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2.
Genome Med ; 11(1): 12, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30819258

RESUMO

BACKGROUND: Neurodevelopmental disorders are genetically and phenotypically heterogeneous encompassing developmental delay (DD), intellectual disability (ID), autism spectrum disorders (ASDs), structural brain abnormalities, and neurological manifestations with variants in a large number of genes (hundreds) associated. To date, a few de novo mutations potentially disrupting TCF20 function in patients with ID, ASD, and hypotonia have been reported. TCF20 encodes a transcriptional co-regulator structurally related to RAI1, the dosage-sensitive gene responsible for Smith-Magenis syndrome (deletion/haploinsufficiency) and Potocki-Lupski syndrome (duplication/triplosensitivity). METHODS: Genome-wide analyses by exome sequencing (ES) and chromosomal microarray analysis (CMA) identified individuals with heterozygous, likely damaging, loss-of-function alleles in TCF20. We implemented further molecular and clinical analyses to determine the inheritance of the pathogenic variant alleles and studied the spectrum of phenotypes. RESULTS: We report 25 unique inactivating single nucleotide variants/indels (1 missense, 1 canonical splice-site variant, 18 frameshift, and 5 nonsense) and 4 deletions of TCF20. The pathogenic variants were detected in 32 patients and 4 affected parents from 31 unrelated families. Among cases with available parental samples, the variants were de novo in 20 instances and inherited from 4 symptomatic parents in 5, including in one set of monozygotic twins. Two pathogenic loss-of-function variants were recurrent in unrelated families. Patients presented with a phenotype characterized by developmental delay, intellectual disability, hypotonia, variable dysmorphic features, movement disorders, and sleep disturbances. CONCLUSIONS: TCF20 pathogenic variants are associated with a novel syndrome manifesting clinical characteristics similar to those observed in Smith-Magenis syndrome. Together with previously described cases, the clinical entity of TCF20-associated neurodevelopmental disorders (TAND) emerges from a genotype-driven perspective.


Assuntos
Anormalidades Craniofaciais/genética , Deficiências do Desenvolvimento/genética , Mutação INDEL , Deficiência Intelectual/genética , Hipotonia Muscular/genética , Síndrome de Smith-Magenis/genética , Fatores de Transcrição/genética , Adolescente , Criança , Pré-Escolar , Anormalidades Craniofaciais/patologia , Deficiências do Desenvolvimento/patologia , Feminino , Humanos , Lactente , Deficiência Intelectual/patologia , Masculino , Hipotonia Muscular/patologia , Síndrome de Smith-Magenis/patologia , Fatores de Transcrição/metabolismo , Adulto Jovem
3.
Am J Obstet Gynecol ; 213(2): 214.e1-5, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25843063

RESUMO

OBJECTIVE: We sought to determine the positive predictive value (PPV) of noninvasive prenatal screening (NIPS) for various aneuploidies based on cases referred for follow-up cytogenetic testing. Secondarily, we wanted to determine the false-negative (FN) rate for those cases with a negative NIPS result. STUDY DESIGN: We compared the cytogenetic findings (primarily from chromosome analysis) from 216 cases referred to our laboratories with either a positive or negative NIPS result, and classified NIPS results as true positive, false positive, true negative, or FN. Diagnostic cytogenetic testing was performed on the following tissue types: amniotic fluid (n = 137), chorionic villi (n = 69), neonatal blood (n = 6), and products of conception (n = 4). RESULTS: The PPV for NIPS were as follows: 93% for trisomy (T)21 (n = 99; 95% confidence interval [CI], 86-97.1%), 58% for T18 (n = 24; 95% CI, 36.6-77.9%), 45% for T13 (n = 11; 95% CI, 16.7-76.6%), 23% for monosomy X (n = 26; 95% CI, 9-43.6%), and 67% for XXY (n = 6; 95% CI, 22.3-95.7%). Of the 26 cases referred for follow-up cytogenetics after a negative NIPS result, 1 (4%) was FN (T13). Two cases of triploidy, a very serious condition but one not claimed to be detectable by the test providers, were among those classified as true negatives. CONCLUSION: T21, which has the highest prevalence of all aneuploidies, demonstrated a high true-positive rate, resulting in a high PPV. However, the other aneuploidies, with their lower prevalence, displayed relatively high false-positive rates and, therefore, lower PPV. Patients and physicians must fully understand the limitations of this screening test and the need in many cases to follow up with appropriate diagnostic testing to obtain an accurate diagnosis.


Assuntos
Aberrações Cromossômicas , Transtornos Cromossômicos/diagnóstico , DNA/sangue , Adulto , Amniocentese , Aneuploidia , Amostra da Vilosidade Coriônica , Transtornos Cromossômicos/genética , Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 18/genética , Estudos de Coortes , Análise Citogenética , Síndrome de Down/diagnóstico , Síndrome de Down/genética , Reações Falso-Negativas , Feminino , Humanos , Síndrome de Klinefelter/diagnóstico , Síndrome de Klinefelter/genética , Valor Preditivo dos Testes , Gravidez , Diagnóstico Pré-Natal , Estudos Retrospectivos , Trissomia/diagnóstico , Trissomia/genética , Síndrome da Trissomia do Cromossomo 13 , Síndrome da Trissomía do Cromossomo 18 , Síndrome de Turner/diagnóstico , Síndrome de Turner/genética
4.
Genet Med ; 17(8): 623-9, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25356966

RESUMO

PURPOSE: Detection of copy-number variation (CNV) is important for investigating many genetic disorders. Testing a large clinical cohort by array comparative genomic hybridization provides a deep perspective on the spectrum of pathogenic CNV. In this context, we describe a bioinformatics approach to extract CNV information from whole-exome sequencing and demonstrate its utility in clinical testing. METHODS: Exon-focused arrays and whole-genome chromosomal microarray analysis were used to test 14,228 and 14,000 individuals, respectively. Based on these results, we developed an algorithm to detect deletions/duplications in whole-exome sequencing data and a novel whole-exome array. RESULTS: In the exon array cohort, we observed a positive detection rate of 2.4% (25 duplications, 318 deletions), of which 39% involved one or two exons. Chromosomal microarray analysis identified 3,345 CNVs affecting single genes (18%). We demonstrate that our whole-exome sequencing algorithm resolves CNVs of three or more exons. CONCLUSION: These results demonstrate the clinical utility of single-exon resolution in CNV assays. Our whole-exome sequencing algorithm approaches this resolution but is complemented by a whole-exome array to unambiguously identify intragenic CNVs and single-exon changes. These data illustrate the next advancements in CNV analysis through whole-exome sequencing and whole-exome array.Genet Med 17 8, 623-629.


Assuntos
Hibridização Genômica Comparativa/métodos , Biologia Computacional/métodos , Variações do Número de Cópias de DNA , Exoma , Algoritmos , Estudos de Coortes , DNA/análise , DNA/sangue , DNA/genética , Variação Genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos
5.
Cancer Genet Cytogenet ; 190(2): 125-30, 2009 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-19380032

RESUMO

We describe the cases of two unrelated patients who exhibited multiple chromosomal abnormalities in donor cells after allogeneic peripheral blood stem cell transplantation (PBSCT). The patients were diagnosed with chronic myeloid leukemia and chronic lymphocytic leukemia, respectively, and both underwent nonmyeloablative conditioning with cyclophosphamide and fludarabine followed by PBSCT from their HLA-matched opposite-sex siblings. Post-transplant bone marrow cytogenetics showed full engraftment, and the early post-transplant studies demonstrated only normal donor metaphases. Subsequent studies of both patients, however, revealed a population of metaphase cells with abnormal, but apparently balanced, donor karyotypes. Chromosome studies performed on peripheral blood cells collected from both donors after transplantation were normal. Both patients remained in clinical remission during follow-up of approximately 8 years in one case, and 6 years in the other case, despite the persistence of the abnormal clones. Chromosomal abnormalities in residual recipient cells after bone marrow or PBSCT are not unusual. In contrast, only rare reports of chromosome abnormalities in donor cells exist, all of which have been associated with post-bone marrow transplant myelodysplastic syndrome or acute leukemias. The present cases demonstrate the rare phenomenon of persistent clonal nonpathogenic chromosome aberrations in cells of donor origin.


Assuntos
Aberrações Cromossômicas , Transplante de Células-Tronco Hematopoéticas , Idoso , Feminino , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/imunologia , Masculino , Pessoa de Meia-Idade
6.
Clin Cancer Res ; 14(2): 388-95, 2008 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-18223213

RESUMO

PURPOSE: Since the identification of PRKAR1A mutations in Carney complex, substitutions and small insertions/deletions have been found in approximately 70% of the patients. To date, no germ-line PRKAR1A deletion and/or insertion exceeded a few base pairs (up to 15). Although a few families map to chromosome 2, it is possible that current sequencing techniques do not detect larger gene changes in PRKAR1A -- mutation-negative individuals with Carney complex. EXPERIMENTAL DESIGN: To screen for gross alterations of the PRKAR1A gene, we applied Southern hybridization analysis on 36 unrelated Carney complex patients who did not have small intragenic mutations or large aberrations in PRKAR1A, including the probands from two kindreds mapping to chromosome 2. RESULTS: We found large PRKAR1A deletions in the germ-line of two patients with Carney complex, both sporadic cases; no changes were identified in the remaining patients, including the two chromosome-2-mapping families. In the first patient, the deletion is expected to lead to decreased PRKAR1A mRNA levels but no other effects on the protein; the molecular phenotype is predicted to be PRKAR1A haploinsufficiency, consistent with the majority of PRKAR1A mutations causing Carney complex. In the second patient, the deletion led to in-frame elimination of exon 3 and the expression of a shorter protein, lacking the primary site for interaction with the catalytic protein kinase A subunit. In vitro transfection studies of the mutant PRKAR1A showed impaired ability to bind cyclic AMP and activation of the protein kinase A enzyme. The patient bearing this mutation had a more-severe-than-average Carney complex phenotype that included the relatively rare psammomatous melanotic schwannoma. CONCLUSIONS: Large PRKAR1A deletions may be responsible for Carney complex in patients that do not have PRKAR1A gene defects identifiable by sequencing. Preliminary data indicate that these patients may have a different phenotype especially if their defect results in an expressed, abnormal version of the PRKAR1A protein.


Assuntos
Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/genética , Deleção de Genes , Neoplasia Endócrina Múltipla/genética , Linhagem Celular , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Éxons , Humanos
7.
Eur J Hum Genet ; 16(1): 79-88, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17667967

RESUMO

Gastrointestinal stromal tumors (GISTs) may be caused by germline mutations of the KIT and platelet-derived growth factor receptor-alpha (PDGFRA) genes and treated by Imatinib mesylate (STI571) or other protein tyrosine kinase inhibitors. However, not all GISTs harbor these genetic defects and several do not respond to STI571 suggesting that other molecular mechanisms may be implicated in GIST pathogenesis. In a subset of patients with GISTs, the lesions are associated with paragangliomas; the condition is familial and transmitted as an autosomal-dominant trait. We investigated 11 patients with the dyad of 'paraganglioma and gastric stromal sarcoma'; in eight (from seven unrelated families), the GISTs were caused by germline mutations of the genes encoding subunits B, C, or D (the SDHB, SDHC and SDHD genes, respectively). In this report, we present the molecular effects of these mutations on these genes and the clinical information on the patients. We conclude that succinate dehydrogenase deficiency may be the cause of a subgroup of GISTs and this offers a therapeutic target for GISTs that may not respond to STI571 and its analogs.


Assuntos
Tumores do Estroma Gastrointestinal/enzimologia , Tumores do Estroma Gastrointestinal/genética , Mutação em Linhagem Germinativa , Proteínas com Ferro-Enxofre/genética , Proteínas de Membrana/genética , Síndromes Neoplásicas Hereditárias/enzimologia , Síndromes Neoplásicas Hereditárias/genética , Paraganglioma/enzimologia , Paraganglioma/genética , Succinato Desidrogenase/genética , Adolescente , Adulto , Alelos , Antineoplásicos/uso terapêutico , Sequência de Bases , Benzamidas , Criança , Primers do DNA/genética , DNA de Neoplasias/genética , Feminino , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/patologia , Genes Dominantes , Heterozigoto , Humanos , Mesilato de Imatinib , Perda de Heterozigosidade , Masculino , Síndromes Neoplásicas Hereditárias/tratamento farmacológico , Síndromes Neoplásicas Hereditárias/patologia , Paraganglioma/tratamento farmacológico , Paraganglioma/patologia , Piperazinas/uso terapêutico , Pirimidinas/uso terapêutico
8.
J Clin Endocrinol Metab ; 93(2): 565-71, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18056771

RESUMO

CONTEXT: Inactivating mutations of PRKAR1A, the regulatory subunit type 1A (RIalpha) of protein kinase A (PKA), are associated with tumor formation. OBJECTIVE: Our objective was to evaluate the role of PKA isozymes on proliferation and cell cycle. METHODS: A cell line with RIalpha haploinsufficiency due to an inactivating PRKAR1A mutation (IVS2+1 G-->A) was transfected with constructs encoding PKA subunits. Genetics, PKA subunit mRNA and protein expression and proliferation, aneuploidy, and cell cycle status were assessed. To identify factors that mediate PKA-associated cell cycle changes, we studied E2F and cyclins expression in transfected cells and E2F's role by small interfering RNA; we also assessed cAMP levels and baseline and stimulated cAMP signaling in transfected cells. RESULTS: Introduction of PKA subunits led to changes in proliferation and cell cycle: a decrease in aneuploidy and G(2)/M for the PRKAR1A-transfected cells and an increase in S phase and aneuploidy for cells transfected with PRKAR2B, a known PRKAR1A mutant (RIalphaP), and the PKA catalytic subunit. There were alterations in cAMP levels, PKA subunit expression, cyclins, and E2F factors; E2F1 was shown to possibly mediate PKA effects on cell cycle by small interfering RNA studies. cAMP levels and constitutive and stimulated cAMP signaling were altered in transfected cells. CONCLUSION: This is the first immortalized cell line with a naturally occurring PRKAR1A-inactivating mutation that is associated in vivo with tumor formation. PKA isozyme balance is critical for the control of cAMP signaling and related cell cycle and proliferation changes. Finally, E2F1 may be a factor that mediates dysregulated PKA's effects on the cell cycle.


Assuntos
Doenças do Córtex Suprarrenal/enzimologia , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Mutação Puntual , Doenças do Córtex Suprarrenal/genética , Neoplasias do Córtex Suprarrenal/enzimologia , Neoplasias do Córtex Suprarrenal/genética , Apoptose/fisiologia , Western Blotting , Ciclo Celular/fisiologia , Processos de Crescimento Celular/fisiologia , Linhagem Celular , AMP Cíclico/metabolismo , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/biossíntese , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/genética , Ciclinas/metabolismo , DNA/química , DNA/genética , Fatores de Transcrição E2F/metabolismo , Feminino , Humanos , Isoenzimas , Ploidias , Análise de Sequência de DNA , Transfecção
9.
J Clin Endocrinol Metab ; 92(9): 3728-32, 2007 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-17566086

RESUMO

CONTEXT: Gastrointestinal stromal tumors (GISTs) may be caused by somatic or germline mutations of the KIT and PDGFRA genes, but most GISTs associated with neuroendocrine tumors (NETs) are not, suggesting that other molecular pathways are implicated in their pathogenesis. OBJECTIVE: In the course of investigating NETs and GIST genetics, we encountered a patient who had a unique combination of multiple fibrous polyps and lipomas of the small intestine and several gastric GISTs. DESIGN: The study included the clinical description of a unique patient, DNA sequencing of germline and tumor DNA, and comparative genomic hybridization (CGH) and allelic marker analysis of tumor DNA. RESULTS: The patient was found to carry a germline PDGFRA mutation (V561D) in the heterozygote state; it has only been seen rarely before and only in the somatic state in sporadic GISTs. CGH identified losses of chromosomal regions 1p33-36, 9q12-24, 11q13, and 16q; loss of the 14q region that is commonly lost in NETs and GISTs was shown by DNA marker analysis. These changes are likely to point to secondary and tertiary genetic hits involved in the formation of these rare tumors. CONCLUSIONS: Multiple GISTs and other tumors may be caused by germline PDGFRA gene mutations; the V561D mutation can occur in the germline state and lead to a syndrome that should not be confused with other genetic conditions associated with a predisposition to NETs and other tumors. A number of chromosomal loci are likely to be involved in the PDGFRA V561D-dependent tumorigenesis, as shown by CGH and other DNA analyses.


Assuntos
Tumores do Estroma Gastrointestinal/genética , Mutação em Linhagem Germinativa , Neoplasias Primárias Múltiplas/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Ácido Aspártico/genética , Sequência de Bases , Análise Mutacional de DNA , Feminino , Humanos , Pessoa de Meia-Idade , Valina/genética
10.
J Clin Endocrinol Metab ; 92(8): 2938-43, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17535989

RESUMO

CONTEXT: Carney triad (CT) describes the association of paragangliomas (PGLs) with gastrointestinal stromal tumors (GISTs) and pulmonary chondromas. Inactivating mutations of the mitochondrial complex II succinate dehydrogenase (SDH) enzyme subunits SDHB, SDHC, and SDHD are found in PGLs, gain-of-function mutations of c-kit (KIT), and platelet-derived growth factor receptor A (PDGFRA) in GISTs. OBJECTIVE: Our objective was to investigate the possibility that patients with CT and/or their tumors may harbor mutations of the SDHB, SDHC, SDHD, KIT, and PDGFRA genes and identify any other genetic alterations in CT tumors. DESIGN: Three males and 34 females with CT were studied retrospectively. We sequenced the stated genes and performed comparative genomic hybridization on a total of 41 tumors. RESULTS: No patient had coding sequence mutations of the investigated genes. Comparative genomic hybridization revealed a number of DNA copy number changes: losses dominated among benign lesions, there were an equal number of gains and losses in malignant lesions, and the average number of alterations in malignant tumors was higher compared with benign lesions. The most frequent and greatest contiguous change was 1q12-q21 deletion, a region that harbors the SDHC gene. Another frequent change was loss of 1p. Allelic losses of 1p and 1q were confirmed by fluorescent in situ hybridization and loss-of-heterozygosity studies. CONCLUSIONS: We conclude that CT is not due to SDH-inactivating or KIT- and PDGFRA-activating mutations. GISTs and PGLs in CT are associated with chromosome 1 and other changes that appear to participate in tumor progression and point to their common genetic cause.


Assuntos
Cromossomos Humanos Par 1/genética , Neoplasias Gastrointestinais/genética , Tumores do Estroma Gastrointestinal/genética , Mutação em Linhagem Germinativa/genética , Neoplasias Pulmonares/genética , Paraganglioma/genética , Adolescente , Adulto , Criança , DNA de Neoplasias/genética , Feminino , Dosagem de Genes , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Perda de Heterozigosidade , Masculino , Hibridização de Ácido Nucleico , Estudos Retrospectivos , Succinato Desidrogenase/genética , Células Tumorais Cultivadas
11.
Nat Genet ; 38(7): 794-800, 2006 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-16767104

RESUMO

Phosphodiesterases (PDEs) regulate cyclic nucleotide levels. Increased cyclic AMP (cAMP) signaling has been associated with PRKAR1A or GNAS mutations and leads to adrenocortical tumors and Cushing syndrome. We investigated the genetic source of Cushing syndrome in individuals with adrenocortical hyperplasia that was not caused by known defects. We performed genome-wide SNP genotyping, including the adrenocortical tumor DNA. The region with the highest probability to harbor a susceptibility gene by loss of heterozygosity (LOH) and other analyses was 2q31-2q35. We identified mutations disrupting the expression of the PDE11A isoform-4 gene (PDE11A) in three kindreds. Tumor tissues showed 2q31-2q35 LOH, decreased protein expression and high cyclic nucleotide levels and cAMP-responsive element binding protein (CREB) phosphorylation. PDE11A codes for a dual-specificity PDE that is expressed in adrenal cortex and is partially inhibited by tadalafil and other PDE inhibitors; its germline inactivation is associated with adrenocortical hyperplasia, suggesting another means by which dysregulation of cAMP signaling causes endocrine tumors.


Assuntos
Glândulas Suprarrenais/enzimologia , Glândulas Suprarrenais/patologia , Mutação , Diester Fosfórico Hidrolases/genética , Adulto , Criança , Cromossomos Humanos Par 2/genética , Síndrome de Cushing/enzimologia , Síndrome de Cushing/genética , Síndrome de Cushing/patologia , Feminino , Humanos , Hiperplasia , Perda de Heterozigosidade , Masculino , Diester Fosfórico Hidrolases/metabolismo , Polimorfismo de Nucleotídeo Único
12.
J Clin Endocrinol Metab ; 91(9): 3626-32, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16772351

RESUMO

CONTEXT: Primary adrenocortical hyperplasias leading to Cushing syndrome include primary pigmented nodular adrenocortical disease and ACTH-independent macronodular adrenal hyperplasia (AIMAH). Inactivating mutations of the 17q22-24-located PRKAR1A gene, coding for the type 1A regulatory subunit of protein kinase A (PKA), cause primary pigmented nodular adrenocortical disease and the multiple endocrine neoplasia syndrome Carney complex. PRKAR1A mutations and 17q22-24 chromosomal losses have been found in sporadic adrenal tumors and are associated with aberrant PKA signaling. OBJECTIVE: The objective of the study was to examine whether somatic 17q22-24 changes, PRKAR1A mutations, and/or PKA abnormalities are present in AIMAH. PATIENTS: We studied fourteen patients with Cushing syndrome due to AIMAH. METHODS: Fluorescent in situ hybridization with a PRKAR1A-specific probe was used for investigating chromosome 17 allelic losses. The PRKAR1A gene was sequenced in all samples, and tissue was studied for PKA activity, cAMP responsiveness, and PKA subunit expression. RESULTS: We found 17q22-24 allelic losses in 73% of the samples. There were no PRKAR1A-coding sequence mutations. The RIIbeta PKA subunit was overexpressed by mRNA, whereas the RIalpha, RIbeta, RIIalpha, and Calpha PKA subunits were underexpressed. These findings were confirmed by immunohistochemistry. Total PKA activity and free PKA activity were higher in AIMAH than normal adrenal glands, consistent with the up-regulation of the RIIbeta PKA subunit. CONCLUSIONS: PRKAR1A mutations are not found in AIMAH. Somatic losses of the 17q22-24 region and PKA subunit and enzymatic activity changes show that PKA signaling is altered in AIMAH in a way that is similar to that of other adrenal tumors with 17q losses or PRKAR1A mutations.


Assuntos
Cromossomos Humanos Par 17/genética , Síndrome de Cushing/genética , Proteínas Quinases Dependentes de AMP Cíclico/genética , Glândulas Suprarrenais/metabolismo , Glândulas Suprarrenais/patologia , Alelos , Síndrome de Cushing/enzimologia , AMP Cíclico/metabolismo , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Humanos , Hiperplasia , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Mutação , RNA/química , RNA/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa
13.
Breast Cancer Res ; 8(1): R10, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16417658

RESUMO

INTRODUCTION: Failure to detect co-expression of estrogen receptor-alpha (ERalpha) and proliferation 'markers' such as Ki67 in human mammary epithelium led to the view that estrogen acts indirectly to stimulate mammary epithelial proliferation. The mitotic index was so low in prior studies, however, that transient co-expression of ERalpha and Ki67 during the cell cycle could have been below detection limits. METHODS: Immunohistochemistry was used on mammary tissue sections from estrogen treated rhesus monkeys to investigate co-expression of ERalpha and the proliferation antigen Ki67. Using the same methods, we investigated the cell localization of proteins involved in estrogen-induced proliferation, including cyclin D1, stromal cell-derived factor (SDF)-1, and MYC. RESULTS: ERalpha was co-expressed with the proliferation marker Ki67 as well as with SDF-1, MYC and cyclin D1 in mammary epithelial cells from estrogen-treated monkeys. CONCLUSION: ERalpha is expressed in proliferating mammary epithelial cells together with the estrogen-induced proteins MYC, cyclin D1 and SDF-1, consistent with a direct mitogenic action by estrogen in primate mammary epithelium.


Assuntos
Receptor alfa de Estrogênio/biossíntese , Estrogênios/fisiologia , Antígeno Ki-67/biossíntese , Glândulas Mamárias Animais/citologia , Animais , Quimiocina CXCL12 , Quimiocinas CXC/biossíntese , Ciclina D1/biossíntese , Células Epiteliais/fisiologia , Receptor alfa de Estrogênio/análise , Feminino , Genes myc , Imuno-Histoquímica , Antígeno Ki-67/análise , Macaca mulatta
14.
Endocrinology ; 146(12): 5086-91, 2005 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16123158

RESUMO

IGF action has been implicated in the promotion of oxidative stress and aging in invertebrate and murine models. However, some in vitro models suggest that IGF-I specifically prevents neuronal oxidative damage. To investigate whether IGF-I promotes or retards brain aging, we evaluated signs of oxidative stress and neuropathological aging in brains from 400-d-old Igf1-/- and wild-type (WT) mice. Lipofuscin pigment accumulation reflects oxidative stress and aging, but we found no difference in lipofuscin deposition in Igf1-/- and WT brains. Likewise, there was no apparent difference in accumulation of nitrotyrosine residues in Igf1-/- and WT brains, except for layer IV/V of the cerebral cortex, where these proteins were about 20% higher in the Igf1-/- brain (P = 0.03). We found no difference in the levels of oxidative stress-related enzymes, neuronal nitric oxide synthase, inducible nitric oxide synthase, and superoxide dismutase in Igf1-/- and WT brains. Tau is a microtubule-associated protein that causes the formation of neurofibrillary tangles and senile plaques as it becomes hyperphosphorylated in the aging brain. Tau phosphorylation was dramatically increased on two specific residues, Ser-396 and Ser-202, both glycogen synthase kinases target sites implicated in neurodegeneration. These observations indicate that IGF-I has a major role in regulating tau phosphorylation in the aging brain, whereas its role in promoting or preventing oxidative stress remains uncertain.


Assuntos
Encéfalo/metabolismo , Fator de Crescimento Insulin-Like I/deficiência , Proteínas tau/metabolismo , Envelhecimento/metabolismo , Animais , Encéfalo/enzimologia , Resíduos de Drogas/metabolismo , Lipofuscina/metabolismo , Masculino , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase/biossíntese , Estresse Oxidativo , Fosforilação , Tirosina/análogos & derivados , Tirosina/metabolismo
15.
Genes Chromosomes Cancer ; 44(2): 204-11, 2005 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16001434

RESUMO

Odontogenic myxomas are rare benign neoplasms affecting the jaw. Myxomas of bones and other sites occur as part of Carney complex (CNC), a multiple neoplasia syndrome caused by mutations in the PRKAR1A gene, which codes for the regulatory subunit of protein kinase A (PKA). In the present study, 17 odontogenic myxomas from patients without CNC were screened for PRKAR1A mutations and PRKAR1A protein expression by immunohistochemistry (IHC). Mutations of the coding region of the PRKAR1A gene were identified in 2 tumors; both these lesions showed no or significantly decreased immunostaining of PRKAR1A in the tumor compared to that in the surrounding normal tissue. One mutation (c.725C>A) led to a nonconservative amino acid substitution in a highly conserved area of the gene (A213D); the other was a single base-pair deletion that led to a frameshift (del774C) and a stop codon 11 amino acids downstream of the mutation site; both tumors were heterozygous for the respective mutations. Of the remaining tumors, 7 of the 15 without mutations showed almost no PRKAR1A in the tumor cells, whereas IHC showed that the protein was abundant in nontumorous cells. We concluded that PRKAR1A may be involved by its down-regulation in the pathogenesis of odontogenic myxomas caused by mutations and/or other genetic mechanisms. Of the sporadic, nonfamilial tumors associated with PRKAR1A mutations, the odontogenic type was the first myxomatous lesion found to harbor somatic PRKAR1A sequence changes.


Assuntos
Mixoma/genética , Tumores Odontogênicos/genética , Proteínas/genética , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico , Proteínas Quinases Dependentes de AMP Cíclico , DNA de Neoplasias/genética , Humanos , Imuno-Histoquímica , Mutação , Mixoma/diagnóstico , Mixoma/metabolismo , Tumores Odontogênicos/diagnóstico , Tumores Odontogênicos/metabolismo , Proteínas/metabolismo
16.
Cancer Res ; 65(11): 4506-14, 2005 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-15930266

RESUMO

Carney complex is an autosomal dominant neoplasia syndrome characterized by spotty skin pigmentation, myxomatosis, endocrine tumors, and schwannomas. This condition may be caused by inactivating mutations in PRKAR1A, the gene encoding the type 1A regulatory subunit of protein kinase A. To better understand the mechanism by which PRKAR1A mutations cause disease, we have developed conventional and conditional null alleles for Prkar1a in the mouse. Prkar1a(+/-) mice developed nonpigmented schwannomas and fibro-osseous bone lesions beginning at approximately 6 months of age. Although genotype-specific cardiac and adrenal lesions were not seen, benign and malignant thyroid neoplasias were observed in older mice. This spectrum of tumors overlaps that seen in Carney complex patients, confirming the validity of this mouse model. Genetic analysis indicated that allelic loss occurred in a subset of tumor cells, suggesting that complete loss of Prkar1a plays a key role in tumorigenesis. Similarly, tissue-specific ablation of Prkar1a from a subset of facial neural crest cells caused the formation of schwannomas with divergent differentiation. These observations confirm the identity of PRKAR1A as a tumor suppressor gene with specific importance to cyclic AMP-responsive tissues and suggest that these mice may be valuable tools not only for understanding endocrine tumorigenesis but also for understanding inherited predispositions for schwannoma formation.


Assuntos
AMP Cíclico/fisiologia , Modelos Animais de Doenças , Neoplasia Endócrina Múltipla/genética , Neurilemoma/genética , Proteínas/genética , Alelos , Animais , Neoplasias Ósseas/enzimologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico , Feminino , Genes Supressores de Tumor , Predisposição Genética para Doença , Masculino , Camundongos , Neoplasia Endócrina Múltipla/enzimologia , Neoplasia Endócrina Múltipla/patologia , Neurilemoma/enzimologia , Neurilemoma/patologia , Osteoblastos/citologia , Osteoblastos/fisiologia , Células de Schwann/citologia , Células de Schwann/fisiologia , Síndrome , Timo/citologia , Timo/fisiologia , Neoplasias da Glândula Tireoide/enzimologia , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia
17.
J Clin Endocrinol Metab ; 90(6): 3773-9, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15741255

RESUMO

Hereditary leiomyomatosis and renal cell cancer (HLRCC) is an autosomal dominant disorder caused by mutations in the fumarate hydratase (FH) gene on chromosome 1q42.3-43. Massive macronodular adrenocortical disease (MMAD) is a heterogeneous condition associated with Cushing syndrome (CS) and bilateral hyperplasia of the adrenal glands. In MMAD, cortisol secretion is often mediated by ectopic, adrenocortical expression of receptors for a variety of substances; however, to date, no consistent genetic defects have been identified. In a patient with HLRCC caused by a germline-inactivating FH mutation, we diagnosed atypical (subclinical) CS due to bilateral, ACTH-independent adrenocortical hyperplasia. A clinical protocol for the detection of ectopic expression of various hormone receptors was employed. Histology was consistent with MMAD. The tumor tissue harbored the germline FH mutation and demonstrated allelic losses of the 1q42.3-43 FH locus. We then searched the National Institutes of Health (NIH) databases of patients with MMAD or HLRCC and found at least three other cases with MMAD that had a history of tumors that could be part of HLRCC; among patients with HLRCC, there were several with some adrenal nodularity noted on computed tomography but none with imaging findings consistent with MMAD. From two of the three MMAD patients, adrenocortical tumor DNA was available and sequenced for coding FH mutations; there were none. We conclude that in a patient with HLRCC, adrenal hyperplasia and CS were due to MMAD. The latter was likely due to the FH germline mutation because in tumor cells, only the mutant allele was retained. However, other patients with MMAD and HLRCC, or HLRCC patients with adrenal imaging findings consistent with MMAD, or MMAD patients with somatic FH mutations were not found among the NIH series. Although a fortuitous association cannot be excluded, HLRCC may be added to the short list of monogenic disorders that have been reported to be associated with the development of adrenal tumors; FH may be considered a candidate gene for MMAD.


Assuntos
Neoplasias do Córtex Suprarrenal/genética , Síndrome de Cushing/genética , Leiomiomatose/genética , Neoplasias do Córtex Suprarrenal/complicações , Neoplasias do Córtex Suprarrenal/patologia , Glândulas Suprarrenais/patologia , Idoso , Mapeamento Cromossômico , Cromossomos Humanos Par 1 , Síndrome de Cushing/complicações , DNA/sangue , DNA/genética , DNA/isolamento & purificação , Feminino , Lateralidade Funcional , Humanos , Hibridização in Situ Fluorescente , Leiomiomatose/cirurgia , Pele/patologia , Resultado do Tratamento
18.
Cancer Res ; 64(24): 8811-5, 2004 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-15604237

RESUMO

Mutations of the human type Ialpha regulatory subunit (RIalpha) of cyclic AMP-dependent protein kinase (PKA; PRKAR1A) lead to altered kinase activity, primary pigmented nodular adrenocortical disease, and tumors of the thyroid and other tissues. To bypass the early embryonic lethality of Prkar1a(-/-) mice, we established transgenic mice carrying an antisense transgene for Prkar1a exon 2 (X2AS) under the control of a tetracycline-responsive promoter. Down-regulation of Prkar1a by up to 70% was achieved in transgenic mouse tissues and embryonic fibroblasts, with concomitant changes in kinase activity and increased cell proliferation, respectively. Mice developed thyroid follicular hyperplasia and adenomas, adrenocortical hyperplasia, and other features reminiscent of primary pigmented nodular adrenocortical disease, histiocytic and epithelial hyperplasias, lymphomas, and other mesenchymal tumors. These were associated with allelic losses of the mouse chromosome 11 Prkar1a locus, an increase in total type II PKA activity, and higher RIIbeta protein levels. This mouse provides a novel, useful tool for the investigation of cyclic AMP, RIalpha, and PKA functions and confirms the critical role of Prkar1a in tumorigenesis in endocrine and other tissues.


Assuntos
Neoplasias do Córtex Suprarrenal/enzimologia , Transtornos Linfoproliferativos/enzimologia , Proteínas/fisiologia , Neoplasias da Glândula Tireoide/enzimologia , Adenoma/enzimologia , Adenoma/genética , Neoplasias do Córtex Suprarrenal/genética , Animais , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico , DNA Antissenso/genética , Regulação para Baixo , Éxons , Feminino , Transtornos Linfoproliferativos/genética , Mesoderma/enzimologia , Mesoderma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Gravidez , Regiões Promotoras Genéticas , Proteínas/genética , Tetraciclina/farmacologia , Neoplasias da Glândula Tireoide/genética , Transativadores/genética
19.
Front Horm Res ; 32: 253-64, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15281351

RESUMO

Carney complex (CNC) is a familial multiple neoplasia and lentiginosis syndrome with features overlapping those of McCune-Albright syndrome (MAS) and other multiple endocrine neoplasia (MEN) syndromes like MEN type 1 (MEN 1). Pituitary tumors have been described in a number of patients with CNC; all have been growth hormone (GH) and prolactin (PRL)-producing. In at least some patients, pituitary gland involvement is manifested by hyperplastic areas; hyperplasia appears to involve somatomammotrophs only and to precede GH-producing tumor formation, in a pathway similar to that seen in MAS-related pituitary tumors (and in oncogenesis in other CNC tissues). One patient with CNC and advanced acromegaly had a GH-producing macroadenoma that showed extensive genetic changes at the chromosomal level. These changes appeared to represent secondary or tertiary genetic 'hits' involved in pituitary oncogenesis and were confirmed at the molecular level. So far, almost half of the patients with CNC have germline-inactivating mutations in the PRKAR1A gene; in their pituitary tumors, the normal allele of the PRKAR1A gene is lost. Loss of heterozygosity suggests that PRKAR1A, which codes for the regulatory subunit type 1alpha of the cAMP-dependent protein kinase A (PKA), may act as a tumor-suppressor gene in pituitary tissue. These data provide evidence for a PKA-induced somatomammotroph hyperplasia in the pituitary tissue of CNC patients; hyperplasia leads to additional genetic changes at the somatic level, which in turn cause the formation of adenomas in some, but not all, patients.


Assuntos
Doenças do Sistema Endócrino/patologia , Mutação em Linhagem Germinativa , Mixoma/patologia , Neurilemoma/patologia , Transtornos da Pigmentação/patologia , Proteínas/genética , Pigmentação da Pele , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico , Proteínas Quinases Dependentes de AMP Cíclico , Doenças do Sistema Endócrino/genética , Humanos , Lentigo/genética , Lentigo/patologia , Neoplasia Endócrina Múltipla/genética , Neoplasia Endócrina Múltipla/patologia , Mixoma/genética , Neurilemoma/genética , Transtornos da Pigmentação/genética , Neoplasias Hipofisárias/genética , Neoplasias Hipofisárias/patologia , Síndrome
20.
J Clin Endocrinol Metab ; 89(7): 3173-82, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15240590

RESUMO

Cushing syndrome is uncommon in childhood and rare in infancy. We report the case of a 3-yr-old child who presented with symptoms of Cushing syndrome beginning shortly after birth. Her hypercortisolemia was cyclical, causing relapsing and remitting symptoms, which eventually led to suspicions of possible Munchausen syndrome by proxy. Investigation at the National Institutes of Health excluded exogenous administration of glucocorticoids and indicated ACTH-independent Cushing syndrome. Paradoxical response to dexamethasone stimulation (Liddle's test) suggested a diagnosis of primary pigmented nodular adrenocortical disease (PPNAD). After bilateral adrenalectomy, both glands showed micronodular adrenocortical hyperplasia, but histology was not consistent with typical PPNAD. DNA analysis of the coding sequences of the PRKAR1A gene (associated with PPNAD and Carney complex) and the GNAS gene (associated with McCune-Albright syndrome) showed no mutations. We conclude that hypercortisolemia in infancy may be caused by micronodular adrenocortical hyperplasia, which can be cyclical and confused with exogenous Cushing syndrome. A paradoxical rise of glucocorticoid excretion during Liddle's test may delineate these patients. Infantile micronodular disease has some features of PPNAD and may represent its early form; however, at least in the case of the patient reported here, micronodular hyperplasia was not caused by coding mutations of the PRKAR1A or GNAS genes or associated with typical histology or any other features of Carney complex or McCune-Albright syndrome and may represent a distinct entity.


Assuntos
Doenças do Córtex Suprarrenal/diagnóstico , Síndrome de Cushing/diagnóstico , Hidrocortisona/sangue , Periodicidade , Doenças do Córtex Suprarrenal/sangue , Doenças do Córtex Suprarrenal/patologia , Doenças do Córtex Suprarrenal/cirurgia , Glândulas Suprarrenais/patologia , Adrenalectomia , DNA de Neoplasias/análise , Diagnóstico Diferencial , Feminino , Seguimentos , Hormônios/sangue , Humanos , Hiperplasia , Recém-Nascido , Microscopia Eletrônica , Tomografia Computadorizada por Raios X
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