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1.
Mol Vis ; 30: 167-174, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38601015

RESUMO

Purpose: To examine whether increased ephrin type-B receptor 1 (EphB1) leads to inflammatory mediators in retinal Müller cells. Methods: Diabetic human and mouse retinal samples were examined for EphB1 protein levels. Rat Müller cells (rMC-1) were grown in culture and treated with EphB1 siRNA or ephrin B1-Fc to explore inflammatory mediators in cells grown in high glucose. An EphB1 overexpression adeno-associated virus (AAV) was used to increase EphB1 in Müller cells in vivo. Ischemia/reperfusion (I/R) was performed on mice treated with the EphB1 overexpression AAV to explore the actions of EphB1 on retinal neuronal changes in vivo. Results: EphB1 protein levels were increased in diabetic human and mouse retinal samples. Knockdown of EphB1 reduced inflammatory mediator levels in Müller cells grown in high glucose. Ephrin B1-Fc increased inflammatory proteins in rMC-1 cells grown in normal and high glucose. Treatment of mice with I/R caused retinal thinning and loss of cell numbers in the ganglion cell layer. This was increased in mice exposed to I/R and treated with the EphB1 overexpressing AAVs. Conclusions: EphB1 is increased in the retinas of diabetic humans and mice and in high glucose-treated Müller cells. This increase leads to inflammatory proteins. EphB1 also enhanced retinal damage in response to I/R. Taken together, inhibition of EphB1 may offer a new therapeutic option for diabetic retinopathy.


Assuntos
Retinopatia Diabética , Efrina-B1 , Doenças Retinianas , Animais , Humanos , Camundongos , Ratos , Retinopatia Diabética/genética , Retinopatia Diabética/metabolismo , Células Ependimogliais/metabolismo , Efrina-B1/genética , Efrina-B1/metabolismo , Glucose/metabolismo , Mediadores da Inflamação/metabolismo , Retina/metabolismo , Doenças Retinianas/metabolismo
2.
Int J Biol Macromol ; 258(Pt 1): 128848, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38114003

RESUMO

The survival benefit for patients with gastric cancer (GC) is modest due to its high transfer potential. Targeted therapy for metastasis-related genes in GC may be a viable approach, however, inhibitors specifically targeting GC are limited. In this study, GC patient-derived xenografts (PDX) with metastatic burden were established via orthotopic transplantation. PCR-Array analysis of primary and metastatic tumors revealed EPH receptor B2 (EPHB2) as the most significantly upregulated gene. The interaction between the EPHB2 receptor and its cognate-specific EFNB1 ligands was high in GC and correlated with a poor prognosis. Fc-EFNB1 treatment increased the invasion and migration abilities of GC cells and induced a high EPHB2 expression. EPHB2 knockdown in GC cells completely abolished the ephrin ligand-induced effects on invasion and migration abilities. Signal transduction analysis revealed Wnt/ß-catenin and FAK as downstream signaling mediators potentially inducing the EPHB2 phenotype. In conclusion, the observed deregulation of EPHB2/EFNB1 expression in GC enhances the invasive phenotype, suggesting a potential role of EPHB2/EFNB1 compound in local tumor cell invasion and the formation of metastasis.


Assuntos
Receptor EphB2 , Neoplasias Gástricas , Humanos , Receptor EphB2/genética , Receptor EphB2/metabolismo , Neoplasias Gástricas/patologia , Efrina-B1/genética , Efrina-B1/metabolismo , beta Catenina/metabolismo , Ligantes , Via de Sinalização Wnt , Movimento Celular/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Proliferação de Células/genética
3.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 40(6): 706-710, 2023 Jun 10.
Artigo em Chinês | MEDLINE | ID: mdl-37212007

RESUMO

OBJECTIVE: To explore the clinical feature and genetic etiology of a patient with Craniofacial nasal syndrome (CNFS). METHODS: A patient with CNFS who had presented at the Guiyang Maternal and Child Health Care Hospital on November 13, 2021 was selected as the study subject. Clinical data of the patient were collected. Peripheral venous blood samples were collected from the patient and her parents and subjected to trio-whole exome sequencing (trio-WES). Candidate variants were verified by Sanger sequencing and bioinformatic analysis. RESULTS: The patient, a 15-year-old female, had predominantly featured forehead bulging, hypertelorism, wide nasal dorsum and bifid nasal tip. Genetic testing revealed that she has harbored a heterozygous missense c.473T>C (p.M158T) variant of the EFNB1 gene, which was detected in either of her parents. By bioinformatic analysis, the variant has not been recorded in the HGMD and ClinVar databases, and no population frequency was recorded in the 1000 Genomes, ExAC, gnomAD and Shenzhou Genome Data Cloud databases. As predicted by the REVEL online software, the variant can confer deleterious effects on the gene or its product. Analysis using UGENE software showed the corresponding amino acid to be highly conserved among various species. Analysis with AlphaFold2 software suggested that the variant may affect the 3D structure and function of the Ephrin-B1 protein. Based on the American College of Medical Genetics and Genomics (ACMG) standards and guidelines and recommendation of Clinical Genome Resource (ClinGen), the variant was rated as pathogenic. CONCLUSION: Combining the patient's clinical features and genetic finding, the diagnosis of CNFS was confirmed. The heterozygous c.473T>C (p.M158T) missense variant of the EFNB1 gene probably underlay the disease in this patient. Above finding has provided a basis for the genetic counseling and prenatal diagnosis for her family.


Assuntos
Biologia Computacional , Efrina-B1 , Humanos , Criança , Feminino , Gravidez , Adolescente , Efrina-B1/genética , China , Família , Mutação
4.
Mol Genet Genomic Med ; 10(7): e1964, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35603962

RESUMO

BACKGROUND: Although B-acute lymphoblastic leukemia (B-ALL) patients' survival has been improved dramatically, some cases still relapse. This study aimed to explore the prognosis-related novel differentially expressed genes (DEGs) for predicting the overall survival (OS) of children and young adults (CAYAs) with B-ALL and analyze the immune-related factors contributing to poor prognosis. METHODS: GSE48558 and GSE79533 from Gene Expression Omnibus (GEO) and clinical sample information and mRNA-seq from Therapeutically Applicable Research to Generate Effective Treatments (TARGET) database were retrieved. Prognosis-related key genes were enrolled to build a Cox proportional model using multivariate Cox regression. Five-year OS of patients, clinical characteristic relevance and clinical independence were assessed based on the model. The mRNA levels of prognosis-related genes were validated in our samples and the difference of immune cells composition between high-risk and low-risk patients were compared. RESULTS: One hundred and twelve DEGs between normal B cells and B-ALL cells were identified based on GSE datasets. They were mainly participated in protein binding and HIF-1 signaling pathway. One hundred and eighty-nine clinical samples were enrolled in the study, both Kaplan-Meier (KM) analysis and univariate Cox regression analysis showed that CYBB, BCL2A1, IFI30, and EFNB1 were associated with prognosis, CYBB, BCL2A1, and EFNB1 were used to construct prognostic risk model. Moreover, compared to clinical indicators, the three-gene signature was an independent prognostic factor for CAYAs with B-ALL. Finally, the mRNA levels of CYBB, BCL2A1, and EFNB1 were significantly lower in B-ALL group as compared to controls. The high-risk group had a significantly higher percentage of infiltrated immune cells. CONCLUSION: We constructed a novel three-gene signature with independent prognostic factor for predicting 5-year OS of CAYAs with B-ALL. Additionally, we discovered the difference of immune cells composition between high-risk and low-risk groups. This study may help to customize individual treatment and improve prognosis of CAYAs with B-ALL.


Assuntos
Biomarcadores Tumorais , Efrina-B1 , Antígenos de Histocompatibilidade Menor , NADPH Oxidase 2 , Leucemia-Linfoma Linfoblástico de Células Precursoras B , Proteínas Proto-Oncogênicas c-bcl-2 , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Criança , Efrina-B1/genética , Efrina-B1/imunologia , Humanos , Sistema Imunitário/imunologia , Sistema Imunitário/patologia , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/imunologia , NADPH Oxidase 2/genética , NADPH Oxidase 2/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Prognóstico , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/imunologia , RNA Mensageiro/genética , Adulto Jovem
5.
Arch Oral Biol ; 137: 105389, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35299000

RESUMO

OBJECTIVE: EFNB1 mutation causes craniofrontonasal dysplasia (CFND), a congenital syndrome associated with craniomaxillofacial anomalies characterised by coronal craniosynostosis, orbital hypertelorism, and midface dysplasia. The aim of this murine study was to investigate the effect of the EfnB1 conditional gene deletion in osteoprogenitor cells on the craniomaxillofacial skeletal morphology. DESIGN: The skulls of male and female mice, in which EfnB1 was deleted by Cre (a site-specific DNA recombinase) under the control of the Osterix (Osx) promoter (EfnB1OB-/-), were compared to those without EfnB1 deletion (Osx:Cre control) at two ages (4 and 8 weeks; n = 6 per group). The three-dimensional micro-computed tomography reconstructions were prepared to calculate 17 linear measurements in the cranial vault (brain box), midface and mandible. Coronal and sagittal sutures from the 8-week-old mice were also subjected to histological examination. RESULTS: EfnB1OB-/- mice displayed significantly larger cranial height, larger interorbital and nasal widths, smaller maxillary width than controls by 8 weeks (p < 0.05), but mandibular size was not significantly different (p > 0.05). Binomial testing showed significantly smaller EfnB1OB-/- skulls at 4 weeks but larger at 8 weeks (p < 0.05). Histological examination revealed increased bony fusion and fibrous connective tissue deposition at the coronal suture of EfnB1OB-/- mice compared with controls. CONCLUSIONS: Craniofacial phenotype of the murine model of EfnB1 deletion in osteoprogenitor cells partially represents the human CFND phenotype, with implications for better understanding mechanisms involved in skeletal morphogenesis and malocclusion.


Assuntos
Efrina-B1 , Desenvolvimento Maxilofacial , Crânio , Animais , Suturas Cranianas/diagnóstico por imagem , Modelos Animais de Doenças , Efrina-B1/genética , Face , Feminino , Masculino , Desenvolvimento Maxilofacial/genética , Camundongos , Fenótipo , Crânio/diagnóstico por imagem , Microtomografia por Raio-X
6.
Dev Dyn ; 251(7): 1138-1155, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35025117

RESUMO

BACKGROUND: Variation in facial shape may arise from the combinatorial or overlapping actions of paralogous genes. Given its many members, and overlapping expression and functions, the EPH receptor family is a compelling candidate source of craniofacial morphological variation. We performed a detailed morphometric analysis of an allelic series of E14.5 Ephb1-3 receptor mutants to determine the effect of each paralogous receptor gene on craniofacial morphology. RESULTS: We found that Ephb1, Ephb2, and Ephb3 genotypes significantly influenced facial shape, but Ephb1 effects were weaker than Ephb2 and Ephb3 effects. Ephb2-/- and Ephb3-/- mutations affected similar aspects of facial morphology, but Ephb3-/- mutants had additional facial shape effects. Craniofacial differences across the allelic series were largely consistent with predicted additive genetic effects. However, we identified a potentially important nonadditive effect where Ephb1 mutants displayed different morphologies depending on the combination of other Ephb paralogs present, where Ephb1+/- , Ephb1-/- , and Ephb1-/- ; Ephb3-/- mutants exhibited a consistent deviation from their predicted facial shapes. CONCLUSIONS: This study provides a detailed assessment of the effects of Ephb receptor gene paralogs on E14.5 mouse facial morphology and demonstrates how the loss of specific receptors contributes to facial dysmorphology.


Assuntos
Efrina-B1 , Desenvolvimento Maxilofacial , Receptor EphB1 , Receptor EphB3 , Receptores da Família Eph , Animais , Efrina-B1/genética , Efrina-B1/metabolismo , Face , Camundongos , Mutação , Receptor EphB1/genética , Receptor EphB2/genética , Receptor EphB3/genética , Receptores da Família Eph/metabolismo
7.
Orphanet J Rare Dis ; 16(1): 286, 2021 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-34174922

RESUMO

BACKGROUND: Craniofrontonasal syndrome (CFNS) is a rare X-linked disorder that results from pathogenic variants in the EFNB1 gene. The syndrome paradoxically presents with greater severity of the symptoms in heterozygous females than hemizygous males. RESULTS: We have recruited and screened a female cohort affected with CFNS. Our primary finding was the description of monozygotic twins, i.e., patients 5 and 6, discordant for the CFNS phenotype. Intriguingly, patient 5 presented classical CFNS gestalt, whereas patient 6 manifested only very subtle craniofacial features, not resembling CFNS. Besides, we have expanded the mutational spectrum of the EFNB1 gene through reporting four novel pathogenic variants-p.(Trp12*), p.(Cys64Phe), p.(Tyr73Metfs*86), p.(Glu210*). All those alterations were found applying either targeted NGS of a custom gene panel or PCR followed by Sanger sequencing and evaluated using in silico predictors. Lastly, we have also expanded the CFNS phenotypic spectrum by describing in patient 3 several novel features of the syndrome, such as bifid hallux, bicornuate uterus, and abnormal right ovary segmented into six parts. CONCLUSIONS: We have described the unreported so far differences of the clinical phenotype in the monozygotic twin patients 5 and 6 harboring an identical p.(Glu210*) variant located in the EFNB1 gene. With our finding, we have pointed to an unusual phenomenon of mildly affected females with CFNS, who may not manifest features suggestive of the syndrome. Consequently, this study may be valuable for geneticists consulting patients with craniofacial disorders.


Assuntos
Anormalidades Craniofaciais , Efrina-B1 , Anormalidades Craniofaciais/genética , Efrina-B1/genética , Feminino , Heterozigoto , Humanos , Masculino , Mutação/genética
8.
Reprod Biol ; 21(3): 100511, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33991765

RESUMO

Ephrin receptors and ligands are membrane-bound molecules that modulate diverse cellular functions such as cell adhesion, epithelial-mesenchymal transition, motility, differentiation and proliferation. We recently reported the co-expression of ephrin-B1 and EphB4 in adult and foetal Leydig cells of the mouse testis, and thus speculated that their co-expression is a common property in gonadal steroidogenic cells. Therefore, in this study we examined the expression and localisation of ephrin-B1 and EphB4 in the naturally cycling mouse ovary, as their expression patterns in the ovary are virtually unknown. We found that ephrin-B1 and EphB4 were co-expressed in steroidogenic cells of all kinds, i.e. granulosa cells and CYP17A1-positive steroidogenic theca cells as well as in 3ß-HSD-positive luteal cells and the interstitial glands; their co-expression potentially serves as a good marker to identify sex steroid-producing cells even in extra-gonadal organs/tissues. We also found that ephrin-B1 and EphB4 expression in granulosa cells was faint and strong, respectively; ephrin-B1 expression in luteal cells was weak in developing and temporally mature corpora lutea (those of the current cycle) and likely strong in regressing corpora lutea (those of the previous cycle) and EphB4 expression in luteal cells was weak in corpora lutea of the current cycle and likely faint/negative in the corpora lutea of the previous cycle. These findings suggest that a luteinising hormone surge triggers the upregulation of ephrin-B1 and downregulation of EphB4, as this expression fluctuation occurs after the surge. Overall, ephrin-B1 and EphB4 expression patterns may represent benchmarks for steroidogenic cells in the ovary.


Assuntos
Efrina-B1/metabolismo , Regulação da Expressão Gênica/fisiologia , Ovário/metabolismo , Receptor EphB4/metabolismo , Animais , Efrina-B1/genética , Feminino , Camundongos , Ovário/citologia , Transporte Proteico , Receptor EphB4/genética
9.
Genes (Basel) ; 12(2)2021 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-33525395

RESUMO

The majority of colorectal cancers harbor loss-of-function mutations in APC, a negative regulator of canonical Wnt signaling, leading to intestinal polyps that are predisposed to malignant progression. Comparable murine APC alleles also evoke intestinal polyps, which are typically confined to the small intestine and proximal colon, but do not progress to carcinoma in the absence of additional mutations. The Cdx transcription factors Cdx1 and Cdx2 are essential for homeostasis of the intestinal epithelium, and loss of Cdx2 has been associated with more aggressive subtypes of colorectal cancer in the human population. Consistent with this, concomitant loss of Cdx1 and Cdx2 in a murine APC mutant background leads to an increase in polyps throughout the intestinal tract. These polyps also exhibit a villous phenotype associated with the loss of EphrinB1. However, the basis for these outcomes is poorly understood. To further explore this, we modeled Cdx2 loss in SW480 colorectal cancer cells. We found that Cdx2 impacted Notch signaling in SW480 cells, and that EphrinB1 is a Notch target gene. As EphrinB1 loss also leads to a villus tumor phenotype, these findings evoke a mechanism by which Cdx2 impacts colorectal cancer via Notch-dependent EphrinB1 signaling.


Assuntos
Fator de Transcrição CDX2/genética , Efrina-B1/genética , Regulação da Expressão Gênica , Mucosa Intestinal/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais , Biomarcadores , Fator de Transcrição CDX2/metabolismo , Linhagem Celular Tumoral , Humanos , Mucosa Intestinal/patologia , Modelos Biológicos , Regiões Promotoras Genéticas
10.
Development ; 148(3)2021 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-33462110

RESUMO

Rab11 family-interacting protein 5 (Rab11fip5) is an adaptor protein that binds to the small GTPase Rab11, which has an important function in endosome recycling and trafficking of cellular proteins to the plasma membrane. Rab11fip5 is involved in many cellular processes, such as cytoskeleton rearrangement, iron uptake and exocytosis in neuroendocrine cells, and is also known as a candidate gene for autism-spectrum disorder. However, the role of Rab11fip5 during early embryonic development is not clearly understood. In this study, we identified Rab11fip5 as a protein that interacts with ephrinB1, a transmembrane ligand for Eph receptors. The PDZ binding motif in ephrinB1 and the Rab-binding domain in Rab11fip5 are necessary for their interaction in a complex. EphrinB1 and Rab11fip5 display overlapping expression in the telencephalon of developing amphibian embryos. The loss of Rab11fip5 function causes a reduction in telencephalon size and a decrease in the expression level of ephrinB1. Moreover, morpholino oligonucleotide-mediated knockdown of Rab11fip5 decreases cell proliferation in the telencephalon. The overexpression of ephrinB1 rescues these defects, suggesting that ephrinB1 recycling by the Rab11/Rab11fip5 complex is crucial for proper telencephalon development.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Efrina-B1/metabolismo , Telencéfalo/crescimento & desenvolvimento , Telencéfalo/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Proliferação de Células , Citoesqueleto , Endossomos/metabolismo , Efrina-B1/genética , Exocitose , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Inativação de Genes , Neurogênese , Telencéfalo/citologia , Xenopus laevis
11.
Proc Natl Acad Sci U S A ; 117(49): 31157-31165, 2020 12 08.
Artigo em Inglês | MEDLINE | ID: mdl-33229577

RESUMO

We combine proximity labeling and single molecule binding assays to discover transmembrane protein interactions in cells. We first screen for candidate binding partners by tagging the extracellular and cytoplasmic regions of a "bait" protein with BioID biotin ligase and identify proximal proteins that are biotin tagged on both their extracellular and intracellular regions. We then test direct binding interactions between proximal proteins and the bait, using single molecule atomic force microscope binding assays. Using this approach, we identify binding partners for the extracellular region of E-cadherin, an essential cell-cell adhesion protein. We show that the desmosomal proteins desmoglein-2 and desmocollin-3, the focal adhesion protein integrin-α2ß1, the receptor tyrosine kinase ligand ephrin-B1, and the classical cadherin P-cadherin, all directly interact with E-cadherin ectodomains. Our data shows that combining extracellular and cytoplasmic proximal tagging with a biophysical binding assay increases the precision with which transmembrane ectodomain interactors can be identified.


Assuntos
Caderinas/genética , Efrina-B1/genética , Ligação Proteica/genética , Mapas de Interação de Proteínas/genética , Caderinas/ultraestrutura , Adesão Celular/genética , Citoplasma/genética , Citoplasma/ultraestrutura , Desmocolinas , Desmogleína 2/genética , Desmogleína 2/ultraestrutura , Desmoplaquinas/genética , Desmoplaquinas/ultraestrutura , Desmossomos/genética , Desmossomos/ultraestrutura , Efrina-B1/ultraestrutura , Humanos , Integrinas/genética , Integrinas/ultraestrutura , Microscopia de Força Atômica , Domínios Proteicos/genética , Imagem Individual de Molécula
12.
J Neurosci ; 40(36): 6854-6871, 2020 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-32801156

RESUMO

Astrocytes are implicated in synapse formation and elimination, which are associated with developmental refinements of neuronal circuits. Astrocyte dysfunctions are also linked to synapse pathologies associated with neurodevelopmental disorders and neurodegenerative diseases. Although several astrocyte-derived secreted factors are implicated in synaptogenesis, the role of contact-mediated glial-neuronal interactions in synapse formation and elimination during development is still unknown. In this study, we examined whether the loss or overexpression of the membrane-bound ephrin-B1 in astrocytes during postnatal day (P) 14-28 period would affect synapse formation and maturation in the developing hippocampus. We found enhanced excitation of CA1 pyramidal neurons in astrocyte-specific ephrin-B1 KO male mice, which coincided with a greater vGlut1/PSD95 colocalization, higher dendritic spine density, and enhanced evoked AMPAR and NMDAR EPSCs. In contrast, EPSCs were reduced in CA1 neurons neighboring ephrin-B1-overexpressing astrocytes. Overexpression of ephrin-B1 in astrocytes during P14-28 developmental period also facilitated evoked IPSCs in CA1 neurons, while evoked IPSCs and miniature IPSC amplitude were reduced following astrocytic ephrin-B1 loss. Lower numbers of parvalbumin-expressing cells and a reduction in the inhibitory VGAT/gephyrin-positive synaptic sites on CA1 neurons in the stratum pyramidale and stratum oriens layers of KO hippocampus may contribute to reduced inhibition and higher excitation. Finally, dysregulation of excitatory/inhibitory balance in KO male mice is most likely responsible for impaired sociability observed in these mice. The ability of astrocytic ephrin-B1 to influence both excitatory and inhibitory synapses during development can potentially contribute to developmental refinement of neuronal circuits.SIGNIFICANCE STATEMENT This report establishes a link between astrocytes and the development of excitatory and inhibitory balance in the mouse hippocampus during early postnatal development. We provide new evidence that astrocytic ephrin-B1 differentially regulates development of excitatory and inhibitory circuits in the hippocampus during early postnatal development using a multidisciplinary approach. The ability of astrocytic ephrin-B1 to influence both excitatory and inhibitory synapses during development can potentially contribute to developmental refinement of neuronal circuits and associated behaviors. Given widespread and growing interest in the astrocyte-mediated mechanisms that regulate synapse development, and the role of EphB receptors in neurodevelopmental disorders, these findings establish a foundation for future studies of astrocytes in clinically relevant conditions.


Assuntos
Astrócitos/metabolismo , Efrina-B1/metabolismo , Potenciais Pós-Sinápticos Excitadores , Hipocampo/metabolismo , Potenciais Pós-Sinápticos Inibidores , Animais , Proteína 4 Homóloga a Disks-Large/metabolismo , Efrina-B1/genética , Hipocampo/citologia , Hipocampo/crescimento & desenvolvimento , Hipocampo/fisiologia , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Células Piramidais/metabolismo , Células Piramidais/fisiologia , Comportamento Social , Proteína Vesicular 1 de Transporte de Glutamato/metabolismo , Proteínas Vesiculares de Transporte de Aminoácidos Inibidores/metabolismo
13.
J Craniofac Surg ; 31(4): e362-e368, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32371695

RESUMO

The purpose of this retrospective study was to assess the genetic and phenotypic features of patients with craniofrontonasal syndrome (CFNS), and the implications of the condition for multidisciplinary management.The subjects were 25 female patients with a mutation of EFNB1, who presented to the Oxford Craniofacial Unit during a 38-year period. Medical records were reviewed for genetic and phenotypic information. Mean duration of follow-up was 12.6 years (range 0-30.7 years).This study examines neurodevelopment in constituent parts, with specific reference to speech, language, and cognition in relation to genotype. Three children had deletions extending beyond the EFNB1 gene; the 2 with available data presented with speech, language, or cognitive delay. The remaining 25 patients had intragenic mutations of EFNB1. Of these 25, those assessed in detail showed variable difficulties with speech and language development; 57% had receptive language difficulties (n = 4/7) and 88% had expressive language difficulties (n = 8/9). 55% presented with speech difficulties (n = 6/11). 2/3 patients with abnormal hearing had speech difficulties; 4/5 with normal hearing had normal speech development. Cognitive assessments indicated that IQ is variable; with full scale IQ ranging from 69 to 100.The complex, multifactorial presentation of patients with CFNS contributed to 41% (n = 7/17) of patients requiring additional educational support.Our results demonstrated significant multidisciplinary input is required, including Speech and Language Therapy, Plastic and Reconstructive Surgery, Genetics, Ear, Nose and Throat, Maxillofacial, Orthodontic, Orthopaedic, Clinical Psychology and Orthoptic teams. The results of this study reinforce the importance of multi-disciplinary long-term follow-up of children with CFNS.


Assuntos
Anormalidades Craniofaciais , Adolescente , Adulto , Criança , Pré-Escolar , Cognição , Anormalidades Craniofaciais/genética , Anormalidades Craniofaciais/terapia , Efrina-B1/genética , Feminino , Humanos , Lactente , Recém-Nascido , Desenvolvimento da Linguagem , Masculino , Mutação , Estudos Retrospectivos , Distúrbios da Fala/terapia , Fonoterapia , Adulto Jovem
14.
J Biol Chem ; 295(22): 7653-7668, 2020 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-32321761

RESUMO

The erythropoietin-producing human hepatocellular receptor EPH receptor B6 (EPHB6) is a receptor tyrosine kinase that has been shown previously to control catecholamine synthesis in the adrenal gland chromaffin cells (AGCCs) in a testosterone-dependent fashion. EPHB6 also has a role in regulating blood pressure, but several facets of this regulation remain unclear. Using amperometry recordings, we now found that catecholamine secretion by AGCCs is compromised in the absence of EPHB6. AGCCs from male knockout (KO) mice displayed reduced cortical F-actin disassembly, accompanied by decreased catecholamine secretion through exocytosis. This phenotype was not observed in AGCCs from female KO mice, suggesting that testosterone, but not estrogen, contributes to this phenotype. Of note, reverse signaling from EPHB6 to ephrin B1 (EFNB1) and a 7-amino acid-long segment in the EFNB1 intracellular tail were essential for the regulation of catecholamine secretion. Further downstream, the Ras homolog family member A (RHOA) and FYN proto-oncogene Src family tyrosine kinase (FYN)-proto-oncogene c-ABL-microtubule-associated monooxygenase calponin and LIM domain containing 1 (MICAL-1) pathways mediated the signaling from EFNB1 to the defective F-actin disassembly. We discuss the implications of EPHB6's effect on catecholamine exocytosis and secretion for blood pressure regulation.


Assuntos
Glândulas Suprarrenais/enzimologia , Catecolaminas/metabolismo , Células Cromafins/enzimologia , Exocitose , Receptor EphB6/metabolismo , Transdução de Sinais , Glândulas Suprarrenais/citologia , Animais , Catecolaminas/genética , Células Cromafins/citologia , Efrina-B1/genética , Efrina-B1/metabolismo , Feminino , Masculino , Camundongos , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-fyn/genética , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Receptor EphB6/genética , Caracteres Sexuais , Proteína rhoA de Ligação ao GTP/genética , Proteína rhoA de Ligação ao GTP/metabolismo
15.
Eur J Med Genet ; 63(6): 103924, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32240825

RESUMO

Craniofrontonasal syndrome (CFNS) is an X-linked disorder caused by mutations in EFNB1. Uncommonly and paradoxically, female patients with CFNS exhibit significantly more severe symptoms than male patients. This is explained by "cellular interference". Nevertheless, there have been a few reports of male patients severely affected with CFNS due to postzygotic mosaicism. Here, we demonstrated a male patient with severe CFNS. Whole exome sequencing showed that he harbored both wild type and nonsense mutation, c.253C > T (p.Gln85Ter), in the EFNB1 gene. Sanger sequencing of his leukocytes, buccal swab, and hair root revealed a variable level of mosaicism. This nonsense mutation is absent in his parents and has never been previously reported. Our findings expand the mutational spectrum of EFNB1 and substantiates that males with severely affected CFNS are mosaic.


Assuntos
Códon sem Sentido , Anormalidades Craniofaciais/genética , Efrina-B1/genética , Pré-Escolar , Anormalidades Craniofaciais/patologia , Humanos , Masculino , Mosaicismo
16.
PLoS Genet ; 16(2): e1008300, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32092051

RESUMO

Craniofrontonasal syndrome (CFNS) is a rare X-linked disorder characterized by craniofacial, skeletal, and neurological anomalies and is caused by mutations in EFNB1. Heterozygous females are more severely affected by CFNS than hemizygous males, a phenomenon called cellular interference that results from EPHRIN-B1 mosaicism. In Efnb1 heterozygous mice, mosaicism for EPHRIN-B1 results in cell sorting and more severe phenotypes than Efnb1 hemizygous males, but how craniofacial dysmorphology arises from cell segregation is unknown and CFNS etiology therefore remains poorly understood. Here, we couple geometric morphometric techniques with temporal and spatial interrogation of embryonic cell segregation in mouse mutant models to elucidate mechanisms underlying CFNS pathogenesis. By generating EPHRIN-B1 mosaicism at different developmental timepoints and in specific cell populations, we find that EPHRIN-B1 regulates cell segregation independently in early neural development and later in craniofacial development, correlating with the emergence of quantitative differences in face shape. Whereas specific craniofacial shape changes are qualitatively similar in Efnb1 heterozygous and hemizygous mutant embryos, heterozygous embryos are quantitatively more severely affected, indicating that Efnb1 mosaicism exacerbates loss of function phenotypes rather than having a neomorphic effect. Notably, neural tissue-specific disruption of Efnb1 does not appear to contribute to CFNS craniofacial dysmorphology, but its disruption within neural crest cell-derived mesenchyme results in phenotypes very similar to widespread loss. EPHRIN-B1 can bind and signal with EPHB1, EPHB2, and EPHB3 receptor tyrosine kinases, but the signaling partner(s) relevant to CFNS are unknown. Geometric morphometric analysis of an allelic series of Ephb1; Ephb2; Ephb3 mutant embryos indicates that EPHB2 and EPHB3 are key receptors mediating Efnb1 hemizygous-like phenotypes, but the complete loss of EPHB1-3 does not fully recapitulate the severity of CFNS-like Efnb1 heterozygosity. Finally, by generating Efnb1+/Δ; Ephb1; Ephb2; Ephb3 quadruple knockout mice, we determine how modulating cumulative receptor activity influences cell segregation in craniofacial development and find that while EPHB2 and EPHB3 play an important role in craniofacial cell segregation, EPHB1 is more important for cell segregation in the brain; surprisingly, complete loss of EPHB1-EPHB3 does not completely abrogate cell segregation. Together, these data advance our understanding of the etiology and signaling interactions underlying CFNS dysmorphology.


Assuntos
Movimento Celular/genética , Anormalidades Craniofaciais/genética , Efrina-B1/genética , Crista Neural/embriologia , Crânio/anormalidades , Animais , Anormalidades Craniofaciais/diagnóstico , Modelos Animais de Doenças , Embrião de Mamíferos , Desenvolvimento Embrionário/genética , Efrina-B1/metabolismo , Feminino , Heterozigoto , Humanos , Masculino , Camundongos , Camundongos Knockout , Mosaicismo , Mutação , Crista Neural/citologia , Fenótipo , Receptores da Família Eph/genética , Receptores da Família Eph/metabolismo , Índice de Gravidade de Doença , Fatores Sexuais , Crânio/embriologia , Cromossomo X/genética
17.
J Biol Chem ; 295(12): 3932-3944, 2020 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-32060095

RESUMO

Eph receptors are a family of receptor tyrosine kinases that control directional cell movement during various biological processes, including embryogenesis, neuronal pathfinding, and tumor formation. The biochemical pathways of Eph receptors are context-dependent in part because of the varied composition of a heterotypic, oligomeric, active Eph receptor complex. Downstream of the Eph receptors, little is known about the essential phosphorylation events that define the context and instruct cell movement. Here, we define a pathway that is required for Eph receptor B2 (EphB2)-mediated cell sorting and is conserved among multiple Eph receptors. Utilizing a HEK293 model of EphB2+/ephrinB1+ cell segregation, we found that the scaffold adaptor protein SH2 domain-containing adaptor protein B (Shb) is essential for EphB2 functionality. Further characterization revealed that Shb interacts with known modulators of cytoskeletal rearrangement and cell mobility, including Nck adaptor protein (Nck), p120-Ras GTPase-activating protein (RasGAP), and the α- and ß-Chimaerin Rac GAPs. We noted that phosphorylation of Tyr297, Tyr246, and Tyr336 of Shb is required for EphB2-ephrinB1 boundary formation, as well as binding of Nck, RasGAP, and the chimaerins, respectively. Similar complexes were formed in the context of EphA4, EphA8, EphB2, and EphB4 receptor activation. These results indicate that phosphotyrosine-mediated signaling through Shb is essential in EphB2-mediated heterotypic cell segregation and suggest a conserved function for Shb downstream of multiple Eph receptors.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Quimerinas/metabolismo , Proteínas Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Receptor EphB2/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/genética , Separação Celular , Proteínas Quimerinas/química , Efrina-B1/genética , Efrina-B1/metabolismo , Células HEK293 , Humanos , Espectrometria de Massas , Proteínas Oncogênicas/química , Fosforilação , Ligação Proteica , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/genética , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Proteínas de Ligação a RNA/química , Receptor EphB2/química , Receptor EphB2/genética , Transdução de Sinais , Domínios de Homologia de src
18.
Am J Med Genet A ; 182(5): 1223-1229, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32022998

RESUMO

We report a female patient with craniofrontonasal syndrome (CFNS) who in addition showed other cranial and extracranial midline defects including partial corpus callosum agenesis, ocular melanocytosis, pigmentary glaucoma, duplex collecting system, uterus didelphys, and septate vagina. She was found to have a novel pathogenic variant in exon 5 of EFNB1, c.646G>T (p.Glu216*) predicted to cause premature protein truncation. From our review, we found at least 39 published CFNS patients with extracranial midline defects, comprising congenital diaphragmatic hernia, congenital heart defects, umbilical hernia, hypospadias, and less frequently, sacrococcygeal teratomas, and internal genital anomalies in females. These findings support that the EFNB1 mutations have systemic consequences disrupting morphogenetic events at the extracranial midline. Though these are not rigorously included as midline defects, we found at least 10 CFNS patients with congenital anomalies of the kidney and urinary tract, all females. Additionally, uterus didelphys and ocular melanocytosis observed in our patient are proposed also as a previously unreported EFNB1-related midline defects. In addition, this case may be useful for considering the intentional search for genitourinary anomalies in future patients with CFNS, which will be helpful to define their frequency in this entity.


Assuntos
Agenesia do Corpo Caloso/genética , Anormalidades Craniofaciais/genética , Efrina-B1/genética , Hérnias Diafragmáticas Congênitas/genética , Agenesia do Corpo Caloso/diagnóstico por imagem , Agenesia do Corpo Caloso/patologia , Criança , Pré-Escolar , Anormalidades Craniofaciais/diagnóstico por imagem , Anormalidades Craniofaciais/patologia , Éxons/genética , Feminino , Hérnias Diafragmáticas Congênitas/diagnóstico por imagem , Hérnias Diafragmáticas Congênitas/patologia , Heterozigoto , Humanos , Lactente , Masculino , Mutação/genética , Crânio/diagnóstico por imagem , Crânio/patologia
19.
Am J Med Genet A ; 182(2): 348-356, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31837199

RESUMO

Craniosynostosis (CS), the premature closure of one or more cranial sutures, occurs both as part of a syndrome or in isolation (nonsyndromic form). Here, we have studied the prevalence and spectrum of genetic alterations associated with coronal suture closure in 100 Scandinavian patients treated at a single craniofacial unit. All patients were phenotypically assessed and analyzed with a custom-designed 63 gene NGS-panel. Most cases (78%) were syndromic forms of CS. Pathogenic and likely pathogenic variants explaining the phenotype were found in 80% of the families with syndromic CS and in 14% of those with nonsyndromic CS. Sixty-five percent of the families had mutations in the CS core genes FGFR2, TWIST1, FGFR3, TCF12, EFNB1, FGFR1, and POR. Five novel pathogenic/likely pathogenic variants in TWIST1, TCF12, and EFNB1 were identified. We also found novel variants in SPECC1L, IGF1R, and CYP26B1 with a possible modulator phenotypic effect. Our findings demonstrate that NGS targeted sequencing is a powerful tool to detect pathogenic mutations in patients with coronal CS and further emphasize the importance of thorough assessment of the patient's phenotype for reliable interpretation of the molecular findings. This is particularly important in patients with complex phenotypes and rare forms of CS.


Assuntos
Anormalidades Craniofaciais/genética , Craniossinostoses/genética , Predisposição Genética para Doença , Sequenciamento de Nucleotídeos em Larga Escala , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Suturas Cranianas/patologia , Anormalidades Craniofaciais/epidemiologia , Anormalidades Craniofaciais/patologia , Craniossinostoses/epidemiologia , Craniossinostoses/patologia , Sistema Enzimático do Citocromo P-450/genética , Efrina-B1/genética , Feminino , Humanos , Masculino , Mutação/genética , Proteínas Nucleares/genética , Fenótipo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Suécia/epidemiologia , Proteína 1 Relacionada a Twist/genética
20.
Am J Pathol ; 190(2): 333-346, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31837290

RESUMO

Ephrin-B1 plays a critical role at slit diaphragm. Partitioning-defective (Par)-6 is down-regulated in podocyte of ephrin-B1 knockout mouse, suggesting that Par-6 is associated with ephrin-B1. Par polarity complex, consisting of Par-6, Par-3, and atypical protein kinase C, is essential for tight junction formation. In this study, the expression of Par-6 was analyzed in the normal and nephrotic syndrome model rats, and the molecular association of Par-6, Par-3, ephrin-B1, and nephrin was assessed with the human embryonic kidney 293 cell expression system. Par-6 was concentrated at slit diaphragm. Par 6 interacted with ephrin-B1 but not with nephrin, and Par-3 interacted with nephrin but not with ephrin-B1. The complexes of Par-6-ephrin-B1 and Par-3-nephrin were linked via extracellular sites of ephrin-B1 and nephrin. The Par-6-ephrin-B1 complex was delinked from the Par-3-nephrin complex, and Par-6 and ephrin-B1 were clearly down-regulated already at early phase of nephrotic model. The alteration of Par-6/ephrin-B1 advanced that of Par-3/nephrin. Stimulation to nephrin phosphorylated not only nephrin but also ephrin-B1, and consequently inhibited the interaction between ephrin-B1 and Par-6. Par-6 appeared at presumptive podocyte of early developmental stage and moved to basal area at capillary loop stage to participate in slit diaphragm formation at the final stage. Par-6-ephrin-B1 interaction is crucial for formation and maintenance of slit diaphragm of podocyte.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Proteínas de Transporte/metabolismo , Efrina-B1/metabolismo , Glomérulos Renais/citologia , Proteínas de Membrana/metabolismo , Síndrome Nefrótica/patologia , Podócitos/citologia , Animais , Animais Recém-Nascidos , Proteínas de Transporte/genética , Diafragma , Efrina-B1/genética , Células HEK293 , Humanos , Glomérulos Renais/metabolismo , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Síndrome Nefrótica/metabolismo , Fosforilação , Podócitos/metabolismo , Ratos , Ratos Wistar
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