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1.
Childs Nerv Syst ; 30(8): 1343-53, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24817375

RESUMO

PURPOSE: Maternal folate intake has reduced the incidence of human neural tube defects by 60-70 %. However, 30-40 % of cases remain nonresponsive to folate intake. The main purpose of this study was to understand the molecular mechanism of folate nonresponsiveness in a mouse model of neural tube defect. METHODS: We used a folate-nonresponsive Fkbp8 knockout mouse model to elucidate the molecular mechanism(s) of folate nonresponsiveness. Neurospheres were grown from neural stem cells isolated from the lumbar neural tube of E9.5 Fkbp8 (-/-) and wild-type embryos. Immunostaining was used to determine the protein levels of oligodendrocyte transcription factor 2 (Olig2), Nkx6.1, class III beta-tubulin (TuJ1), O4, glial fibrillary acidic protein (GFAP), histone H3 Lys27 trimethylation (H3K27me3), ubiquitously transcribed tetratricopeptide repeat (UTX), and Msx2, and quantitative real-time (RT)-PCR was used to determine the message levels of Olig2, Nkx6.1, Msx2, and noggin in neural stem cells differentiated in the presence and absence of folic acid. RESULTS: Fkbp8 (-/-)-derived neural stem cells showed (i) increased noggin expression; (ii) decreased Msx2 expression; (iii) premature differentiation--neurogenesis, oligodendrogenesis (Olig2 expression), and gliogenesis (GFAP expression); and (iv) increased UTX expression and decreased H3K27me3 polycomb modification. Exogenous folic acid did not reverse these markers. CONCLUSIONS: Folate nonresponsiveness could be attributed in part to increased noggin expression in Fkbp8 (-/-) embryos, resulting in decreased Msx2 expression. Folate treatment further increases Olig2 and noggin expression, thereby exacerbating ventralization.


Assuntos
Proteínas de Transporte/metabolismo , Ácido Fólico/efeitos adversos , Regulação da Expressão Gênica no Desenvolvimento/genética , Defeitos do Tubo Neural , Proteínas de Ligação a Tacrolimo/deficiência , Animais , Proteínas de Transporte/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Modelos Animais de Doenças , Embrião de Mamíferos , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Histona Desmetilases com o Domínio Jumonji , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , Defeitos do Tubo Neural/induzido quimicamente , Defeitos do Tubo Neural/genética , Defeitos do Tubo Neural/metabolismo , Gravidez , Proteínas de Ligação a Tacrolimo/genética
2.
Sci Rep ; 2: 980, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23243496

RESUMO

Folic acid (FA) has traditionally been associated with prevention of neural tube defects; more recent work suggests that it may also be involved in in the prevention of adult onset diseases. As the role of FA in human health and disease expands, it also becomes more critical to understand the mechanisms behind FA action. In this work we examined the hypothesis that folate receptor alpha (FRα) acts as a transcription factor. FRα is a GPI-anchored protein and a component of the caveolae fraction. The work described here shows that FRα translocates to the nucleus, where it binds to cis-regulatory elements at promoter regions of Fgfr4 and Hes1, and regulates their expression. The FRα recognition domain mapped to AT rich regions on the promoters. Until this time FRα has only been considered as a folate transporter, these studies describe a novel role for FRα as a transcription factor.


Assuntos
Núcleo Celular/metabolismo , Receptor 1 de Folato/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Sequência Consenso , Ácido Fólico/farmacologia , Humanos , Camundongos , Regiões Promotoras Genéticas , Ligação Proteica , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Elementos Reguladores de Transcrição , Transfecção
3.
Stem Cells Dev ; 21(2): 321-30, 2012 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-21521032

RESUMO

Although maternal intake of folic acid (FA) prevents neural tube defects in 70% of the population, the exact mechanism of prevention has not been elucidated. We hypothesized that FA affects neural stem cell (NSC) proliferation and differentiation. This hypothesis was examined in a folate-responsive spina bifida mouse model, Splotch (Sp(-/-)), which has a homozygous loss-of-function mutation in the Pax3 gene. Neurospheres were generated with NSCs from the lower lumbar neural tube of E10.5 wild-type (WT) and Sp(-/-) embryos, in the presence and absence of FA. In the absence of FA, the number of neurospheres generated from Sp(-/-) embryos compared with WT was minimal (P<0.05). Addition of FA to Sp(-/-) cultures increased the expression of a Pax3 downstream target, fgfr4, and rescued NSC proliferative potential, as demonstrated by a significant increase in neurosphere formation (P<0.01). To ascertain if FA affected cell differentiation, FA-stimulated Sp(-/-) neurospheres were allowed to differentiate in the continued presence or absence of FA. Neurospheres from both conditions expressed multi-potent stem cell characteristics and the same differentiation potential as WT. Further, multiple neurospheres from both WT and FA-stimulated Sp(-/-) cell cultures formed extensive synaptic connections. On the whole, FA-mediated rescue of neural tube defects in Sp(-/-) embryos promotes NSC proliferation at an early embryonic stage. FA-stimulated Sp(-/-) neurospheres differentiate and form synaptic connections, comparable to WT.


Assuntos
Células-Tronco Fetais/efeitos dos fármacos , Ácido Fólico/farmacologia , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Neurais/efeitos dos fármacos , Tubo Neural/efeitos dos fármacos , Disrafismo Espinal/tratamento farmacológico , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Embrião de Mamíferos , Células-Tronco Fetais/patologia , Feto , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Técnicas de Inativação de Genes , Homozigoto , Camundongos , Camundongos Knockout , Células-Tronco Neurais/patologia , Tubo Neural/embriologia , Tubo Neural/patologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Fator de Transcrição PAX3 , Fatores de Transcrição Box Pareados/deficiência , Fatores de Transcrição Box Pareados/genética , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Disrafismo Espinal/embriologia , Disrafismo Espinal/genética , Disrafismo Espinal/patologia , Sinapses/efeitos dos fármacos , Sinapses/fisiologia
4.
Vitam Horm ; 87: 143-73, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22127242

RESUMO

Maternal folic acid (FA) intake has beneficial effects in preventing neural tube defects and may also play a role in the prevention of adult onset diseases such as Alzheimer's disease, dementia, neuropsychiatric disorders, cardiovascular diseases, and cerebral ischemia. This review will focus on the effects of maternal FA intake on neural crest stem cell proliferation and differentiation. Although FA is generally considered beneficial, it has the potential of promoting cell proliferation at the expense of differentiation. In some situations, this may lead to miscarriage or postnatal developmental abnormalities. Therefore, a blind approach such as "FA for everyone" is not necessarily the best course of action. Ultimately, the best approach for FA supplementation, and potentially other nutritional supplements, will include customized patient genomic profiles for determining dose and duration.


Assuntos
Células-Tronco Embrionárias/citologia , Ácido Fólico/administração & dosagem , Fenômenos Fisiológicos da Nutrição Materna , Crista Neural/citologia , Animais , Diferenciação Celular , Proliferação de Células , Anormalidades Congênitas/metabolismo , Anormalidades Congênitas/patologia , Anormalidades Congênitas/prevenção & controle , Células-Tronco Embrionárias/metabolismo , Células-Tronco Embrionárias/patologia , Feminino , Desenvolvimento Fetal , Ácido Fólico/uso terapêutico , Humanos , Crista Neural/metabolismo , Crista Neural/patologia , Defeitos do Tubo Neural/metabolismo , Defeitos do Tubo Neural/patologia , Defeitos do Tubo Neural/prevenção & controle , Gravidez
5.
Epigenetics ; 6(10): 1207-16, 2011 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-21931278

RESUMO

The epigenetic mechanism of folic acid (FA) action on dorsal root ganglion (DRG) cell proliferation and sensory neuron differentiation is not well understood. In this study, the ND7 cell line, derived from DRG cells, was used to elucidate this mechanism. In ND7 cells differentiated with dbcAMP and NGF, Hes1 and Pax3 levels decreased, whereas Neurog2 levels showed a modest increase. Chromatin immunoprecipitation (ChIP) assays examining epigenetic marks at the Hes1 promoter showed that FA favored increased H3K9 and H3K19 acetylation and decreased H3K27 methylation. Hence, FA plays a positive role in cell proliferation. In differentiated ND7 cells, H3K27 methylation decreased, whereas H3K9 and H3K18 acetylation increased at the Neurog2 promoter. FA did not favor this phenotypic outcome. Additionally, in differentiated ND7 Neurog2 associated with the NeuroD1 promoter, FA decreased this association. The results suggest that the switch from proliferation to sensory neuron differentiation in DRG cells is regulated by alterations in epigenetic marks, H3K9/18 acetylation and H3K27 methylation, at Hes1 and Neurog2 promoters, as well as by Neurog2 association with NeuroD1 promoter. FA although positive for proliferation, does not appear to play a role in differentiation.


Assuntos
Epigênese Genética/efeitos dos fármacos , Ácido Fólico/farmacologia , Gânglios Espinais/citologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Neurogênese/genética , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular , Cromatina/química , Cromatina/metabolismo , Cromatina/fisiologia , Histonas/química , Histonas/metabolismo , Histonas/fisiologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Regiões Promotoras Genéticas , Ratos
6.
Mol Biol Cell ; 22(4): 503-12, 2011 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-21169561

RESUMO

Pax3 plays a role in regulating Hes1 and Neurog2 activity and thereby stem cell maintenance and neurogenesis. A mechanism for Pax3 regulation of these two opposing events, during caudal neural tube development, is examined in this study. Pax3 acetylation on C-terminal lysine residues K437 and K475 may be critical for proper regulation of Hes1 and Neurog2. Removal of these lysine residues increased Hes1 but decreased Neurog2 promoter activity. SIRT1 deacetylase may be a key component in regulating Pax3 acetylation. Chromatin immunoprecipitation assays showed that SIRT1 is associated with Hes1 and Neurog2 promoters during murine embryonic caudal neural tube development at E9.5, but not at E12.5. Overexpression of SIRT1 decreased Pax3 acetylation, Neurog2 and Brn3a positive staining. Conversely, siRNA-mediated silencing of SIRT1 increased these factors. These studies suggest that Pax3 acetylation results in decreased Hes1 and increased Neurog2 activity, thereby promoting sensory neuron differentiation.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neurogênese/genética , Fatores de Transcrição Box Pareados/metabolismo , Células Receptoras Sensoriais/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Acetilação , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina/métodos , Proteínas de Homeodomínio/genética , Humanos , Camundongos , Proteínas do Tecido Nervoso/genética , Tubo Neural/metabolismo , Fator de Transcrição PAX3 , Fatores de Transcrição Box Pareados/genética , Regiões Promotoras Genéticas/genética , Processamento de Proteína Pós-Traducional/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , Fatores de Transcrição HES-1
7.
J Biol Chem ; 285(47): 36922-32, 2010 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-20833714

RESUMO

The mechanism(s) behind folate rescue of neural tube closure are not well understood. In this study we show that maternal intake of folate prior to conception reverses the proliferation potential of neural crest stem cells in homozygous Splotch embryos (Sp(-/-)) via epigenetic mechanisms. It is also shown that the pattern of differentiation seen in these cells is similar to wild-type (WT). Cells from open caudal neural tubes of Sp(-/-) embryos exhibit increased H3K27 methylation and decreased expression of KDM6B possibly due to up-regulation of KDM6B targeting micro-RNAs such as miR-138, miR-148a, miR-185, and miR-339-5p. In our model, folate reversed these epigenetic marks in folate-rescued Sp(-/-) embryos. Using tissue from caudal neural tubes of murine embryos we also examined H3K27me2 and KDM6B association with Hes1 and Neurog2 promoters at embryonic day E10.5, the proliferative stage, and E12.5, when neural differentiation begins. In Sp(-/-) embryos compared with WT, levels of H3K27me2 associated with the Hes1 promoter were increased at E10.5, and levels associated with the Neurog2 promoter were increased at E12.5. KDM6B association with Hes1 and Neurog2 promoters was inversely related to H3K27me2 levels. These epigenetic changes were reversed in folate-rescued Sp(-/-) embryos. Thus, one of the mechanisms by which folate may rescue the Sp(-/-) phenotype is by increasing the expression of KDM6B, which in turn decreases H3K27 methylation marks on Hes1 and Neurog2 promoters thereby affecting gene transcription.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Ácido Fólico/administração & dosagem , Proteínas de Homeodomínio/genética , Proteínas do Tecido Nervoso/genética , Tubo Neural/efeitos dos fármacos , Tubo Neural/embriologia , Regiões Promotoras Genéticas/genética , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Western Blotting , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sistema Nervoso Central/embriologia , Montagem e Desmontagem da Cromatina/genética , Imunoprecipitação da Cromatina , Embrião de Mamíferos/citologia , Embrião de Mamíferos/efeitos dos fármacos , Embrião de Mamíferos/metabolismo , Epigenômica , Feminino , Imunofluorescência , Ácido Fólico/farmacologia , Histonas/genética , Histonas/metabolismo , Proteínas de Homeodomínio/metabolismo , Técnicas Imunoenzimáticas , Imunoprecipitação , Histona Desmetilases com o Domínio Jumonji/antagonistas & inibidores , Histona Desmetilases com o Domínio Jumonji/genética , Histona Desmetilases com o Domínio Jumonji/metabolismo , Luciferases/metabolismo , Masculino , Metilação , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , Defeitos do Tubo Neural/prevenção & controle , Fator de Transcrição PAX3 , Fatores de Transcrição Box Pareados/fisiologia , Regiões Promotoras Genéticas/efeitos dos fármacos , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição HES-1 , Complexo Vitamínico B/administração & dosagem , Complexo Vitamínico B/farmacologia
8.
Childs Nerv Syst ; 25(7): 801-6, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19326126

RESUMO

INCIDENCE: Worldwide, the incidence of neural tube defects (NTDs) varies from 0.17 to 6.39 per 1,000 live births. The declining prevalence of myelomeningocele, the most common NTD, is secondary to several factors including folic acid fortification, prenatal diagnosis with termination of affected fetuses, and unknown factors. IMPACT OF CHANGES: Of those born with myelomeningocele, survival during infancy and preschool years has improved over the last 25 years (Bowman et al., Pediatr Neurosurg 34:114-120). Fewer newborns today require shunt placement, which will hopefully improve the long-term mortality associated with this disease (Chakraborty et al., J Neurosurg Pediatr 1(5):361-365, unpublished data). Of a cohort born in 1975-1979 and treated at a single US institution, 74% have survived into young adulthood. CLINICAL IMPLICATIONS: One of the greatest challenges facing these young adults is the transitioning of their medical care into an adult medical community.


Assuntos
Meningomielocele/epidemiologia , Meningomielocele/cirurgia , Neurocirurgia/tendências , Pediatria/tendências , Deficiência de Ácido Fólico/complicações , Deficiência de Ácido Fólico/dietoterapia , Predisposição Genética para Doença , Humanos , Incidência , Recém-Nascido , Meningomielocele/diagnóstico , Mortalidade , Prognóstico
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