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1.
Vestn Otorinolaringol ; 85(2): 14-20, 2020.
Artigo em Russo | MEDLINE | ID: mdl-32476383

RESUMO

OBJECTIVE: The description of a clinical picture and audiological features at the hearing loss caused by changes of a STRC gene, coding protein stereocillin (MIM: 606440). Mutations in the numerous genes responsible for the inner ear proteins are the reason for congenital sensorineural hearing loss. The main cause of congenital bilateral sensorineural hearing loss in the Russian Federation are mutations in GJB2 gene it reaches up 68% of cases identified in infancy. GJB2 gene tests already became routine around the world. Possibilities of new methods based on sequencing of new generation (NGS, next generation sequencing) allow to conduct a research of more rare genes connected with a hearing impairment. The most often among GJB2 negative patients reveal mutations and deletion of a gene of STRC. PATIENTS AND METHODS: Full audiological examination of 5 children and one adult with a hearing loss from 2 unrelated families is provided. Mutations in STRC gene were identified. All children are examined aged before 8 years, and 3 children failed universal audiological screening in maternity hospital, to two children screening was not carried out as they were born till 2009. RESULTS: The children with the sensorineural hearing loss connected with mutations and deletion of STRC gene failed hearing screening in maternity hospital because of the OAE is not registered, what indicates the congenital nature of a hearing loss. Recently it could not be noticed earlier because of slight increase of hearing thresholds and was regarded only as the early onset. Our data emphasize that the of thresholds from 35 to 60 dB in frequencies 0,5-4 kHz is common for mutations/deletions of STRC gene. CONCLUSION: The development of molecular genetics methods confirms the hereditary causes of GJB2-negative patients and expands indications for family counseling. Special approach for child with hearing loss so early revealed is necessary and the consultation of parents frightened of screening results is very important.


Assuntos
Surdez , Perda Auditiva Neurossensorial , Perda Auditiva , Adulto , Criança , Conexina 26 , Conexinas/genética , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Proteínas de Membrana/genética , Mutação , Gravidez , Federação Russa
2.
Nat Struct Mol Biol ; 27(4): 373-381, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32231289

RESUMO

The plasma membrane adenosine triphosphate (ATP) release channel pannexin 1 (PANX1) has been implicated in many physiological and pathophysiological processes associated with purinergic signaling, including cancer progression, apoptotic cell clearance, inflammation, blood pressure regulation, oocyte development, epilepsy and neuropathic pain. Here we present near-atomic-resolution structures of human and frog PANX1 determined by cryo-electron microscopy that revealed a heptameric channel architecture. Compatible with ATP permeation, the transmembrane pore and cytoplasmic vestibule were exceptionally wide. An extracellular tryptophan ring located at the outer pore created a constriction site, potentially functioning as a molecular sieve that restricts the size of permeable substrates. The amino and carboxyl termini, not resolved in the density map, appeared to be structurally dynamic and might contribute to narrowing of the pore during channel gating. In combination with functional characterization, this work elucidates the previously unknown architecture of pannexin channels and establishes a foundation for understanding their unique channel properties.


Assuntos
Trifosfato de Adenosina/química , Membrana Celular/ultraestrutura , Conexinas/ultraestrutura , Microscopia Crioeletrônica , Proteínas do Tecido Nervoso/ultraestrutura , Trifosfato de Adenosina/genética , Animais , Anuros/genética , Membrana Celular/química , Membrana Celular/genética , Conexinas/química , Conexinas/genética , Humanos , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Conformação Proteica , Transdução de Sinais/genética
3.
Invest Ophthalmol Vis Sci ; 61(3): 25, 2020 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-32182330

RESUMO

Purpose: To investigate the underlying mechanisms for how the mouse Cx50-R205G point mutation, a homologue of the human Cx50-R198W mutation that is linked to cataract-microcornea syndrome, affects proper lens growth and fiber cell differentiation to lead to severe lens phenotypes. Methods: EdU labeling, immunostaining, confocal imaging analysis, and primary lens epithelial cell culture were performed to characterize the lens epithelial cell (LEC) proliferation and fiber cell differentiation in wild-type and Cx50-R205G mutant lenses in vivo and in vitro. Results: The Cx50-R205G mutation severely disrupts the lens size and transparency. Heterozygous and homozygous Cx50-R205G mutant and Cx50 knockout lenses all show decreased central epithelium proliferation while only the homozygous Cx50-R205G mutant lenses display obviously decreased proliferating LECs in the germinative zone of neonatal lenses. Cultured Cx50-R205G lens epithelial cells reveal predominantly reduced Cx50 gap junction staining but no change of the endoplasmic reticulum stress marker BiP. The heterozygous Cx50-R205G lens fibers show moderately disrupted Cx50 and Cx46 gap junctions while the homozygous Cx50-R205G lens fibers have drastically reduced Cx50 and Cx46 gap junctions with severely altered fiber cell shape in vivo. Conclusions: The Cx50-R205G mutation inhibits both central and equatorial lens epithelial cell proliferation to cause small lenses. This mutation also disrupts the assembly and functions of both Cx50 and Cx46 gap junctions in lens fibers to alter fiber cell differentiation and shape to lead to severe lens phenotypes.


Assuntos
Catarata/genética , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Conexinas/genética , Doenças da Córnea/genética , Células Epiteliais/patologia , Cristalino/patologia , Mutação Puntual , Animais , Animais Recém-Nascidos , Catarata/congênito , Catarata/patologia , Células Cultivadas , Doenças da Córnea/patologia , Técnica Indireta de Fluorescência para Anticorpo , Regulação da Expressão Gênica/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Confocal , Microscopia de Fluorescência
4.
Artigo em Chinês | MEDLINE | ID: mdl-32086913

RESUMO

Objective:The aim of this study is to explore the genotype and hearing phenotype of deaf infants with mutation of GJB2 gene. Method:Subjects were 121 infants with GJB2 gene mutations who were treated in the Children's Hearing Diagnosis Center of Beijing Tongren hospital. All subjects were accepted to undertake the universal newborns hearing screening(UNHS) and series of objective audiometry, including auditory brainstem response, distortion product otoacoustic emission, auditory steady-state response and other audiological tests. All subjects were screened for nine pathogenic variants in four genes or all exons of the GJB2 gene, and then were diagnosed as infants with GJB2 gene mutations. Initially, analyzing their genotypes and hearing phenotypes generally. Then, the subjects were divided into two groups according to the genotypes: T/T group(truncated/truncated mutations, 89 cases) and T/NT group(truncated/non-truncated mutations, 32 cases). Chi-square test was used to analyze the results of UNHS, hearing degree, audiogram patterns and symmetry/asymmetry of binaural hearing phenotype. Eventually, analyzing the results of UNHS. Result:The most common truncated mutation was c.235delC(64.88%, 157/242) and the most common non-truncated mutation was c.109G>A(11.16%, 27/242). The homozygous mutation of c.235delC/c.235delC was the dominant in T/T group(38.84%, 47/121), and the compound heterozygous mutation of c.235delC/c.109G>A was the dominant in T/NT group(18.18%, 22/121). 81.82%(99/121) of subjects failed in UNHS, including 74.38%(90/121) with bilateral reference, 7.44%(9/121) with a single pass. The refer rate of UNHS of group T/T and T/NT were 86.52%(77/89) and 68.75%, respectively. There was a statistically significant difference between the two groups(P<0.05). 85.95%(104/121) of subjects were diagnosed as hearing loss and 14.05%(17/121) of subjects were diagnosed as normal hearing. The degree of hearing loss: profound, severe, moderate and mild were 31.40%(38/121), 19.01%(23/121), 24.79%(30/121) and 10.74%(13/121), respectively. There was no subjects with normal hearing in T/T group and individuals with severe and profound hearing loss accounted for the highest proportion(65.17%, 58/89), while in T/NT group, normal hearing accounted for 53.13%(17/32) and mild and moderate hearing loss accounted for the highest proportion(37.5%, 12/32). There was statistically significant difference between the two groups(P<0.05). Of 104 patients(208 ears) with hearing loss, the audiogram patterns: flat, descending, ascending, residual, Valley and other types were 49.03%(102/208), 12.02%(25/208), 8.65%(18/208), 7.69%(16/204), 3.36%(7/204) and 19.23%(40/204), respectively. The two most common types in T/T group were flat(47.19%, 84/178) and other types(20.22%, 36/178), while in T/NT group were flat(60.00%, 18/30) and ascending(20.00%, 6/30). There was statistically significant difference between the two groups(P<0.05). There were 50 cases(48.07%) with symmetrical hearing phenotype and 54 cases(51.93%) with asymmetrical hearing phenotype. Asymmetry was predominant in T/T group(53.93%, 48/89), and symmetry was predominant in T/NT group(60.00%, 9/15). There was no statistically significant difference between the two groups(P>0.05). Conclusion:In this study, c.235delC/c.235delC homozygous mutation was dominant in T/T group and c.235delC/c.109G>A heterozygous mutation was dominant in T/NT Group. The hearing phenotypes in T/T group were mostly bilateral asymmetric severe hearing loss, and those in T/NT Group were bilateral symmetric mild to moderate hearing loss, special attention should be paid to the audiological characteristics of different genotypes.


Assuntos
Conexinas/genética , Surdez/genética , Análise Mutacional de DNA , Genótipo , Humanos , Lactente , Recém-Nascido , Mutação , Fenótipo
5.
Artigo em Chinês | MEDLINE | ID: mdl-32086922

RESUMO

Objective:To detect 20 common deafness gene mutations in non- syndromic deafness patients in China using PCR- RDB, and analyze and summarize the mutation data to explore the clinical value of this method. Method:The PCR- RDB and Sanger sequencing were used to detect 20 common mutations of four deafness genes(GJB2, GJB3, SLC26A4 and mtDNA) in 500 patients with non- syndromic hearing loss . The Sanger sequencing was used to compare the sensitivity, specificity, positive predictive value, negative predictive value, and total coincidence rate of the deafness mutation detected by PCR- RDB. Result:A total of 500 samples were detected. 147 wild- type samples, 81 homozygous mutant samples, 240 heterozygous mutant samples, 32 composite heterozygous mutant samples were detected using the PCR- RDB within the range of 20 gene mutations, which were identical to the Sanger sequencing results. GJB2 c.235delC and SLC26A4 c.919- 2 A>G are the most common hotspot mutations in this study, followed by mtDNA m. 1555 A>G. Compared with the Sanger sequencing method, the sensitivity, specificity, positive predictive value, negative predictive value, and total coincidence rate of the real- time fluorescence PCR melting curve method were 100%, and the Kappa value was one. Conclusion:PCR reverse dot-blot hybridization is a simple, rapid, sensitive and specific method for detecting 20 mutations of 4 common deafness genes in Chinese population, it is expected to be used in clinical detection of deafness genes in the future.


Assuntos
Análise Mutacional de DNA , Surdez/genética , China , Conexinas/genética , DNA Mitocondrial/genética , Humanos , Mutação , Reação em Cadeia da Polimerase , Transportadores de Sulfato/genética
6.
Arterioscler Thromb Vasc Biol ; 40(4): e87-e104, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32078368

RESUMO

OBJECTIVE: Impaired ALK1 (activin receptor-like kinase-1)/Endoglin/BMP9 (bone morphogenetic protein 9) signaling predisposes to arteriovenous malformations (AVMs). Activation of SMAD1/5 signaling can be enhanced by shear stress. In the genetic disease hereditary hemorrhagic telangiectasia, which is characterized by arteriovenous malformations, the affected receptors are those involved in the activation of mechanosensitive SMAD1/5 signaling. To elucidate how genetic and mechanical signals interact in AVM development, we sought to identify targets differentially regulated by BMP9 and shear stress. Approach and Results: We identify Cx37 (Connexin37) as a differentially regulated target of ligand-induced and mechanotransduced SMAD1/5 signaling. We show that stimulation of endothelial cells with BMP9 upregulated Cx37, whereas shear stress inhibited this expression. This signaling was SMAD1/5-dependent, and in the absence of SMAD1/5, there was an inversion of the expression pattern. Ablated SMAD1/5 signaling alone caused AVM-like vascular malformations directly connecting the dorsal aorta to the inlet of the heart. In yolk sacs of mouse embryos with an endothelial-specific compound heterozygosity for SMAD1/5, addition of TNFα (tumor necrosis factor-α), which downregulates Cx37, induced development of these direct connections bypassing the yolk sac capillary bed. In wild-type embryos undergoing vascular remodeling, Cx37 was globally expressed by endothelial cells but was absent in regions of enlarging vessels. TNFα and endothelial-specific compound heterozygosity for SMAD1/5 caused ectopic regions lacking Cx37 expression, which correlated to areas of vascular malformations. Mechanistically, loss of Cx37 impairs correct directional migration under flow conditions. CONCLUSIONS: Our data demonstrate that Cx37 expression is differentially regulated by shear stress and SMAD1/5 signaling, and that reduced Cx37 expression is permissive for capillary enlargement into shunts.


Assuntos
Malformações Arteriovenosas/genética , Conexinas/genética , Regulação para Baixo , Mecanotransdução Celular , Proteína Smad1/genética , Proteína Smad5/genética , Regulação para Cima , Receptores de Activinas Tipo II/metabolismo , Animais , Malformações Arteriovenosas/metabolismo , Malformações Arteriovenosas/patologia , Capilares/patologia , Células Cultivadas , Conexinas/metabolismo , Embrião de Mamíferos , Endoglina/metabolismo , Células Endoteliais/metabolismo , Feminino , Fator 2 de Diferenciação de Crescimento/metabolismo , Humanos , Masculino , Camundongos Knockout , Proteína Smad1/metabolismo , Proteína Smad5/metabolismo , Remodelação Vascular
7.
J Clin Neurosci ; 73: 311-313, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31987637

RESUMO

Charcot-Marie-Tooth (CMT) disease is the most common hereditary peripheral neuropathy. X-linked Charcot-Marie-Tooth disease in the GJB1 gene is known as CMTX1. We report a 14 years-old young man with walked unstably, bilateral strephenopodia, severe alopecia and paroxysmal bilateral upper limbs tremor without obvious muscle atrophy. Diagnostic whole-exome sequencing revealed a hemizygote missense mutation c.278 T > A in exon 2 of the GJB1 gene, with lysine at position 93 of the mature protein (p.M93K). This is the first CMT case with alopecia areata reported in the world.


Assuntos
Doença de Charcot-Marie-Tooth/diagnóstico , Conexinas/genética , Fenótipo , Adolescente , Doença de Charcot-Marie-Tooth/genética , Testes Genéticos , Humanos , Masculino , Mutação de Sentido Incorreto , Sequenciamento Completo do Exoma
8.
Proc Natl Acad Sci U S A ; 117(4): 2160-2169, 2020 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-31932428

RESUMO

In multiple sclerosis plaques, oligodendroglial connexin (Cx) 47 constituting main gap junction channels with astroglial Cx43 is persistently lost. As mice with Cx47 single knockout exhibit no demyelination, the roles of Cx47 remain undefined. We aimed to clarify the effects of oligodendroglia-specific Cx47 inducible conditional knockout (icKO) on experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein peptide (MOG35-55) in PLP/CreERT;Cx47fl/fl mice at 14 d after tamoxifen injection. Cx47 icKO mice demonstrated exacerbation of acute and chronic relapsing EAE with more pronounced demyelination than Cx47 flox (fl)/fl littermates. CD3+ T cells more abundantly infiltrated the spinal cord in Cx47 icKO than in Cx47 fl/fl mice throughout the acute to chronic phases. CXCR3-CCR6+CD4+ and IL17+IFNγ-CD4+ helper T (Th) 17 cells isolated from spinal cord and brain tissues were significantly increased in Cx47 icKO mice compared with Cx47 fl/fl mice, while MOG35-55-specific proliferation and proinflammatory cytokine production of splenocytes were unaltered. Microarray analysis of isolated microglia revealed stronger microglial activation toward proinflammatory and injury-response phenotypes with increased expressions of chemokines that can attract Th17 cells, including Ccl2, Ccl3, Ccl4, Ccl7, and Ccl8, in Cx47 icKO mice compared with Cx47 fl/fl mice. In Cx47 icKO mice, NOS2+ and MHC class II+ microglia were more enriched immunohistochemically, and A1-specific astroglial gene expressions and astroglia immunostained for C3, a representative A1 astrocyte marker, were significantly increased at the acute phase, compared with Cx47 fl/fl mice. These findings suggest that oligodendroglia-specific Cx47 ablation induces severe inflammation upon autoimmune demyelination, underscoring a critical role for Cx47 in regulating neuroinflammation.


Assuntos
Conexinas/imunologia , Esclerose Múltipla/imunologia , Oligodendroglia/imunologia , Animais , Quimiocinas/genética , Quimiocinas/imunologia , Conexinas/genética , Doenças Desmielinizantes , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Camundongos Knockout , Esclerose Múltipla/genética , Bainha de Mielina/genética , Bainha de Mielina/imunologia , Medula Espinal/imunologia , Células Th17/imunologia
9.
Curr Pharm Biotechnol ; 21(1): 79-88, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31580250

RESUMO

BACKGROUND: Large-scale population studies showed that the SNP rs1764391 of Connexin37 gene also known as Cx37 gene may play a pivotal role in the occurrence and development of acute myocardial infarction (AMI). Published results, however, are highly controversial. OBJECTIVE: This study aimed to examine the association between SNP rs1764391 of Cx37 and diseasesusceptibility, several risk factors, and gene-environment interactions of AMI in Guangxi Han Chinese population. METHODS: In this study, 344 healthy controls and 344 AMI patients of Han Chinese population were enrolled. The TaqMan assay was implemented to identify genotypes of Cx37 and allele frequencies of SNP rs1764391 in both the AMI and control groups. RESULTS: Significant differences were detected in TT genotype frequencies of SNP rs1764391 between the AMI and control groups (P < 0.05). In the context of gender stratification, the result was also statistically different in women (P < 0.05). Each variable such as age, BMI, diabetes, high blood pressure, smoking and TC was a risk factor and correlated significantly (P < 0.05) with the development of AMI. HDL-C correlated negatively with the risk of AMI (P < 0.001). BMI, smoking or alcohol consumed interacts significantly (P < 0.017) with the presence of the SNP rs1764391 CC genotype. CONCLUSION: Evidences were presented that Cx37 rs1764391 variation may contribute to the risk for AMI, especially in women and this genetic variant may prove to be a potential biomarker for AMI risk stratification and may prove to be a useful target for therapeutic intervention to further improve prognosis in high-risk patients.


Assuntos
Conexinas/genética , Infarto do Miocárdio/genética , Doença Aguda , Adulto , Idoso , Idoso de 80 Anos ou mais , China , Feminino , Frequência do Gene , Interação Gene-Ambiente , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Fatores de Risco
10.
Medicine (Baltimore) ; 98(50): e18253, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31852093

RESUMO

RATIONALE: Molecular mechanism underlying the autosomal recessive non-syndromic hearing loss (ARNSHL) is still plausible. Pathogenic mutations of the gap junction beta 2 protein (GJB2) are reported to be the primary causes of ARNSHL. PATIENT CONCERNS: A propositus was diagnosed as ARNSHL with bilateral congenital profound hearing loss. DIAGNOSIS: With microarray and target gene sequencing testing methods, a novel GJB2 mutant was found to be associated with ARNSHL in this Han Chinese family. INTERVENTIONS/OUTCOMES: Based on the finding in this research, prenatal screening of GJB2 mutation and genetic counseling are recommended to this family for their next pregnancy. Our interventions allow the family to plan informatively. LESSONS: In this family, we discovered 2 heterozygous carriers of c.113T>C variation in the GJB2 gene. The propositus, who had profound hearing loss, had inherited the c.113T>C variation from his normal mother and the c.235delC from his father.


Assuntos
Conexinas/genética , DNA/genética , Surdez/genética , Grupos Étnicos , Mutação , Adulto , China/epidemiologia , Conexinas/metabolismo , Análise Mutacional de DNA , Surdez/diagnóstico , Surdez/etnologia , Feminino , Humanos , Lactente , Masculino , Emissões Otoacústicas Espontâneas/fisiologia , Linhagem , Prevalência
11.
BMC Neurol ; 19(1): 325, 2019 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-31842800

RESUMO

BACKGROUND: The X-linked form of Charcot-Marie-Tooth disease type 1 (CMTX1) is an inherited peripheral neuropathy that arises in patients with mutations in the gap-junction beta-1 gene (GJB1). CASE PRESENTATION: Three young male patients from Southern China with pes cavus experienced multiple episodes of transient central nervous system (CNS) dysfunction. Three patients all had reversible posterior leukoencephalopathy as detected by brain diffusion-weighted magnetic resonance imaging (MRI-DWI). Nerve conduction velocity (NCV) showed sensorimotor polyneuropathy with mixed demyelinating and axonal features. Genetic testing indicated a c.425G > A (p.Arg142Glu) or c.563 C > T (p.Thr188Ile) or c.103G > C (p.Val35Leu) mutation in GJB1. The unique feature of this report is the identification of two novel mutations: c.563 C > T and sc.103G > C of the GJB1 gene detected in two families respectively. Another unique feature is that peripheral neuropathy symptoms in the three patients were insidious and found at the onset of CNS symptoms. CONCLUSIONS: Posterior leukoencephalopathy is involved in CMTX1 patients. The white matter changes in MRI of CMTX1 patients are reversible and recover later than CNS symptoms.


Assuntos
Doença de Charcot-Marie-Tooth/complicações , Doença de Charcot-Marie-Tooth/genética , Conexinas/genética , Leucoencefalopatias/genética , Adolescente , China , Testes Genéticos , Humanos , Leucoencefalopatias/patologia , Masculino , Mutação , Linhagem
12.
Behav Neurol ; 2019: 1630718, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31871493

RESUMO

The advent of cochlear implants has enormously improved the quality of sensory perception in deaf children. Notwithstanding these advantages, the current literature shows a substantial variability in language proficiency among implanted children. This case series explores the variability of language acquisition in congenitally deaf children with cochlear implants. We report 4 prelingually deaf children (mean age = 10.5; SD = 1.08), affected by a genetically determined bilateral deafness, due to GJB2 gene mutation Cx26. Each implanted child underwent a systematic assessment of speech perception and production, as well as of lexical, morphologic, and syntactic skills in both comprehension and production. Notwithstanding similar clinical histories and similarly good postimplant pure-tone audiometry, two of the four children fared very poorly in speech audiometry, whereas the other two children gained very good results. We suggest that the language impairment detected in (some) implanted children may not be fully accounted for by pure auditory thresholds and that may be the outcome of concomitant damage to core components of the child's linguistic brain.


Assuntos
Desenvolvimento Infantil/fisiologia , Desenvolvimento da Linguagem , Linguística/métodos , Audiometria da Fala/métodos , Criança , Implante Coclear/métodos , Implantes Cocleares/psicologia , Conexinas/genética , Conexinas/metabolismo , Surdez/fisiopatologia , Feminino , Humanos , Masculino , Percepção da Fala/fisiologia
13.
Integr Biol (Camb) ; 11(10): 373-383, 2019 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-31851358

RESUMO

Behavioral responses of zebrafish larvae to environmental cues are important functional readouts that should be evoked on-demand and studied phenotypically in behavioral, genetical and developmental investigations. Very recently, it was shown that zebrafish larvae execute a voluntary and oriented movement toward the positive electrode of an electric field along a microchannel. Phenotypic characterization of this response was not feasible due to larva's rapid movement along the channel. To overcome this challenge, a microfluidic device was introduced to partially immobilize the larva's head while leaving its mid-body and tail unrestrained in a chamber to image motor behaviors in response to electric stimulation, hence achieving quantitative phenotyping of the electrically evoked movement in zebrafish larvae. The effect of electric current on the tail-beat frequency and response duration of 5-7 days postfertilization zebrafish larvae was studied. Investigations were also performed on zebrafish exposed to neurotoxin 6-hydroxydopamine and larvae carrying a pannexin1a (panx1a) gene knockout, as a proof of principle applications to demonstrate on-demand movement behavior screening in chemical and mutant assays. We demonstrated for the first time that 6-hydroxydopamine leads to electric response impairment, levodopa treatment rescues the response and panx1a is involved in the electrically evoked movement of zebrafish larvae. We envision that our technique is broadly applicable as a screening tool to quantitatively examine zebrafish larvae's movements in response to physical and chemical stimulations in investigations of Parkinson's and other neurodegenerative diseases, and as a tool to combine recent advances in genome engineering of model organisms to uncover the biology of electric response.


Assuntos
Análise Mutacional de DNA , Estimulação Elétrica , Mutação , Peixe-Zebra/embriologia , Animais , Animais Geneticamente Modificados , Comportamento Animal , Conexinas/genética , Dispositivos Lab-On-A-Chip , Levodopa/farmacologia , Oxidopamina/toxicidade , Fenótipo , Fatores de Risco , Proteínas de Peixe-Zebra/genética
14.
J Int Adv Otol ; 15(3): 373-378, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31846914

RESUMO

OBJECTIVES: The aim of the present study was to investigate the presence of GJB2, GJB3, and GJB6 gene mutations in non-syndromic sensorineural hearing loss (NSHL) cases living in Sivas region, to provide appropriate genetic counseling for cases who were found to have mutation, and to contribute to decrease the frequency of mutant allele in the next generation and plan treatment and rehabilitation with early diagnosis. MATERIALS AND METHODS: The study included 53 unrelated cases that were diagnosed with congenital NSHL between June 2009 and March 2010. Multiplex ligation-dependent probe amplification method was used for genotyping of GJB2, GJB3, and GJB6 gene mutations. RESULTS: Heterozygous 35delG variant was determined in 1.9% (n=1) of cases, homozygous 35delG in 15.1% (n=8), heterozygous IVS1+1G>A mutation in 1.9% (n=1), compound heterozygous in 3.8% (n=2), and homozygous IVS1+1G>A variant in 3.8% (n=2). None of the cases had mutation in GJB3 and GJB6 genes. Mutated allele frequencies in the present study were found to be 17.9% for 35delG and 6.6% for IVS1+1G>A. CONCLUSION: The present study showed that 35delG mutation is the most common variant in the Sivas region, and that IVS1+1G>A mutation should be investigated in hearing loss. Another result of the present study was that genetic analyzes would allow early diagnosis of hearing impairments particularly when infants whose parents have consanguinity do not pass the newborn hearing screening.


Assuntos
Conexina 30/genética , Conexinas/genética , Perda Auditiva Neurossensorial/genética , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Consanguinidade , Feminino , Frequência do Gene , Genótipo , Heterozigoto , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Turquia , Adulto Jovem
15.
Adv Exp Med Biol ; 1190: 301-321, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31760652

RESUMO

Charcot-Marie-Tooth (CMT) disease is the most common hereditary neuropathy and genetically heterogeneous. CMT1 and CMTX are autosomal dominant and X-linked demyelinating neuropathies, respectively. CMT1A, CMT1B, and CMTX1 are the common forms of CMT, which are attributed to the genes encoding the myelin or gap junction proteins expressed in the myelinating Schwann cells. CMT4 is a rare autosomal recessive demyelinating neuropathy that usually shows an early-onset severe phenotype. Twelve genes have been described as CMT4, which encodes many kinds of proteins including mitochondrial proteins, phosphatases in the endosomal pathway, endocytic recycling proteins, and trafficking proteins. The genes responsible for CMT4 are expressed in Schwan cells and necessary for the development and maintenance in the peripheral nervous system. However, CMT1, CMT4, and CMTX1 are primarily demyelinating neuropathies, axonal degeneration is necessary for symptoms to develop. Schwann cell-axon interactions are impaired in the pathogenesis of demyelinating CMT.


Assuntos
Doença de Charcot-Marie-Tooth/fisiopatologia , Doenças Desmielinizantes/fisiopatologia , Células de Schwann/patologia , Axônios/patologia , Doença de Charcot-Marie-Tooth/classificação , Conexinas/genética , Doenças Desmielinizantes/classificação , Humanos , Mutação , Proteínas da Mielina/genética
16.
Genes (Basel) ; 10(11)2019 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-31731535

RESUMO

This study aimed to investigate GJB2 (connexin 26) and GJB6 (connexin 30) mutations associated with familial non-syndromic childhood hearing impairment (HI) in Cameroon. We selected only families segregating HI, with at least two affected individuals and with strong evidence of non-environmental causes. DNA was extracted from peripheral blood, and the entire coding region of GJB2 was interrogated using Sanger sequencing. Multiplex PCR and Sanger sequencing were used to analyze the prevalence of the GJB6-D3S1830 deletion. A total of 93 patients, belonging to 41 families, were included in the analysis. Hearing impairment was sensorineural in 51 out of 54 (94.4%) patients. Pedigree analysis suggested autosomal recessive inheritance in 85.4% (35/41) of families. Hearing impairment was inherited in an autosomal dominant and mitochondrial mode in 12.2% (5/41) and 2.4% (1/41) of families, respectively. Most HI participants were non-syndromic (92.5%; 86/93). Four patients from two families presented with type 2 Waardenburg syndrome, and three cases of type 2 Usher syndrome were identified in one family. No GJB2 mutations were found in any of the 29 families with non-syndromic HI. Additionally, the GJB6-D3S1830 deletion was not identified in any of the HI patients. This study confirms that mutations in the GJB2 gene and the del(GJB6-D13S1830) mutation do not contribute to familial HI in Cameroon.


Assuntos
Conexina 30/genética , Conexinas/genética , Perda Auditiva Neurossensorial/genética , Mutação , Adolescente , Adulto , Idoso , Camarões , Criança , Pré-Escolar , Feminino , Perda Auditiva Neurossensorial/epidemiologia , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Linhagem
19.
Mol Med Rep ; 20(6): 5091-5099, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31638216

RESUMO

C­type natriuretic peptide (CNP), from the family of natriuretic peptides (NPs), has been shown to induce antihypertrophic and antifibrotic effects in cardiomyocytes. However, the roles of CNP in the atrial dysregulation of connexin (Cx)40 and Cx43 remain to be elucidated. The present study aimed to investigate the effects of CNP on angiotensin (Ang) II­induced Cx40 and Cx43 dysregulation in isolated perfused beating rat left atria. A rat isolated perfused beating atrial model was used and the protein levels were determined via western blotting. Ang II significantly upregulated NF­κB, activator protein­1, transforming growth factor­ß1 (TGF­ß1), collagen I and matrix metalloproteinase 2, leading to atrial fibrosis, and downregulated expression of Cx40 and Cx43. The changes in Cx40 and Cx43 induced by Ang II were abolished by CNP through upregulation of phosphorylated AMP­activated kinase a1 (AMPK) and downregulation of TGF­ß1. The effects of CNP on AMPK and TGF­ß1 levels were inhibited by KT5823 and pertussis toxin, inhibitors of protein kinase G (PKG) and NP receptor type C (NPR­C), respectively. Thus, CNP can prevent Ang II­induced dysregulation of Cx40 and Cx43 through activation of AMPK via the CNP­PKG and CNP­NPR­C pathways in isolated beating rat atria. The present findings suggested that CNP may be therapeutically useful for clinical conditions involving cardiac dysregulation of Cx expression­related diseases.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Angiotensina II/metabolismo , Conexina 43/genética , Conexinas/genética , Átrios do Coração/metabolismo , Peptídeo Natriurético Tipo C/metabolismo , Transdução de Sinais/genética , Angiotensina II/farmacologia , Animais , Biomarcadores , Conexina 43/metabolismo , Conexinas/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose , Regulação da Expressão Gênica/efeitos dos fármacos , Átrios do Coração/efeitos dos fármacos , Átrios do Coração/patologia , Masculino , Peptídeo Natriurético Tipo C/farmacologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo
20.
Am J Physiol Renal Physiol ; 317(6): F1649-F1655, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31630543

RESUMO

The severity of polycystic kidney diseases (PKD) depends on the counterbalancing of genetic predisposition and environmental factors exerting permissive or protective influence on cyst development. One poorly characterized phenomenon in the cystic epithelium is abnormal purinergic signaling. Earlier experimental studies revealed the high importance of the ionotropic P2X receptors (particularly, P2X7) in the pathophysiology of the cyst wall. To study mechanisms of P2X7 involvement in cyst growth and aspects of targeting these receptors in PKD treatment we performed a CRISPR/SpCas9-mediated global knockout of the P2rx7 gene in PCK rats, a model of autosomal recessive PKD (ARPKD). A single base insertion in exon 2 of the P2rx7 gene in the renal tissues of homozygous mutant animals leads to lack of P2X7 protein that did not affect their viability or renal excretory function. However, PCK.P2rx7 rats demonstrated slower cyst growth (but not formation of new cysts) compared with heterozygous and PCK.P2rx7+ littermates. P2X7 receptors are known to activate pannexin-1, a plasma channel capable of releasing ATP, and we found here that pannexin-1 expression in the cystic epithelium is significantly higher than in nondilated tubules. P2X7 deficiency reduces renal pannexin-1 protein expression and daily urinary ATP excretion. Patch-clamp analysis revealed that lack of P2X7 increases epithelial sodium channel activity in renal tissues and restores impaired channel activity in cysts. Interpretation of our current data in the context of earlier studies strongly suggests that P2X7 contributes to cyst growth by increasing pannexin-1-dependent pathogenic ATP release into the lumen and reduction of sodium reabsorption across the cyst walls.


Assuntos
Cistos/patologia , Nefropatias/patologia , Rim Policístico Autossômico Recessivo/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Trifosfato de Adenosina/urina , Animais , Sistemas CRISPR-Cas , Conexinas/biossíntese , Conexinas/genética , Cistos/genética , Canais Epiteliais de Sódio/metabolismo , Feminino , Técnicas de Inativação de Genes , Nefropatias/genética , Mutagênese Insercional , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Rim Policístico Autossômico Recessivo/genética , Gravidez , Ratos , Receptores Purinérgicos P2X7/genética , Sódio/metabolismo
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