Nanette H. Bishopric, M.D., F.A.C.C., F.A.H.A., is Professor of Oncology at Georgetown University and a faculty member in the MedStar Heart Institute in Washington, DC. Her research focuses on molecular effectors of the response to stress, and how stress affects cell death, growth and renewal. In the myocardium. Her recent work has explored the role of microenvironmental factors in cancer cell growth and metabolism. She has received multiple grants from the National Institutes of Health, Fondation Leducq (Paris) the Miami Heart Research Institute and the American Heart Association. Dr. Bishopric has authored more than 100 peer-reviewed journal articles as well as many reviews, editorials and book chapters. Dr. Bishopric attended Duke University Medical School and was a Howard Hughes fellow in the laboratory of Dr. Robert Lefkowitz before training in Internal Medicine at University of Florida and in Cardiology at UC San Francisco. Research honors include the Merck/ACC Fellowship Award, the SRI International Technical Excellence Award, the Established Investigator Award from the American Heart Association, the American College of Cardiology 2010 Distinguished Scientist award, and election to the Association of University Cardiologists in 2014.
The skeletal alpha-actin gene encodes a major component of the embryonic cardiac sarcomere that i... more The skeletal alpha-actin gene encodes a major component of the embryonic cardiac sarcomere that is strongly and selectively re-induced during beta-adrenoceptor-mediated hypertrophy in neonatal rat cardiac myocytes. We present evidence that beta-adrenergic induction of this gene is mediated, not by cAMP, but by a calcium-dependent pathway involving ryanodine-sensitive calcium stores. Nifedipine-induced blockade of the plasma membrane L-type calcium entry channel prevented induction of skeletal alpha-actin mRNA by isoproterenol. Activation of calcium entry by the dihydropyridine agonist Bay K8644 independently induced skeletal alpha-actin mRNA, as did cholera toxin-mediated activation of Gs. Induction of skeletal alpha-actin mRNA by compounds that directly elevate cAMP was weak relative to their effects on other cAMP-dependent phenomena and required calcium entry. In addition, selective inhibition of protein kinase A with KT5720 did not block beta-adrenergic induction of skeletal alpha-actin. Calcium ionophore A23187 did not induce skeletal actin, but prevented its induction by isoproterenol. Ryanodine had bimodal effects: 10(-10) M ryanodine induced skeletal alpha-actin mRNA, whereas 10(-6) M ryanodine prevented skeletal actin induction by beta-adrenergic stimuli. We postulate that beta-adrenergic stimulation of skeletal alpha-actin mRNA requires G-protein-coupled calcium channel activation and compartmentalized calcium release in a manner independent of the cAMP/protein kinase A signal pathway.
In recent years genome-wide association studies (GWAS) have uncovered a plethora of chromosomal l... more In recent years genome-wide association studies (GWAS) have uncovered a plethora of chromosomal loci associated with a variety of cardiac traits. As in most GWAS, a considerable portion of identified signals occur at inter-genic regions of the genome. Trait-associated single nucleotide polymorphisms (SNPs) located in these non-coding regions likely exert their effect through modulation of gene expression. Thus interrogating these variants for association with differential transcript abundance by expression quantitative trait locus (eQTL) analysis is a highly relevant means to unravel the causal genes at loci identified by GWAS. This will in turn further our understanding of the molecular mechanisms underlying cardiac traits. In this study, for the first time, we carried out an eQTL analysis in human heart. A total of 129 human left ventricle samples were collected at four collaborating centers. Samples were acquired from non-diseased hearts that were considered suitable for transplantation, yet not used for logistical reasons. All individuals were of Western European descent. Genome-wide transcript abundance and genotypes were assessed using the Illumina HumanHT-12 and HumanOmniExpress microarrays, respectively. After pre-processing and stringent quality control of transcript and genotypic data, each transcript was tested for association with genotypes genome-wide using linear modelling in R, correcting for age, gender and center effects. We identified 770 independent cis-eQTLs (SNPs within 1 Mb of transcript) mapping to 457 unique transcripts (FDR < 1%). Overlaying these eQTLs with cardiac GWAS loci identified strong candidate genes for functional studies. One of these involves an eQTL effect of rs9912468, a robust modifier of QRS, on PRKCA (p = 1.52E-11). Our findings provide evidence for the role of PRKCA in mediating the effect of the rs9912468-tagged haplotype on QRS duration. Additionally, we identified several trans-eQTL hotspots in the vicinity of transcription factors, regulating targets predominantly involved in immune response, signalling and cardiac muscle structure related processes, all known to impact on the heart function. An on-going effort entails overlaying the eQTLs with known cardiac regulatory regions, such as binding regions of cardiac specific transcription factors (TBX3, TBX5, NKX2-5, GATA4, SRF) and enhancers, identified through ChIP-sequencing in the ENCODE project and other studies. Our study illustrates the power of integrating gene expression, phenotype and genotype data in a systems genetics approach, in identifying novel causal genes for human cardiac phenotypes.
Since apoptosis was described as a process distinct from necrosis, there have been many studies o... more Since apoptosis was described as a process distinct from necrosis, there have been many studies of programmed cell death in diseases, especially immunological diseases. Because cardiac myocytes are terminally differentiated cells, they have typically been assumed to die exclusively by necrosis. However, during the last six to seven years this view has been challenged by several studies demonstrating that a significant number of myocytes undergo apoptosis in myocardial infarction, heart failure, myocarditis, arrhythmogen right ventricular dysplasia, and immune rejection after cardiac transplantation, as well as in other conditions of stress. These are potentially very important observations, because apoptosis--unlike necrosis--can be blocked or reversed at early stages. The tracking of cytoprotective and apoptotic signal transduction pathways has proceeded rapidly with important new insights into the roles of mitochondria-dependent pathway, Bcl-2 protein family, p38 mitogen-activated...
Proceedings of the National Academy of Sciences, 1992
The cardiac alpha-actin gene is expressed in both heart and skeletal muscle. In skeletal myogenic... more The cardiac alpha-actin gene is expressed in both heart and skeletal muscle. In skeletal myogenic cells, the 177-base-pair promoter of the human cardiac alpha-actin (HCA) gene requires three transcription factors for activation: Sp1, serum response factor (SRF), and MyoD. However, MyoD is undetectable in heart. To search for a functional equivalent of MyoD, we analyzed the transcriptional regulation of the HCA promoter in primary cultures of rat cardiac myocytes. The same DNA sequence elements recognized by SRF, Sp1, and MyoD and required for HCA transcription in skeletal muscle cells were also found to be necessary for expression in cardiomyocytes. Overexpression of Id, a negative regulator of basic helix-loop-helix proteins, selectively attenuated expression of the HCA promoter. Cardiomyocyte nuclei contain a protein complex that specifically interacts with the same required sequence (E box) in the HCA promoter that is bound by MyoD in skeletal myogenic cells. Furthermore, these c...
Proceedings of the National Academy of Sciences, 1991
The skeletal alpha-actin gene is expressed in fetal rat heart and is induced during norepinephrin... more The skeletal alpha-actin gene is expressed in fetal rat heart and is induced during norepinephrine (NE)-stimulated hypertrophy in cultures of neonatal rat cardiac myocytes. Here we report that NE positively regulates the human skeletal alpha-actin gene promoter in transiently transfected neonatal rat cardiac myocytes. NE increased expression from the full-length promoter by 2.4-fold. A DNA region required for NE responsiveness but not for tissue-specific expression was located between base pair -2000 and base pair -1300. Distinct regions required for cardiac myocyte expression were located between -1300 to -710 and -153 to -87. None of these elements separately conferred tissue specificity or adrenergic responsiveness on a heterologous promoter, although the intact promoter from -2000 to -36 conferred both when cloned in its correct position and orientation. Additional elements in the basal promoter (-87 to +187) were required for maximal NE responsiveness. The NE induction was medi...
Journal of Molecular and Cellular Cardiology, 1995
Short periods of myocardial ischemia appear to provide protection against subsequent prolonged is... more Short periods of myocardial ischemia appear to provide protection against subsequent prolonged ischemic episodes in experimental animals and in man. This phenomenon, known as ischemic preconditioning, has not yet been characterized at the cellular or molecular levels; however, tissue hypoxia appears to be required. In this study, we used a previously developed method for hypoxic cardiac myocyte culture in order to establish a model for ischemic (or hypoxic) preconditioning in cell culture. We demonstrate that cultured neonatal rat cardiac myocytes preconditioned by 25 min of exposure to hypoxia followed by reoxygenation were protected against membrane damage for up to 6 h of prolonged severe hypoxia, as determined by arachidonic acid release and contractile recovery. In contrast, non-preconditioned myocytes exhibited significant hypoxic damage after 2-4 h. Pretreatment of cells with PMA, a tumor-promoting phorbol ester, mimicked the protective effects of hypoxic preconditioning; pretreatment with the muscarinic cholinergic agonist carbachol had no effect. Our data suggests that isolated myocytes in culture remain competent to be preconditioned by hypoxia, through a pathway that may involve the activation of protein kinase C.
Journal of Molecular and Cellular Cardiology, 1993
Positive inotropic agents that increase the sensitivity of myofilaments to calcium have recently ... more Positive inotropic agents that increase the sensitivity of myofilaments to calcium have recently been described (Kitada et al., 1987; Cottney et al., 1990; Ferroni et al., 1991; Lee and Allen, 1991; Beier et al., 1992). These drugs appear to augment contractility independently of cAMP or calcium, and thus may have fewer of the adverse side effects seen with other currently available agents (Katz, 1986; Packer 1989). The clinical utility of &quot;calcium-sensitizers&quot; has been questioned on the theoretical grounds that such agents may interfere with relaxation and impair diastolic function (Hajjar and Gwathmey, 1991). Previous studies have shown a small but significant negative lusitropic effect of the calcium sensitizer EMD 53998 in ferret papillary muscle, although this effect was considered to be outweighed by powerful augmentation of contractility. Modelling studies have suggested that the impairment of relaxation by calcium-sensitizers may be even more severe when myocardial calcium is abnormally elevated, such as in hypoxia (Allen and Orchard, 1987; Lodge and Gelband, 1988) and end-stage heart failure (Hajjar and Gwathmey, 1991). We have examined the effects of EMD 53998 and milrinone on contractility and calcium flux in a cell culture model of myocardial hypoxia. The results indicate that increased calcium sensitivity results in marked impairment of relaxation under hypoxic conditions, possibly due to the impaired calcium sequestration and increased calcium availability exhibited by hypoxic myocytes. These studies show that the effects of calcium sensitizers can be strongly influenced by the prevailing status of intracellular calcium handling, and may be deleterious in the diseased or ischemic myocardium.
The skeletal alpha-actin gene encodes a major component of the embryonic cardiac sarcomere that i... more The skeletal alpha-actin gene encodes a major component of the embryonic cardiac sarcomere that is strongly and selectively re-induced during beta-adrenoceptor-mediated hypertrophy in neonatal rat cardiac myocytes. We present evidence that beta-adrenergic induction of this gene is mediated, not by cAMP, but by a calcium-dependent pathway involving ryanodine-sensitive calcium stores. Nifedipine-induced blockade of the plasma membrane L-type calcium entry channel prevented induction of skeletal alpha-actin mRNA by isoproterenol. Activation of calcium entry by the dihydropyridine agonist Bay K8644 independently induced skeletal alpha-actin mRNA, as did cholera toxin-mediated activation of Gs. Induction of skeletal alpha-actin mRNA by compounds that directly elevate cAMP was weak relative to their effects on other cAMP-dependent phenomena and required calcium entry. In addition, selective inhibition of protein kinase A with KT5720 did not block beta-adrenergic induction of skeletal alpha-actin. Calcium ionophore A23187 did not induce skeletal actin, but prevented its induction by isoproterenol. Ryanodine had bimodal effects: 10(-10) M ryanodine induced skeletal alpha-actin mRNA, whereas 10(-6) M ryanodine prevented skeletal actin induction by beta-adrenergic stimuli. We postulate that beta-adrenergic stimulation of skeletal alpha-actin mRNA requires G-protein-coupled calcium channel activation and compartmentalized calcium release in a manner independent of the cAMP/protein kinase A signal pathway.
In recent years genome-wide association studies (GWAS) have uncovered a plethora of chromosomal l... more In recent years genome-wide association studies (GWAS) have uncovered a plethora of chromosomal loci associated with a variety of cardiac traits. As in most GWAS, a considerable portion of identified signals occur at inter-genic regions of the genome. Trait-associated single nucleotide polymorphisms (SNPs) located in these non-coding regions likely exert their effect through modulation of gene expression. Thus interrogating these variants for association with differential transcript abundance by expression quantitative trait locus (eQTL) analysis is a highly relevant means to unravel the causal genes at loci identified by GWAS. This will in turn further our understanding of the molecular mechanisms underlying cardiac traits. In this study, for the first time, we carried out an eQTL analysis in human heart. A total of 129 human left ventricle samples were collected at four collaborating centers. Samples were acquired from non-diseased hearts that were considered suitable for transplantation, yet not used for logistical reasons. All individuals were of Western European descent. Genome-wide transcript abundance and genotypes were assessed using the Illumina HumanHT-12 and HumanOmniExpress microarrays, respectively. After pre-processing and stringent quality control of transcript and genotypic data, each transcript was tested for association with genotypes genome-wide using linear modelling in R, correcting for age, gender and center effects. We identified 770 independent cis-eQTLs (SNPs within 1 Mb of transcript) mapping to 457 unique transcripts (FDR < 1%). Overlaying these eQTLs with cardiac GWAS loci identified strong candidate genes for functional studies. One of these involves an eQTL effect of rs9912468, a robust modifier of QRS, on PRKCA (p = 1.52E-11). Our findings provide evidence for the role of PRKCA in mediating the effect of the rs9912468-tagged haplotype on QRS duration. Additionally, we identified several trans-eQTL hotspots in the vicinity of transcription factors, regulating targets predominantly involved in immune response, signalling and cardiac muscle structure related processes, all known to impact on the heart function. An on-going effort entails overlaying the eQTLs with known cardiac regulatory regions, such as binding regions of cardiac specific transcription factors (TBX3, TBX5, NKX2-5, GATA4, SRF) and enhancers, identified through ChIP-sequencing in the ENCODE project and other studies. Our study illustrates the power of integrating gene expression, phenotype and genotype data in a systems genetics approach, in identifying novel causal genes for human cardiac phenotypes.
Since apoptosis was described as a process distinct from necrosis, there have been many studies o... more Since apoptosis was described as a process distinct from necrosis, there have been many studies of programmed cell death in diseases, especially immunological diseases. Because cardiac myocytes are terminally differentiated cells, they have typically been assumed to die exclusively by necrosis. However, during the last six to seven years this view has been challenged by several studies demonstrating that a significant number of myocytes undergo apoptosis in myocardial infarction, heart failure, myocarditis, arrhythmogen right ventricular dysplasia, and immune rejection after cardiac transplantation, as well as in other conditions of stress. These are potentially very important observations, because apoptosis--unlike necrosis--can be blocked or reversed at early stages. The tracking of cytoprotective and apoptotic signal transduction pathways has proceeded rapidly with important new insights into the roles of mitochondria-dependent pathway, Bcl-2 protein family, p38 mitogen-activated...
Proceedings of the National Academy of Sciences, 1992
The cardiac alpha-actin gene is expressed in both heart and skeletal muscle. In skeletal myogenic... more The cardiac alpha-actin gene is expressed in both heart and skeletal muscle. In skeletal myogenic cells, the 177-base-pair promoter of the human cardiac alpha-actin (HCA) gene requires three transcription factors for activation: Sp1, serum response factor (SRF), and MyoD. However, MyoD is undetectable in heart. To search for a functional equivalent of MyoD, we analyzed the transcriptional regulation of the HCA promoter in primary cultures of rat cardiac myocytes. The same DNA sequence elements recognized by SRF, Sp1, and MyoD and required for HCA transcription in skeletal muscle cells were also found to be necessary for expression in cardiomyocytes. Overexpression of Id, a negative regulator of basic helix-loop-helix proteins, selectively attenuated expression of the HCA promoter. Cardiomyocyte nuclei contain a protein complex that specifically interacts with the same required sequence (E box) in the HCA promoter that is bound by MyoD in skeletal myogenic cells. Furthermore, these c...
Proceedings of the National Academy of Sciences, 1991
The skeletal alpha-actin gene is expressed in fetal rat heart and is induced during norepinephrin... more The skeletal alpha-actin gene is expressed in fetal rat heart and is induced during norepinephrine (NE)-stimulated hypertrophy in cultures of neonatal rat cardiac myocytes. Here we report that NE positively regulates the human skeletal alpha-actin gene promoter in transiently transfected neonatal rat cardiac myocytes. NE increased expression from the full-length promoter by 2.4-fold. A DNA region required for NE responsiveness but not for tissue-specific expression was located between base pair -2000 and base pair -1300. Distinct regions required for cardiac myocyte expression were located between -1300 to -710 and -153 to -87. None of these elements separately conferred tissue specificity or adrenergic responsiveness on a heterologous promoter, although the intact promoter from -2000 to -36 conferred both when cloned in its correct position and orientation. Additional elements in the basal promoter (-87 to +187) were required for maximal NE responsiveness. The NE induction was medi...
Journal of Molecular and Cellular Cardiology, 1995
Short periods of myocardial ischemia appear to provide protection against subsequent prolonged is... more Short periods of myocardial ischemia appear to provide protection against subsequent prolonged ischemic episodes in experimental animals and in man. This phenomenon, known as ischemic preconditioning, has not yet been characterized at the cellular or molecular levels; however, tissue hypoxia appears to be required. In this study, we used a previously developed method for hypoxic cardiac myocyte culture in order to establish a model for ischemic (or hypoxic) preconditioning in cell culture. We demonstrate that cultured neonatal rat cardiac myocytes preconditioned by 25 min of exposure to hypoxia followed by reoxygenation were protected against membrane damage for up to 6 h of prolonged severe hypoxia, as determined by arachidonic acid release and contractile recovery. In contrast, non-preconditioned myocytes exhibited significant hypoxic damage after 2-4 h. Pretreatment of cells with PMA, a tumor-promoting phorbol ester, mimicked the protective effects of hypoxic preconditioning; pretreatment with the muscarinic cholinergic agonist carbachol had no effect. Our data suggests that isolated myocytes in culture remain competent to be preconditioned by hypoxia, through a pathway that may involve the activation of protein kinase C.
Journal of Molecular and Cellular Cardiology, 1993
Positive inotropic agents that increase the sensitivity of myofilaments to calcium have recently ... more Positive inotropic agents that increase the sensitivity of myofilaments to calcium have recently been described (Kitada et al., 1987; Cottney et al., 1990; Ferroni et al., 1991; Lee and Allen, 1991; Beier et al., 1992). These drugs appear to augment contractility independently of cAMP or calcium, and thus may have fewer of the adverse side effects seen with other currently available agents (Katz, 1986; Packer 1989). The clinical utility of &quot;calcium-sensitizers&quot; has been questioned on the theoretical grounds that such agents may interfere with relaxation and impair diastolic function (Hajjar and Gwathmey, 1991). Previous studies have shown a small but significant negative lusitropic effect of the calcium sensitizer EMD 53998 in ferret papillary muscle, although this effect was considered to be outweighed by powerful augmentation of contractility. Modelling studies have suggested that the impairment of relaxation by calcium-sensitizers may be even more severe when myocardial calcium is abnormally elevated, such as in hypoxia (Allen and Orchard, 1987; Lodge and Gelband, 1988) and end-stage heart failure (Hajjar and Gwathmey, 1991). We have examined the effects of EMD 53998 and milrinone on contractility and calcium flux in a cell culture model of myocardial hypoxia. The results indicate that increased calcium sensitivity results in marked impairment of relaxation under hypoxic conditions, possibly due to the impaired calcium sequestration and increased calcium availability exhibited by hypoxic myocytes. These studies show that the effects of calcium sensitizers can be strongly influenced by the prevailing status of intracellular calcium handling, and may be deleterious in the diseased or ischemic myocardium.
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Papers by Nanette H Bishopric