than males, there were no sex differences when data were normalized to body size. Systolic ventricular function was similar in both sexes at all time points, but differences in diastolic function were evident from 6â months onwards. Wild-type fish of both sexes showed a reliance on atrial contraction for ventricular diastolic filling. Echocardiographic evaluation of adult zebrafish with diphtheria toxin-induced myocarditis or anemia-induced volume overload accurately identified ventricular dilation and altered contraction, with suites of B-mode, ventricular strain, pulsed-wave Doppler and tissue Doppler indices showing concordant changes indicative of myocardial hypocontractility or hypercontractility, respectively. Repeatability, intra-observer and inter-observer correlations for echocardiographic measurements were high. We demonstrate that high frequency echocardiography allows reliable in vivo cardiac assessment in adult zebrafish and make recommendations for optimizing data acquisition and analysis. This enabling technology reveals new insights into zebrafish cardiac physiology and provides an imaging platform for zebrafish-based translational research.
Asunto(s)
Envejecimiento/fisiología , Ecocardiografía/normas , Cardiopatías/diagnóstico por imagen , Cardiopatías/fisiopatología , Pruebas de Función Cardíaca/normas , Pez Cebra/fisiología , Puntos Anatómicos de Referencia , Anemia/patología , Anestesia , Animales , Tamaño Corporal , Toxina Diftérica , Modelos Animales de Enfermedad , Estudios de Factibilidad , Femenino , Ventrículos Cardíacos/diagnóstico por imagen , Ventrículos Cardíacos/patología , Ventrículos Cardíacos/fisiopatología , Masculino , Contracción Miocárdica , Miocarditis/diagnóstico por imagen , Miocarditis/patología , Miocarditis/fisiopatología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Tamaño de los Órganos , Estándares de Referencia , Reproducibilidad de los Resultados , Función Ventricular IzquierdaRESUMEN
Mutations in the giant sarcomeric protein titin (TTN) are a major cause for inherited forms of dilated cardiomyopathy (DCM). We have previously developed a mouse model that imitates a TTN truncation mutation we found in a large pedigree with DCM. While heterozygous Ttn knock-in mice do not display signs of heart failure under sedentary conditions, they recapitulate the human phenotype when exposed to the pharmacological stressor angiotensin II or isoproterenol. In this study we investigated the effects of pressure overload by transverse aortic constriction (TAC) in heterozygous (Het) Ttn knock-in mice. Two weeks after TAC, Het mice developed marked impairment of left ventricular ejection fraction (p < 0.05), while wild-type (WT) TAC mice did not. Het mice also trended toward increased ventricular end diastolic pressure and volume compared to WT littermates. We found an increase in histologically diffuse cardiac fibrosis in Het compared to WT in TAC mice. This study shows that a pattern of DCM can be induced by TAC-mediated pressure overload in a TTN-truncated mouse model. This model enlarges our arsenal of cardiac disease models, adding a valuable tool to understand cardiac pathophysiological remodeling processes and to develop therapeutic approaches to combat heart failure.
Asunto(s)
Estenosis de la Válvula Aórtica/fisiopatología , Cardiomiopatía Dilatada/fisiopatología , Modelos Animales de Enfermedad , Hipertrofia Ventricular Izquierda/fisiopatología , Proteínas Quinasas/genética , Animales , Estenosis de la Válvula Aórtica/complicaciones , Estenosis de la Válvula Aórtica/diagnóstico por imagen , Cardiomiopatía Dilatada/diagnóstico por imagen , Cardiomiopatía Dilatada/etiología , Técnicas de Sustitución del Gen/métodos , Hipertrofia Ventricular Izquierda/diagnóstico por imagen , Hipertrofia Ventricular Izquierda/etiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , UltrasonografíaRESUMEN
BACKGROUND: The aim of this study was to determine the efficacy of cariporide (a sodium-hydrogen exchanger inhibitor), BMS180448 (a pharmacologic ischemic preconditioning agent), and the combination thereof, as adjuvant therapies for extended cardiac allograft preservation. METHODS: A porcine model of donor brain death and orthotopic heart transplantation was used. All hearts were arrested and stored for 14 hr in an extracellular preservation solution. Control hearts (CON; n=3) did not receive any additional treatment. Treated hearts received BMS180448 alone (BMS; n=3), cariporide alone (CAR; n=6), or both BMS180448 and cariporide (B+C; n=6). Donors of BMS180448-treated hearts received 2 mg/kg, 15 min before explantation. Donors and recipients of cariporide-treated hearts received 2 mg/kg, 15 min before explantation and reperfusion, respectively. RESULTS: The CON and BMS arms of the study were terminated after three transplantations because initial results in these groups were poor. Significantly, none of the control hearts could be weaned successfully from bypass, whereas all of the treated hearts were weaned successfully (CAR vs. CON and B+C vs. CON: P=0.012). The rate of troponin I release during the first 3 hr after reperfusion was significantly lower in CAR (P=0.0180) and B+C (P=0.0154) recipients than in CON recipients. Mean plasma troponin I levels (microg/mL) 3 hr after reperfusion were as follows: CON 633+/-177, BMS 576+/-110, CAR 346+/-93, and B+C 296+/-97. CONCLUSION: In this porcine model of extended cardiac allograft preservation, cariporide was more effective than BMS180448 as an adjuvant to our usual preservation solution. There was no additional benefit from the combination of the two therapies.
Asunto(s)
Antiarrítmicos/farmacología , Benzopiranos/farmacología , Cardiotónicos/farmacología , Guanidinas/farmacología , Trasplante de Corazón , Precondicionamiento Isquémico/métodos , Intercambiadores de Sodio-Hidrógeno/antagonistas & inhibidores , Sulfonas/farmacología , Animales , Puente Cardiopulmonar , Quimioterapia Combinada , Porcinos , Trasplante Homólogo , Troponina I/sangreRESUMEN
BACKGROUND: Acute graft dysfunction caused by ischemia-reperfusion injury is recognized as a major source of morbidity and mortality following adult heart transplantation. The aim of this study was to determine whether treating the donor and recipient with cariporide, an inhibitor of the sodium-hydrogen exchanger, could reduce ischemia-reperfusion injury. METHODS: A porcine model of donor brain death, hypothermic ischemic preservation, and orthotopic cardiac transplantation was used. Allografts in both the control group (CON, n=6) and treatment group (CAR, n=6) were arrested and stored for 4 hours in the extracellular crystalloid cardioplegia currently used in the clinical transplantation program at our institution. In addition, both the donor and recipient animals in the CAR group received a single intravenous dose of cariporide (2 mg/kg) 15 minutes before harvesting and reperfusion, respectively. RESULTS: The initial rate of troponin I release was significantly lower in recipients of CAR hearts than in recipients of CON hearts (P =0.020). All hearts were weaned successfully from bypass. More CAR hearts were weaned successfully at the first attempt, at 1 hour post-reperfusion, than CON hearts (6 of 6 vs 3 of 6), but this did not achieve statistical significance. Left ventricular contractility (preload recruitable stroke-work relationship) and left ventricular compliance (end-diastolic pressure-volume relationship) were significantly better preserved in CAR hearts than CON hearts (both P <0.0001). CONCLUSIONS: Myocardial injury was reduced, and contractile function was better preserved in allografts that received cariporide, compared with allografts that received conventional preservation alone.
Asunto(s)
Criopreservación , Guanidinas/farmacología , Trasplante de Corazón , Corazón/efectos de los fármacos , Sulfonas/farmacología , Animales , Presión Sanguínea , Volumen Sanguíneo , Grupos Control , Contracción Miocárdica , Isquemia Miocárdica/fisiopatología , Daño por Reperfusión Miocárdica/prevención & control , Intercambiadores de Sodio-Hidrógeno/antagonistas & inhibidores , Volumen Sistólico , Porcinos , Trasplante Homólogo , Troponina I/sangre , Función Ventricular IzquierdaRESUMEN
BACKGROUND: Acute brain death from increased intracranial pressure results in a transient increase in myocardial adenosine and lactate, which indicates that oxygen demand exceeds oxygen delivery during the sympathetic "storm". The aim of this study was to determine the functional significance of this period of ischemia. METHODS: Brain death was inflicted on 40 Westran pigs (36.5-68.0 kg) by inflating a 21-ml subdural balloon over 3 minutes. In 38 animals, micromanometry and sonomicrometry were used to obtain left ventricular pressure-volume loops to determine the preload recruitable stroke work (PRSW) relationship. Data files were recorded before and at 15-minute intervals after beginning balloon inflation. Plasma troponin I was measured before and 60 minutes after beginning balloon inflation in the 38 instrumented and 2 non-instrumented animals. RESULTS: All animals experienced the classical sympathetic storm. The slope of the PRSW relationship decreased, and the volume-axis intercept shifted to the right 15 minutes after beginning balloon inflation (p < 0.0001). Progressive incremental recovery (leftward shift) occurred between subsequent time points (p < or = 0.0018). In the instrumented animals, the mean plasma troponin I level increased from 1.4 +/- 1.6 microg/liter to 2.8 +/- 2.3 microg/liter (p < 0.001). However, troponin I was not detected before or after induction of brain death in the plasma of either non-instrumented animal (p = 0.001). CONCLUSIONS: The sympathetic storm produced transient contractile dysfunction, consistent with ischemic injury. However, troponin I release reflected surgical instrumentation and not brain death.
Asunto(s)
Presión Sanguínea/fisiología , Muerte Encefálica/fisiopatología , Frecuencia Cardíaca/fisiología , Hipertensión Intracraneal/fisiopatología , Isquemia Miocárdica/fisiopatología , Sistema Nervioso Simpático/fisiopatología , Animales , Cadáver , Circulación Coronaria/fisiología , Modelos Animales de Enfermedad , Trasplante de Corazón , Hipertensión Intracraneal/complicaciones , Contracción Miocárdica/fisiología , Isquemia Miocárdica/etiología , Porcinos , Troponina I/sangreRESUMEN
BACKGROUND: U74389G (16-desmethyl tirilazad), a 21-aminosteroid or "lazaroid," inhibits lipid peroxidation, which is an important element of ischemia-reperfusion injury. The aim of this study was to determine whether the addition of U74389G to the cardioplegic preservation solution could improve early cardiac allograft function. METHODS: A porcine model of donor brain death and orthotopic cardiac transplantation was used. Hearts were arrested and preserved for 6 hours in an aspartate-enriched extracellular cardioplegia that had been supplemented with either U74389G and its carrier (n = 7) or the carrier alone (n = 9). Epicardial sonomicrometry and transmyocardial micromanometry were used to obtain pressure-volume loops before and after transplantation. Left ventricular wall volume was measured by volume displacement. RESULTS: A higher proportion of U74389G-treated hearts were weaned successfully from cardiopulmonary bypass, but this difference did not achieve statistical significance (86% [6 of 7] vs 56% [5 of 9]; p = 0.308). In the hearts that were weaned successfully, preservation of left ventricular contractility, as judged by the pre-load recruitable stroke work relationship, was significantly better in the U74389G-treated hearts (p = 0.0271). In contrast, left ventricular compliance, as judged by the end-diastolic pressure-volume relationship, was significantly better preserved in the control group (p < 0.0001). U74389G-treated hearts developed less myocardial edema, as judged by the post-transplant left ventricular wall volume/baseline steady-state epicardial end-diastolic volume ratio (64 +/- 9% vs 76 +/- 11%; p = 0.045). CONCLUSIONS: The benefit obtained from U74389G-supplemented cardioplegic preservation solution was marginal for hearts stored for 6 hours. After longer ischemic times, the benefit may be clearer.
Asunto(s)
Antioxidantes/farmacología , Soluciones Cardiopléjicas , Trasplante de Corazón , Soluciones Preservantes de Órganos , Pregnatrienos/farmacología , Animales , Puente Cardiopulmonar , Corazón , Trasplante de Corazón/fisiología , Porcinos , Factores de Tiempo , Trasplante HomólogoRESUMEN
OBJECTIVE: To determine if the initial rate of troponin I release post-reperfusion reflects the effectiveness of myocardial protection during cardiac allograft preservation. METHODS: A porcine model of orthotopic heart transplantation was used. Data from two control groups (CON(4) and CON(14)) and two treatment groups (CAR(4) and CAR(14)) were analysed. Hearts in CON(4) (n=6) and CAR(4) (n=6) were subjected to 4 h of ischaemia while hearts in CON(14) (n=3) and CAR(14) (n=6) were subjected to 14 h of ischaemia. All hearts were arrested and stored in the same extracellular preservation solution. Both donor and recipient animals in the CAR(4) and CAR(14) groups received a single intravenous dose of cariporide (2 mg/kg), prior to explantation and reperfusion, respectively. RESULTS: Mean (SEM) plasma troponin I levels (microg/ml) 3 h post-reperfusion were: CON(4) 210+/-52, CAR(4) 68+/-21, CON(14) 633+/-177, CAR(14) 346+/-93. On multiple linear regression analysis, the rate of troponin I release over the first 3 h post-reperfusion was significantly lower in hearts stored for 4 h compared to hearts stored for 14 h (P<0.0001) and in hearts treated with cariporide compared to control hearts (P=0.0017). Early graft function was superior in hearts treated with cariporide, when compared to control hearts stored for the same period of time. All of the CAR(14) hearts could be weaned from cardiopulmonary bypass whereas none of the CON(14) could be weaned (6/6 vs. 0/3; P=0.012). While all hearts stored for 4 h could be weaned, contractility, as measured by the preload recruitable stroke work (PRSW) relationship, was significantly better preserved in CAR(4) hearts than in CON(4) hearts (P<0.0001). CONCLUSIONS: The initial rate of troponin I release post-reperfusion is determined by the duration of cardiac allograft ischaemia. Altering the myocardial preservation strategy can reduce the rate of release. Such reductions are associated with improvements in early graft function. These findings validate the initial rate of troponin I release post-reperfusion as an end-point when comparing cardiac allograft preservation strategies. In addition, the present study provides indirect evidence that troponin I degradation during ischaemia-reperfusion is related to the accumulation of intracellular calcium.