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1.
Adv Exp Med Biol ; 1164: 207-224, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31576551

RESUMO

Prostate cancers have a justified reputation as one of the most heterogeneous human tumours. Indeed, there are some who consider that advanced and castration-resistant prostate cancers are incurable, as a direct result of this heterogeneity. However, tumour heterogeneity can be defined in different ways. To a clinician, prostate cancer is a number of different diseases, the treatments for which remain equally heterogeneous and uncertain. To the pathologist, the histopathological appearances of the tumours are notoriously heterogeneous. Indeed, the genius of Donald Gleason in the 1960s was to devise a classification system designed to take into account the heterogeneity of the tumours both individually and in the whole prostate context. To the cell biologist, a prostate tumour consists of multiple epithelial cell types, inter-mingled with various fibroblasts, neuroendocrine cells, endothelial cells, macrophages and lymphocytes, all of which interact to influence treatment responses in a patient-specific manner. Finally, genetic analyses of prostate cancers have been compromised by the variable gene rearrangements and paucity of activating mutations observed, even in large numbers of patient tumours with consistent clinical diagnoses and/or outcomes. Research into familial susceptibility has even generated the least tractable outcome of such studies: the genetic loci are of low penetrance and are of course heterogeneous. By fractionating the tumour (and patient-matched non-malignant tissues) heterogeneity can be resolved, revealing homogeneous markers of patient outcomes.


Assuntos
Células Endoteliais , Neoplasias da Próstata , Células Endoteliais/citologia , Heterogeneidade Genética , Humanos , Masculino , Mutação , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Neoplasias da Próstata/terapia
3.
Medicine (Baltimore) ; 98(34): e16916, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31441872

RESUMO

BACKGROUND: Colorectal Cancer (CRC) is a highly heterogeneous disease. RNA profiles of bulk tumors have enabled transcriptional classification of CRC. However, such ways of sequencing can only target a cell colony and obscure the signatures of distinct cell populations. Alternatively, single-cell RNA sequencing (scRNA-seq), which can provide unbiased analysis of all cell types, opens the possibility to map cellular heterogeneity of CRC unbiasedly. METHODS: In this study, we utilized scRNA-seq to profile cells from cancer tissue of a CRC patient. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to understand the roles of genes within the clusters. RESULTS AND CONCLUSION: The 2824 cells were analyzed and categorized into 5 distinct clusters by scRNA-seq. For every cluster, specific cell markers can be applied, indicating each 1 of them different from another. We discovered that the tumor of CRC displayed a clear sign of heterogenicity, while genes within each cluster serve different functions. GO term analysis also stated that different cluster's relatedness towards the tumor of CRC differs. Three clusters participate in peripheral works in cells, including, energy transport, extracellular matrix generation, etc; Genes in other 2 clusters participate more in immunology processes. Lastly, trajectory plot analysis also supports the viewpoint, in that some clusters present in different states and pseudo-time, while others present in a single state or pseudo time. Our analysis provides more insight into the heterogeneity of CRC, which can provide assistance to further researches on this topic.


Assuntos
Neoplasias Colorretais/genética , Perfilação da Expressão Gênica/métodos , Heterogeneidade Genética , Análise de Sequência de RNA/métodos , Idoso , Biomarcadores Tumorais/genética , Feminino , Humanos
5.
World J Emerg Surg ; 14: 24, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31164913

RESUMO

Background: Trauma leads to a complex inflammatory cascade that induces both immune activation and a refractory immune state in parallel. Although both components are deemed necessary for recovery, the balance is tight and easily lost. Losing the balance can lead to life-threatening infectious complications as well as long-term immunosuppression with recurrent infections. Neutrophils are known to play a key role in these processes. Therefore, this review focuses on neutrophil characteristics and function after trauma and how these features can be used to identify trauma patients at risk for infectious complications. Results: Distinct neutrophil subtypes exist that play their own role in the recovery and/or development of infectious complications after trauma. Furthermore, the refractory immune state is related to the risk of infectious complications. These findings change the initial concepts of the immune response after trauma and give rise to new biomarkers for monitoring and predicting inflammatory complications in severely injured patients. Conclusion: For early recognition of patients at risk, the immune system should be monitored. Several neutrophil biomarkers show promising results and analysis of these markers has become accessible to such extent that they can be used for point-of-care decision making after trauma.


Assuntos
Doenças Transmissíveis/classificação , Heterogeneidade Genética , Neutrófilos/classificação , Doenças Transmissíveis/fisiopatologia , Humanos , Integrina alfa4beta1/genética , Integrina alfa4beta1/metabolismo , Ferimentos e Lesões/complicações
6.
Nat Genet ; 51(6): 1060-1066, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31152164

RESUMO

Modulation of chromatin structure via histone modification is a major epigenetic mechanism and regulator of gene expression. However, the contribution of chromatin features to tumor heterogeneity and evolution remains unknown. Here we describe a high-throughput droplet microfluidics platform to profile chromatin landscapes of thousands of cells at single-cell resolution. Using patient-derived xenograft models of acquired resistance to chemotherapy and targeted therapy in breast cancer, we found that a subset of cells within untreated drug-sensitive tumors share a common chromatin signature with resistant cells, undetectable using bulk approaches. These cells, and cells from the resistant tumors, have lost chromatin marks-H3K27me3, which is associated with stable transcriptional repression-for genes known to promote resistance to treatment. This single-cell chromatin immunoprecipitation followed by sequencing approach paves the way to study the role of chromatin heterogeneity, not just in cancer but in other diseases and healthy systems, notably during cellular differentiation and development.


Assuntos
Neoplasias da Mama/genética , Imunoprecipitação da Cromatina , Cromatina/genética , Heterogeneidade Genética , Sequenciamento de Nucleotídeos em Larga Escala , Análise de Célula Única , Cromatina/metabolismo , Biologia Computacional/métodos , Epigênese Genética , Feminino , Histonas/metabolismo , Humanos , Técnicas Analíticas Microfluídicas , Análise de Célula Única/métodos , Células Estromais , Fluxo de Trabalho
7.
BMC Cancer ; 19(1): 434, 2019 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-31077166

RESUMO

BACKGROUND: Chromosomal rearrangements are the typical phenomena in cancer genomes causing gene disruptions and fusions, corruption of regulatory elements, damage to chromosome integrity. Among the factors contributing to genomic instability are non-B DNA structures with stem-loops and quadruplexes being the most prevalent. We aimed at investigating the impact of specifically these two classes of non-B DNA structures on cancer breakpoint hotspots using machine learning approach. METHODS: We developed procedure for machine learning model building and evaluation as the considered data are extremely imbalanced and it was required to get a reliable estimate of the prediction power. We built logistic regression models predicting cancer breakpoint hotspots based on the densities of stem-loops and quadruplexes, jointly and separately. We also tested Random Forest models varying different resampling schemes (leave-one-out cross validation, train-test split, 3-fold cross-validation) and class balancing techniques (oversampling, stratification, synthetic minority oversampling). RESULTS: We performed analysis of 487,425 breakpoints from 2234 samples covering 10 cancer types available from the International Cancer Genome Consortium. We showed that distribution of breakpoint hotspots in different types of cancer are not correlated, confirming the heterogeneous nature of cancer. It appeared that stem-loop-based model best explains the blood, brain, liver, and prostate cancer breakpoint hotspot profiles while quadruplex-based model has higher performance for the bone, breast, ovary, pancreatic, and skin cancer. For the overall cancer profile and uterus cancer the joint model shows the highest performance. For particular datasets the constructed models reach high predictive power using just one predictor, and in the majority of the cases, the model built on both predictors does not increase the model performance. CONCLUSION: Despite the heterogeneity in breakpoint hotspots' distribution across different cancer types, our results demonstrate an association between cancer breakpoint hotspots and stem-loops and quadruplexes. Approximately for half of the cancer types stem-loops are the most influential factors while for the others these are quadruplexes. This fact reflects the differences in regulatory potential of stem-loops and quadruplexes at the tissue-specific level, which yet to be discovered at the genome-wide scale. The performed analysis demonstrates that influence of stem-loops and quadruplexes on breakpoint hotspots formation is tissue-specific.


Assuntos
Pontos de Quebra do Cromossomo , DNA/química , Neoplasias/genética , DNA/genética , Feminino , Heterogeneidade Genética , Instabilidade Genômica , Humanos , Modelos Logísticos , Aprendizado de Máquina , Masculino , Conformação de Ácido Nucleico , Especificidade de Órgãos
8.
BMC Cancer ; 19(1): 403, 2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31035962

RESUMO

BACKGROUND: Modern cancer treatment strategies aim to target tumour specific genetic (or epigenetic) alterations. Treatment response improves if these alterations are clonal, i.e. present in all cancer cells within tumours. However, the identification of truly clonal alterations is impaired by the tremendous intra-tumour genetic heterogeneity and unavoidable sampling biases. METHODS: Here, we investigate the underlying causes of these spatial sampling biases and how the distribution and sizes of biopsies in sampling protocols can be optimised to minimize such biases. RESULTS: We find that in the ideal case, less than a handful of samples can be enough to infer truly clonal mutations. The frequency of the largest sub-clone at diagnosis is the main factor determining the accuracy of truncal mutation estimation in structured tumours. If the first sub-clone is dominating the tumour, higher spatial dispersion of samples and larger sample size can increase the accuracy of the estimation. In such an improved sampling scheme, fewer samples will enable the detection of truly clonal alterations with the same probability. CONCLUSIONS: Taking spatial tumour structure into account will decrease the probability to misclassify a sub-clonal mutation as clonal and promises better informed treatment decisions.


Assuntos
Heterogeneidade Genética , Mutação , Neoplasias/genética , Algoritmos , Contagem de Células , Células Clonais/metabolismo , Humanos , Modelos Teóricos , Neoplasias/patologia
9.
BMC Med Genet ; 20(1): 80, 2019 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-31088393

RESUMO

BACKGROUND: Intellectual disability/developmental delay is a complex condition with extraordinary heterogeneity. A large proportion of patients lacks a specific diagnosis. Next generation sequencing, enabling identification of genetic variations in multiple genes, has become an efficient strategy for genetic analysis in intellectual disability/developmental delay. METHODS: Clinical data of 112 Chinese families with unexplained intellectual disability/developmental delay was collected. Targeted next generation sequencing of 454 genes related to intellectual disability/developmental delay was performed for all 112 index patients. Patients with promising variants and their other family members underwent Sanger sequencing to validate the authenticity and segregation of the variants. RESULTS: Fourteen promising variants in genes EFNB1, MECP2, ATRX, NAA10, ANKRD11, DHCR7, LAMA1, NFIX, UBE3A, ARID1B and PTPRD were identified in 11 of 112 patients (11/112, 9.82%). Of 14 variants, eight arose de novo, and 13 are novel. Nine patients (9/112, 8.03%) got definite molecular diagnoses. It is the first time to report variants in EFNB1, NAA10, DHCR7, LAMA1 and NFIX in Chinese intellectual disability/developmental delay patients and first report about variants in NAA10 and LAMA1 in affected individuals of Asian ancestry. CONCLUSIONS: Targeted next generation sequencing of 454 genes is an effective test strategy for patients with unexplained intellectual disability/developmental delay. Genetic heterogenicity is significant in this Chinese cohort and de novo variants play an important role in the diagnosis. Findings of this study further delineate the corresponding phenotypes, expand the mutation spectrum and support the involvement of PTPRD in the disease.


Assuntos
Deficiências do Desenvolvimento/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Deficiência Intelectual/genética , Mutação , Adolescente , Criança , Pré-Escolar , China , Cromossomos Humanos X , Feminino , Genes Dominantes , Genes Recessivos , Heterogeneidade Genética , Humanos , Lactente , Masculino , Linhagem , Fenótipo
10.
BMC Med Genet ; 20(1): 90, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31126250

RESUMO

BACKGROUND: Hereditary spherocytosis (HS) is a common type of hereditary hemolytic anemia. According to the current diagnostic criteria of HS, patients with a family history of HS, typical clinical features and laboratory investigations could be diagnosed without the requirement of any additional tests, including genetic analysis. However, the clinical heterogeneities incur difficulties in HS diagnosis. We therefore aimed to investigate the application of genetic diagnosis in a family-based cohort. CASE PRESENTATION: In the present Chinese family, two probands sharing similar clinical manifestations, including jaundice, cholelithiasis, splenomegaly and spherocytes, while the clinical features of other family members were inconclusive. Whole-exome sequencing (WES) unexpectedly unveiled two separate disease-causing mutations in the two probands. SPTB R1625X mutation detected in proband D was a de novo mutation; while proband W inherited the SLC4A1 c.G1469A mutation from her mother, which was also inherited by her brother. However, the clinical features of proband W and her mother and brother were discrepant: proband W suffered from significant splenomegaly, jaundice and cholelithiasis, which resulted in cholecystectomy and splenectomy; while her mother and brother's HS were not complicated by cholelithiasis, and their splenomegaly and elevated serum bilirubin were moderate. In addition, additional genomic defects involved with HS-related symptoms have not been detected in this family. CONCLUSIONS: Both genotypes and phenotypes could be heterogeneous in the same HS family. The analysis of pathogenic gene mutations may endeavor to play an indispensable role in the accurate diagnosis and genetic consultation of HS individuals and their family members.


Assuntos
Saúde da Família , Heterogeneidade Genética , Mutação , Esferocitose Hereditária/genética , Adulto , Grupo com Ancestrais do Continente Asiático/genética , China , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Esferocitose Hereditária/etnologia , Esferocitose Hereditária/patologia , Sequenciamento Completo do Exoma/métodos
11.
BMC Med Genet ; 20(1): 89, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31126253

RESUMO

BACKGROUND: Cystic fibrosis has been largely under-diagnosed and thus, limited data is available on the incidence of cystic fibrosis in Sri Lanka. Our aim is to describe the phenotypic and genotypic spectrum of children with cystic fibrosis in Sri Lanka. CASE PRESENTATION: This report describes 10 unrelated cystic fibrosis cases with phenotypic features of cystic fibrosis and abnormal or intermediate sweat tests. The most common phenotypic features in this sample of symptomatic patients were persistent or recurrent lower respiratory tract infections, failure to thrive and Pseudo-Bartter syndrome. Altogether 7 cystic fibrosis causing mutations were identified in 10 patients. Except delta F508 which is the commonest mutation worldwide all the other mutations detected in Sri Lankan patients are rare mutations. 1161delC and V456A detected in our patients are South Asian mutations. The other mutations such as [C.1282C > G; C.2738A > G], C.53 + 1G > C, 2184insA and a deletion encompassing exons 4 to 11 have been reported previously from European patients with cystic fibrosis. CONCLUSION: These cases highlight the importance of considering the diagnosis of cystic fibrosis in children and young adults presenting with persistent respiratory tract infections associated with severe malnutrition and Pseudo-Bartter syndrome, especially in low income countries where newborn screening for cystic fibrosis is not available. The spectrum of CFTR mutations in Sri Lanka is heterogeneous and possibly linked to genetic flow from Indian subcontinent and Europe. The common mutations should be identified by sequencing the entire CFTR gene in adequate number of cystic fibrosis patients in order to design a mutation panel for common regional mutations.


Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/genética , Fibrose Cística/genética , Heterogeneidade Genética , Mutação , Criança , Pré-Escolar , Fibrose Cística/patologia , Feminino , Frequência do Gene , Genótipo , Humanos , Lactente , Masculino , Fenótipo , Sri Lanka , Adulto Jovem
12.
MBio ; 10(2)2019 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-30940709

RESUMO

Biofilms are multicellular bacterial communities encased in a self-secreted extracellular matrix comprised of polysaccharides, proteinaceous fibers, and DNA. Organization of these components lends spatial organization to the biofilm community such that biofilm residents can benefit from the production of common goods while being protected from exogenous insults. Spatial organization is driven by the presence of chemical gradients, such as oxygen. Here we show that two quinol oxidases found in Escherichia coli and other bacteria organize along the biofilm oxygen gradient and that this spatially coordinated expression controls architectural integrity. Cytochrome bd, a high-affinity quinol oxidase required for aerobic respiration under hypoxic conditions, is the most abundantly expressed respiratory complex in the biofilm community. Depletion of the cytochrome bd-expressing subpopulation compromises biofilm complexity by reducing the abundance of secreted extracellular matrix as well as increasing cellular sensitivity to exogenous stresses. Interrogation of the distribution of quinol oxidases in the planktonic state revealed that ∼15% of the population expresses cytochrome bd at atmospheric oxygen concentration, and this population dominates during acute urinary tract infection. These data point toward a bet-hedging mechanism in which heterogeneous expression of respiratory complexes ensures respiratory plasticity of E. coli across diverse host niches.IMPORTANCE Biofilms are multicellular bacterial communities encased in a self-secreted extracellular matrix comprised of polysaccharides, proteinaceous fibers, and DNA. Organization of these components lends spatial organization in the biofilm community. Here we demonstrate that oxygen gradients in uropathogenic Escherichia coli (UPEC) biofilms lead to spatially distinct expression programs for quinol oxidases-components of the terminal electron transport chain. Our studies reveal that the cytochrome bd-expressing subpopulation is critical for biofilm development and matrix production. In addition, we show that quinol oxidases are heterogeneously expressed in planktonic populations and that this respiratory heterogeneity provides a fitness advantage during infection. These studies define the contributions of quinol oxidases to biofilm physiology and suggest the presence of respiratory bet-hedging behavior in UPEC.


Assuntos
Biofilmes/crescimento & desenvolvimento , Variação Biológica da População , Heterogeneidade Genética , Oxirredutases/metabolismo , Oxigênio/metabolismo , Escherichia coli Uropatogênica/fisiologia , Aerobiose , Anaerobiose , Oxirredução , Oxirredutases/genética
13.
Gene ; 705: 109-112, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31028865

RESUMO

Variants of KCNQ4 are one of the most common causes of dominantly inherited nonsyndromic hearing loss. We investigated a consanguineous family in which two individuals had prelignual hearing loss, apparently inherited in a recessive mode. Whole-exome sequencing analyses demonstrated genetic heterogeneity as variants in two different genes segregated with the phenotype in two branches of the family. Members in one branch were homozygous for a pathogenic variant of TMC1. The other two affected individuals were homozygous for a missense pathogenic variant in KCNQ4 c.872C>T; p.(Pro291Leu). These two individuals had prelingual, progressive moderate to severe hearing loss, while a heterozygous carrier had late onset mild hearing loss. Our work demonstrates that p.Pro291L variant is semi-dominantly inherited. This is the first report of semi-dominance of a KCNQ4 variant.


Assuntos
Surdez/genética , Canais de Potássio KCNQ/genética , Mutação de Sentido Incorreto , Sequenciamento Completo do Exoma/métodos , Idade de Início , Consanguinidade , Feminino , Heterogeneidade Genética , Predisposição Genética para Doença , Humanos , Leucina/genética , Masculino , Linhagem , Prolina/genética
14.
Int J Mol Sci ; 20(8)2019 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-31003536

RESUMO

Mesenchymal stem cells (MSCs) possess a broad spectrum of therapeutic applications and have been used in clinical trials. MSCs are mainly retrieved from adult or fetal tissues. However, there are many obstacles with the use of tissue-derived MSCs, such as shortages of tissue sources, difficult and invasive retrieval methods, cell population heterogeneity, low purity, cell senescence, and loss of pluripotency and proliferative capacities over continuous passages. Therefore, other methods to obtain high-quality MSCs need to be developed to overcome the limitations of tissue-derived MSCs. Pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), are considered potent sources for the derivation of MSCs. PSC-derived MSCs (PSC-MSCs) may surpass tissue-derived MSCs in proliferation capacity, immunomodulatory activity, and in vivo therapeutic applications. In this review, we will discuss basic as well as recent protocols for the production of PSC-MSCs and their in vitro and in vivo therapeutic efficacies. A better understanding of the current advances in the production of PSC-MSCs will inspire scientists to devise more efficient differentiation methods that will be a breakthrough in the clinical application of PSC-MSCs.


Assuntos
Reprogramação Celular/genética , Heterogeneidade Genética , Células-Tronco Mesenquimais/citologia , Células-Tronco Pluripotentes/citologia , Diferenciação Celular/genética , Linhagem da Célula/genética , Proliferação de Células/genética , Células-Tronco Embrionárias/citologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Medicina Regenerativa
15.
Diabetes Res Clin Pract ; 151: 96-105, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30954511

RESUMO

AIMS: To conduct a meta-analysis of statin-associated type 2 diabetes mellitus (T2D) risk among randomized controlled trials (RCTs) and observational studies (OBSs), excluding studies conducted among secondary prevention populations. METHODS: Studies were identified by searching PubMed (1994-present) and EMBASE (1994-present). Articles had to meet the following criteria: (1) follow-up >one year; (2) >50% of participants free of clinically diagnosed ASCVD; (3) adult participants ≥30 years old; (4) reported statin-associated T2D effect estimates; and (5) quantified precision using 95% confidence interval. Data were pooled using random-effects model. RESULTS: We identified 23 studies (35% RCTs) of n = 4,012,555 participants. OBS participants were on average younger (mean difference = 6.2 years) and had lower mean low-density lipoprotein cholesterol (LDL-C, mean difference = 20.6 mg/dL) and mean fasting plasma glucose (mean difference = 5.2 mg/dL) compared to RCT participants. There was little evidence for publication bias (P > 0.1). However, evidence of heterogeneity was observed overall and among OBSs and RCTs (PCochran = <0.05). OBS designs, younger baseline mean ages, lower LDL-C concentrations, and high proportions of never or former smokers were significantly associated with increased statin-associated T2D risk. CONCLUSIONS: Potentially elevated statin-associated T2D risk in younger populations with lower LDL-C merits further investigation in light of evolving statin guidelines targeting primary prevention populations.


Assuntos
Diabetes Mellitus Tipo 2/induzido quimicamente , Inibidores de Hidroximetilglutaril-CoA Redutases/efeitos adversos , Adulto , Idoso , Heterogeneidade Genética , Humanos , Masculino , Pessoa de Meia-Idade , Ensaios Clínicos Controlados Aleatórios como Assunto
16.
Pediatr Cardiol ; 40(5): 950-957, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30993396

RESUMO

Dilated cardiomyopathy (DCM) is characterized by left or bilateral ventricular dilation and systolic dysfunction without rational conditions, which can lead to progressive heart failure and sudden cardiac death. Most of the pathogenic genes have been reported in adult population by locus mapping in familial cases and animal model studies. However, it still remains challenging to decipher the role of genetics in the etiology of pediatric DCM. We applied whole-exome sequencing (WES) for 30 sporadic pediatric DCM subjects and 100 non-DCM local controls. We identified the pathogenic mutations using bioinformatics tools based on genomic strategies synergistically and confirmed mutations by Sanger sequencing. We identified compound heterozygous nonsense mutations in DSP (c.3799C > T, p.R1267X; c.4444G > T, p.E1482X). In sporadic cases, the two heterozygous mutations in XIRP2 were identified. Then we performed an exome-wide association study with 30 case and 100 control subjects. Interestingly, we could not identify TTN truncating variants in all cases. Collectively, we observed a significant risk signal between carriers of TTN deleterious missense variants and DCM risk (odds ratio 4.0, 95% confidence interval 1.1-22.2, p = 3.12 × 10-2). Our observations expanded the spectrum of mutations and were valuable in the pre- and postnatal screening and genetic diagnosis for DCM.


Assuntos
Cardiomiopatia Dilatada/genética , Heterogeneidade Genética , Sequenciamento Completo do Exoma/métodos , Cardiomiopatia Dilatada/diagnóstico , Estudos de Casos e Controles , Criança , Pré-Escolar , China , Códon sem Sentido , Feminino , Humanos , Masculino , Linhagem , Mutação Puntual
18.
Food Microbiol ; 81: 89-96, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30910091

RESUMO

Bacterial spores are ubiquitous in nature and can withstand both chemical and physical stresses. Spores can survive food preservation processes and upon outgrowth cause food spoilage as well as safety risks. The heterogeneous germination and outgrowth behavior of isogenic spore populations exacerbates this risk. A major unknown factor of spores is likely to be the inherently heterogeneous spore protein composition. The proteomics methods discussed here help in broadening the knowledge about spore structure and identification of putative target proteins from spores of different spore formers. Approaches to synchronize Bacillus subtilis spore formation, and to analyze spore proteins as well as the physiology of spore germination and outgrowth are also discussed. Live-imaging and fluorescence microscopy techniques discussed here allow analysis, at single cell level, of the 'germinosome', the process of spore germination itself, spore outgrowth and the spore intracellular pH dynamics. For the latter, a recently published improved pHluorin (IpHluorin) under control of the ptsG promoter is applicable. While the data obtained from such tools offers novel insight in the mechanisms of bacterial spore awakening, it may also be used to probe candidate antimicrobial compounds for inhibitory effects on spore germination and strengthen microbial risk assessment.


Assuntos
Farmacorresistência Bacteriana , Microbiologia de Alimentos , Microscopia/métodos , Proteômica/métodos , Esporos Bacterianos/crescimento & desenvolvimento , Esporos Bacterianos/metabolismo , Bacillus subtilis/genética , Bacillus subtilis/crescimento & desenvolvimento , Bacillus subtilis/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Manipulação de Alimentos , Conservação de Alimentos , Heterogeneidade Genética , Concentração de Íons de Hidrogênio , Cinética , Modelos Teóricos , Sistema Fosfotransferase de Açúcar do Fosfoenolpiruvato/genética , Regiões Promotoras Genéticas , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Esporos Bacterianos/citologia , Esporos Bacterianos/genética , Estresse Fisiológico
19.
Proc Natl Acad Sci U S A ; 116(13): 6244-6249, 2019 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-30867282

RESUMO

An approach to the coalescent, the fractional coalescent (f-coalescent), is introduced. The derivation is based on the discrete-time Cannings population model in which the variance of the number of offspring depends on the parameter α. This additional parameter α affects the variability of the patterns of the waiting times; values of [Formula: see text] lead to an increase of short time intervals, but occasionally allow for very long time intervals. When [Formula: see text], the f-coalescent and the Kingman's n-coalescent are equivalent. The distribution of the time to the most recent common ancestor and the probability that n genes descend from m ancestral genes in a time interval of length T for the f-coalescent are derived. The f-coalescent has been implemented in the population genetic model inference software Migrate Simulation studies suggest that it is possible to accurately estimate α values from data that were generated with known α values and that the f-coalescent can detect potential environmental heterogeneity within a population. Bayes factor comparisons of simulated data with [Formula: see text] and real data (H1N1 influenza and malaria parasites) showed an improved model fit of the f-coalescent over the n-coalescent. The development of the f-coalescent and its inclusion into the inference program Migrate facilitates testing for deviations from the n-coalescent.


Assuntos
Teorema de Bayes , Heterogeneidade Genética , Genética Populacional/métodos , Modelos Genéticos , Simulação por Computador , Meio Ambiente , Genoma Humano , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Malária , Modelos Estatísticos , Mutação , Crescimento Demográfico , Seleção Genética , Software
20.
Mol Vis ; 25: 144-154, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30820150

RESUMO

Purpose: Primary congenital glaucoma (PCG) is a clinically and genetically heterogeneous disease. The present study was undertaken to find the genetic causes of PCG segregating in 36 large consanguineous Pakistani families. Methods: Ophthalmic examination including fundoscopy, or slit-lamp microscopy was performed to clinically characterize the PCG phenotype. Genomic nucleotide sequences of the CYP1B1 and LTBP2 genes were analyzed with either Sanger or whole exome sequencing. In silico prediction programs were used to assess the pathogenicity of identified alleles. ClustalW alignments were performed to determine evolutionary conservation, and three-dimensional (3D) modeling was performed using HOPE and Phyre2 software. Results: Among the known loci, mutations in CYP1B1 and LTBP2 are the common causes of PCG. Therefore, we analyzed the genomic nucleotide sequences of CYP1B1 and LTBP2, and detected probable pathogenic variants cosegregating with PCG in 14 families. These included the three novel (c.542T>A, c.1436A>G, and c.1325delC) and five known (c.868dupC, c.1168C>T, c.1169G>A, c.1209InsTCATGCCACC, and c.1310C>T) variants in CYP1B1. Two of the novel variants are missense substitutions [p.(Leu181Gln), p.(Gln479Arg)], which replaced evolutionary conserved amino acids, and are predicted to be pathogenic by various in silico programs, while the third variant (c.1325delC) is predicted to cause reading frameshift and premature truncation of the protein. A single mutation, p.(Arg390His), causes PCG in six (~43%) of the 14 CYP1B1 mutations harboring families, and thus, is the most common variant in this cohort. Surprisingly, we did not find any LTBP2 pathogenic variants in the families, which further supports the genetic heterogeneity of PCG in the Pakistani population. Conclusions: In conclusion, results of the present study enhance our understanding of the genetic basis of PCG, support the notion of a genetic modifier of CYP1B1, and contribute to the development of genetic testing protocols and genetic counseling for PCG in Pakistani families.


Assuntos
Citocromo P-450 CYP1B1/genética , Heterogeneidade Genética , Glaucoma/genética , Mutação , Adolescente , Adulto , Idoso , Alelos , Sequência de Aminoácidos , Substituição de Aminoácidos , Criança , Pré-Escolar , Consanguinidade , Análise Mutacional de DNA , Feminino , Expressão Gênica , Frequência do Gene , Glaucoma/congênito , Glaucoma/patologia , Glaucoma/cirurgia , Humanos , Lactente , Proteínas de Ligação a TGF-beta Latente/genética , Masculino , Pessoa de Meia-Idade , Paquistão , Linhagem , Alinhamento de Sequência , Trabeculectomia/métodos
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