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1.
Orv Hetil ; 160(37): 1447-1454, 2019 Sep.
Artigo em Húngaro | MEDLINE | ID: mdl-31495190

RESUMO

Myotonic dystrophy is one of the most common, autosomal dominantly inherited adult-onset muscle disorders. Two types of the disease are known: type 1 is characterized by distal weakness and myotonia, but type 2 is associated with proximal weakness and milder clinical course. It is also called as Steinert Disease, which affects the heart conduction system, the internal secretional glands, the ocular lens as well as carbohydrate-, fat metabolism and gonadal functions. These systemic symptoms have high impact on the quality of life and might impact on patients' survival. Here we would like to emphasize these clinical conditions and the diagnostic possibilities. We hope our recommendations can help neurologists and general practitioners to achieve an optimal and individual care for patients suffering from this muscle disease. Orv Hetil. 2019; 160(37): 1447-1454.


Assuntos
Sistema de Condução Cardíaco/fisiopatologia , Distrofia Miotônica/diagnóstico , Adulto , Catarata/genética , Humanos , Resistência à Insulina/genética , Músculos/fisiopatologia , Distrofia Miotônica/complicações , Distrofia Miotônica/genética , Doenças do Sistema Nervoso , Qualidade de Vida , Expansão das Repetições de Trinucleotídeos
2.
Adv Exp Med Biol ; 1155: 155-162, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31468394

RESUMO

Fragile X syndrome is an X-linked dominant disorder and the most common cause of inherited mental retardation. It is caused by trinucleotide repeat expansion in the fragile X mental retardation 1 gene (FMR1) at the Xq27.3. The expansion blocks expression of the gene product, Fragile X Mental Retardation Protein (FMRP). The syndrome includes mild to moderate mental retardation and behavioral manifestations such as tactile defensiveness, gaze avoidance, repetitive motor mannerisms, perseverative (repetitive) speech, hyperarousal and it frequently includes seizures. This behavioral phenotype overlaps significantly with autism spectrum disorder. The knockout mice lack normal Fmr1 protein and show macro-orchidism, learning deficits, and hyperactivity. Consequently, this knockout mouse may serve as a valuable tool in the elucidation of the physiological role of FMR1 and the mechanisms involved in macroorchidism, abnormal behavior, abnormalities comparable to those of human fragile X patients. In this study we evaluated the effects of taurine on the testicular physiology to better understand the cellular mechanisms underlying macro-orchidism. We found that there was a significant decrease in the number of Leydig cells in the testis of fragile X mouse. Furthermore, the expression of somatostatin was drastically decreased and differential expression pattern of CDK5 in fragile X mouse testis. In the control testis, CDK is expressed in primary and secondary spermatids whereas in the Fmr1 ko mice CDK 5 is expressed mainly in spermatogonia. Taurine supplementation led to an increase in CDK5 expression in both controls and Ko mice. CDKs (Cyclin-dependent kinases) are a group of serine/threonine protein kinases activated by binding to a regulatory subunit cyclin. Over 20 functionally diverse proteins involved in cytoskeleton dynamics, cell adhesion, transport, and membrane trafficking act as CDK5 substrates elucidating the molecular mechanisms of CDK5 function. CDK5 phosphorylates a diverse list of substrates, implicating it in the regulation of a range of cellular processes. CDK5 is expressed in Leydig cells, Sertoli cells, spermatogonia and peritubular cells indicating a role in spermatogenesis. In this study we examined the expression levels of CDK5 and how it is affected by taurine supplementation in the testes and found that taurine plays an important role in testicular physiology and corrected some of the pathophysiology observed in the fragile x mouse testis.


Assuntos
Quinase 5 Dependente de Ciclina/metabolismo , Proteína do X Frágil de Retardo Mental/genética , Síndrome do Cromossomo X Frágil/genética , Taurina/farmacologia , Testículo/fisiopatologia , Animais , Suplementos Nutricionais , Modelos Animais de Doenças , Síndrome do Cromossomo X Frágil/fisiopatologia , Masculino , Camundongos , Camundongos Knockout , Expansão das Repetições de Trinucleotídeos
3.
Molecules ; 24(9)2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31075983

RESUMO

Synthetic acyclic receptors, composed of two arms connected with a spacer enabling molecular recognition, have been intensively explored in host-guest chemistry in the past decades. They fall into the categories of molecular tweezers, clefts and clips, depending on the geometry allowing the recognition of various guests. The advances in synthesis and mechanistic studies have pushed them forward to pharmaceutical applications, such as neurodegenerative disorders, infectious diseases, cancer, cardiovascular disease, diabetes, etc. In this review, we provide a summary of the synthetic molecular tweezers, clefts and clips that have been reported for pharmaceutical applications. Their structures, mechanism of action as well as in vitro and in vivo results are described. Such receptors were found to selectively bind biological guests, namely, nucleic acids, sugars, amino acids and proteins enabling their use as biosensors or therapeutics. Particularly interesting are dynamic molecular tweezers which are capable of controlled motion in response to an external stimulus. They proved their utility as imaging agents or in the design of controlled release systems. Despite some issues, such as stability, cytotoxicity or biocompatibility that still need to be addressed, it is obvious that molecular tweezers, clefts and clips are promising candidates for several incurable diseases as therapeutic agents, diagnostic or delivery tools.


Assuntos
Indústria Farmacêutica , Conformação Molecular , Sistemas de Liberação de Medicamentos , Liberação Controlada de Fármacos , Humanos , Ácidos Nucleicos/química , Expansão das Repetições de Trinucleotídeos
4.
Biochimie ; 163: 21-32, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31075282

RESUMO

Huntington's diseases (HD) is a very devastating disease caused by r(CAG) expansion in HTT gene, encoding the huntingtin protein. r(CAG) expansion causes disease via multiple pathways including, 1) loss of normal protein function like sequestration of RNA binding protein such as Muscleblind-like (MBNL) and nucleolin, 2) Gain of function for mutant proteins and 3) repeat-associated non-ATG (RAN) translation; in which expanded r(CAG) translates into toxic poly glu, poly ser, or poly ala without the use of any canonical start codon. Herein, we have rationally designed and synthesized a unique class of pyridocoumarin derivatives that target the r(CAG)exp involved in HD and spinocerebellar ataxia (SCA) pathogenesis. Notably, compounds 3 and 15 showed higher affinity (nanomolar Kd) and selectivity for diseased r(CAG)exp RNA compared to regular duplex AU-paired RNA. Interestingly, both scaffolds are cell permeable, exhibit low toxicity to healthy fibroblast cells and are also capable of reducing the level of poly Q aggregation in cellular models. Indeed, our current study offers promising facet for selectively targeting repeats containing RNAs that cause severe diseases like HD and SCAs.


Assuntos
Cumarínicos/química , Doença de Huntington/tratamento farmacológico , Proteínas Mutantes/genética , RNA Mensageiro/química , Ataxias Espinocerebelares/tratamento farmacológico , Células Cultivadas , Cumarínicos/farmacologia , Cumarínicos/uso terapêutico , Desenho de Drogas , Humanos , Proteína Huntingtina , Doença de Huntington/metabolismo , Cinética , Simulação de Acoplamento Molecular , Conformação de Ácido Nucleico , RNA Mensageiro/efeitos dos fármacos , Ataxias Espinocerebelares/metabolismo , Expansão das Repetições de Trinucleotídeos
5.
Cell Physiol Biochem ; 52(5): 970-983, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30977983

RESUMO

BACKGROUND/AIMS: Regulation of mRNA translation is central to protein homeostasis and is optimized for speed and accuracy. Spontaneous recoding events occur virtually at any codon but at very low frequency and are commonly assumed to increase as the cell ages. METHODS: Here, we leveraged the polyglutamine(polyQ)-frameshifting model of huntingtin exon 1 with CAG repeat length in the pathological range (Htt51Q), which undergoes enhanced non-programmed translational -1 frameshifting. RESULTS: In body muscle cells of Caenorhabditis elegans, -1 frameshifting occured at the onset of expression of the zero-frame product, correlated with mRNA level of the non-frameshifted expression and formed aggregates correlated with reduced motility in C. elegans. Spontaneous frameshifting was modulated by IFG-1, the homologue of the nutrient-responsive eukaryotic initiation factor 4G (eIF4G), under normal growth conditions and NSUN-5, a conserved ribosomal RNA methyltransferase, under osmotic stress. CONCLUSION: Our results suggest that frameshifting and aggregation occur at even early stages of development and, because of their intrinsic stability, may persist and accelerate the onset of age-related proteinopathies.


Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Mutação da Fase de Leitura , Proteína Huntingtina , Doença de Huntington , Expansão das Repetições de Trinucleotídeos , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Modelos Animais de Doenças , Éxons , Humanos , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Doença de Huntington/genética , Doença de Huntington/metabolismo , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/metabolismo
6.
Cornea ; 38(7): 799-805, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30973406

RESUMO

PURPOSE: To investigate single nucleotide polymorphisms (SNPs) and trinucleotide repeat (TNR) expansion in the transcription factor 4 (TCF4) gene in a large cohort of German patients with Fuchs endothelial corneal dystrophy (FECD). METHODS: Genomic DNA was obtained from 398 patients with FECD and from 58 non-FECD controls. Thirty-seven previously reported SNPs were evaluated by genotyping. The 398 FECD samples were analyzed for TNR expansions by short tandem repeat assays and Southern blotting. The possible associations between the TNR length and clinical parameters (age, sex, visual acuity, and central corneal thickness) were analyzed in 132 patients. RESULTS: The SNPs in COL8A2, TCF8, LOXHD1, and AGBL1 showed no heterogeneity in 36 cases, although SLCA411 showed 3 nonsense mutations. SNPs were detected for TCF4 (rs613872, rs2123392, rs17089887, rs1452787, and rs1348047), but only rs613872 showed a significant association with FECD (P = 9.93 × 10). Overall, 315/398 (79%) patients harbored TNR lengths >50, whereas no non-FECD controls harbored TNR lengths >50. The TCF4 SNP rs613872 genotype was TT: 39 (67%), TG: 18 (31%), and GG: 1 (2%) in non-FECD controls; TT: 39 (47%), TG: 38 (46%), and GG: 6 (7%) in FECD cases harboring TNR <50; and TT: 23 (8%), TG: 224 (79%), and GG: 38 (13%) in FECD cases harboring TNR >50 (P = 2.93 × 10). No significant association was detected between the TNR length and clinical parameters. CONCLUSIONS: Our large German cohort demonstrated a significant association between the risk allele G in rs613872 and FECD, irrespective of TNR expansion, although this risk allele was more frequent in FECD cases with TNR expansion than without.


Assuntos
Distrofia Endotelial de Fuchs/genética , Predisposição Genética para Doença , Fator de Transcrição 4/genética , Expansão das Repetições de Trinucleotídeos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Adulto Jovem
7.
Methods Mol Biol ; 1942: 49-59, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30900174

RESUMO

Knowledge of the CGG•CCG-repeat number, AGG interruption status, and the extent of DNA methylation of the FMR1 gene are vital for both diagnosis of the fragile X-related disorders and for basic research into disease mechanisms. We describe here assays that we use in our laboratory to assess these parameters. Our assays are PCR-based and include one for repeat size that can also be used to assess the extent of methylation and a related assay that allows the AGG interruption pattern to be reliably determined even in women. A second more quantitative methylation assay is also described. We also describe our method for cloning of repeats to generate the reference standards necessary for the accurate determination of repeat number and AGG interruption status.


Assuntos
Metilação de DNA , Proteína do X Frágil de Retardo Mental/genética , Síndrome do Cromossomo X Frágil/diagnóstico , Síndrome do Cromossomo X Frágil/genética , Expansão das Repetições de Trinucleotídeos , Humanos
8.
Methods Mol Biol ; 1972: 199-210, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30847793

RESUMO

Fragile X mental retardation 1 (FMR1) CGG repeat expansions cause fragile X syndrome-the leading monogenic form of intellectual disability-and increase the risk for fragile X-associated tremor ataxia syndrome and fragile X-associated primary ovarian insufficiency. Southern blot (SB) analysis is the current gold standard test for FMR1 molecular diagnosis. Several polymerase chain reaction (PCR)-based methods are now available for sizing FMR1 CGG repeat expansions. These methods offer higher diagnostic sensitivity and specificity compared to SB analysis, significantly reduce the turnaround time and increase throughput. In this chapter, we describe a triplet-repeat primed PCR protocol that employs capillary electrophoresis to resolve the derived amplicon products, enabling precise determination of the FMR1 genotypes in both males and females and characterization of the CGG repeat structure.


Assuntos
Primers do DNA/metabolismo , Eletroforese Capilar/métodos , Proteína do X Frágil de Retardo Mental/genética , Reação em Cadeia da Polimerase/métodos , Expansão das Repetições de Trinucleotídeos/genética , Repetições de Trinucleotídeos/genética , Humanos , Mutação/genética
9.
Biometals ; 32(2): 307-315, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30874991

RESUMO

Friedreich's ataxia (FRDA), a progressive neurodegenerative disorder caused by trinucleotide (GAA) repeat expansion in frataxin (fxn) gene which results in decreased levels of frataxin protein. Insufficient frataxin levels leads to iron and copper deposits in the brain and cardiac cells. A total of hundred and twenty patients, suspected of FRDA were screened for the (GAA) repeats in the fxn gene and only confirmed patients (n = 25) were recruited in the study. The total Iron and total copper concentrations were measured in blood plasma using Nitro PAPS and Dibrom PAESA method, respectively both in patients and age, sex matched healthy controls. The iron levels mean ± SD (6.2 ± 3.8) in plasma of FRDA patients were found to be significantly decreased as compared to healthy controls mean ± SD (15.2 ± 4.2). A similar trend was observed in case of plasma copper levels in FRDA patient (8.15 ± 4.6) as compared to controls (17.5 ± 3.40). Present results clearly prove abnormal distribution of extra-cellular iron in FRDA patients, which is in accordance with the well established fact of intracellular iron overload, which is the key feature of the pathogenesis of this disease. This can be of importance in understanding the pathophysiology of the disease in association with frataxin/iron. It appears that intracellular sequestration of trace metals in FRDA patients (due to low frataxin) results in their sub-optimal levels in blood plasma (extra-cellular) an observation that can find prognostic application in clinical trials.


Assuntos
Cobre/sangue , Ataxia de Friedreich/sangue , Ataxia de Friedreich/patologia , Ferro/sangue , Ataxia de Friedreich/genética , Humanos , Expansão das Repetições de Trinucleotídeos/genética
10.
Mol Neurodegener ; 14(1): 9, 2019 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-30767771

RESUMO

BACKGROUND: A G4C2 hexanucleotide repeat expansion in the noncoding region of C9orf72 is the major genetic cause of frontotemporal dementia and amyotrophic lateral sclerosis (c9FTD/ALS). Putative disease mechanisms underlying c9FTD/ALS include toxicity from sense G4C2 and antisense G2C4 repeat-containing RNA, and from dipeptide repeat (DPR) proteins unconventionally translated from these RNA products. METHODS: Intracerebroventricular injections with adeno-associated virus (AAV) encoding 2 or 149 G4C2 repeats were performed on postnatal day 0, followed by assessment of behavioral and neuropathological phenotypes. RESULTS: Relative to control mice, gliosis and neurodegeneration accompanied by cognitive and motor deficits were observed in (G4C2)149 mice by 6 months of age. Recapitulating key pathological hallmarks, we also demonstrate that sense and antisense RNA foci, inclusions of poly(GA), poly(GP), poly(GR), poly(PR), and poly(PA) DPR proteins, and inclusions of endogenous phosphorylated TDP-43 (pTDP-43) developed in (G4C2)149 mice but not control (G4C2)2 mice. Notably, proteins that play a role in the regulation of stress granules - RNA-protein assemblies that form in response to translational inhibition and that have been implicated in c9FTD/ALS pathogenesis - were mislocalized in (G4C2)149 mice as early as 3 months of age. Specifically, we observed the abnormal deposition of stress granule components within inclusions immunopositive for poly(GR) and pTDP-43, as well as evidence of nucleocytoplasmic transport defects. CONCLUSIONS: Our in vivo model of c9FTD/ALS is the first to robustly recapitulate hallmark features derived from both sense and antisense C9orf72 repeat-associated transcripts complete with neurodegeneration and behavioral impairments. More importantly, the early appearance of persistent pathological stress granules prior to significant pTDP-43 deposition implicates an aberrant stress granule response as a key disease mechanism driving TDP-43 proteinopathy in c9FTD/ALS.


Assuntos
Proteína C9orf72/genética , Proteínas de Choque Térmico/metabolismo , Degeneração Neural/patologia , Proteinopatias TDP-43/patologia , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/metabolismo , Esclerose Amiotrófica Lateral/patologia , Animais , Demência Frontotemporal/genética , Demência Frontotemporal/metabolismo , Demência Frontotemporal/patologia , Camundongos , Degeneração Neural/genética , Degeneração Neural/metabolismo , Proteinopatias TDP-43/genética , Proteinopatias TDP-43/metabolismo , Expansão das Repetições de Trinucleotídeos
11.
Invest Ophthalmol Vis Sci ; 60(2): 779-786, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30811544

RESUMO

Purpose: CTG trinucleotide repeat (TNR) expansion is frequently found in transcription factor 4 (TCF4) in Fuchs' endothelial corneal dystrophy (FECD), though the effect of TNR expansion on FECD pathophysiology remains unclear. The purpose of this study was to evaluate the effect of TNR expansion on TCF4 expression in corneal endothelium of patients with FECD. Methods: Peripheral blood DNA and Descemet membrane with corneal endothelium were obtained from 203 German patients with FECD. The CTG TNR repeat length in TCF4 was determined by short tandem repeat (STR) assays and Southern blotting using genomic DNA. Genotyping of rs613872 in TCF4 was performed by PCR. TCF4 mRNA levels in corneal endothelium were evaluated by quantitative PCR using three different probes. Control corneal endothelial samples were obtained from 35 non-FECD subjects. Results: The STR assay and Southern blotting showed that 162 of the 203 patients with FECD (80%) harbored CTG trinucleotide repeat lengths larger than 50. Quantitative PCR using all three probes demonstrated that TCF4 mRNA is significantly upregulated in the corneal endothelium of patients with FECD, regardless of the presence of TNR expansion. However, the length of the TNR tended to show a positive correlation with TCF4 expression level. No correlation was shown between the genotype of TCF4 SNP, rs613872, and the level of TCF4 expression. Conclusions: Our findings showed that TCF4 mRNA is upregulated in the corneal endothelium of patients with FECD. Further studies on the effects of TCF4 upregulation on corneal endothelial cell function will aid in understanding the pathophysiology of FECD.


Assuntos
Distrofia Endotelial de Fuchs/genética , Regulação da Expressão Gênica/fisiologia , RNA Mensageiro/genética , Fator de Transcrição 4/genética , Expansão das Repetições de Trinucleotídeos , Adulto , Idoso , Idoso de 80 Anos ou mais , Southern Blotting , Feminino , Técnicas de Genotipagem , Humanos , Masculino , Repetições de Microssatélites , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Adulto Jovem
12.
Stem Cell Res ; 34: 101361, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30611021

RESUMO

Spinocerebellar Ataxia Type 2 (SCA2) is an autosomal dominant disease characterized by progressive degeneration of the cerebellum, brain stem, and spinal cord. SCA2 is caused by spontaneous misfolding and aggregate formation from abnormal CAG trinucleotide repeat expansion in the coding region of the ATXN2 gene. Here we describe the generation of two distinct iPSC lines from patients with SCA2.


Assuntos
Ataxina-2/genética , Técnicas de Cultura de Células/métodos , Células-Tronco Pluripotentes Induzidas/patologia , Fases de Leitura Aberta/genética , Ataxias Espinocerebelares/genética , Expansão das Repetições de Trinucleotídeos/genética , Adulto , Animais , Linhagem Celular , Humanos , Camundongos , Pessoa de Meia-Idade
13.
PLoS One ; 14(1): e0210996, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30682148

RESUMO

PURPOSE: To determine if CTG18.1 TNR expansion length prognosticates the clinical progression of Fuchs' Endothelial Corneal Dystrophy (FECD). METHODS: This was a prospective cohort study. A total of 51 patients with newly diagnosed FECD were recruited and followed-up over a period of 12 years, from November 2004 to April 2016. Baseline clinical measurements included central corneal thickness (CCT), endothelial cell density (ECD) and CTG18.1 TNR expansion length from peripheral leukocytes, with yearly repeat measurements of CCT and ECD. A patient was defined to have experienced significant clinical progression and to have developed Threshold Disease if any of these criteria were fulfilled in either eye: a) CCT increased to >700µm, b) ECD decreased to <700 cells/mm2, or c) underwent keratoplasty for treatment of FECD. RESULTS: Patients were categorized as having at least one allele whose maximum allele length was equal to or greater than 40 repeats (L≥40, n = 22, 43.1%), or having both alleles shorter than 40 repeats (L<40). Threshold Disease rates at the 5-year time point were 87.5% for the L≥40 group and 47.8% for the L<40 group (p = 0.012). This difference narrowed and was no longer statistically significant at the 8-years (92.9% vs 78.9%, p = 0.278) and 10-years (92.9% vs 84.2%, p = 0.426) time points. CONCLUSIONS: L≥40 patients are at greater risk of FECD progression and development of Threshold Disease within the first 5 years following diagnosis.


Assuntos
Distrofia Endotelial de Fuchs/genética , Fator de Transcrição 4/genética , Expansão das Repetições de Trinucleotídeos , Idoso , Cromossomos Humanos Par 18/genética , Estudos de Coortes , Paquimetria Corneana , Progressão da Doença , Células Endoteliais/patologia , Feminino , Distrofia Endotelial de Fuchs/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos
14.
Am J Chin Med ; 47(1): 63-95, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30612452

RESUMO

Nine autosomal dominant spinocerebellar ataxias (SCAs) are caused by an abnormal expansion of CAG trinucleotide repeats that encodes a polyglutamine (polyQ) tract within different genes. Accumulation of aggregated mutant proteins is a common feature of polyQ diseases, leading to progressive neuronal dysfunction and degeneration. SCA type 3 (SCA3), the most common form of SCA worldwide, is characterized by a CAG triplet expansion in chromosome 14q32.1 ATXN3 gene. As accumulation of the mutated polyQ protein is a possible initial event in the pathogenic cascade, clearance of aggregated protein by ubiquitin proteasome system (UPS) has been proposed to inhibit downstream detrimental events and suppress neuronal cell death. In this study, Chinese herbal medicine (CHM) extracts were studied for their proteasome-activating, polyQ aggregation-inhibitory and neuroprotective effects in GFPu and ATXN3/Q 75 -GFP 293/SH-SY5Y cells. Among the 14 tested extracts, 8 displayed increased proteasome activity, which was confirmed by 20S proteasome activity assay and analysis of ubiquitinated and fused GFP proteins in GFPu cells. All the eight extracts displayed good aggregation-inhibitory potential when tested in ATXN3/Q 75 -GFP 293 cells. Among them, neuroprotective effects of five selected extracts were shown by analyses of polyQ aggregation, neurite outgrowth, caspase 3 and proteasome activities, and ATXN3-GFP, ubiquitin, BCL2 and BAX protein levels in neuronal differentiated ATXN3/Q 75 -GFP SH-SY5Y cells. Finally, enhanced proteasome function, anti-oxidative activity and neuroprotection of catalpol, puerarin and daidzein (active constituents of Rehmannia glutinosa and Pueraria lobata) were demonstrated in GFPu and/or ATXN3/Q 75 -GFP 293/SH-SY5Y cells. This study may have therapeutic implication in polyQ-mediated disorders.


Assuntos
Antioxidantes , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Doença de Machado-Joseph/tratamento farmacológico , Doença de Machado-Joseph/genética , Fármacos Neuroprotetores , Peptídeos/genética , Peptídeos/metabolismo , Fitoterapia , Complexo de Endopeptidases do Proteassoma/metabolismo , Agregação Patológica de Proteínas/metabolismo , Ubiquitina/metabolismo , Células Cultivadas , Células HEK293 , Humanos , Glucosídeos Iridoides/farmacologia , Glucosídeos Iridoides/uso terapêutico , Isoflavonas/farmacologia , Isoflavonas/uso terapêutico , Terapia de Alvo Molecular , Mutação , Agregação Patológica de Proteínas/prevenção & controle , Pueraria/química , Rehmannia/química , Expansão das Repetições de Trinucleotídeos/genética
15.
Am J Hum Genet ; 104(1): 35-44, 2019 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-30554721

RESUMO

Baratela-Scott syndrome (BSS) is a rare, autosomal-recessive disorder characterized by short stature, facial dysmorphisms, developmental delay, and skeletal dysplasia caused by pathogenic variants in XYLT1. We report clinical and molecular investigation of 10 families (12 individuals) with BSS. Standard sequencing methods identified biallelic pathogenic variants in XYLT1 in only two families. Of the remaining cohort, two probands had no variants and six probands had only a single variant, including four with a heterozygous 3.1 Mb 16p13 deletion encompassing XYLT1 and two with a heterozygous truncating variant. Bisulfite sequencing revealed aberrant hypermethylation in exon 1 of XYLT1, always in trans with the sequence variant or deletion when present; both alleles were methylated in those with no identified variant. Expression of the methylated XYLT1 allele was severely reduced in fibroblasts from two probands. Southern blot studies combined with repeat expansion analysis of genome sequence data showed that the hypermethylation is associated with expansion of a GGC repeat in the XYLT1 promoter region that is not present in the reference genome, confirming that BSS is a trinucleotide repeat expansion disorder. The hypermethylated allele accounts for 50% of disease alleles in our cohort and is not present in 130 control subjects. Our study highlights the importance of investigating non-sequence-based alterations, including epigenetic changes, to identify the missing heritability in genetic disorders.


Assuntos
Anormalidades Múltiplas/genética , Metilação de DNA/genética , Epigênese Genética/genética , Éxons/genética , Mutação , Pentosiltransferases/genética , Expansão das Repetições de Trinucleotídeos/genética , Alelos , Southern Blotting , Estudos de Coortes , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Linhagem , Sulfitos/metabolismo , Síndrome
16.
J Biol Chem ; 294(6): 1846-1859, 2019 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-30552117

RESUMO

Friedreich ataxia (FRDA) is a neurodegenerative disorder caused by transcriptional silencing of the frataxin (FXN) gene, resulting in loss of the essential mitochondrial protein frataxin. Based on the knowledge that a GAA·TTC repeat expansion in the first intron of FXN induces heterochromatin, we previously showed that 2-aminobenzamide-type histone deacetylase inhibitors (HDACi) increase FXN mRNA levels in induced pluripotent stem cell (iPSC)-derived FRDA neurons and in circulating lymphocytes from patients after HDACi oral administration. How the reduced expression of frataxin leads to neurological and other systemic symptoms in FRDA patients remains unclear. Similar to other triplet-repeat disorders, it is unknown why FRDA affects only specific cell types, primarily the large sensory neurons of the dorsal root ganglia and cardiomyocytes. The combination of iPSC technology and genome-editing techniques offers the unique possibility to address these questions in a relevant cell model of FRDA, obviating confounding effects of variable genetic backgrounds. Here, using "scarless" gene-editing methods, we created isogenic iPSC lines that differ only in the length of the GAA·TTC repeats. To uncover the gene expression signatures due to the GAA·TTC repeat expansion in FRDA neuronal cells and the effect of HDACi on these changes, we performed RNA-seq-based transcriptomic analysis of iPSC-derived central nervous system (CNS) and isogenic sensory neurons. We found that cellular pathways related to neuronal function, regulation of transcription, extracellular matrix organization, and apoptosis are affected by frataxin loss in neurons of the CNS and peripheral nervous system and that these changes are partially restored by HDACi treatment.


Assuntos
Ataxia de Friedreich/genética , Inibidores de Histona Desacetilases/farmacologia , Neurônios/patologia , Transcriptoma , Células Cultivadas , Ataxia de Friedreich/patologia , Edição de Genes/métodos , Perfilação da Expressão Gênica , Humanos , Células-Tronco Pluripotentes Induzidas/química , Proteínas de Ligação ao Ferro/genética , Proteínas de Ligação ao Ferro/metabolismo , Neurônios/química , Expansão das Repetições de Trinucleotídeos/genética
17.
Int J Mol Sci ; 19(12)2018 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-30567354

RESUMO

Myotonic dystrophy type 1 (DM1), the most common cause of adult-onset muscular dystrophy, is autosomal dominant, multisystemic disease with characteristic symptoms including myotonia, heart defects, cataracts and testicular atrophy. DM1 disease is being successfully modelled in Drosophila allowing to identify and validate new pathogenic mechanisms and potential therapeutic strategies. Here we provide an overview of insights gained from fruit fly DM1 models, either: (i) fundamental with particular focus on newly identified gene deregulations and their link with DM1 symptoms; or (ii) applied via genetic modifiers and drug screens to identify promising therapeutic targets.


Assuntos
Músculo Esquelético/fisiopatologia , Distrofia Miotônica/genética , Distrofia Miotônica/terapia , Miotonina Proteína Quinase/genética , Animais , Modelos Animais de Doenças , Drosophila melanogaster/genética , Avaliação Pré-Clínica de Medicamentos , Humanos , Terapia de Alvo Molecular , Distrofia Miotônica/patologia , Miotonina Proteína Quinase/antagonistas & inibidores , Expansão das Repetições de Trinucleotídeos/genética
18.
PLoS One ; 13(12): e0209309, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30576349

RESUMO

Low FMR1 variants (CGGn<26) have been associated with premature ovarian aging, female infertility and poor IVF treatment success. Until now, there is little published information concerning possible molecular mechanisms for this effect. We wished to examine whether relative expression of RNA and the FMR1 gene's fragile X mental retardation protein (FMRP) RNA isoforms differ in women with various FMR1 sub-genotypes (normal, low CGGn<26 and/or high CGGn≥34). This prospective cohort study was conducted between 2014 and 2017 in a clinical research unit of the Center for Human Reproduction in New York City. The study involved a total of 98 study subjects, including 18 young oocyte donors and 80 older infertility patients undergoing routine in vitro fertilization (IVF) cycles. The main outcome measure was RNA expression in human luteinized granulosa cells of 5 groups of FMRP isoforms. The relative expression of FMR1 RNA in human luteinized granulosa cells was measured by real-time PCR and a possible association with CGGn was explored. All 5 groups of FMRP RNA isoforms examined were found to be differentially expressed in human luteinized granulosa cells. The relative expression of four FMR1 RNA isoforms showed significant differences among 6 FMR1 sub-genotypes. Women with at least one low allele expressed significantly lower levels of all 5 sets of FRMP isoforms in comparison to the non-low group. While it would be of interest to see whether FMRP is also decreased in the low-group we recognize that in recent years it has been increasingly documented that information flow of genetics may be regulated by non-coding RNA, that is, without translation to a protein product. We, thus, conclude that various CGG expansions of FMR1 allele may lead to changes of RNA levels and ratios of distinct FMRP RNA isoforms, which could regulate the translation and/or cellular localization of FMRP, affect the expression of steroidogenic enzymes and hormonal receptors, or act in some other epigenetic process and therefore result in the ovarian dysfunction in infertility.


Assuntos
Proteína do X Frágil de Retardo Mental/genética , Infertilidade Feminina/genética , Insuficiência Ovariana Primária/genética , Regiões 5' não Traduzidas , Adulto , Alelos , Sequência de Aminoácidos , Sequência de Bases , Estudos de Coortes , Feminino , Fertilização In Vitro , Proteína do X Frágil de Retardo Mental/metabolismo , Síndrome do Cromossomo X Frágil/genética , Expressão Gênica , Células da Granulosa/metabolismo , Humanos , Infertilidade Feminina/etiologia , Infertilidade Feminina/metabolismo , Reserva Ovariana/genética , Insuficiência Ovariana Primária/etiologia , Insuficiência Ovariana Primária/metabolismo , Estudos Prospectivos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , RNA/genética , RNA/metabolismo , Homologia de Sequência de Aminoácidos , Expansão das Repetições de Trinucleotídeos
19.
Artigo em Inglês | MEDLINE | ID: mdl-30191086

RESUMO

Background: Access to medical care in many regions is limited by socioeconomic status, at both the individual and the community level. This report describes the diagnostic process of a family residing on an underserved Caribbean island where routine neurological care is typically addressed by general practitioners, and genetic diagnosis is not available through regular medical channels. The diagnosis and management of neurodegenerative disorders is especially challenging in this setting. Case Report: We diagnosed a family with spinocerebellar ataxia type 3 (SCA3) in an underdeveloped nation with limited access to genetic medicine and no full-time neurologist. Discussion: Molecular diagnosis of the SCAs can be challenging, even in developed countries. In the Caribbean, genetic testing is generally only available at a small number of academic centers. Diagnosis in this family was ultimately made by utilizing an international, pro bono, research-based collaborative process. Although access to appropriate resources, such as speech, physical, and occupational therapies, is limited on this island because of economic and geographical factors, the provision of a diagnosis appeared to be ultimately beneficial for this family. Identification of affected families highlights the need for access to genetic diagnosis in all communities, and can help direct resources where needed.


Assuntos
Gerenciamento Clínico , Saúde da Família , Ataxias Espinocerebelares/diagnóstico , Adulto , Ataxina-3/genética , Feminino , Testes Genéticos , Humanos , Pessoa de Meia-Idade , Proteínas Repressoras/genética , Ataxias Espinocerebelares/complicações , Ataxias Espinocerebelares/genética , Ataxias Espinocerebelares/terapia , Expansão das Repetições de Trinucleotídeos/genética , Índias Ocidentais
20.
Neurol Sci ; 39(11): 1935-1943, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30094526

RESUMO

The aims of this study were to investigate the correlations of tri-nucleotide (CTG) repeat length with detailed echocardiography (ECHO) parameters that represent myocardial function and to find a relationship between heart function and CTG repeat length in adult-onset myotonic dystrophy type 1 (DM1). In this study, clinical data for patients with DM1, including age, onset age, CTG repeat length, Medical Research Council sum score (MRCSS), and 6-min walking test (6MWT), were recorded. In addition, ECHO parameters and cardiac conduction abnormalities were evaluated. Among the cardiac parameters, the EA ratio and left ventricular end-diastolic dimension (LVEDD) were significantly correlated with the CTG repeat length (p < 0.05). Interventricular septal thickness at end-diastole was also significantly correlated with the 6MWT in a multivariate linear regression model (p < 0.05). In conclusion, motor function (MRCSS and 6MWT) and CTG repeat length significantly correlated with LV diastolic dysfunction in patients with DM1. More emphasis should be given to diastolic dysfunction, which is currently under-recognized, when evaluating patients with DM1 with no abnormalities in routine electrocardiography studies. Lastly, well-designed and longitudinal studies are warranted to characterize and understand the pathophysiology of diastolic dysfunction in DM1.


Assuntos
Cardiopatias/etiologia , Distrofia Miotônica/complicações , Distrofia Miotônica/genética , Miotonina Proteína Quinase/genética , Expansão das Repetições de Trinucleotídeos/genética , Adulto , Idoso , Cardiomiopatias/etiologia , Cardiomiopatias/genética , Ecocardiografia , Extremidades/fisiopatologia , Feminino , Marcha/fisiologia , Coração/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Distrofia Miotônica/fisiopatologia , Reprodutibilidade dos Testes , Índice de Gravidade de Doença , Caminhada/fisiologia , Adulto Jovem
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