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1.
Mol Genet Genomic Med ; : e962, 2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31595705

RESUMO

BACKGROUND: Maturity-onset diabetes of the young (MODY) is a form of monogenic diabetes with autosomal dominant inheritance. To date, mutations in 11 genes have been frequently associated with this phenotype. In Brazil, few cohorts have been screened for MODY, all using a candidate gene approach, with a high prevalence of undiagnosed cases (MODY-X). METHODS: We conducted a next-generation sequencing target panel (tNGS) study to investigate, for the first time, a Brazilian cohort of MODY patients with a negative prior genetic analysis. One hundred and two patients were selected, of which 26 had an initial clinical suspicion of MODY-GCK and 76 were non-GCK MODY. RESULTS: After excluding all benign and likely benign variants and variants of uncertain significance, we were able to assign a genetic cause for 12.7% (13/102) of the probands. Three rare MODY subtypes were identified (PDX1/NEUROD1/ABCC8), and eight variants had not been previously described/mapped in genomic databases. Important clinical findings were evidenced in some cases after genetic diagnosis, such as MODY-PDX1/HNF1B. CONCLUSION: A multiloci genetic approach allowed the identification of rare MODY subtypes, reducing the large percentage of MODY-X in Brazilian cases and contributing to a better clinical, therapeutic, and prognostic characterization of these rare phenotypes.

2.
J Esthet Restor Dent ; 2019 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-31622014

RESUMO

OBJECTIVE: To evaluate the influence of the degree of dentin moisture on interfacial ultramorphology and bond strength (µTBS) of universal adhesives. MATERIALS AND METHODS: Futurabond U (FBU), Scotchbond Universal (SBU), Adhese Universal (ADU), and Prime&Bond active (PBA) were used. After acid-etching, moist or over-dried dentin surfaces were tested. Teeth were restored for scanning and transmission electron microscopy (n = 3) and µTBS evaluation (n = 5). µTBS results were analyzed by two-way ANOVA and Tukey. RESULTS: For moist dentin, a well-formed hybrid layer (HL) was observed. However, when applied to over-dried dentin, remarkable differences were observed. Defects, gaps, and reduced HL thickness were observed mainly for ADU and FBU. When applied to wet dentin, µTBS values were similar for all adhesives, except for FBU, which was significantly lower. When applied to over-dried dentin, PBA presented the highest µTBS values, followed by SBU, ADU, and FBU. ADU presented significantly lower µTBS when applied to over-dried dentin. CONCLUSION: PBA, SBU, and FBU µTBS values were not sensitive to the degree of moisture. Even though application to over-dried dentin revealed defects, gaps and reduced HL thickness for SBU, ADU, and FBU, µTBS analysis only revealed a significant reduction for ADU. CLINICAL SIGNIFICANCE: Universal adhesives can be applied in either self-etching or etch-and-rinse mode. However, clinicians are not aware which universal adhesives should be strictly applied on a moist dentin for bonding in the etch-and-rinse mode.

3.
Ecotoxicol Environ Saf ; 186: 109747, 2019 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-31634660

RESUMO

Cadmium (Cd) contamination has generated an environmental problem worldwide, leading to harmful effects on human health and damages to plant metabolism. Selenium (Se) is non essential for plants, however it can improve plant growth and reduce the adverse effects of abiotic stress. In addition, ethylene may interplay the positive effects of Se in plants. In order to investigate the role of ethylene in Se-modulation of antioxidant defence system in response to Cd-stress, we tested the hormonal mutant Epinastic (epi) with a subset of constitutive activation of the ethylene response and Micro-Tom (MT) plants. For this purpose, Se mineral uptake, Cd and Se concentrations, pigments, malondialdeyde (MDA) and hydrogen peroxide (H2O2) contents, ethylene production, glutathione (GSH) compound, and superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR) and glutathione peroxidase (GSH-Px) activities were analysed in MT and epi plants submitted to 0.5 mM CdCl2 and 1 µM of selenate or selenite. MT plants treated with both Se forms increased growth in the presence or not of 0.5 mM CdCl2, but not change epi growth. Both Se forms reduced Cd uptake in MT plants and cause reverse effect in epi plants. P, Mg, S, K and Zn uptake increased in epi plants with Se application, irrespective to Cd exposure. Chlorophylls and carotenoids contents decreased in both genotypes under Cd exposure, in contrast to what was observed in epi leaves in the presence of Se. When antioxidant enzymes activities were concerned, Se application increased Mn-SOD, Fe-SOD and APX activities. In the presence of Cd, MT and epi plants exhibited decreased SOD activity and increased CAT, APX and GR activities. MT and epi plants with Se supply exhibited increased APX and GR activities in the presence of Cd. Overall, these results suggest that ethylene may be involved in Se induced-defence responses, that triggers a positive response of the antioxidant system and improve growth under Cd stress. These results showed integrative roles of ethylene and Se in regulating the cell responses to stressful-conditions and, the cross-tolerance to stress could be used to manipulate ethylene regulated gene expression to induce heavy metal tolerance.

4.
Eur J Med Res ; 24(1): 32, 2019 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-31521205

RESUMO

BACKGROUND: Growing demand for risk-reducing surgery in individuals with inherited susceptibility to cancer leads to the question whether these procedures are cost effective for the executing hospitals. This study compared the clinical costs for bilateral risk-reducing mastectomy (BRRM) with and without different types of reconstruction, risk-reducing salpingo-oophorectomy (RRSO), and their combinations with corresponding reimbursements in the statutory health-care system in Germany. PATIENTS AND METHODS: Real total costs of care for BRRM with and without reconstruction, RRSO, and their combinations were calculated as the sum of all personnel and technical costs. These costs calculated in a German University hospital were compared with the sum of all reimbursements in the German DRG-based health-care system. RESULTS: While sole RRSO, BRRM without reconstruction, and BRRM with secondary DIEP (deep inferior epigastric perforator)-reconstruction still result in a small benefit, we even found shortfalls for the hospital with all other prophylactic operations under consideration. The calculated deficits were especially high for BRRM with implant-based breast reconstruction and for combined operations when the risk reduction is achieved with a minimum of separate operations. CONCLUSIONS: Risk-reducing surgery in BRCA-mutation carriers is frequently not cost-covering for the executing hospitals in the German health-care system. Thus, appropriate concepts are required to ensure a nationwide care.

5.
Prenat Diagn ; 2019 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-31498910

RESUMO

OBJECTIVE: 17q12 microdeletions containing HNF1B and intragenic variants within this gene are associated with variable developmental, endocrine, and renal anomalies, often already noted prenatally as hyperechogenic/cystic kidneys. Here, we describe prenatal and postnatal phenotypes of seven individuals with HNF1B aberrations and compare their clinical and genetic data to those of previous studies. METHODS: Prenatal sequencing and postnatal chromosomal microarray analysis were performed in seven individuals with renal and/or neurodevelopmental phenotypes. We evaluated HNF1B-related clinical features from 82 studies and reclassified 192 reported intragenic HNF1B variants. RESULTS: In a prenatal case, we identified a novel in-frame deletion p.(Gly239del) within the HNF1B DNA-binding domain, a mutational hot spot as demonstrated by spatial clustering analysis and high computational prediction scores. The six postnatally diagnosed individuals harbored 17q12 microdeletions. Literature screening revealed variable reporting of HNF1B-associated clinical traits. Overall, both mutation groups showed a high phenotypic heterogeneity. The reclassification of all previously reported intragenic HNF1B variants provided an up-to-date overview of the mutational spectrum. CONCLUSIONS: We highlight the value of prenatal HNF1B screening in renal developmental diseases. Standardized clinical reporting and systematic classification of HNF1B variants are necessary for a more accurate risk quantification of prenatal and postnatal clinical features, improving genetic counseling and prenatal decision making.

6.
Sci Total Environ ; 687: 1219-1231, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31412457

RESUMO

Arsenic (As) in native soils of the Amazon rainforest is a concern due to its likely origin from the Andean rivers, which transport loads of sediments containing substantial amounts of trace elements coming from the cordilleras. Yet, unveiling soil As baseline concentrations in the Amazon basin is still a need because most studies in Brazil have been performed in areas with predominantly high concentrations and cannot express a real baseline value for the region. In this study, 414 soil samples (0-20, 20-40 and 40-60 cm layers) were collected from different sites throughout the Amazon basin - including native Amazon rainforest and minimally disturbed areas - and used to determine total and extractable (soluble + available) As concentrations along with relevant soil physicochemical properties. Descriptive statistics of the data was performed and Pearson correlation supported by a Principal Component Analysis (PCA) provided an improved understanding of where and how As concentrations are influenced by soil attributes. Total As concentration ranged from 0.98 to 41.71 mg kg-1 with values usually increasing from the topsoil (0-20 cm) to the deepest layer (40-60 cm) in all sites studied. Considering the proportional contribution given by each fraction (soluble and available) on extractable As concentration, it is noticeable that KH2PO4-extractable As represents the most important fraction, with >70% of the As extracted on average in all the sites studied. Still, the extractable fractions (soluble + available) correspond to ~0.24% of the total As, on average. Total, available, and soluble As fractions were strongly and positively correlated with soil Al3+. The PCA indicated that soil pH in combination with CEC might be the key factors controlling soil As concentrations and the occurrence of each arsenic fraction in the soil layers.

7.
Ann Clin Transl Neurol ; 6(7): 1263-1272, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31353855

RESUMO

OBJECTIVE: To analyze clinical phenotypes associated with KCNC1 variants other than the Progressive Myoclonus Epilepsy-causing variant p.Arg320His, determine the electrophysiological functional impact of identified variants and explore genotype-phenotype-physiological correlations. METHODS: Ten cases with putative pathogenic variants in KCNC1 were studied. Variants had been identified via whole-exome sequencing or gene panel testing. Clinical phenotypic data were analyzed. To determine functional impact of variants detected in the Kv 3.1 channel encoded by KCNC1, Xenopus laevis oocyte expression system and automated two-electrode voltage clamping were used. RESULTS: Six unrelated patients had a Developmental and Epileptic Encephalopathy and a recurrent de novo variant p.Ala421Val (c.1262C > T). Functional analysis of p.Ala421Val revealed loss of function through a significant reduction in whole-cell current, but no dominant-negative effect. Three patients had a contrasting phenotype of Developmental Encephalopathy without seizures and different KCNC1 variants, all of which caused loss of function with reduced whole-cell currents. Evaluation of the variant p.Ala513Val (c.1538C > T) in the tenth case, suggested it was a variant of uncertain significance. INTERPRETATION: These are the first reported cases of Developmental and Epileptic Encephalopathy due to KCNC1 mutation. The spectrum of phenotypes associated with KCNC1 is now broadened to include not only a Progressive Myoclonus Epilepsy, but an infantile onset Developmental and Epileptic Encephalopathy, as well as Developmental Encephalopathy without seizures. Loss of function is a key feature, but definitive electrophysiological separation of these phenotypes has not yet emerged.

8.
Molecules ; 24(12)2019 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-31238580

RESUMO

The objective of this study, for the first time, was to optimize Amazonian cyanobacterial culture conditions for improving cell productivity and lipid content, by analyzing the effect of light intensity and nitrogen concentration, for empirically evaluating biodiesel quality parameters. The strains Synechocystis sp. CACIAM05, Microcystis aeruginosa CACIAM08, Pantanalinema rosaneae CACIAM18, and Limnothrix sp. CACIAM25, were previously identified by morphological and molecular analysis (16S rRNA) and were selected based on their production of chlorophyll a and dry cell weight. Then, factorial planning (22) with central points was applied, with light intensity and NaNO3 concentration as independent variables. As response variables, cell productivity and lipid content were determined. Statistical analysis indicated that for all strains, the independent variables were statistically significant for cell productivity. Analysis of the fatty acid composition demonstrated diversity in the composition of the fatty acid profile from the experimental planning assays of each strain. The Biodiesel Analyzer software predicted the biodiesel quality parameters. CACIAM05 and CACIAM25 obtained better parameters with low levels of light intensity and NaNO3 concentration, whereas CACIAM08 and CACIAM18 obtained better parameters with low NaNO3 concentrations and high luminous intensity.

9.
J Sci Food Agric ; 99(13): 5969-5983, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31215030

RESUMO

BACKGROUND: Selenium (Se) is a nutrient for animals and humans, and is considered beneficial to higher plants. Selenium concentrations are low in most soils, which can result in a lack of Se in plants, and consequently in human diets. Phytic acid (PA) is the main storage form of phosphorus in seeds, and it is able to form insoluble complexes with essential minerals in the monogastric gut. This study aimed to establish optimal levels of Se application to cowpea, with the aim of increasing Se concentrations. The efficiency of agronomic biofortification was evaluated by the application of seven levels of Se (0, 2.5, 5, 10, 20, 40, and 60 g ha-1 ) from two sources (selenate and selenite) to the soil under field conditions in 2016 and 2017. RESULTS: Application of Se as selenate led to greater plant Se concentrations than application as selenite in both leaves and grains. Assuming human cowpea consumption of 54.2 g day-1 , Se application of 20 g ha-1 in 2016 or 10 g ha-1 in 2017 as selenate would have provided a suitable daily intake of Se (between 20 and 55 µg day-1 ) for humans. Phytic acid showed no direct response to Se application. CONCLUSION: Selenate provides greater phytoavailability than selenite. The application of 10 g Se ha-1 of selenate to cowpea plants could provide sufficient seed Se to increase daily human intake by 13-14 µg d-1 . © 2019 Society of Chemical Industry.


Assuntos
Biofortificação/métodos , Ácido Fítico/análise , Ácido Selênico/análise , Ácido Selenioso/análise , Selênio/análise , Vigna/química , Fertilizantes/análise , Ácido Fítico/metabolismo , Folhas de Planta/química , Folhas de Planta/metabolismo , Sementes/química , Sementes/metabolismo , Vigna/metabolismo
10.
J Adhes Dent ; 21(3): 281-286, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31165107

RESUMO

PURPOSE: To evaluate the bond strength to dentin produced by experimental adhesives formulated with an elastomeric methacrylate monomer (EMM) and an alternative initiator system based on a Thioxanthone derivative (QTX). MATERIALS AND METHODS: A self-etching primer was used. For the bonding resin, a model adhesive (G1) was formulated containing bis-GMA/TEG-DMA/HEMA (co-monomeric blend) + CQ/EDAB (initiator system). The other groups were formulated by adding to this formulation: EMM only (G2), QTX (G3), or EMM and QTX (G4). Clearfil SE Bond was used as the commercial control group. Fifty bovine teeth (n = 5) were restored with each one of the five adhesives. After restorative procedures, half of the specimens were stored in distilled water at 37°C for 24 h. The other half was fixed on a metal stub and subjected to 200,000 mechanical (50 N loading at 2 Hz frequency) and 1000 thermal cycles (5°C and 55°C). Afterwards, specimens were serially sectioned into beams and tested in tension until fracture. Bond strengths were statistically analyzed by two-way ANOVA and Tukey's test (α = 5%). RESULTS: After 24 h, significantly higher µTBS was observed for the formulation containing EMM and QTX (G4) when compared to Clearfil SE Bond (p < 0.05). No significant differences in µTBS were detected among the experimental groups after 24 h (p>0.05). After thermomechanical cycling, no significant differences were observed among groups. CONCLUSION: The addition of EMM and QTX can be considered as possible alternative in dental adhesive formulations.


Assuntos
Colagem Dentária , Animais , Bis-Fenol A-Glicidil Metacrilato , Bovinos , Resinas Compostas , Cimentos Dentários , Dentina , Adesivos Dentinários , Teste de Materiais , Metacrilatos , Cimentos de Resina , Resistência à Tração
11.
Arch. endocrinol. metab. (Online) ; 63(3): 250-257, May-June 2019. tab, graf
Artigo em Inglês | LILACS-Express | ID: biblio-1011159

RESUMO

ABSTRACT Objective To verify the presence of variants in HNF1B in a sample of the Brazilian population selected according to the presence of renal cysts associated with hyperglycemia. Subjects and methods We evaluated 28 unrelated patients with clinical suspicion of HNF1B mutation because of the concomitant presence of diabetes mellitus (DM) or prediabetes and renal cysts. Genotyping was accomplished using Sanger sequencing or multiplex ligation-dependent probe amplification (MLPA). In positive cases, available relatives were recruited. Results We found two patients with HNF1B mutations. The first presented the variant p.Pro328Leufs*48(c.983delC) and had DM, renal cysts, and hypomagnesemia. The second presented a heterozygous whole gene deletion in HNF1B, DM, renal cysts, body and tail pancreatic agenesis, and hypomagnesemia; this alteration was also found in his two siblings and his father. Conclusion The recruitment of suspected cases of HNF1B gene mutations in Brazilians due to hyperglycemia and renal cysts presents two positive cases. Our cases contribute to the annotation of clinical and biochemical phenotypes of this rare form of maturity-onset diabetes of the young (MODY).

12.
Nat Protoc ; 14(7): 2119-2151, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31217595

RESUMO

Next-generation sequencing (NGS) has been widely adopted to identify genetic variants and investigate their association with disease. However, the analysis of sequencing data remains challenging because of the complexity of human genetic variation and confounding errors introduced during library preparation, sequencing and analysis. We have developed a set of synthetic DNA spike-ins-termed 'sequins' (sequencing spike-ins)-that are directly added to DNA samples before library preparation. Sequins can be used to measure technical biases and to act as internal quantitative and qualitative controls throughout the sequencing workflow. This step-by-step protocol explains the use of sequins for both whole-genome and targeted sequencing of the human genome. This includes instructions regarding the dilution and addition of sequins to human DNA samples, followed by the bioinformatic steps required to separate sequin- and sample-derived sequencing reads and to evaluate the diagnostic performance of the assay. These practical guidelines are accompanied by a broader discussion of the conceptual and statistical principles that underpin the design of sequin standards. This protocol is suitable for users with standard laboratory and bioinformatic experience. The laboratory steps require ~1-4 d and the bioinformatic steps (which can be performed with the provided example data files) take an additional day.

13.
Arch Endocrinol Metab ; 63(3): 250-257, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31066763

RESUMO

OBJECTIVE: To verify the presence of variants in HNF1B in a sample of the Brazilian population selected according to the presence of renal cysts associated with hyperglycemia. SUBJECTS AND METHODS: We evaluated 28 unrelated patients with clinical suspicion of HNF1B mutation because of the concomitant presence of diabetes mellitus (DM) or prediabetes and renal cysts. Genotyping was accomplished using Sanger sequencing or multiplex ligation-dependent probe amplification (MLPA). In positive cases, available relatives were recruited. RESULTS: We found two patients with HNF1B mutations. The first presented the variant p.Pro328Leufs*48(c.983delC) and had DM, renal cysts, and hypomagnesemia. The second presented a heterozygous whole gene deletion in HNF1B, DM, renal cysts, body and tail pancreatic agenesis, and hypomagnesemia; this alteration was also found in his two siblings and his father. CONCLUSION: The recruitment of suspected cases of HNF1B gene mutations in Brazilians due to hyperglycemia and renal cysts presents two positive cases. Our cases contribute to the annotation of clinical and biochemical phenotypes of this rare form of maturity-onset diabetes of the young (MODY).


Assuntos
Nefropatias Diabéticas/genética , Fator 1-beta Nuclear de Hepatócito/genética , Hiperglicemia/genética , Doenças Renais Císticas/genética , Mutação , Adulto , Brasil , Estudos de Coortes , Nefropatias Diabéticas/complicações , Deleção de Genes , Humanos , Hiperglicemia/complicações , Doenças Renais Císticas/complicações , Pessoa de Meia-Idade , Fenótipo , Polimorfismo Genético/genética
14.
BMC Cancer ; 19(1): 435, 2019 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-31077186

RESUMO

BACKGROUND: Several subunits of the SWI/SNF chromatin remodeling complex are implicated in both cancer and neurodevelopmental disorders (NDD). Though there is no clinical evidence for an increased tumor risk in individuals with NDDs due to germline mutations in most of these genes so far, this has been repeatedly proposed and discussed. A young woman with NDD due to a de novo mutation in ARID1B now presented with a large renal (> 19 cm in diameter) and multiple hepatic angiomyolipomas (AMLs) but no other signs of tuberous sclerosis complex. METHODS: We analyzed tumor and healthy tissue samples with exome and panel sequencing. RESULTS: Additionally to the previously known, germline ARID1B variant we identified a post-zygotic truncating TSC2 variant in both renal and hepatic AMLs but not in any of the healthy tissues. We did not detect any further, obvious tumor driver events. The identification of a passenger variant in SIPA1L3 in both AMLs points to a common clonal origin. Metastasis of the renal AML into the liver is unlikely on the basis of discordant histopathological features. Our findings therefore point to very low-grade mosaicism for the TSC2 variant, possibly in a yet unknown mesenchymal precursor cell that expanded clonally during tumor development. A possible contribution of the germline ARID1B variant to the tumorigenesis remains unclear but cannot be excluded given the absence of any other evident tumor drivers in the AMLs. CONCLUSION: This unique case highlights the blurred line between tumor genetics and post-zygotic events that can complicate exact molecular diagnoses in patients with rare manifestations. It also demonstrates the relevance of multiple disorders in a single individual, the challenges of detecting low-grade mosaicisms, and the importance of proper diagnosis for treatment and surveillance.


Assuntos
Angiomiolipoma/genética , Deficiência Intelectual/complicações , Neoplasias Renais/genética , Neoplasias Hepáticas/genética , Proteína 2 do Complexo Esclerose Tuberosa/genética , Proteínas de Ligação a DNA/genética , Feminino , Mutação em Linhagem Germinativa , Humanos , Deficiência Intelectual/genética , Mosaicismo , Fatores de Transcrição/genética , Sequenciamento Completo do Exoma , Adulto Jovem
16.
PLoS Genet ; 15(4): e1008088, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-31034465

RESUMO

PIK3C2A is a class II member of the phosphoinositide 3-kinase (PI3K) family that catalyzes the phosphorylation of phosphatidylinositol (PI) into PI(3)P and the phosphorylation of PI(4)P into PI(3,4)P2. At the cellular level, PIK3C2A is critical for the formation of cilia and for receptor mediated endocytosis, among other biological functions. We identified homozygous loss-of-function mutations in PIK3C2A in children from three independent consanguineous families with short stature, coarse facial features, cataracts with secondary glaucoma, multiple skeletal abnormalities, neurological manifestations, among other findings. Cellular studies of patient-derived fibroblasts found that they lacked PIK3C2A protein, had impaired cilia formation and function, and demonstrated reduced proliferative capacity. Collectively, the genetic and molecular data implicate mutations in PIK3C2A in a new Mendelian disorder of PI metabolism, thereby shedding light on the critical role of a class II PI3K in growth, vision, skeletal formation and neurological development. In particular, the considerable phenotypic overlap, yet distinct features, between this syndrome and Lowe's syndrome, which is caused by mutations in the PI-5-phosphatase OCRL, highlight the key role of PI metabolizing enzymes in specific developmental processes and demonstrate the unique non-redundant functions of each enzyme. This discovery expands what is known about disorders of PI metabolism and helps unravel the role of PIK3C2A and class II PI3Ks in health and disease.


Assuntos
Doenças do Desenvolvimento Ósseo/genética , Catarata/genética , Transtornos da Motilidade Ciliar/genética , Nanismo/genética , Mutação , Fosfatidilinositol 3-Quinases/genética , Adolescente , Adulto , Criança , Consanguinidade , Feminino , Fibroblastos/metabolismo , Humanos , Masculino , Linhagem , Fenótipo , Adulto Jovem
17.
Genet Med ; 21(10): 2345-2354, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31000793

RESUMO

PURPOSE: Primary open-angle glaucoma (POAG) is the leading cause of irreversible blindness worldwide and mutations in known genes can only explain 5-6% of POAG. This study was conducted to identify novel POAG-causing genes and explore the pathogenesis of this disease. METHODS: Exome sequencing was performed in a Han Chinese cohort comprising 398 sporadic cases with POAG and 2010 controls, followed by replication studies by Sanger sequencing. A heterozygous Ramp2 knockout mouse model was generated for in vivo functional study. RESULTS: Using exome sequencing analysis and replication studies, we identified pathogenic variants in receptor activity-modifying protein 2 (RAMP2) within three genetically diverse populations (Han Chinese, German, and Indian). Six heterozygous RAMP2 pathogenic variants (Glu39Asp, Glu54Lys, Phe103Ser, Asn113Lysfs*10, Glu143Lys, and Ser171Arg) were identified among 16 of 4763 POAG patients, whereas no variants were detected in any exon of RAMP2 in 10,953 control individuals. Mutant RAMP2s aggregated in transfected cells and resulted in damage to the AM-RAMP2/CRLR-cAMP signaling pathway. Ablation of one Ramp2 allele led to cAMP reduction and retinal ganglion cell death in mice. CONCLUSION: This study demonstrated that disruption of RAMP2/CRLR-cAMP axis could cause POAG and identified a potential therapeutic intervention for POAG.

18.
Hum Mol Genet ; 2019 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-30986821

RESUMO

LOXL1 (lysyl oxidase-like 1) has been identified as the major effect locus in pseudoexfoliation (PEX) syndrome, a fibrotic disorder of the extracellular matrix and frequent cause of chronic open-angle glaucoma. However, all known PEX-associated common variants show allele effect reversal in populations of different ancestry, casting doubt on their biological significance. Based on extensive LOXL1 deep sequencing, we report here the identification of a common noncoding sequence variant, rs7173049A>G, located downstream of LOXL1, consistently associated with a decrease in PEX risk (OR=0.63, p=6.33x10-31) in nine different ethnic populations. We provide experimental evidence for a functional enhancer-like regulatory activity of the genomic region surrounding rs7173049 influencing expression levels of ISLR2 (immunoglobulin superfamily containing leucine-rich repeat protein 2) and STRA6 (stimulated by retinoic acid receptor 6), apparently mediated by allele-specific binding of the transcription factor THRß (thyroid hormone receptor beta). We further show that the protective rs7173049-G allele correlates with increased tissue expression levels of ISLR2 and STRA6 and that both genes are significantly downregulated in tissues of PEX patients together with other key components of the STRA6 receptor-driven retinoic acid signaling pathway. siRNA-mediated downregulation of retinoic acid signaling induces upregulation of LOXL1 and PEX-associated matrix genes in PEX-relevant cell types. These data indicate that dysregulation of STRA6 and impaired retinoid metabolismare involved in the pathophysiology of PEX syndrome and that the variant rs7173049-G, which represents the first common variant at the broad LOXL1 locus without allele effect reversal, mediates a protective effect through upregulation of STRA6 in ocular tissues.

19.
Hum Mol Genet ; 2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31035284

RESUMO

The SOXC transcription factors Sox4, Sox11, and Sox12, are critical neurodevelopmental regulators that are thought to function in a highly redundant fashion. Surprisingly, heterozygous missense mutations or deletions of SOX11 were recently detected in patients with Coffin-Siris syndrome like syndrome (CSSLS), a neurodevelopmental disorder associated with intellectual disability, demonstrating that in humans SOX11 haploinsufficiency cannot be compensated and raising the question of the function of SOX11 in human neurodevelopment. Here, we describe the generation of SOX11+/- heterozygous human embryonic stem cell (hESC) lines by CRISPR/Cas9 genome engineering. SOX11 haploinsufficiency impaired the generation of neurons and resulted in a proliferation/differentiation imbalance of neural precursor cells and enhanced neuronal cell death. Using the SOX11+/- hESC model we provide for the first time experimental evidence that SOX11 haploinsufficiency is sufficient to impair key processes of human neurodevelopment, giving a first insight into the pathophysiology of CSSLS and SOX11 function in human neurodevelopment.

20.
Sensors (Basel) ; 19(7)2019 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-30959784

RESUMO

We improved a magnetic scanning microscope for measuring the magnetic properties of minerals in thin sections of geological samples at submillimeter scales. The microscope is comprised of a 200 µm diameter Hall sensor that is located at a distance of 142 µm from the sample; an electromagnet capable of applying up to 500 mT DC magnetic fields to the sample over a 40 mm diameter region; a second Hall sensor arranged in a gradiometric configuration to cancel the background signal applied by the electromagnet and reduce the overall noise in the system; a custom-designed electronics system to bias the sensors and allow adjustments to the background signal cancelation; and a scanning XY stage with micrometer resolution. Our system achieves a spatial resolution of 200 µm with a noise at 6.0 Hz of 300 nTrms/(Hz)1/2 in an unshielded environment. The magnetic moment sensitivity is 1.3 × 10-11 Am². We successfully measured the representative magnetization of a geological sample using an alternative model that takes the sample geometry into account and identified different micrometric characteristics in the sample slice.

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