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Biomedical subjects

K R Chien

Publications and source records attributed to K R Chien.

At least 73 records · Page 4Linked to original sources
PubMed1997

Report of the National Heart, Lung, and Blood Institute Special Emphasis Panel on Heart Failure Research.

The SEP identified priorities to support in future basic and clinical research and pointed out directions likely to result in advances against heart failure. The list is not intended to be all-encompassing and does not address, for example, exciting lines of work already under way. Rather, the recommendations are designed to point out gaps in current knowledge not being adequately addressed and highly promising new directions. Although the incidence of heart failure continues to grow, emerging lines of research provide hope that research advances will eventually lead to more effective treatment and ultimately to prevention. This research will be well served by bringing the latest multidisciplinary approaches and the best investigators to focus on the problems of heart failure. It is hoped the efforts of distinguished expert entities such as the task force and SEP will be a useful guide in addressing the needs of the biomedical community and assisting in its success.

Animals↗
PubMed1997

MLP-deficient mice exhibit a disruption of cardiac cytoarchitectural organization, dilated cardiomyopathy, and heart failure.

MLP is a LIM-only protein of terminally differentiated striated muscle cells, where it accumulates at actin-based structures involved in cytoarchitecture organization. To assess its role in muscle differentiation, we disrupted the MLP gene in mice. MLP (-/-) mice developed dilated cardiomyopathy with hypertrophy and heart failure after birth. Ultrastructural analysis revealed dramatic disruption of cardiomyocyte cytoarchitecture. At birth, these hearts were not hypertrophic, but already abnormally soft, with cell-autonomous and MLP-sensitive alterations in cytoarchitecture. Thus, MLP promotes proper cardiomyocyte cytoarchitecture, whose perturbation can lead to dilated cardiomyopathy. In vivo analysis revealed that MLP-deficient mice reproduce the morphological and clinical picture of dilated cardiomyopathy and heart failure in humans, providing the first model for this condition in a genetically manipulatable organism.

Animals↗
PubMed1997

Developmental expression of the murine spliceosome-associated protein mSAP49.

We have isolated the mouse homologue of human spliceosome-associated protein SAP49, mSAP49. mSAP49 contains two RNA recognition motifs (RRM) in the N terminus of the predicted amino acid sequence, and a highly basic C terminus rich in glycine/proline. mSAP49 displayed a plastic of expression in cardiac development. In the adult mouse, mSAP49 is widely distributed, although it was found at relatively lower levels in the heart. In situ hybridization analysis of mSAP49 mRNA distribution in staged mouse embryos showed that mSAP49 onset occurs later in the heart than in other embryonic tissues. While mSAP49 expression was found at day 10.0 postconception (pc) in the optic eminence, optic vesicle, hindbrain, and somites, it was not in cardiac structures. mSAP49 was detected in the ventricles at day 11.5, and at day 13.5 it was also detected in the atria. Northern analysis showed that mSAP49 mRNA displayed a peak of expression in the heart at days 14.0-15.0 pc, and its abundance decayed in the adult. This dynamic pattern of cardiac expression suggests that mSAP49 may be contributing to a change in the ratio of spliceosome components during cardiac growth and development, which may have consequences for tissue-specific splicing, RNA stabilization, or translation.

Amino Acid Sequence↗
PubMed1997

A novel A/T-rich element mediates ANF gene expression during cardiac myocyte hypertrophy.

The induction of the atrial natriuretic factor (ANF) gene during alpha 1-adrenergic stimulation of neonatal rat ventricular myocytes has served as a model for gene expression during cardiac muscle cell hypertrophy. This study describes and identifies a single regulatory element that mediates expression of the ANF gene. Deletional mutations were generated in a 639-bp fragment of the ANF promoter that confers alpha 1-adrenergic inducibility to a luciferase reporter gene in transient transfection assays in ventricular myocytes. The results of gel mobility shift and diethylpyrocarbonate (DEPC) interference studies with nuclear cardiac cell extracts identified the nucleotide contract points for a novel A/T-rich element (ANF-AT) at positions -582/-575 that partially mediates alpha 1-adrenergic inducibility. Mutations in the ANF-AT element reduced alpha-adrenergic inducibility of an ANF-TK-luciferase fusion gene in cardiac cells by 35% but had no effect on expression in other muscle and non-muscle cells tested. Gel mobility supershift assays with antibodies directed against the MEF-2 protein, the homeobox protein MHox, or the zinc finger protein HF-1b, document that these factors are not major components of the endogenous ANF-AT binding activity in cardiac muscle cells. The current study provides evidence for a role for a novel A/T-rich element in the regulation of ANF gene expression in cardiac ventricular myocytes.

Animals↗
PubMed1997

Cardiotrophin-1 and the role of gp130-dependent signaling pathways in cardiac growth and development.

The reactivation of an embryonic pattern of gene expression is a central feature common to virtually all forms of cardiac hypertrophy. Unraveling the regulatory mechanisms, growth factors and cytokines controlling gene expression and cell fate during cardiac development may therefore have implications for our understanding of cardiac hypertrophy in the adult. Along this line, a cDNA expression library was established from an embryonic stem cell-based in vitro model of cardiogenesis, and screened for clones that would induce an increase in cell size in cultured cardiomyocytes. This experimental strategy resulted in the isolation of a novel cytokine, cardiotrophin-1 (CT-1), that activates several features of cardiomyocyte hypertrophy in vitro, including sarcomeric organization and embryonic gene expression. CT-1 displays structural similarities to the interleukin (IL)-6 related cytokines. Furthermore, receptor binding studies and functional studies reveal that CT-1 shares the signal transducing receptor components gp130 and LIFR with the previously identified members of the IL-6 cytokine family. CT-1 rapidly activates gp130 and LIFR tyrosine phosphorylation in cultured cardiac myocytes. The growth promoting effects of CT-1 therefore indicate that signaling pathways emanating from gp130 and LIFR are coupled to cardiomyocyte hypertrophy. In support of this notion, the simultaneous overexpression of IL-6 and the IL-6 receptor in transgenic mice has been shown to result in a constitutive tyrosine phosphorylation of gp130 in the myocardium and cardiac hypertrophy. The striking phenotype of gp130 null-mutant mice, generated by homologous recombination, implies gp130 in cardiac development as well: mutant mice exhibit severe ventricular hypoplasia, suggesting a role for gp130-dependent signaling pathways in the expansion of the compact layer of the ventricular myocardium. CT-1 is expressed at high levels in the myocardium during the course of cardiogenesis, and promotes the proliferation and survival of embryonic cardiomyocytes. CT-1 may therefore represent a candidate cytokine to activate gp130 during cardiac development. In summary, cytokines signaling through gp130 are emerging as potent regulators of embryonic heart development and adult cardiac hypertrophy.

Animals↗
PubMed1997

Physiological assessment of complex cardiac phenotypes in genetically engineered mice.

The recent development of techniques for surgical manipulation and for the assessment of cardiac physiology in genetically engineered mice has allowed scientists to address some of the most fundamental questions related to congenital and acquired forms of human heart disease. This review discusses recent advances in the techniques for studying cardiac disease using the mouse as a model system. Because cardiac overload is one of the most important stimuli for development of hypertrophy and heart failure in humans, various models of cardiac pressure and volume overload, as well as myocardial ischemia, have been developed and characterized. Moreover, it is possible to reliably examine murine cardiac physiology in vivo with microtransducers, echocardiography, and other miniaturized techniques. Sophisticated methods have also been developed to enable an examination of single-cell phenotypes of isolated cardiomyocytes derived from genetically engineered mice. These physiological assessments, coupled with conventional histology and molecular markers, have allowed the characterization of several gene-targeted and transgenic mouse models of hypertrophy and dilated cardiomyopathy, as well as mouse models of cardiac developmental defects. Such mouse models of heart disease will ultimately allow the molecular dissection of the interplay between the various factors leading to heart disease, and they may serve as a guide to appropriate therapeutic strategies for human heart disease.

Animals↗
PubMed1997

Left ventricular volumes and function in the embryonic mouse heart.

This study describes miniaturized technology for the in vivo analysis of the volume and function of the embryonic mouse heart and the application of this technology to study the normal embryonic left ventricle (LV) at two stages of development. With the use of microsurgical techniques, embryos from embryonic day (ED) 10.5 (ED10.5) to ED16 were delivered individually from litters of normal dams, and cardiac visualization was achieved with the use of intravital microscopy by transillumination, with the umbilical circulation intact. At ED10.5-11, the heart could be imaged in color in the intact embryo, whereas at ED12.5 it was necessary to open the chest; at ED13.5-14.5, fluorescent imaging with the use of microinjection of fluorescein-conjugated albumin was necessary to visualize the LV chamber. At ED10.5-11, the LV end-diastolic volumes averaged 0.16 microliter (n = 14), and at ED13.5-14.5, they averaged 0.57 microliter (n = 16). At both ages there was a positive linear relationship between the LV end-diastolic volume and the stroke volume despite substantial variations in individual heart rates, reflecting the relative uniformity of the LV ejection fractions within each age group. The average of the individual ejection fractions was 27.4% at ED10.5-11 and 58.4% at ED13.5-14.5, the latter being within the normal range for the adult rodent heart. These methods will be useful for assessing in vivo cardiac function at ED10.5 and older murine embryos in litters of transgenic or gene-targeted mice when the mutation leads to later embryonic lethality.

Animals↗
PubMed1997

Fashioning the vertebrate heart: earliest embryonic decisions.

Our goal here is to set out the types of unitary decisions made by heart progenitor cells, from their appearance in the heart field until they form the simple heart tube. This provides a context to evaluate cell fate, lineage and, finally, morphogenetic decisions that configure global heart form and function. Some paradigms for cellular differentiation and for pattern generation may be borrowed from invertebrates, but neither Drosophila nor Caenorhabditis elegans suffice to unravel higher order decisions. Genetic analyses in mouse and zebrafish may provide one entrance to these pathways.

Animals↗
PubMed1997

CARP, a cardiac ankyrin repeat protein, is downstream in the Nkx2-5 homeobox gene pathway.

To identify the molecular pathways that guide cardiac ventricular chamber specification, maturation and morphogenesis, we have sought to characterize factors that regulate the expression of the ventricular myosin light chain-2 gene, one of the earliest markers of ventricular regionalization during mammalian cardiogenesis. Previously, our laboratory identified a 28 bp HF-la/MEF-2 element in the MLC-2v promoter region, which confers cardiac ventricular chamber-specific gene expression during murine cardiogenesis, and showed that the ubiquitous transcription factor YB-1 binds to the HF-la site in conjunction with a co-factor. In a search for interacting co-factors, a nuclear ankyrin-like repeat protein CARP (cardiac ankyrin repeat protein) was isolated from a rat neonatal heart cDNA library by yeast two-hybrid screening, using YB-1 as the bait. Co-immunoprecipitation and GST-CARP pulldown studies reveal that CARP forms a physical complex with YB-1 in cardiac myocytes and immunostaining shows that endogenous CARP is localized in the cardiac myocyte nucleus. Co-transfection assays indicate that CARP can negatively regulate an HF-1-TK minimal promoter in an HF-1 sequence-dependent manner in cardiac myocytes, and CARP displays a transcriptional inhibitory activity when fused to a GAL4 DNA-binding domain in both cardiac and noncardiac cell context. Northern analysis revealed that carp mRNA is highly enriched in the adult heart, with only trace levels in skeletal muscle. During murine embryogenesis, endogenous carp expression was first clearly detected as early as E8.5 specifically in heart and is regulated temporally and spatially in the myocardium. Nkx2-5, the murine homologue of Drosophila gene tinman was previously shown to be required for heart tube looping morphogenesis and ventricular chamber-specific myosin light chain-2 expression during mammalian heart development. In Nkx2-5(-/-)embryos, carp expression was found to be significantly and selectively reduced as assessed by both whole-mount in situ hybridizations and RNase protection assays, suggesting that carp is downstream of the homeobox gene Nkx2-5 in the cardiac regulatory network. Co-transfection assays using a dominant negative mutant Nkx2-5 construct with CARP promoter-luciferase reporter constructs in cardiac myocytes confirms that Nkx2-5 either directly or indirectly regulates carp at the transcriptional level. Finally, a carp promoter-lacZ transgene, which displays cardiac-specific expression in wild-type and Nkx2-5(+/-) background, was also significantly reduced in Nkx2-5(-/-) embryos, indicating that Nkx2-5 either directly or indirectly regulates carp promoter activity during in vivo cardiogenesis as well as in cultured cardiac myocytes. Thus, CARP is a YB-1 associated factor and represents the first identified cardiac-restricted downstream regulatory gene in the homeobox gene Nkx2-5 pathway and may serve as a negative regulator of HF-1-dependent pathways for ventricular muscle gene expression.

Amino Acid Sequence↗
PubMed1997

Toward molecular strategies for heart disease--past, present, future.

The past two decades of cardiovascular biology and medicine have been based largely upon the consideration of the heart and vasculature as an integrated physiological system, a view that has resulted in major therapeutic advances. With the advent of developments of gene transfer, mouse and human genetics, genetic engineering of intact animals, and molecular and cellular technology, cardiovascular medicine is now on the threshold of a molecular therapeutic era. Major steps have been taken toward unraveling the molecular determinants of complex, integrative, and polygenic cardiovascular disease states, including atherogenesis, hypertension, cardiac hypertrophy and failure, congenital heart disease, and coronary restenosis following balloon angioplasty. Our improved understanding of the fundamental basis of these important cardiovascular disease processes has established a scientific foundation for diagnostic, prognostic, and therapeutic advances in the mainstream of cardiovascular medicine.

Animals↗
PubMed1996

Rho is required for Galphaq and alpha1-adrenergic receptor signaling in cardiomyocytes. Dissociation of Ras and Rho pathways.

G protein-coupled receptor agonists initiate a cascade of signaling events in neonatal rat ventricular myocytes that culminates in changes in gene expression and cell growth characteristic of hypertrophy. These responses have been previously shown to be dependent on Gq and Ras. Rho, a member of the Ras superfamily of GTPases, regulates cytoskeletal rearrangement and transcriptional activation of the c-fos serum response element. Immunofluorescence staining of cardiomyocytes shows that Rho is present and predominantly cytosolic. We used two inhibitors of Rho function, dominant negative N19RhoA and Clostridium botulinum C3 transferase, to examine the possible requirement for Rho in alpha1-adrenergic receptor-mediated hypertrophy. Both inhibitors markedly attenuated atrial natriuretic factor (ANF) reporter gene expression induced by alpha1-adrenergic receptor stimulation with phenylephrine, and virtually abolished the increase in ANF reporter gene expression induced by GTPase-deficient Galphaq. These effects were reproduced with the myosin light chain-2 reporter gene. Notably, N19RhoA did not block the ability of activated Ras to induce ANF and myosin light chain-2 reporter gene expression. Furthermore, activation of the extracellular signal-regulated kinase by phenylephrine was not blocked by N19RhoA, nor was it stimulated by an activated mutant of RhoA. Since activated RhoA and Ras produce a large synergistic effect on ANF-luciferase gene expression, we conclude that Rho functions in a pathway separate from but complementary to Ras. Our results provide direct evidence that Rho is an effector of Galphaq signaling and suggest for the first time that a low molecular weight GTPase other than Ras is involved in regulating myocardial cell growth and gene expression in response to heterotrimeric G protein-linked receptor activation.

ADP Ribose Transferases↗
PubMed1996

Elevated blood pressure and enhanced myocardial contractility in mice with severe IGF-1 deficiency.

To circumvent the embryonic lethality of a complete deficiency in insulin-like growth factor 1 (IGF-1), we generated mice homozygous for a site-specific insertional event that created a mutant IGF-1 allele (igf1m). These mice have IGF-1 levels 30% of wild type yet survive to adulthood, thereby allowing physiological analysis of the phenotype. Miniaturized catheterization technology revealed elevated conscious blood pressure in IGF-1(m/m) mice, and measurements of left ventricular contractility were increased. Adenylyl cyclase activity was enhanced in IGF-1(m/m) hearts, without an increase in beta-adrenergic receptor density, suggesting that crosstalk between IGF-1 and beta-adrenergic signaling pathways may mediate the increased contractility. The hypertrophic response of the left ventricular myocardium in response to aortic constriction, however, was preserved in IGF-1(m/m) mice. We conclude that chronic alterations in IGF-1 levels can selectively modulate blood pressure and left ventricular function, while not affecting adaptive myocardial hypertrophy in vivo.

Adenylyl Cyclases↗
PubMed1996

RXR alpha deficiency confers genetic susceptibility for aortic sac, conotruncal, atrioventricular cushion, and ventricular muscle defects in mice.

Retinoid-dependent pathways play a central role in regulating cardiac morphogenesis. Recently, we characterized gene-targeted RXR alpha -/- embryos, which display an atrial-like ventricular phenotype with the development of heart failure and lethality at embryonic day 14.5. To quantitate the frequency and complexity of cardiac morphogenic defects, we now use microdissection and scanning electron microscopy to examine 107 wild-type, heterozygous, and homozygous embryos at embryonic day 13.5, 14.5, and 15.5. RXR alpha -/- embryos display complex defects, including ventricular septal, atrioventricular cushion, and conotruncal ridge defects, with double outlet right ventricle, aorticopulmonary window, and persistent truncus arteriosus. In addition, heterozygous RXR alpha embryos display a predisposition for trabecular and papillary muscle defects, ventricular septal defects, conotruncal ridge defects, atrioventricular cushion defects, and pulmonic stenosis. Lastly, we show that the intermediate anatomic phenotype displayed by heterozygous embryos is mirrored in the molecular marker MLC-2a. The intermediate phenotype of RXR alpha heterozygous embryos documents a gene dosage effect for RXR alpha in maintaining normal cardiac morphogenesis. In addition, some defects in RXR alpha mutant mice are phenocopies of human congenital heart defects, thereby suggesting that a relative deficiency in RXR alpha or molecules downstream in its signaling pathway may represent congenital heart disease-susceptibility genes.

Animals↗
PubMed1996

Transthoracic echocardiography in models of cardiac disease in the mouse.

BACKGROUND: Transthoracic echocardiography (M-mode and Doppler) offers a noninvasive approach for in vivo evaluation of the mouse heart. The present study examines its usefulness for assessing the morphological/functional phenotype of the left ventricle (LV) in several transgenic and surgical murine models of cardiac disease. METHODS AND RESULTS: Observations were made in 83 intact, anesthetized mice. In mice with a surgical arteriovenous fistula, volume overload and LV dilation were detected. In normal mice, echocardiographic indexes of increased contractility (dobutamine) were confirmed by LV dP/dtmax. In transgenic mice with overexpression of the beta 2-adrenergic receptor, heart rate and mean velocity of circumferential fiber shortening were increased, indicating enhanced contractility. In colony screening of transgenic mice overexpressing the H-ras gene, 45% had increased LV wall thickness (> 0.9 mm), and those showing a striking increase were selected for breeding. In mice with LV hypertrophy (aortic constriction) and normal mice, the actual LV mass determined by echocardiography correlated well (r = .93), and 95% confidence limits were determined. The maximum intraobserver and interobserver coefficients of variation for M-mode data were 0.03 +/- 0.29 mm (+/- 2 SD), < 10% for LV internal dimensions but 27% to 30% for wall thickness. CONCLUSIONS: These studies provide the first application of transthoracic echocardiography for morphological/functional characterization of the cardiac phenotype in transgenic and surgical murine models, including (1) high reliability for detecting LV chamber dilation and function; (2) reliability (and its limits) for determining abnormal LV wall thickness and LV mass; (3) identification of marked, sometimes asymmetrical, hypertrophy in a transgenic model of hypertrophic cardiomyopathy; and (4) usefulness for transgenic colony screening to identify markedly abnormal phenotypes.

Anesthesia↗
PubMed1996

Cardiovascular effects of insulin-like growth factor-1 and growth hormone in chronic left ventricular failure in the rat.

BACKGROUND: Insulin-like growth factor-1 (IGF-1) appears to have favorable cardiac effects associated with left ventricular remodeling early after myocardial infarction in the rat. The present study was designed to determine whether IGF-1 combined with growth hormone would be beneficial later as well, when infarct healing and cardiac remodeling have occurred. METHODS AND RESULTS: Four weeks after coronary occlusion, 36 rats were randomized to IGF-1 (3 mg.kg-1.d-1) plus growth hormone (0.1 mg BID) or to placebo for 4 weeks. Treated rats had significant increases in body weight (22%), while the ratio of heart weight to body weight was unchanged. Under anesthesia, cardiac output (fluorescent microspheres) increased 46%, and systemic vascular resistance decreased by 21% (P < .001) in the treated group; a significant (22%) increase of the cardiac index was limited to treated rats with large myocardial infarctions. Small increases in the reduced left ventricular ejection fractions and left ventricular dP/dt(max) values with treatment were not significant. Treated rats showed a borderline (16%) increase in left ventricular end-diastolic volume (angiography), whereas the ratio of left ventricular end-diastolic volume to body weight was reduced in the treated group. CONCLUSIONS: IGF-1 plus growth hormone administered to rats with left ventricular failure starting 1 month after MI was associated with substantial body growth, decreased systemic vascular resistance, and increased cardiac output. The failing heart also underwent treatment-induced increases in left and right ventricular weights in proportion to body growth, but left ventricular remodeling was minor, and a decrease in the ratio of left ventricular end-diastolic volume to body weight reflected relatively less chamber dilation compared with controls. A significant interaction between size of the myocardial infarction and treatment was observed for several variables, and IGF-1 and growth hormone increased the cardia index (P < .035) in rats with a large myocardial infarction.

Animals↗
PubMed1996

Cardiotrophin-1 activates a distinct form of cardiac muscle cell hypertrophy. Assembly of sarcomeric units in series VIA gp130/leukemia inhibitory factor receptor-dependent pathways.

Cardiotrophin-1 (CT-1) was recently isolated by expression cloning based on its ability to induce an increase in cell size in neonatal rat ventricular cardiomyocytes. Sequence similarity data suggested that CT-1 is a novel member of a family of structurally related cytokines sharing the receptor component gp130. The present study documents that gp130 is required for CT-1 signaling in cardiomyocytes, by demonstrating that a monoclonal anti-gp130 antibody completely inhibits c-fos induction by CT-1. Similarly, a leukemia inhibitory factor receptor subunit beta (LIFRbeta) antagonist effectively blocks the CT-1 induction of c-fos, indicating a requirement for LIFRbeta in the hypertrophic response, as well. Upon stimulation with CT-1, both gpl30 and the LIFRbeta are tyrosine-phosphorylated, providing further evidence that CT-1 signals through the gp130/LIFRbeta heterodimer in cardiomyocytes. CT-1 induces a hypertrophic response in cardiomyocytes that is distinct from the phenotype seen after alpha-adrenergic stimulation, both with regard to cell morphology and gene expression pattern. Stimulation with CT-1 results in an increase in cardiac cell size that is characterized by an increase in cell length but no significant change in cell width. Confocal laser microscopy of CT-1 stimulated cells reveals the assembly of sarcomeric units in series rather than in parallel, as seen after alpha-adrenergic stimulation. CT-1 induces a distinct pattern of immediate early genes, and up-regulates the atrial natriuretic factor (ANF) gene, but does not affect skeletal alpha-actin or myosin light chain-2v expression. As evidenced by nuclear run-on transcription assays, both CT-1 and alpha-adrenergic stimulation lead to an increase in ANF gene transcription. Transient transfection analyses document that, in contrast to alpha-adrenergic stimulation, the CT-1 responsive cis-regulatory elements are located outside of the proximal 3 kilobase pairs of the ANF 5'-flanking region. These studies indicate that CT-1 can activate a distinct form of myocardial cell hypertrophy, characterized by the promotion of sarcomere assembly in series, via gpl30/LIFRbeta-dependent signaling pathways.

Actins↗