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1.
Am J Hum Genet ; 104(3): 530-541, 2019 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-30827496

RESUMO

Acetylation of the lysine residues in histones and other DNA-binding proteins plays a major role in regulation of eukaryotic gene expression. This process is controlled by histone acetyltransferases (HATs/KATs) found in multiprotein complexes that are recruited to chromatin by the scaffolding subunit transformation/transcription domain-associated protein (TRRAP). TRRAP is evolutionarily conserved and is among the top five genes intolerant to missense variation. Through an international collaboration, 17 distinct de novo or apparently de novo variants were identified in TRRAP in 24 individuals. A strong genotype-phenotype correlation was observed with two distinct clinical spectra. The first is a complex, multi-systemic syndrome associated with various malformations of the brain, heart, kidneys, and genitourinary system and characterized by a wide range of intellectual functioning; a number of affected individuals have intellectual disability (ID) and markedly impaired basic life functions. Individuals with this phenotype had missense variants clustering around the c.3127G>A p.(Ala1043Thr) variant identified in five individuals. The second spectrum manifested with autism spectrum disorder (ASD) and/or ID and epilepsy. Facial dysmorphism was seen in both groups and included upslanted palpebral fissures, epicanthus, telecanthus, a wide nasal bridge and ridge, a broad and smooth philtrum, and a thin upper lip. RNA sequencing analysis of skin fibroblasts derived from affected individuals skin fibroblasts showed significant changes in the expression of several genes implicated in neuronal function and ion transport. Thus, we describe here the clinical spectrum associated with TRRAP pathogenic missense variants, and we suggest a genotype-phenotype correlation useful for clinical evaluation of the pathogenicity of the variants.

2.
J Pediatr Genet ; 7(4): 164-173, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30430034

RESUMO

Wolf-Hirschhorn syndrome (WHS) is caused by partial deletion of the short arm of chromosome 4 and is characterized by dysmorphic facies, congenital heart defects, intellectual/developmental disability, and increased risk for congenital diaphragmatic hernia (CDH). In this report, we describe a stillborn girl with WHS and a large CDH. A literature review revealed 15 cases of WHS with CDH, which overlap a 2.3-Mb CDH critical region. We applied a machine-learning algorithm that integrates large-scale genomic knowledge to genes within the 4p16.3 CDH critical region and identified FGFRL1 , CTBP1 , NSD2 , FGFR3 , CPLX1 , MAEA , CTBP1-AS2 , and ZNF141 as genes whose haploinsufficiency may contribute to the development of CDH.

3.
Dis Model Mech ; 11(9)2018 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-30061196

RESUMO

Deletions of chromosome 1p36 are associated with a high incidence of congenital heart defects (CHDs). The arginine-glutamic acid dipeptide repeats gene (RERE) is located in a critical region for CHD on chromosome 1p36 and encodes a cardiac-expressed nuclear receptor co-regulator. Mutations affecting RERE cause atrial and ventricular septal defects (VSDs) in humans, and RERE-deficient mice also develop VSDs. During cardiac development, mesenchymal cells destined to form part of the atrioventricular (AV) septum are generated when endocardial cells in the AV canal undergo epithelial-to-mesenchymal transition (EMT) and migrate into the space between the endocardium and the myocardium. These newly generated mesenchymal cells then proliferate to fill the developing AV endocardial cushions. Here, we demonstrate that RERE-deficient mouse embryos have reduced numbers of mesenchymal cells in their AV endocardial cushions owing to decreased levels of EMT and mesenchymal cell proliferation. In the endocardium, RERE colocalizes with GATA4, a transcription factor required for normal levels of EMT and mesenchymal cell proliferation. Using a combination of in vivo and in vitro studies, we show that Rere and Gata4 interact genetically in the development of CHDs, RERE positively regulates transcription from the Gata4 promoter and GATA4 levels are reduced in the AV canals of RERE-deficient embryos. Tissue-specific ablation of Rere in the endocardium leads to hypocellularity of the AV endocardial cushions, defective EMT and VSDs, but does not result in decreased GATA4 expression. We conclude that RERE functions in the AV canal to positively regulate the expression of GATA4, and that deficiency of RERE leads to the development of VSDs through its effects on EMT and mesenchymal cell proliferation. However, the cell-autonomous role of RERE in promoting EMT in the endocardium must be mediated by its effects on the expression of proteins other than GATA4.This article has an associated First Person interview with the first author of the paper.

4.
Hum Mol Genet ; 27(12): 2064-2075, 2018 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-29618029

RESUMO

Congenital diaphragmatic hernia (CDH) has been reported twice in individuals with a clinical diagnosis of Fraser syndrome, a genetic disorder that can be caused by recessive mutations affecting FREM2 and FRAS1. In the extracellular matrix, FREM2 and FRAS1 form a self-stabilizing complex with FREM1, a protein whose deficiency causes sac CDH in humans and mice. By sequencing FREM2 and FRAS1 in a CDH cohort, and searching online databases, we identified five individuals who carried recessive or double heterozygous, putatively deleterious variants in these genes which may represent susceptibility alleles. Three of these alleles were significantly enriched in our CDH cohort compared with ethnically matched controls. We subsequently demonstrated that 8% of Frem2ne/ne and 1% of Fras1Q1263*/Q1263* mice develop the same type of anterior sac CDH seen in FREM1-deficient mice. We went on to show that development of sac hernias in FREM1-deficient mice is preceded by failure of anterior mesothelial fold progression resulting in the persistence of an amuscular, poorly vascularized anterior diaphragm that is abnormally adherent to the underlying liver. Herniation occurs in the perinatal period when the expanding liver protrudes through this amuscular region of the anterior diaphragm that is juxtaposed to areas of muscular diaphragm. Based on these data, we conclude that deficiency of FREM2, and possibly FRAS1, are associated with an increased risk of developing CDH and that loss of the FREM1/FREM2/FRAS1 complex, or its function, leads to anterior sac CDH development through its effects on mesothelial fold progression.

5.
J Med Genet ; 54(1): 47-53, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27550220

RESUMO

BACKGROUND: The non-POU domain containing octamer-binding gene (NONO) is located on chromosome Xq13.1 and encodes a member of a small family of RNA-binding and DNA-binding proteins that perform a variety of tasks involved in RNA synthesis, transcriptional regulation and DNA repair. Loss-of-function variants in NONO have been described as a cause of intellectual disability in males but have not been described in association with congenital heart defects or cardiomyopathy. In this article, we seek to further define the phenotypic consequences of NONO depletion in human subjects. METHODS: We searched a clinical database of over 6000 individuals referred for exome sequencing and over 60 000 individuals referred for CNV analysis. RESULTS: We identified two males with atrial and ventricular septal defects, left ventricular non-compaction (LVNC), developmental delay and intellectual disability, who harboured de novo, loss-of-function variants in NONO. We also identified a male infant with developmental delay, congenital brain anomalies and severe LVNC requiring cardiac transplantation, who inherited a single-gene deletion of NONO from his asymptomatic mother. CONCLUSIONS: We conclude that in addition to global developmental delay and intellectual disability, males with loss-of-function variants in NONO may also be predisposed to developing congenital heart defects and LVNC with the penetrance of these cardiac-related problems being influenced by genetic, epigenetic, environmental or stochastic factors. Brain imaging of males with NONO deficiency may reveal structural defects with abnormalities of the corpus callosum being the most common. Although dysmorphic features vary between affected individuals, relative macrocephaly is a common feature.


Assuntos
Cardiopatias Congênitas/genética , Ventrículos do Coração/patologia , Proteínas Associadas à Matriz Nuclear/genética , Fatores de Transcrição de Octâmero/genética , Proteínas de Ligação a RNA/genética , Criança , Pré-Escolar , Proteínas de Ligação a DNA/genética , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/patologia , Exoma/genética , Cardiopatias Congênitas/patologia , Humanos , Lactente , Masculino
6.
Am J Hum Genet ; 98(5): 963-970, 2016 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-27087320

RESUMO

Deletions of chromosome 1p36 affect approximately 1 in 5,000 newborns and are associated with developmental delay, intellectual disability, and defects involving the brain, eye, ear, heart, and kidney. Arginine-glutamic acid dipeptide repeats (RERE) is located in the proximal 1p36 critical region. RERE is a widely-expressed nuclear receptor coregulator that positively regulates retinoic acid signaling. Animal models suggest that RERE deficiency might contribute to many of the structural and developmental birth defects and medical problems seen in individuals with 1p36 deletion syndrome, although human evidence supporting this role has been lacking. In this report, we describe ten individuals with intellectual disability, developmental delay, and/or autism spectrum disorder who carry rare and putatively damaging changes in RERE. In all cases in which both parental DNA samples were available, these changes were found to be de novo. Associated features that were recurrently seen in these individuals included hypotonia, seizures, behavioral problems, structural CNS anomalies, ophthalmologic anomalies, congenital heart defects, and genitourinary abnormalities. The spectrum of defects documented in these individuals is similar to that of a cohort of 31 individuals with isolated 1p36 deletions that include RERE and are recapitulated in RERE-deficient zebrafish and mice. Taken together, our findings suggest that mutations in RERE cause a genetic syndrome and that haploinsufficiency of RERE might be sufficient to cause many of the phenotypes associated with proximal 1p36 deletions.


Assuntos
Anormalidades Múltiplas/etiologia , Proteínas de Transporte/genética , Transtornos Cromossômicos/etiologia , Deficiências do Desenvolvimento/etiologia , Haploinsuficiência/genética , Mutação/genética , Animais , Criança , Pré-Escolar , Deleção Cromossômica , Cromossomos Humanos Par 1 , Feminino , Humanos , Lactente , Masculino , Camundongos , Fenótipo , Prognóstico
7.
Am J Med Genet A ; 167A(4): 831-6, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25736269

RESUMO

Congenital diaphragmatic hernia (CDH) is a relatively common, life--threatening birth defect. We present a family with recurrent CDH--paraesophageal and central--for whom exome sequencing (ES) revealed a frameshift mutation (c.4969_4970insA, p.Ile1657Asnfs*30) in the fibrillin 1 gene (FBN1) that causes Marfan syndrome. A diagnosis of Marfan syndrome had not been considered previously in this family. However, a review of the literature demonstrated that FBN1 mutations have an unusual pattern of CDH in which paraesophageal hernias are particularly common. Subsequent clinical evaluations revealed evidence for ectopia lentis in affected family members supporting a clinical diagnosis of Marfan syndrome. Since only two other cases of familial CDH have been described in association with FBN1 mutations, we investigated an oligogenic hypothesis by examining ES data for deleterious sequence changes in other CDH-related genes. This search revealed putatively deleterious sequence changes in four other genes that have been shown to cause diaphragm defects in humans and/or mice--FREM1, DES, PAX3 and MET. It is unclear whether these changes, alone or in aggregate, are contributing to the development of CDH in this family. However, their individual contribution is likely to be small compared to that of the frameshift mutation in FBN1. We conclude that ES can be used to identify both major and minor genetic factors that may contribute to CDH. These results also suggest that ES should be considered in the diagnostic evaluation of individuals and families with CDH, particularly when other diagnostic modalities have failed to reveal a molecular etiology.


Assuntos
Hérnias Diafragmáticas Congênitas/diagnóstico , Síndrome de Marfan/diagnóstico , Proteínas dos Microfilamentos/genética , Adulto , Pré-Escolar , Análise Mutacional de DNA , Exoma , Feminino , Fibrilina-1 , Fibrilinas , Mutação da Fase de Leitura , Estudos de Associação Genética , Hérnias Diafragmáticas Congênitas/genética , Humanos , Masculino , Síndrome de Marfan/genética , Linhagem
8.
Am J Physiol Gastrointest Liver Physiol ; 307(11): G1073-87, 2014 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-25301185

RESUMO

Extracellular nucleotides via activation of P2 purinergic receptors influence hepatocyte proliferation and liver regeneration in response to 70% partial hepatectomy (PH). Adult hepatocytes express multiple P2Y (G protein-coupled) and P2X (ligand-gated ion channels) purinergic receptor subtypes. However, the identity of key receptor subtype(s) important for efficient hepatocyte proliferation in regenerating livers remains unknown. To evaluate the impact of P2Y2 purinergic receptor-mediated signaling on hepatocyte proliferation in regenerating livers, wild-type (WT) and P2Y2 purinergic receptor knockout (P2Y2-/-) mice were subjected to 70% PH. Liver tissues were analyzed for activation of early events critical for hepatocyte priming and subsequent cell cycle progression. Our findings suggest that early activation of p42/44 ERK MAPK (5 min), early growth response-1 (Egr-1) and activator protein-1 (AP-1) DNA-binding activity (30 min), and subsequent hepatocyte proliferation (24-72 h) in response to 70% PH were impaired in P2Y2-/- mice. Interestingly, early induction of cytokines (TNF-α, IL-6) and cytokine-mediated signaling (NF-κB, STAT-3) were intact in P2Y2-/- remnant livers, uncovering the importance of cytokine-independent and nucleotide-dependent early priming events critical for subsequent hepatocyte proliferation in regenerating livers. Hepatocytes isolated from the WT and P2Y2-/- mice were treated with ATP or ATPγS for 5-120 min and 12-24 h. Extracellular ATP alone, via activation of P2Y2 purinergic receptors, was sufficient to induce ERK phosphorylation, Egr-1 protein expression, and key cyclins and cell cycle progression of hepatocytes in vitro. Collectively, these findings highlight the functional significance of P2Y2 purinergic receptor activation for efficient hepatocyte priming and proliferation in response to PH.


Assuntos
Hepatectomia , Hepatócitos/efeitos dos fármacos , Agonistas do Receptor Purinérgico P2Y/farmacologia , Receptores Purinérgicos P2Y2/efeitos dos fármacos , Trifosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/farmacologia , Animais , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ciclinas/farmacologia , Proteína 1 de Resposta de Crescimento Precoce/biossíntese , Proteína 1 de Resposta de Crescimento Precoce/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores Purinérgicos P2Y2/genética
9.
PLoS One ; 9(1): e85600, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24454898

RESUMO

Cardiovascular malformations and cardiomyopathy are among the most common phenotypes caused by deletions of chromosome 1p36 which affect approximately 1 in 5000 newborns. Although these cardiac-related abnormalities are a significant source of morbidity and mortality associated with 1p36 deletions, most of the individual genes that contribute to these conditions have yet to be identified. In this paper, we use a combination of clinical and molecular cytogenetic data to define five critical regions for cardiovascular malformations and two critical regions for cardiomyopathy on chromosome 1p36. Positional candidate genes which may contribute to the development of cardiovascular malformations associated with 1p36 deletions include DVL1, SKI, RERE, PDPN, SPEN, CLCNKA, ECE1, HSPG2, LUZP1, and WASF2. Similarly, haploinsufficiency of PRDM16-a gene which was recently shown to be sufficient to cause the left ventricular noncompaction-SKI, PRKCZ, RERE, UBE4B and MASP2 may contribute to the development of cardiomyopathy. When treating individuals with 1p36 deletions, or providing prognostic information to their families, physicians should take into account that 1p36 deletions which overlie these cardiac critical regions may portend to cardiovascular complications. Since several of these cardiac critical regions contain more than one positional candidate gene-and large terminal and interstitial 1p36 deletions often overlap more than one cardiac critical region-it is likely that haploinsufficiency of two or more genes contributes to the cardiac phenotypes associated with many 1p36 deletions.


Assuntos
Anormalidades Cardiovasculares/genética , Doenças Cardiovasculares/genética , Cromossomos Humanos Par 1 , Deleção de Genes , Humanos
10.
PLoS One ; 8(2): e57460, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23451234

RESUMO

Individuals with terminal and interstitial deletions of chromosome 1p36 have a spectrum of defects that includes eye anomalies, postnatal growth deficiency, structural brain anomalies, seizures, cognitive impairment, delayed motor development, behavior problems, hearing loss, cardiovascular malformations, cardiomyopathy, and renal anomalies. The proximal 1p36 genes that contribute to these defects have not been clearly delineated. The arginine-glutamic acid dipeptide (RE) repeats gene (RERE) is located in this region and encodes a nuclear receptor coregulator that plays a critical role in embryonic development as a positive regulator of retinoic acid signaling. Rere-null mice die of cardiac failure between E9.5 and E11.5. This limits their usefulness in studying the role of RERE in the latter stages of development and into adulthood. To overcome this limitation, we created an allelic series of RERE-deficient mice using an Rere-null allele, om, and a novel hypomorphic Rere allele, eyes3 (c.578T>C, p.Val193Ala), which we identified in an N-ethyl-N-nitrosourea (ENU)-based screen for autosomal recessive phenotypes. Analyses of these mice revealed microphthalmia, postnatal growth deficiency, brain hypoplasia, decreased numbers of neuronal nuclear antigen (NeuN)-positive hippocampal neurons, hearing loss, cardiovascular malformations-aortic arch anomalies, double outlet right ventricle, and transposition of the great arteries, and perimembranous ventricular septal defects-spontaneous development of cardiac fibrosis and renal agenesis. These findings suggest that RERE plays a critical role in the development and function of multiple organs including the eye, brain, inner ear, heart and kidney. It follows that haploinsufficiency of RERE may contribute-alone or in conjunction with other genetic, environmental, or stochastic factors-to the development of many of the phenotypes seen in individuals with terminal and interstitial deletions that include the proximal region of chromosome 1p36.


Assuntos
Anormalidades Múltiplas/genética , Deleção Cromossômica , Transtornos Cromossômicos/genética , Desenvolvimento Embrionário/genética , Proteínas do Tecido Nervoso/genética , Proteínas Repressoras/genética , Alelos , Animais , Peso Corporal/efeitos dos fármacos , Peso Corporal/genética , Doenças Cardiovasculares/genética , Cromossomos/efeitos dos fármacos , Cromossomos/genética , Cromossomos Humanos Par 1/genética , Desenvolvimento Embrionário/efeitos dos fármacos , Etilnitrosoureia , Perda Auditiva/genética , Hipocampo/efeitos dos fármacos , Hipocampo/embriologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Fenótipo
11.
PLoS One ; 8(3): e58830, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23536828

RESUMO

The FRAS1-related extracellular matrix 1 (FREM1) gene encodes an extracellular matrix protein that plays a critical role in the development of multiple organ systems. In humans, recessive mutations in FREM1 cause eye defects, congenital diaphragmatic hernia, renal anomalies and anorectal malformations including anteriorly placed anus. A similar constellation of findings-microphthalmia, cryptophthalmos, congenital diaphragmatic hernia, renal agenesis and rectal prolapse-have been described in FREM1-deficient mice. In this paper, we identify a homozygous Frem1 missense mutation (c.1687A>T, p.Ile563Phe) in an N-ethyl-N-nitrosourea (ENU)-derived mouse strain, crf11, with microphthalmia, cryptophthalmos, renal agenesis and rectal prolapse. This mutation affects a highly conserved residue in FREM1's third CSPG domain. The p.Ile563Phe change is predicted to be deleterious and to cause decreased FREM1 protein stability. The crf11 allele also fails to complement the previously described eyes2 allele of Frem1 (p.Lys826*) providing further evidence that the crf11 phenotype is due to changes affecting Frem1 function. We then use mice bearing the crf11 and eyes2 alleles to identify lung lobulation defects and decreased anogenital distance in males as novel phenotypes associated with FREM1 deficiency in mice. Due to phenotypic overlaps between FREM1-deficient mice and mice that are deficient for the retinoic acid-responsive transcription factor GATA4 and the extracellular matrix protein SLIT3, we also perform experiments to look for in vivo genetic interactions between the genes that encode these proteins. These experiments reveal that Frem1 interacts genetically with Gata4 in the development of lung lobulation defects and with Slit3 in the development of renal agenesis. These results demonstrate that FREM1-deficient mice faithfully recapitulate many of the phenotypes seen in individuals with FREM1 deficiency and that variations in GATA4 and SLIT3 expression modulate some FREM1-related phenotypes in mice.


Assuntos
Epistasia Genética , Proteínas da Matriz Extracelular/genética , Fator de Transcrição GATA4/genética , Proteínas de Membrana/genética , Fenótipo , Anormalidades Múltiplas/genética , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Anormalidades Congênitas/genética , Proteínas da Matriz Extracelular/química , Proteínas da Matriz Extracelular/deficiência , Proteínas da Matriz Extracelular/metabolismo , Fator de Transcrição GATA4/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Estudos de Associação Genética , Haploinsuficiência , Homozigoto , Rim/anormalidades , Nefropatias/congênito , Nefropatias/genética , Pulmão/embriologia , Pulmão/metabolismo , Pulmão/patologia , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Alinhamento de Sequência
12.
Hum Mol Genet ; 21(18): 4115-25, 2012 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-22723016

RESUMO

Recurrent microdeletions of 8p23.1 that include GATA4 and SOX7 confer a high risk of both congenital diaphragmatic hernia (CDH) and cardiac defects. Although GATA4-deficient mice have both CDH and cardiac defects, no humans with cardiac defects attributed to GATA4 mutations have been reported to have CDH. We were also unable to identify deleterious GATA4 sequence changes in a CDH cohort. This suggested that haploinsufficiency of another 8p23.1 gene may contribute, along with GATA4, to the development of CDH. To determine if haploinsufficiency of SOX7-another transcription factor encoding gene-contributes to the development of CDH, we generated mice with a deletion of the second exon of Sox7. A portion of these Sox7(Δex2/+) mice developed retrosternal diaphragmatic hernias located in the anterior muscular portion of the diaphragm. Anterior CDH is also seen in Gata4(+/-) mice and has been described in association with 8p23.1 deletions in humans. Immunohistochemistry revealed that SOX7 is expressed in the vascular endothelial cells of the developing diaphragm and may be weakly expressed in some diaphragmatic muscle cells. Sox7(Δex2/Δex2) embryos die prior to diaphragm development with dilated pericardial sacs and failure of yolk sac remodeling suggestive of cardiovascular failure. Similar to our experience screening GATA4, no clearly deleterious SOX7 sequence changes were identified in our CDH cohort. We conclude that haploinsufficiency of Sox7 or Gata4 is sufficient to produce anterior CDH in mice and that haploinsufficiency of SOX7 and GATA4 may each contribute to the development of CDH in individuals with 8p23.1 deletions.


Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 8/genética , Hérnias Diafragmáticas Congênitas , Fatores de Transcrição SOXF/genética , Animais , Sequência de Bases , Análise Mutacional de DNA , Diafragma/metabolismo , Diafragma/patologia , Modelos Animais de Doenças , Feminino , Fator de Transcrição GATA4/genética , Fator de Transcrição GATA4/metabolismo , Genes Letais , Estudos de Associação Genética , Haploinsuficiência , Hérnia Diafragmática/genética , Hérnia Diafragmática/patologia , Humanos , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fatores de Transcrição SOXF/deficiência
14.
J Hepatol ; 52(1): 54-62, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19914731

RESUMO

BACKGROUND & AIMS: Paracrine interactions are critical to liver physiology, particularly during regeneration, although physiological involvement of extracellular ATP, a crucial intercellular messenger, remains unclear. The physiological release of ATP into extracellular milieu and its impact on regeneration after partial hepatectomy were investigated in this study. METHODS: Hepatic ATP release after hepatectomy was examined in the rat and in human living donors for liver transplantation. Quinacrine was used for in vivo staining of ATP-enriched compartments in rat liver sections and isolated hepatocytes. Rats were treated with an antagonist for purinergic receptors (Phosphate-6-azo(benzene-2,4-disulfonic acid), PPADS), and liver regeneration after hepatectomy was analyzed. RESULTS: A robust and transient ATP release due to acute portal hyperpressure was observed immediately after hepatectomy in rats and humans. Clodronate liposomal pre-treatment partly inhibited ATP release in rats. Quinacrine-stained vesicles, co-labeled with a lysosomal marker in liver sections and isolated hepatocytes, were predominantly detected in periportal areas. These vesicles significantly disappeared after hepatectomy, in parallel with a decrease in liver ATP content. PPADS treatment inhibited hepatocyte cell cycle progression after hepatectomy, as revealed by a reduction in bromodeoxyuridine incorporation, phosphorylated histone 3 immunostaining, cyclin D1 and A expression and immediate early gene induction. CONCLUSION: Extracellular ATP is released immediately after hepatectomy from hepatocytes and Kupffer cells under mechanical stress and promotes liver regeneration in the rat. We suggest that in hepatocytes, ATP is released from a lysosomal compartment. Finally, observations made in living donors suggest that purinergic signalling could be critical for human liver regeneration.


Assuntos
Trifosfato de Adenosina/metabolismo , Hepatectomia/métodos , Regeneração Hepática/fisiologia , Fígado/metabolismo , Fígado/cirurgia , Adulto , Animais , Matriz Extracelular/metabolismo , Feminino , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Macrófagos do Fígado/citologia , Macrófagos do Fígado/efeitos dos fármacos , Macrófagos do Fígado/metabolismo , Transplante de Fígado , Lisossomos/metabolismo , Masculino , Modelos Animais , Antagonistas do Receptor Purinérgico P2 , Fosfato de Piridoxal/análogos & derivados , Fosfato de Piridoxal/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores Purinérgicos P2/metabolismo , Estresse Mecânico , Doadores de Tecidos
15.
Reprod Toxicol ; 19(2): 149-54, 2004 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15501379

RESUMO

Serum lead, cadmium and zinc levels from 31 newborns with neural tube defects (NTD), and 54 healthy controls living in a polluted area in Mexico were estimated using atomic absorption spectrophotometry (AAS). NTD family history was found to be of greater importance in the case group (OR 6.95, 95% CI 1.51-36.3, p=0.002). In 25% of the children, serum lead concentrations were above the admissible maximum level (AML) of 10 microg/dL within 24 h of extra-uterine life. Cadmium concentrations were below the AML. Zinc deficiency was found in nine (29%) of the cases and four (9.3%) of the controls (p=0.04). The logistic regression multivariate analysis showed no correlation between NTD and high levels of any of these metals; however, a positive correlation was found to zinc deficiency (OR 5.0, 95% CI 1.07-23.00, p=0.04). These results focus attention to the surrounding nutritional and maternal health factors of major importance in disease etiology.


Assuntos
Cádmio/sangue , Poluição Ambiental , Chumbo/sangue , Defeitos do Tubo Neural/sangue , Zinco/sangue , Estudos de Casos e Controles , Feminino , Humanos , Recém-Nascido , Modelos Logísticos , Masculino , México/epidemiologia , Defeitos do Tubo Neural/epidemiologia , Defeitos do Tubo Neural/etiologia , Espectrofotometria Atômica
16.
Mol Ther ; 6(3): 342-8, 2002 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-12231170

RESUMO

Immune responses against adenoviral vectors may influence the toxicity and therapeutic effectiveness of adenovirus-mediated gene transfer and may be a limiting factor in adenovirus-mediated gene therapy. The purpose of this study was to determine the effects of preimmunization on intratumoral adenoviral transduction and systemic spread. The hypothesis was that increased doses of adenoviral vectors could overcome local neutralization without added systemic toxicity. The level and duration of gene expression were assessed as a function of time and dose after intratumoral delivery of adenoviral vector (AdV) encoding the luciferase reporter gene (AdV-luc) in a subcutaneous mouse mammary tumor model. Preimmunization resulted in significantly decreased gene expression in tumor and normal tissues (P < 0.01). The decrease was significantly greater in liver than in tumor. Increased AdV doses could be used to overcome the intratumoral inhibition without a concomitant increase in liver transduction. However, preimmunized animals showed greater toxicity than nai;ve animals (P < 0.001). The preimmunized group developed histologic evidence of grade 2-3 hepatic toxicity and increases in the average values of hepatic enzymes. In addition, there was a significant increase in mortality (P < 0.01) in the preimmunized group (12 of 20 animals) compared with the naive group (3 of 20 animals). These findings suggest that although preimmunity can inhibit systemic expression from adenoviral vectors, at high vector doses it may potentiate hepatotoxicity.


Assuntos
Dependovirus/genética , Terapia Genética , Vetores Genéticos/imunologia , Neoplasias Mamárias Experimentais/terapia , Vacinação , Vacinas Virais , Animais , Dependovirus/efeitos dos fármacos , Dependovirus/imunologia , Dependovirus/patogenicidade , Genes Reporter , Vetores Genéticos/administração & dosagem , Vetores Genéticos/efeitos dos fármacos , Vetores Genéticos/farmacologia , Vetores Genéticos/toxicidade , Hepatite/patologia , Imunidade , Fígado/efeitos dos fármacos , Fígado/enzimologia , Fígado/patologia , Neoplasias Mamárias Experimentais/imunologia , Camundongos , Vacinas Virais/administração & dosagem , Vacinas Virais/imunologia
17.
Rev Invest Clin ; 54(1): 57-67, 2002 Jan-Feb.
Artigo em Espanhol | MEDLINE | ID: mdl-11995408

RESUMO

Gene therapy is a new modality of treatment in which a gene is used to modify or add new biochemical properties to a patient's target cells with therapeutics purposes. Currently, this experimental therapy is under intensive development as an alternative to treat cancer, because it is possible that this therapy may generate a higher antineoplastic activity, more tissue selectivity and less contralateral effects than conventional therapy. After a decade of preclinical and clinical assays, still there are several obstacles that impose limits to the antineoplastic efficacy of this therapy. However, with the advances in molecular biology and related fields, there is a promise to improve, expand and strength the powerful antineoplastic arsenal of gene therapy.


Assuntos
Terapia Genética , Neoplasias/terapia , Ensaios Clínicos como Assunto , Vetores Genéticos , Humanos , Imunoterapia , Neoplasias/genética , Transdução Genética
18.
Rev. invest. clín ; 54(1): 57-67, 2002 Jan-Feb.
Artigo em Espanhol | LILACS | ID: lil-332947

RESUMO

Gene therapy is a new modality of treatment in which a gene is used to modify or add new biochemical properties to a patient's target cells with therapeutics purposes. Currently, this experimental therapy is under intensive development as an alternative to treat cancer, because it is possible that this therapy may generate a higher antineoplastic activity, more tissue selectivity and less contralateral effects than conventional therapy. After a decade of preclinical and clinical assays, still there are several obstacles that impose limits to the antineoplastic efficacy of this therapy. However, with the advances in molecular biology and related fields, there is a promise to improve, expand and strength the powerful antineoplastic arsenal of gene therapy.


Assuntos
Humanos , Terapia Genética , Neoplasias , Ensaios Clínicos como Assunto , Vetores Genéticos , Imunoterapia , Neoplasias , Transdução Genética
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