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1.
Nature ; 588(7838): 466-472, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32971526

RESUMO

Cardiovascular disease is the leading cause of death worldwide. Advanced insights into disease mechanisms and therapeutic strategies require a deeper understanding of the molecular processes involved in the healthy heart. Knowledge of the full repertoire of cardiac cells and their gene expression profiles is a fundamental first step in this endeavour. Here, using state-of-the-art analyses of large-scale single-cell and single-nucleus transcriptomes, we characterize six anatomical adult heart regions. Our results highlight the cellular heterogeneity of cardiomyocytes, pericytes and fibroblasts, and reveal distinct atrial and ventricular subsets of cells with diverse developmental origins and specialized properties. We define the complexity of the cardiac vasculature and its changes along the arterio-venous axis. In the immune compartment, we identify cardiac-resident macrophages with inflammatory and protective transcriptional signatures. Furthermore, analyses of cell-to-cell interactions highlight different networks of macrophages, fibroblasts and cardiomyocytes between atria and ventricles that are distinct from those of skeletal muscle. Our human cardiac cell atlas improves our understanding of the human heart and provides a valuable reference for future studies.


Assuntos
Miocárdio/citologia , Análise de Célula Única , Transcriptoma , Adipócitos/classificação , Adipócitos/metabolismo , Adulto , Enzima de Conversão de Angiotensina 2/análise , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/metabolismo , Células Epiteliais/classificação , Células Epiteliais/metabolismo , Epitélio , Feminino , Fibroblastos/classificação , Fibroblastos/metabolismo , Perfilação da Expressão Gênica , Estudo de Associação Genômica Ampla , Átrios do Coração/anatomia & histologia , Átrios do Coração/citologia , Átrios do Coração/inervação , Ventrículos do Coração/anatomia & histologia , Ventrículos do Coração/citologia , Ventrículos do Coração/inervação , Homeostase/imunologia , Humanos , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Músculo Esquelético/citologia , Músculo Esquelético/metabolismo , Miócitos Cardíacos/classificação , Miócitos Cardíacos/metabolismo , Neurônios/classificação , Neurônios/metabolismo , Pericitos/classificação , Pericitos/metabolismo , Receptores de Coronavírus/análise , Receptores de Coronavírus/genética , Receptores de Coronavírus/metabolismo , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidade , Células Estromais/classificação , Células Estromais/metabolismo
2.
Am J Hum Genet ; 109(5): 961-966, 2022 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-35397206

RESUMO

The well-established manifestation of mitochondrial mutations in functional cardiac disease (e.g., mitochondrial cardiomyopathy) prompted the hypothesis that mitochondrial DNA (mtDNA) sequence and/or copy number (mtDNAcn) variation contribute to cardiac defects in congenital heart disease (CHD). MtDNAcns were calculated and rare, non-synonymous mtDNA mutations were identified in 1,837 CHD-affected proband-parent trios, 116 CHD-affected singletons, and 114 paired cardiovascular tissue/blood samples. The variant allele fraction (VAF) of heteroplasmic variants in mitochondrial RNA from 257 CHD cardiovascular tissue samples was also calculated. On average, mtDNA from blood had 0.14 rare variants and 52.9 mtDNA copies per nuclear genome per proband. No variation with parental age at proband birth or CHD-affected proband age was seen. mtDNAcns in valve/vessel tissue (320 ± 70) were lower than in atrial tissue (1,080 ± 320, p = 6.8E-21), which were lower than in ventricle tissue (1,340 ± 280, p = 1.4E-4). The frequency of rare variants in CHD-affected individual DNA was indistinguishable from the frequency in an unaffected cohort, and proband mtDNAcns did not vary from those of CHD cohort parents. In both the CHD and the comparison cohorts, mtDNAcns were significantly correlated between mother-child, father-child, and mother-father. mtDNAcns among people with European (mean = 52.0), African (53.0), and Asian haplogroups (53.5) were calculated and were significantly different for European and Asian haplogroups (p = 2.6E-3). Variant heteroplasmic fraction (HF) in blood correlated well with paired cardiovascular tissue HF (r = 0.975) and RNA VAF (r = 0.953), which suggests blood HF is a reasonable proxy for HF in heart tissue. We conclude that mtDNA mutations and mtDNAcns are unlikely to contribute significantly to CHD risk.


Assuntos
DNA Mitocondrial , Cardiopatias Congênitas , Variações do Número de Cópias de DNA/genética , DNA Mitocondrial/genética , Cardiopatias Congênitas/genética , Humanos , Mitocôndrias/genética , Mutação/genética
3.
Proc Natl Acad Sci U S A ; 119(21): e2203928119, 2022 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-35584116

RESUMO

Microtia is a congenital malformation that encompasses mild hypoplasia to complete loss of the external ear, or pinna. Although the contribution of genetic variation and environmental factors to microtia remains elusive, Amerindigenous populations have the highest reported incidence. Here, using both transmission disequilibrium tests and association studies in microtia trios (parents and affected child) and microtia cohorts enrolled in Latin America, we map an ∼10-kb microtia locus (odds ratio = 4.7; P = 6.78e-18) to the intergenic region between Roundabout 1 (ROBO1) and Roundabout 2 (ROBO2) (chr3: 78546526 to 78555137). While alleles at the microtia locus significantly increase the risk of microtia, their penetrance is low (<1%). We demonstrate that the microtia locus contains a polymorphic complex repeat element that is expanded in affected individuals. The locus is located near a chromatin loop region that regulates ROBO1 and ROBO2 expression in induced pluripotent stem cell­derived neural crest cells. Furthermore, we use single nuclear RNA sequencing to demonstrate ROBO1 and ROBO2 expression in both fibroblasts and chondrocytes of the mature human pinna. Because the microtia allele is enriched in Amerindigenous populations and is shared by some East Asian subjects with craniofacial malformations, we propose that both populations share a mutation that arose in a common ancestor prior to the ancient migration of Eurasian populations into the Americas and that the high incidence of microtia among Amerindigenous populations reflects the population bottleneck that occurred during the migration out of Eurasia.


Assuntos
Indígena Americano ou Nativo do Alasca , Microtia Congênita , Microtia Congênita/genética , Orelha Externa , Efeito Fundador , Humanos , Mutação , Proteínas do Tecido Nervoso/genética , Receptores Imunológicos/genética , Indígena Americano ou Nativo do Alasca/genética , Proteínas Roundabout
4.
Proc Natl Acad Sci U S A ; 119(28): e2204174119, 2022 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-35787042

RESUMO

Myocardial fibrosis is a key pathologic feature of hypertrophic cardiomyopathy (HCM). However, the fibrotic pathways activated by HCM-causing sarcomere protein gene mutations are poorly defined. Because lysophosphatidic acid is a mediator of fibrosis in multiple organs and diseases, we tested the role of the lysophosphatidic acid pathway in HCM. Lysphosphatidic acid receptor 1 (LPAR1), a cell surface receptor, is required for lysophosphatidic acid mediation of fibrosis. We bred HCM mice carrying a pathogenic myosin heavy-chain variant (403+/-) with Lpar1-ablated mice to create mice carrying both genetic changes (403+/- LPAR1 -/-) and assessed development of cardiac hypertrophy and fibrosis. Compared with 403+/- LPAR1WT, 403+/- LPAR1 -/- mice developed significantly less hypertrophy and fibrosis. Single-nucleus RNA sequencing of left ventricular tissue demonstrated that Lpar1 was predominantly expressed by lymphatic endothelial cells (LECs) and cardiac fibroblasts. Lpar1 ablation reduced the population of LECs, confirmed by immunofluorescence staining of the LEC markers Lyve1 and Ccl21a and, by in situ hybridization, for Reln and Ccl21a. Lpar1 ablation also altered the distribution of fibroblast cell states. FB1 and FB2 fibroblasts decreased while FB0 and FB3 fibroblasts increased. Our findings indicate that Lpar1 is expressed predominantly by LECs and fibroblasts in the heart and is required for development of hypertrophy and fibrosis in an HCM mouse model. LPAR1 antagonism, including agents in clinical trials for other fibrotic diseases, may be beneficial for HCM.


Assuntos
Cardiomiopatia Hipertrófica , Receptores de Ácidos Lisofosfatídicos/genética , Animais , Cardiomiopatia Hipertrófica/genética , Cardiomiopatia Hipertrófica/patologia , Proteínas de Transporte , Modelos Animais de Doenças , Células Endoteliais/patologia , Fibrose , Hipertrofia/patologia , Camundongos
5.
Circulation ; 146(22): 1674-1693, 2022 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-36321451

RESUMO

BACKGROUND: ALPK3 encodes α-kinase 3, a muscle-specific protein of unknown function. ALPK3 loss-of-function variants cause cardiomyopathy with distinctive clinical manifestations in both children and adults, but the molecular functions of ALPK3 remain poorly understood. METHODS: We explored the putative kinase activity of ALPK3 and the consequences of damaging variants using isogenic human induced pluripotent stem cell-derived cardiomyocytes, mice, and human patient tissues. RESULTS: Multiple sequence alignment of all human α-kinase domains and their orthologs revealed 4 conserved residues that were variant only in ALPK3, demonstrating evolutionary divergence of the ALPK3 α-kinase domain sequence. Phosphoproteomic evaluation of both ALPK3 kinase domain inhibition and overexpression failed to detect significant changes in catalytic activity, establishing ALPK3 as a pseudokinase. Investigations into alternative functions revealed that ALPK3 colocalized with myomesin proteins (MYOM1, MYOM2) at both the nuclear envelope and the sarcomere M-band. ALPK3 loss-of-function variants caused myomesin proteins to mislocalize and also dysregulated several additional M-band proteins involved in sarcomere protein turnover, which ultimately impaired cardiomyocyte structure and function. CONCLUSIONS: ALPK3 is an essential cardiac pseudokinase that inserts in the nuclear envelope and the sarcomere M-band. Loss of ALPK3 causes mislocalization of myomesins, critical force-buffering proteins in cardiomyocytes, and also dysregulates M-band proteins necessary for sarcomere protein turnover. We conclude that ALPK3 cardiomyopathy induces ventricular dilatation caused by insufficient myomesin-mediated force buffering and hypertrophy by impairment of sarcomere proteostasis.


Assuntos
Cardiomiopatias , Células-Tronco Pluripotentes Induzidas , Proteínas Musculares , Proteínas Quinases , Adulto , Animais , Criança , Humanos , Camundongos , Cardiomiopatias/genética , Cardiomiopatias/metabolismo , Conectina/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Proteínas Musculares/genética , Miócitos Cardíacos/metabolismo , Sarcômeros/metabolismo , Proteínas Quinases/genética
6.
Genet Med ; 25(1): 143-150, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36260083

RESUMO

PURPOSE: Craniofacial microsomia (CFM) represents a spectrum of craniofacial malformations, ranging from isolated microtia with or without aural atresia to underdevelopment of the mandible, maxilla, orbit, facial soft tissue, and/or facial nerve. The genetic causes of CFM remain largely unknown. METHODS: We performed genome sequencing and linkage analysis in patients and families with microtia and CFM of unknown genetic etiology. The functional consequences of damaging missense variants were evaluated through expression of wild-type and mutant proteins in vitro. RESULTS: We studied a 5-generation kindred with microtia, identifying a missense variant in FOXI3 (p.Arg236Trp) as the cause of disease (logarithm of the odds = 3.33). We subsequently identified 6 individuals from 3 additional kindreds with microtia-CFM spectrum phenotypes harboring damaging variants in FOXI3, a regulator of ectodermal and neural crest development. Missense variants in the nuclear localization sequence were identified in cases with isolated microtia with aural atresia and found to affect subcellular localization of FOXI3. Loss of function variants were found in patients with microtia and mandibular hypoplasia (CFM), suggesting dosage sensitivity of FOXI3. CONCLUSION: Damaging variants in FOXI3 are the second most frequent genetic cause of CFM, causing 1% of all cases, including 13% of familial cases in our cohort.


Assuntos
Microtia Congênita , Síndrome de Goldenhar , Micrognatismo , Humanos , Síndrome de Goldenhar/genética , Microtia Congênita/genética , Orelha/anormalidades , Face
9.
Hum Mol Genet ; 28(10): 1682-1693, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30649309

RESUMO

Sex differences exist in the prevalence, presentation and outcomes of ischemic heart disease (IHD). Females have higher risk of heart failure post-myocardial infarction relative to males and are two to three times more likely to die after coronary artery bypass grafting surgery. We examined sex differences in human myocardial gene expression in response to ischemia. Left ventricular biopsies from 68 male/46 female patients undergoing aortic valve replacement surgery were obtained at baseline and after a median 74 min of cold cardioplegic arrest/ischemia. Transcriptomes were quantified by RNA-sequencing. Cell-type enrichment analysis was used to estimate the identity and relative proportions of different cell types in each sample. A sex-specific response to ischemia was observed for 271 genes. Notably, the expression FAM5C, PLA2G4E and CYP1A1 showed an increased expression in females compared to males due to ischemia and DIO3, MT1G and CMA1 showed a decreased expression in females compared to males due to ischemia. Functional annotation analysis revealed sex-specific modulation of the oxytocin signaling pathway and common pathway of fibrin clot formation. Expression quantitative trait locus (eQTL) analysis identified variant-by-sex interaction eQTLs, indicative of sex differences in the genotypic effects on gene expression. Cell-type enrichment analysis showed sex-bias in proportion of specific cell types. Common lymphoid progenitor cells and M2 macrophages were found to increase in female samples from pre- to post-ischemia, but no change was observed in male samples. These differences in response to myocardial ischemia provide insight into the sexual dimorphism of IHD and may aid in the development of sex-specific therapies that reduce myocardial injury.


Assuntos
Ventrículos do Coração/metabolismo , Isquemia Miocárdica/genética , Miocárdio/metabolismo , Caracteres Sexuais , Idoso , Idoso de 80 Anos ou mais , Valva Aórtica/fisiopatologia , Valva Aórtica/cirurgia , Procedimentos Cirúrgicos Cardíacos , Ponte de Artéria Coronária , Citocromo P-450 CYP1A1/genética , Proteínas de Ligação a DNA/genética , Feminino , Regulação da Expressão Gênica/genética , Fosfolipases A2 do Grupo IV/genética , Ventrículos do Coração/patologia , Humanos , Masculino , Isquemia Miocárdica/patologia , Isquemia Miocárdica/cirurgia , Miocárdio/patologia , Análise de Sequência de RNA
10.
Proc Natl Acad Sci U S A ; 115(35): E8143-E8152, 2018 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-30104387

RESUMO

Mutations in ß-cardiac myosin, the predominant motor protein for human heart contraction, can alter power output and cause cardiomyopathy. However, measurements of the intrinsic force, velocity, and ATPase activity of myosin have not provided a consistent mechanism to link mutations to muscle pathology. An alternative model posits that mutations in myosin affect the stability of a sequestered, super relaxed state (SRX) of the protein with very slow ATP hydrolysis and thereby change the number of myosin heads accessible to actin. Here we show that purified human ß-cardiac myosin exists partly in an SRX and may in part correspond to a folded-back conformation of myosin heads observed in muscle fibers around the thick filament backbone. Mutations that cause hypertrophic cardiomyopathy destabilize this state, while the small molecule mavacamten promotes it. These findings provide a biochemical and structural link between the genetics and physiology of cardiomyopathy with implications for therapeutic strategies.


Assuntos
Benzilaminas/química , Uracila/análogos & derivados , Miosinas Ventriculares/química , Animais , Benzilaminas/farmacologia , Cardiomegalia/enzimologia , Cardiomegalia/genética , Humanos , Músculo Esquelético/enzimologia , Mutação , Suínos , Porco Miniatura , Uracila/química , Uracila/farmacologia , Miosinas Ventriculares/genética , Miosinas Ventriculares/metabolismo
11.
Circulation ; 140(1): 31-41, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-30987448

RESUMO

BACKGROUND: Cancer therapy-induced cardiomyopathy (CCM) is associated with cumulative drug exposures and preexisting cardiovascular disorders. These parameters incompletely account for substantial interindividual susceptibility to CCM. We hypothesized that rare variants in cardiomyopathy genes contribute to CCM. METHODS: We studied 213 patients with CCM from 3 cohorts: retrospectively recruited adults with diverse cancers (n=99), prospectively phenotyped adults with breast cancer (n=73), and prospectively phenotyped children with acute myeloid leukemia (n=41). Cardiomyopathy genes, including 9 prespecified genes, were sequenced. The prevalence of rare variants was compared between CCM cohorts and The Cancer Genome Atlas participants (n=2053), healthy volunteers (n=445), and an ancestry-matched reference population. Clinical characteristics and outcomes were assessed and stratified by genotypes. A prevalent CCM genotype was modeled in anthracycline-treated mice. RESULTS: CCM was diagnosed 0.4 to 9 years after chemotherapy; 90% of these patients received anthracyclines. Adult patients with CCM had cardiovascular risk factors similar to the US population. Among 9 prioritized genes, patients with CCM had more rare protein-altering variants than comparative cohorts ( P≤1.98e-04). Titin-truncating variants (TTNtvs) predominated, occurring in 7.5% of patients with CCM versus 1.1% of The Cancer Genome Atlas participants ( P=7.36e-08), 0.7% of healthy volunteers ( P=3.42e-06), and 0.6% of the reference population ( P=5.87e-14). Adult patients who had CCM with TTNtvs experienced more heart failure and atrial fibrillation ( P=0.003) and impaired myocardial recovery ( P=0.03) than those without. Consistent with human data, anthracycline-treated TTNtv mice and isolated TTNtv cardiomyocytes showed sustained contractile dysfunction unlike wild-type ( P=0.0004 and P<0.002, respectively). CONCLUSIONS: Unrecognized rare variants in cardiomyopathy-associated genes, particularly TTNtvs, increased the risk for CCM in children and adults, and adverse cardiac events in adults. Genotype, along with cumulative chemotherapy dosage and traditional cardiovascular risk factors, improves the identification of patients who have cancer at highest risk for CCM. CLINICAL TRIAL REGISTRATION: URL: https://www.clinicaltrials.gov . Unique identifiers: NCT01173341; AAML1031; NCT01371981.


Assuntos
Antineoplásicos/efeitos adversos , Cardiomiopatias/induzido quimicamente , Cardiomiopatias/genética , Variação Genética/genética , Neoplasias/tratamento farmacológico , Neoplasias/genética , Adulto , Idoso , Animais , Cardiomiopatias/epidemiologia , Estudos de Coortes , Feminino , Variação Genética/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pessoa de Meia-Idade , Neoplasias/epidemiologia , Estudos Prospectivos , Estudos Retrospectivos
12.
Proc Natl Acad Sci U S A ; 114(43): E9096-E9104, 2017 10 24.
Artigo em Inglês | MEDLINE | ID: mdl-29073106

RESUMO

Comparative analyses of transcriptional profiles from humans and mice with cardiovascular pathologies revealed consistently elevated expression of MICU2, a regulatory subunit of the mitochondrial calcium uniporter complex. To determine if MICU2 expression was cardioprotective, we produced and characterized Micu2-/- mice. Mutant mice had left atrial enlargement and Micu2-/- cardiomyocytes had delayed sarcomere relaxation and cytosolic calcium reuptake kinetics, indicating diastolic dysfunction. RNA sequencing (RNA-seq) of Micu2-/- ventricular tissues revealed markedly reduced transcripts encoding the apelin receptor (Micu2-/- vs. wild type, P = 7.8 × 10-40), which suppresses angiotensin II receptor signaling via allosteric transinhibition. We found that Micu2-/- and wild-type mice had comparable basal blood pressures and elevated responses to angiotensin II infusion, but that Micu2-/- mice exhibited systolic dysfunction and 30% lethality from abdominal aortic rupture. Aneurysms and rupture did not occur with norepinephrine-induced hypertension. Aortic tissue from Micu2-/- mice had increased expression of extracellular matrix remodeling genes, while single-cell RNA-seq analyses showed increased expression of genes related to reactive oxygen species, inflammation, and proliferation in fibroblast and smooth muscle cells. We concluded that Micu2-/- mice recapitulate features of diastolic heart disease and define previously unappreciated roles for Micu2 in regulating angiotensin II-mediated hypertensive responses that are critical in protecting the abdominal aorta from injury.


Assuntos
Canais de Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Cálcio/metabolismo , Cardiomiopatia Hipertrófica Familiar/genética , Angiotensina Amida/genética , Angiotensina II/farmacologia , Animais , Aorta Abdominal/patologia , Canais de Cálcio/genética , Proteínas de Ligação ao Cálcio/genética , Cardiomiopatia Hipertrófica Familiar/patologia , Eletrocardiografia , Regulação da Expressão Gênica , Homeostase/efeitos dos fármacos , Homeostase/fisiologia , Humanos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Mitocôndrias Hepáticas/fisiologia , Miócitos Cardíacos/patologia , Miócitos Cardíacos/fisiologia
13.
Proc Natl Acad Sci U S A ; 114(29): 7689-7694, 2017 07 18.
Artigo em Inglês | MEDLINE | ID: mdl-28679633

RESUMO

Genetic variants that cause haploinsufficiency account for many autosomal dominant (AD) disorders. Gene-based diagnosis classifies variants that alter canonical splice signals as pathogenic, but due to imperfect understanding of RNA splice signals other variants that may create or eliminate splice sites are often clinically classified as variants of unknown significance (VUS). To improve recognition of pathogenic splice-altering variants in AD disorders, we used computational tools to prioritize VUS and developed a cell-based minigene splicing assay to confirm aberrant splicing. Using this two-step procedure we evaluated all rare variants in two AD cardiomyopathy genes, lamin A/C (LMNA) and myosin binding protein C (MYBPC3). We demonstrate that 13 LMNA and 35 MYBPC3 variants identified in cardiomyopathy patients alter RNA splicing, representing a 50% increase in the numbers of established damaging splice variants in these genes. Over half of these variants are annotated as VUS by clinical diagnostic laboratories. Familial analyses of one variant, a synonymous LMNA VUS, demonstrated segregation with cardiomyopathy affection status and altered cardiac LMNA splicing. Application of this strategy should improve diagnostic accuracy and variant classification in other haploinsufficient AD disorders.


Assuntos
Proteínas de Transporte/genética , Lamina Tipo A/genética , Mutação , Splicing de RNA , Adulto , Idoso , Alelos , Cardiomiopatias/genética , Biologia Computacional , Feminino , Variação Genética , Genótipo , Células HEK293 , Haploinsuficiência , Cardiopatias/genética , Transplante de Coração , Humanos , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Marca-Passo Artificial , Linhagem , Sítios de Splice de RNA , Análise de Sequência de DNA , Adulto Jovem
14.
N Engl J Med ; 375(18): 1749-1755, 2016 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-27806233

RESUMO

Immune checkpoint inhibitors have improved clinical outcomes associated with numerous cancers, but high-grade, immune-related adverse events can occur, particularly with combination immunotherapy. We report the cases of two patients with melanoma in whom fatal myocarditis developed after treatment with ipilimumab and nivolumab. In both patients, there was development of myositis with rhabdomyolysis, early progressive and refractory cardiac electrical instability, and myocarditis with a robust presence of T-cell and macrophage infiltrates. Selective clonal T-cell populations infiltrating the myocardium were identical to those present in tumors and skeletal muscle. Pharmacovigilance studies show that myocarditis occurred in 0.27% of patients treated with a combination of ipilimumab and nivolumab, which suggests that our patients were having a rare, potentially fatal, T-cell-driven drug reaction. (Funded by Vanderbilt-Ingram Cancer Center Ambassadors and others.).


Assuntos
Anticorpos Monoclonais/efeitos adversos , Imunoterapia/efeitos adversos , Miocardite/etiologia , Miocárdio/patologia , Idoso , Anticorpos Monoclonais/uso terapêutico , Arritmias Cardíacas/induzido quimicamente , Eletrocardiografia/efeitos dos fármacos , Evolução Fatal , Feminino , Glucocorticoides/uso terapêutico , Bloqueio Cardíaco/diagnóstico , Bloqueio Cardíaco/etiologia , Humanos , Ipilimumab , Masculino , Melanoma/complicações , Melanoma/tratamento farmacológico , Pessoa de Meia-Idade , Miocardite/tratamento farmacológico , Miocardite/patologia , Miosite/induzido quimicamente , Nivolumabe
15.
Hum Mutat ; 39(6): 870-881, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29527824

RESUMO

Multiple tools have been developed to identify copy number variants (CNVs) from whole exome (WES) and whole genome sequencing (WGS) data. Current tools such as XHMM for WES and CNVnator for WGS identify CNVs based on changes in read depth. For WGS, other methods to identify CNVs include utilizing discordant read pairs and split reads and genome-wide local assembly with tools such as Lumpy and SvABA, respectively. Here, we introduce a new method to identify deletion CNVs from WES and WGS trio data based on the clustering of Mendelian errors (MEs). Using our Mendelian Error Method (MEM), we identified 127 deletions (inherited and de novo) in 2,601 WES trios from the Pediatric Cardiac Genomics Consortium, with a validation rate of 88% by digital droplet PCR. MEM identified additional de novo deletions compared with XHMM, and a significant enrichment of 15q11.2 deletions compared with controls. In addition, MEM identified eight cases of uniparental disomy, sample switches, and DNA contamination. We applied MEM to WGS data from the Genome In A Bottle Ashkenazi trio and identified deletions with 97% specificity. MEM provides a robust, computationally inexpensive method for identifying deletions, and an orthogonal approach for verifying deletions called by other tools.


Assuntos
Variações do Número de Cópias de DNA/genética , Análise Mutacional de DNA/métodos , Genoma Humano/genética , Deleção de Sequência/genética , Mapeamento Cromossômico , Exoma/genética , Feminino , Cardiopatias Congênitas/genética , Humanos , Masculino , Sequenciamento do Exoma , Sequenciamento Completo do Genoma
16.
Hum Genet ; 137(2): 183-193, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29417219

RESUMO

Mosaicism due to somatic mutations can cause multiple diseases including cancer, developmental and overgrowth syndromes, neurodevelopmental disorders, autoinflammatory diseases, and atrial fibrillation. With the increased use of next generation sequencing technology, multiple tools have been developed to identify low-frequency variants, specifically from matched tumor-normal tissues in cancer studies. To investigate whether mosaic variants are implicated in congenital heart disease (CHD), we developed a pipeline using the cancer somatic variant caller MuTect to identify mosaic variants in whole-exome sequencing (WES) data from a cohort of parent/affected child trios (n = 715) and a cohort of healthy individuals (n = 416). This is a novel application of the somatic variant caller designed for cancer to WES trio data. We identified two cases with mosaic KMT2D mutations that are likely pathogenic for CHD, but conclude that, overall, mosaicism detectable in peripheral blood or saliva does not account for a significant portion of CHD etiology.


Assuntos
Sequenciamento do Exoma , Variação Genética , Cardiopatias Congênitas/genética , Mosaicismo , Criança , Exoma/genética , Cardiopatias Congênitas/fisiopatologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mutação , Software
17.
Proc Natl Acad Sci U S A ; 112(29): 9046-51, 2015 Jul 21.
Artigo em Inglês | MEDLINE | ID: mdl-26153423

RESUMO

Homozygous cardiac myosin binding protein C-deficient (Mybpc(t/t)) mice develop dramatic cardiac dilation shortly after birth; heart size increases almost twofold. We have investigated the mechanism of cardiac enlargement in these hearts. Throughout embryogenesis myocytes undergo cell division while maintaining the capacity to pump blood by rapidly disassembling and reforming myofibrillar components of the sarcomere throughout cell cycle progression. Shortly after birth, myocyte cell division ceases. Cardiac MYBPC is a thick filament protein that regulates sarcomere organization and rigidity. We demonstrate that many Mybpc(t/t) myocytes undergo an additional round of cell division within 10 d postbirth compared with their wild-type counterparts, leading to increased numbers of mononuclear myocytes. Short-hairpin RNA knockdown of Mybpc3 mRNA in wild-type mice similarly extended the postnatal window of myocyte proliferation. However, adult Mybpc(t/t) myocytes are unable to fully regenerate the myocardium after injury. MYBPC has unexpected inhibitory functions during postnatal myocyte cytokinesis and cell cycle progression. We suggest that human patients with homozygous MYBPC3-null mutations develop dilated cardiomyopathy, coupled with myocyte hyperplasia (increased cell number), as observed in Mybpc(t/t) mice. Human patients, with heterozygous truncating MYBPC3 mutations, like mice with similar mutations, have hypertrophic cardiomyopathy. However, the mechanism leading to hypertrophic cardiomyopathy in heterozygous MYBPC3(+/-) individuals is myocyte hypertrophy (increased cell size), whereas the mechanism leading to cardiac dilation in homozygous Mybpc3(-/-) mice is primarily myocyte hyperplasia.


Assuntos
Proteínas de Transporte/metabolismo , Citocinese , Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Animais Recém-Nascidos , Aurora Quinases/metabolismo , Biomarcadores/metabolismo , Cálcio/metabolismo , Contagem de Células , Diferenciação Celular , Proliferação de Células , Dependovirus/metabolismo , Células Endoteliais/metabolismo , Regulação da Expressão Gênica , Ventrículos do Coração/metabolismo , Histonas/metabolismo , Humanos , Indóis/metabolismo , Camundongos , Modelos Biológicos , Miocárdio/citologia , Miócitos Cardíacos/citologia , Fosforilação , RNA Interferente Pequeno/metabolismo
18.
Cardiovasc Diabetol ; 16(1): 87, 2017 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-28687077

RESUMO

BACKGROUND: Omentin-1, also known as Intelectin-1 (ITLN1), is an adipokine with plasma levels associated with diabetes, obesity, and coronary artery disease. Recent studies suggest that ITLN1 can mitigate myocardial ischemic injury but the expression of ITLN1 in the heart itself has not been well characterized. The purpose of this study is to discern the relationship between the expression pattern of ITLN1 RNA in the human heart and the level of circulating ITLN1 protein in plasma from the same patients following myocardial ischemia. METHODS: A large cohort of patients (n = 140) undergoing elective cardiac surgery for aortic valve replacement were enrolled in this study. Plasma and left ventricular biopsy samples were taken at the beginning of cardiopulmonary bypass and after an average of 82 min of ischemic cross clamp time. The localization of ITLN1 in epicardial adipose tissue (EAT) was also further characterized with immunoassays and cell fate transition studies. RESULTS: mRNA expression of ITLN1 decreases in left ventricular tissue after acute ischemia in human patients (mean difference 280.48, p = 0.001) whereas plasma protein levels of ITLN1 increase (mean difference 5.24, p < 0.001). Immunohistochemistry localized ITLN1 to the mesothelium or visceral pericardium of EAT. Epithelial to mesenchymal transition in mesothelial cells leads to a downregulation of ITLN1 expression. CONCLUSIONS: Myocardial injury leads to a decrease in ITLN1 expression in the heart and a corresponding increase in plasma levels. These changes may in part be due to an epithelial to mesenchymal transition of the cells that express ITLN1 following ischemia. Trial Registration Clinicaltrials.gov ID: NCT00985049.


Assuntos
Doença da Artéria Coronariana/metabolismo , Citocinas/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Lectinas/metabolismo , Isquemia Miocárdica/metabolismo , Pericárdio/metabolismo , Adipocinas/metabolismo , Tecido Adiposo/metabolismo , Idoso , Idoso de 80 Anos ou mais , Valva Aórtica/metabolismo , Feminino , Proteínas Ligadas por GPI/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade
19.
Hum Mol Genet ; 23(1): 209-25, 2014 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-23975679

RESUMO

Recent human genetic studies have provided evidences that sporadic or inherited missense mutations in four-and-a-half LIM domain protein 1 (FHL1), resulting in alterations in FHL1 protein expression, are associated with rare congenital myopathies, including reducing body myopathy and Emery-Dreifuss muscular dystrophy. However, it remains to be clarified whether mutations in FHL1 cause skeletal muscle remodeling owing to gain- or loss of FHL1 function. In this study, we used FHL1-null mice lacking global FHL1 expression to evaluate loss-of-function effects on skeletal muscle homeostasis. Histological and functional analyses of soleus, tibialis anterior and sternohyoideus muscles demonstrated that FHL1-null mice develop an age-dependent myopathy associated with myofibrillar and intermyofibrillar (mitochondrial and sarcoplasmic reticulum) disorganization, impaired muscle oxidative capacity and increased autophagic activity. A longitudinal study established decreased survival rates in FHL1-null mice, associated with age-dependent impairment of muscle contractile function and a significantly lower exercise capacity. Analysis of primary myoblasts isolated from FHL1-null muscles demonstrated early muscle fiber differentiation and maturation defects, which could be rescued by re-expression of the FHL1A isoform, highlighting that FHL1A is necessary for proper muscle fiber differentiation and maturation in vitro. Overall, our data show that loss of FHL1 function leads to myopathy in vivo and suggest that loss of function of FHL1 may be one of the mechanisms underlying muscle dystrophy in patients with FHL1 mutations.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/patologia , Distrofias Musculares/patologia , Miofibrilas/patologia , Fatores Etários , Animais , Diferenciação Celular , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Atividade Motora , Músculo Esquelético/metabolismo , Distrofias Musculares/genética , Distrofia Muscular de Emery-Dreifuss/patologia , Mioblastos Esqueléticos/metabolismo , Mioblastos Esqueléticos/patologia , Miofibrilas/metabolismo
20.
J Mol Cell Cardiol ; 79: 234-43, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25463273

RESUMO

Mutations in MYBPC3, the gene encoding cardiac myosin binding protein-C (cMyBP-C), account for ~40% of hypertrophic cardiomyopathy (HCM) cases. Most pathological MYBPC3 mutations encode truncated protein products not found in tissue. Reduced protein levels occur in symptomatic heterozygous human HCM carriers, suggesting haploinsufficiency as an underlying mechanism of disease. However, we do not know if reduced cMyBP-C content results from, or initiates the development of HCM. In previous studies, heterozygous (HET) mice with a MYBPC3 C'-terminal truncation mutation and normal cMyBP-C levels show altered contractile function prior to any overt hypertrophy. Therefore, this study aimed to test whether haploinsufficiency occurs, with decreased cMyBP-C content, following cardiac stress and whether the functional impairment in HET MYBPC3 hearts leads to worsened disease progression. To address these questions, transverse aortic constriction (TAC) was performed on three-month-old wild-type (WT) and HET MYBPC3-truncation mutant mice and then characterized at 4 and 12weeks post-surgery. HET-TAC mice showed increased hypertrophy and reduced ejection fraction compared to WT-TAC mice. At 4weeks post-surgery, HET myofilaments showed significantly reduced cMyBP-C content. Functionally, HET-TAC cardiomyocytes showed impaired force generation, higher Ca(2+) sensitivity, and blunted length-dependent increase in force generation. RNA sequencing revealed several differentially regulated genes between HET and WT groups, including regulators of remodeling and hypertrophic response. Collectively, these results demonstrate that haploinsufficiency occurs in HET MYBPC3 mutant carriers following stress, causing, in turn, reduced cMyBP-C content and exacerbating the development of dysfunction at myofilament and whole-heart levels.


Assuntos
Cardiomiopatia Hipertrófica/genética , Cardiomiopatia Hipertrófica/patologia , Proteínas de Transporte/metabolismo , Progressão da Doença , Haploinsuficiência/genética , Alelos , Animais , Aorta/metabolismo , Aorta/patologia , Cardiomiopatia Hipertrófica/fisiopatologia , Cardiomiopatia Hipertrófica/cirurgia , Constrição Patológica , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Heterozigoto , Humanos , Camundongos , Mutação/genética , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Pressão , Análise de Sequência de RNA , Sístole
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