RESUMO
Alternative splicing is prevalent in the mammalian brain. To interrogate the functional role of alternative splicing in neural development, we analyzed purified neural progenitor cells (NPCs) and neurons from developing cerebral cortices, revealing hundreds of differentially spliced exons that preferentially alter key protein domains-especially in cytoskeletal proteins-and can harbor disease-causing mutations. We show that Ptbp1 and Rbfox proteins antagonistically govern the NPC-to-neuron transition by regulating neuron-specific exons. Whereas Ptbp1 maintains apical progenitors partly through suppressing a poison exon of Flna in NPCs, Rbfox proteins promote neuronal differentiation by switching Ninein from a centrosomal splice form in NPCs to a non-centrosomal isoform in neurons. We further uncover an intronic human mutation within a PTBP1-binding site that disrupts normal skipping of the FLNA poison exon in NPCs and causes a brain-specific malformation. Our study indicates that dynamic control of alternative splicing governs cell fate in cerebral cortical development.
Assuntos
Processamento Alternativo , Córtex Cerebral/embriologia , Células-Tronco Neurais/citologia , Neurogênese/genética , Neurônios/citologia , Animais , Centrossomo/metabolismo , Córtex Cerebral/anormalidades , Córtex Cerebral/citologia , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Éxons , Ribonucleoproteínas Nucleares Heterogêneas/genética , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Humanos , Camundongos , Células-Tronco Neurais/metabolismo , Neurônios/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Proteína de Ligação a Regiões Ricas em Polipirimidinas/metabolismo , Domínios Proteicos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Fatores de Processamento de RNARESUMO
Differential polyadenylation sites (PAs) critically regulate gene expression, but their cell type-specific usage and spatial distribution in the brain have not been systematically characterized. Here, we present Infernape, which infers and quantifies PA usage from single-cell and spatial transcriptomic data and show its application in the mouse brain. Infernape uncovers alternative intronic PAs and 3'-UTR lengthening during cortical neurogenesis. Progenitor-neuron comparisons in the excitatory and inhibitory neuron lineages show overlapping PA changes in embryonic brains, suggesting that the neural proliferation-differentiation axis plays a prominent role. In the adult mouse brain, we uncover cell type-specific PAs and visualize such events using spatial transcriptomic data. Over two dozen neurodevelopmental disorder-associated genes such as Csnk2a1 and Mecp2 show differential PAs during brain development. This study presents Infernape to identify PAs from scRNA-seq and spatial data, and highlights the role of alternative PAs in neuronal gene regulation.
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Regulação da Expressão Gênica , Poliadenilação , Camundongos , Animais , Neurônios/metabolismo , Regiões 3' não Traduzidas/genética , EncéfaloRESUMO
The genetic identification of skeletal remains from Chinese People's Volunteers (CPVs) of the Korean War has been challenging because of the degraded DNA samples and the lack of living close relatives. This study established a workflow for identifying CPVs by combining Y-chromosome short tandem repeats (Y-STRs), mitochondrial DNA (mtDNA) hypervariable regions I and II, autosomal STRs (aSTRs), and identity-informative SNPs (iiSNPs). A total of 20 skeletal remains of CPVs and 46 samples from their alleged relatives were collected. The success rate of DNA extraction from human remains was 100%. Based on Y-STRs, six remains shared the same male lineages with their alleged relatives. Meanwhile, mtDNA genotyping supports two remains sharing the same maternal lineages with their alleged relatives. Likelihood ratios (LRs) were further obtained from 27 aSTRs and 94 iiSNPs or 1936 iiSNPs to confirm their relationship. All joint pedigree LRs were >100. Finally, six remains were successfully identified. This pilot study for the systematic genetic identification of CPVs from the Korean War can be applied for the large-scale identification of CPVs in the future.
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Povo Asiático , Cromossomos Humanos Y , DNA Mitocondrial , Guerra da Coreia , Repetições de Microssatélites , Polimorfismo de Nucleotídeo Único , Humanos , DNA Mitocondrial/genética , DNA Mitocondrial/análise , Masculino , Repetições de Microssatélites/genética , Cromossomos Humanos Y/genética , Povo Asiático/genética , Restos Mortais , Genética Forense/métodos , Feminino , Linhagem , Projetos PilotoRESUMO
In the mammalian neocortex, projection neuron types are sequentially generated by the same pool of neural progenitors. How neuron type specification is related to developmental timing remains unclear. To determine whether temporal gene expression in neural progenitors correlates with neuron type specification, we performed single-cell RNA sequencing (scRNA-Seq) analysis of the developing mouse neocortex. We uncovered neuroepithelial cell enriched genes such as Hmga2 and Ccnd1 when compared to radial glial cells (RGCs). RGCs display dynamic gene expression over time; for instance, early RGCs express higher levels of Hes5, and late RGCs show higher expression of Pou3f2 Interestingly, intermediate progenitor cell marker gene Eomes coexpresses temporally with known neuronal identity genes at different developmental stages, though mostly in postmitotic cells. Our results delineate neural progenitor cell diversity in the developing mouse neocortex and support that neuronal identity genes are transcriptionally evident in Eomes-positive cells.
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Diferenciação Celular , Neocórtex/embriologia , Células-Tronco Neurais/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteína HMGA2/metabolismo , Camundongos , Neocórtex/citologia , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/citologia , Neuroglia/citologia , Neuroglia/metabolismo , Fatores do Domínio POU/metabolismo , Proteínas Repressoras/metabolismo , Proteínas com Domínio T/metabolismoRESUMO
Chronic Hepatitis B virus (CHB) infection is a global health challenge, causing damage ranging from hepatitis to cirrhosis and hepatocellular carcinoma. In our study, single-cell RNA sequencing (scRNA-seq) analysis was performed in livers from mice models with chronic inflammation induced by CHB infection and we found that endothelial cells (ECs) exhibited the largest number of differentially expressed genes (DEGs) among all ten cell types. NF-κB signaling was activated in ECs to induce cell dysfunction and subsequent hepatic inflammation, which might be mediated by the interaction of macrophage-derived and cholangiocyte-derived VISFATIN/Nampt signaling. Moreover, we divided ECs into three subclusters, including periportal ECs (EC_Z1), midzonal ECs (EC_Z2), and pericentral ECs (EC_Z3) according to hepatic zonation. Functional analysis suggested that pericentral ECs and midzonal ECs, instead of periportal ECs, were more vulnerable to HBV infection, as the VISFATIN/Nampt- NF-κB axis was mainly altered in these two subpopulations. Interestingly, pericentral ECs showed increasing communication with macrophages and cholangiocytes via the Nampt-Insr and Nampt-Itga5/Itgb1 axis upon CHB infection, which contribute to angiogenesis and vascular capillarization. Additionally, ECs, especially pericentral ECs, showed a close connection with nature killer (NK) cells and T cells via the Cxcl6-Cxcr6 axis, which is involved in shaping the microenvironment in CHB mice livers. Thus, our study described the heterogeneity and functional alterations of three subclusters in ECs. We revealed the potential role of VISFATIN/Nampt signaling in modulating ECs characteristics and related hepatic inflammation, and EC-derived chemokine Cxcl16 in shaping NK and T cell recruitment, providing key insights into the multifunctionality of ECs in CHB-associated pathologies.
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Células Endoteliais , Hepatite B Crônica , Análise de Célula Única , Animais , Hepatite B Crônica/virologia , Hepatite B Crônica/genética , Hepatite B Crônica/metabolismo , Camundongos , Células Endoteliais/metabolismo , Células Endoteliais/virologia , Análise de Sequência de RNA , Vírus da Hepatite B/genética , Vírus da Hepatite B/fisiologia , Transdução de Sinais , Fígado/metabolismo , Fígado/virologia , Fígado/patologia , NF-kappa B/metabolismo , Masculino , Nicotinamida Fosforribosiltransferase/metabolismo , Nicotinamida Fosforribosiltransferase/genética , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , HumanosRESUMO
Baicalin (7-d-glucuronic acid-5, 6-dihydroxyflavone) derived from the root of Scutellaria baicalensis used as Traditional Chinese Medicine (TCM) has been revealed to exert potential antiviral activity via various pathways, while the molecular mechanisms have not been fully understood. Pyroptosis, an inflammatory form of programmed cell death (PCD), is reported to play a crucial role in host cell fate during viral infection. In this study, transcriptome analysis of mice lung tissue reveals that baicalin reverses the alterations of the mRNA levels of PCD-associated genes upon H1N1 challenge, with a concomitant decrease in the population of H1N1-induced propidium iodide (PI)+ and Annexin â ¤+ cells. Intriguingly, we find that baicalin contributes to the survival of infected lung alveolar epithelial cells partly through its inhibition of H1N1-induced cell pyroptosis, which is manifested by reduced bubble-like protrusion cells and lactate dehydrogenase (LDH) release. Moreover, the antipyroptosis effect of baicalin in response to H1N1 infection is found to be mediated by its repression on caspase-3/Gasdermin E (GSDME) pathway. Cleaved caspase-3 and N-terminal fragment of GSDME (GSDME-N) are detected in H1N1-infected cell lines and mice lung tissues, which are markedly reversed by baicalin treatment. Furthermore, inhibition of caspase-3/GSDME pathway by caspase-3 inhibitor or siRNA exerts an antipyroptosis effect equal to that of baicalin treatment in infected A549 and BEAS-2B cells, indicating a pivotal role of caspase-3 in the antiviral activities of baicalin. Conclusively, for the first time, we demonstrate that baicalin could effectively suppress H1N1-induced pyroptosis of lung alveolar epithelial cells via caspase-3/GSDME pathway both in vitro and in vivo.
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Vírus da Influenza A Subtipo H1N1 , Influenza Humana , Animais , Camundongos , Humanos , Piroptose , Células Epiteliais Alveolares/metabolismo , Influenza Humana/tratamento farmacológico , Caspase 3/genética , Caspase 3/metabolismo , Pulmão/metabolismo , Antivirais/farmacologiaRESUMO
PURPOSE: Adducins interconnect spectrin and actin filaments to form polygonal scaffolds beneath the cell membranes and form ring-like structures in neuronal axons. Adducins regulate mouse neural development, but their function in the human brain is unknown. METHODS: We used exome sequencing to uncover ADD1 variants associated with intellectual disability (ID) and brain malformations. We studied ADD1 splice isoforms in mouse and human neocortex development with RNA sequencing, super resolution imaging, and immunoblotting. We investigated 4 variant ADD1 proteins and heterozygous ADD1 cells for protein expression and ADD1-ADD2 dimerization. We studied Add1 functions in vivo using Add1 knockout mice. RESULTS: We uncovered loss-of-function ADD1 variants in 4 unrelated individuals affected by ID and/or structural brain defects. Three additional de novo copy number variations covering the ADD1 locus were associated with ID and brain malformations. ADD1 is highly expressed in the neocortex and the corpus callosum, whereas ADD1 splice isoforms are dynamically expressed between cortical progenitors and postmitotic neurons. Human variants impair ADD1 protein expression and/or dimerization with ADD2. Add1 knockout mice recapitulate corpus callosum dysgenesis and ventriculomegaly phenotypes. CONCLUSION: Our human and mouse genetics results indicate that pathogenic ADD1 variants cause corpus callosum dysgenesis, ventriculomegaly, and/or ID.
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Hidrocefalia , Deficiência Intelectual , Agenesia do Corpo Caloso/genética , Agenesia do Corpo Caloso/patologia , Animais , Variações do Número de Cópias de DNA , Humanos , Hidrocefalia/genética , Deficiência Intelectual/genética , Camundongos , FenótipoRESUMO
Accurately predicting the binding affinity of protein-ligand pairs is an essential part of drug discovery. Since wet laboratory experiments to determine the binding affinity are expensive and time-consuming, several computational methods for binding affinity prediction have been proposed. In the representation of compounds, most methods only focus on the structural properties such as SMILES and ignore the bioactive properties. In this study, we proposed a novel model named PLA-MoRe to predict protein-ligand binding affinity, which represents compounds based on both structural and bioactive properties and mainly contains three feature extractors. First, a structure feature extractor based on the graph isomorphism network was constructed to learn the representations of the molecular graphs. Second, we designed an Autoencoder-based bioactive feature extractor to integrate the multisource bioactive information including chemical, target, network, cellular, and clinical. The above two parts aimed to learn representations of compounds in terms of structures and bioactivities, respectively. Then, we constructed a sequence feature extractor to learn embeddings for protein sequences. The output of the three extractors was concatenated and fed into a fully connected network for affinity prediction. We compared PLA-MoRe with three state-of-the-art methods, and an ablation study was conducted to test the role of each part of the model. Further attention visualization showed that our model had the potential to locate the binding sites, which might help explain the mechanism of interaction. These results prove that PLA-MoRe is competitive and reliable. The resource codes are freely available at the GitHub repository https://github.com/QingyuLiaib/PLA-MoRe.
Assuntos
Poliésteres , Proteínas , Sequência de Aminoácidos , Sítios de Ligação , Ligantes , Proteínas/químicaRESUMO
PURPOSE: The exocyst complex is a conserved protein complex that mediates fusion of intracellular vesicles to the plasma membrane and is implicated in processes including cell polarity, cell migration, ciliogenesis, cytokinesis, autophagy, and fusion of secretory vesicles. The essential role of these genes in human genetic disorders, however, is unknown. METHODS: We performed homozygosity mapping and exome sequencing of consanguineous families with recessively inherited brain development disorders. We modeled an EXOC7 splice variant in vitro and examined EXOC7 messenger RNA (mRNA) expression in developing mouse and human cortex. We modeled exoc7 loss-of-function in a zebrafish knockout. RESULTS: We report variants in exocyst complex members, EXOC7 and EXOC8, in a novel disorder of cerebral cortex development. In EXOC7, we identified four independent partial loss-of-function (LOF) variants in a recessively inherited disorder characterized by brain atrophy, seizures, and developmental delay, and in severe cases, microcephaly and infantile death. In EXOC8, we found a homozygous truncating variant in a family with a similar clinical disorder. We modeled exoc7 deficiency in zebrafish and found the absence of exoc7 causes microcephaly. CONCLUSION: Our results highlight the essential role of the exocyst pathway in normal cortical development and how its perturbation causes complex brain disorders.
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Encefalopatias , Microcefalia , Animais , Proliferação de Células/genética , Homozigoto , Humanos , Camundongos , Microcefalia/genética , Peixe-Zebra/genéticaRESUMO
OBJECTIVE: To explore current status of knowledge, attitude and practice of salt reduction among adults in four counties of Shandong and Jiangsu Provinces, and analyze the relevant influencing factors. METHODS: In 2013, multi-stage cluster sampling was used to select the adults(aged 18-69) in Gaomi City of Weifang City and Fushan Distract of Yantai City, Shandong Province, and Xinyi City of Xuzhou City and Ganyu County of Lianyungang City, Jiangsu Province. A total of 9573 subjects were included. Knowledge, attitude and practice of anti-hypertension by low salt was collected using questionnaire and physical examination was conducted. Then the level of knowledge, attitude and practice of salt reduction was calculated, and multivariate Logistic regression model was used to analyze the influencing factors. RESULTS: A total of 3214 people had hypertension(33.57%), and the proportion of overweight and obesity was 36.20% and 19.48%, respectively. The awareness rate of salt reduction knowledge was between 22.50% and 51.35%; the attitude of salt reduction was between 82.24% and 93.01%, and the rate of salt reduction was between 10.86% and 46.16%. According to Logistic analysis, the degree of education had the greatest impact on knowledge(junior high school vs. elementary school and below: OR=2.30, 95%CI 2.06-2.57, high school and above vs. elementary school and below: OR=5.00, 95%CI 4.35-5.76). The knowledge level had the greatest impact on attitude(OR=3.10, 95%CI 2.80-3.43) and practice(OR=4.66, 95%CI 4.29-5.07). Those aged 45-69 years old had lower knowledge level(OR=0.64, 95%CI 0.58-0.71) but higher practice(OR=1.18, 95%CI 1.09-1.29). The knowledge(OR=3.66, 95%CI 3.34-4.01) and attitude(OR=1.84, 95%CI 1.69-2.02) of salt reduction among residents in Shandong were higher than those in Jiangsu. CONCLUSION: The awareness rate of knowledge is lower, the attitude support rate is higher, and the practice rate is lower among residents in Shandong and Jiangsu Provinces. More attention should be paid to the practical application of salt reduction.
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Hipertensão , Cloreto de Sódio na Dieta , Adolescente , Adulto , Idoso , China , Cidades , Conhecimentos, Atitudes e Prática em Saúde , Humanos , Pessoa de Meia-Idade , Inquéritos e Questionários , Adulto JovemRESUMO
Progressive microcephaly is a heterogeneous condition with causes including mutations in genes encoding regulators of neuronal survival. Here, we report the identification of mutations in QARS (encoding glutaminyl-tRNA synthetase [QARS]) as the causative variants in two unrelated families affected by progressive microcephaly, severe seizures in infancy, atrophy of the cerebral cortex and cerebellar vermis, and mild atrophy of the cerebellar hemispheres. Whole-exome sequencing of individuals from each family independently identified compound-heterozygous mutations in QARS as the only candidate causative variants. QARS was highly expressed in the developing fetal human cerebral cortex in many cell types. The four QARS mutations altered highly conserved amino acids, and the aminoacylation activity of QARS was significantly impaired in mutant cell lines. Variants p.Gly45Val and p.Tyr57His were located in the N-terminal domain required for QARS interaction with proteins in the multisynthetase complex and potentially with glutamine tRNA, and recombinant QARS proteins bearing either substitution showed an over 10-fold reduction in aminoacylation activity. Conversely, variants p.Arg403Trp and p.Arg515Trp, each occurring in a different family, were located in the catalytic core and completely disrupted QARS aminoacylation activity in vitro. Furthermore, p.Arg403Trp and p.Arg515Trp rendered QARS less soluble, and p.Arg403Trp disrupted QARS-RARS (arginyl-tRNA synthetase 1) interaction. In zebrafish, homozygous qars loss of function caused decreased brain and eye size and extensive cell death in the brain. Our results highlight the importance of QARS during brain development and that epilepsy due to impairment of QARS activity is unusually severe in comparison to other aminoacyl-tRNA synthetase disorders.
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Aminoacil-tRNA Sintetases/genética , Encefalopatias/genética , Predisposição Genética para Doença , Microcefalia/genética , Mutação , Convulsões/genética , Aminoacilação , Animais , Pré-Escolar , Feminino , Humanos , Imageamento por Ressonância Magnética , Masculino , Microcefalia/patologia , Linhagem , Peixe-ZebraRESUMO
BACKGROUND: Although there is increasing recognition of the role of somatic mutations in genetic disorders, the prevalence of somatic mutations in neurodevelopmental disease and the optimal techniques to detect somatic mosaicism have not been systematically evaluated. METHODS: Using a customized panel of known and candidate genes associated with brain malformations, we applied targeted high-coverage sequencing (depth, ≥200×) to leukocyte-derived DNA samples from 158 persons with brain malformations, including the double-cortex syndrome (subcortical band heterotopia, 30 persons), polymicrogyria with megalencephaly (20), periventricular nodular heterotopia (61), and pachygyria (47). We validated candidate mutations with the use of Sanger sequencing and, for variants present at unequal read depths, subcloning followed by colony sequencing. RESULTS: Validated, causal mutations were found in 27 persons (17%; range, 10 to 30% for each phenotype). Mutations were somatic in 8 of the 27 (30%), predominantly in persons with the double-cortex syndrome (in whom we found mutations in DCX and LIS1), persons with periventricular nodular heterotopia (FLNA), and persons with pachygyria (TUBB2B). Of the somatic mutations we detected, 5 (63%) were undetectable with the use of traditional Sanger sequencing but were validated through subcloning and subsequent sequencing of the subcloned DNA. We found potentially causal mutations in the candidate genes DYNC1H1, KIF5C, and other kinesin genes in persons with pachygyria. CONCLUSIONS: Targeted sequencing was found to be useful for detecting somatic mutations in patients with brain malformations. High-coverage sequencing panels provide an important complement to whole-exome and whole-genome sequencing in the evaluation of somatic mutations in neuropsychiatric disease. (Funded by the National Institute of Neurological Disorders and Stroke and others.).
Assuntos
Córtex Cerebral/anormalidades , Análise Mutacional de DNA/métodos , Malformações do Desenvolvimento Cortical/genética , Mutação , Lissencefalias Clássicas e Heterotopias Subcorticais em Banda/genética , Humanos , Lisencefalia/genética , Imageamento por Ressonância Magnética , Malformações do Desenvolvimento Cortical/patologia , Heterotopia Nodular Periventricular/genéticaRESUMO
Environmental stresses and nutrition availability critically affect animal development. Numerous animal species across multiple phyla enter developmental arrest for long-term survival in unfavorable environments and resume development upon stress removal. Here we show that compromising overall microRNA (miRNA) functions or mutating certain individual miRNAs impairs the long-term survival of nematodes during starvation-induced L1 diapause. We provide evidence that miRNA miR-71 is not required for the animals' entry into L1 diapause, but plays a critical role in long-term survival by repressing the expression of insulin receptor/PI3K pathway genes and genes acting downstream or in parallel to the pathway. Furthermore, miR-71 plays a prominent role in developmental recovery from L1 diapause partly through repressing the expression of certain heterochronic genes. The presented results indicate that interactions between multiple miRNAs and likely a large number of their mRNA targets in multiple pathways regulate the response to starvation-induced L1 diapause.
Assuntos
Caenorhabditis elegans/fisiologia , MicroRNAs/fisiologia , Inanição , Regiões 3' não Traduzidas , Animais , Caenorhabditis elegans/citologia , Caenorhabditis elegans/genética , Divisão Celular , MutaçãoRESUMO
OBJECTIVE: To assess the implementation of World Health Organization Framework Convention on Tobacco Control (WHO FCTC) by organizations in seven provinces/municipalities in China. METHODS: A total of 901 organizations, including hospitals, schools, government departments, health administrative departments and public transportation facilities, were selected by multistage sampling method in five provinces and two municipalities in China, 2010 and 2012. Key informant interview and observation survey were conducted to collect data on implementation of WHO FCTC. Analysis was performed among three clusters of indexes, which were establishment of smoke-free environment, education and training on tobacco control, and tobacco cessation measures. RESULTS: The five types of organizations performed differently in creating smoke-free environment. The ratios of conducting complete smoke-free policy in hospitals, health administrative departments and schools were separately 83.3% (111/192), 81.6% (146/179) and 66.5% (121/182) in 2012, which were comparatively higher than those in governments (32.4%, 33/102) and public transport facilities (25.0%, 27/108) (χ(2) = 174.93, P < 0.01) . As for promotion and training programs of tobacco control information, the ratio of health administrative departments raised from 78.1% (150/192) to 100.0% (192/192), and the difference showed statistical significance (χ(2) = 42.00, P < 0.01). But those departments who provide training to social media only accounted for 11.8% (22/187), which was substantially lower than the percentage of those providing training to themselves (67.7%, 128/189) (χ(2) = 413.99, P < 0.01). Three indexes of tobacco cessation related work--offering help in tobacco use quitting, providing health workers with training on tobacco cessation skills and establishing tobacco cessation clinics, only reached as low as 41.1% (312/760) , 55.6% (104/187) and 45.9% (89/194), respectively. Among the seven provinces/municipalities, Shanghai did better than the others on most of the indexes. 27 out of 28 schools in Shanghai implemented complete smoke-free policy, which was the best among the seven, while only 3 out of 26 in Jiangxi, as the poorest (χ(2) = 47.63, P < 0.01). Meanwhile, all of the 24 schools in Shanghai had health education classes on tobacco control, followed by Zhejiang (23 out of 29), while Jiangxi was also the poorest, only 12 out of 26 (χ(2) = 17.95, P < 0.01). CONCLUSION: The implementation of WHO FCTC by the five types of organizations in the seven provinces/municipalities has improved to various degrees. Nevertheless, further actions should be taken to promote smoke-free environment especially in certain circumstances such as public transport facilities, to strengthen training programs of tobacco control for social media, and to enhance hospitals' abilities in providing tobacco cessation services.
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Educação em Saúde , Política de Saúde , Abandono do Hábito de Fumar , Poluição por Fumaça de Tabaco , China , Cidades , Coleta de Dados , Órgãos Governamentais , Hospitais , Humanos , Instituições Acadêmicas , Fumar , Nicotiana , Organização Mundial da SaúdeRESUMO
Pediatric neurological disorders are frequently devastating and present unmet needs for effective medicine. The successful treatment of spinal muscular atrophy with splice-switching antisense oligonucleotides (SSO) indicates a feasible path to targeting neurological disorders by redirecting pre-mRNA splicing. One direct outcome is the development of SSOs to treat haploinsufficient disorders by targeting naturally occurring non-productive splice isoforms. The development of personalized SSO treatment further inspired the therapeutic exploration of rare diseases. This review will discuss the recent advances that utilize SSOs to treat pediatric neurological disorders.
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In spite of the development of diagnostic tests for Mycobacterium tuberculosis (M. tuberculosis), the etiological agent of tuberculosis, there has remained a gap between the established methods and an easily accessible diagnostic test, particularly in developing and resource-poor areas. By combining isothermal amplification of IS6110 as the target gene and recognition by DNA-functionalized Au nanoparticles (DNA-AuNPs), we develop a colorimetric LAMP assay for convenient in vitro diagnostics of tuberculosis with a quick (≤50 min) "yes" or "no" readout. The DNA-AuNPs not only tolerate the interference in the complex LAMP system but also afford in situ identification of the amplicon, allowing for colloidal dispersion via steric effect depending on DNA grafting density. The target-induced stabilization and red appearance of the DNA-AuNPs contrast with the occurrence of gray aggregates in a negative sample. Furthermore, the DNA-AuNPs demonstrate excellent performance after long-term (≥7 months) storage while preserving the unsacrificed sensitivity. The high specificity of the DNA-AuNPs is further demonstrated in the naked-eye LAMP assay of M. tuberculosis in patients' sputum samples. Given the rapidity, cost-effectiveness, and instrument-free characteristics, the naked-eye LAMP assay is particularly beneficial for tuberculosis diagnosis in urgent situations and resource-limited settings and can potentially expedite patient care and treatment initiation.
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Ouro , Nanopartículas Metálicas , Técnicas de Diagnóstico Molecular , Mycobacterium tuberculosis , Técnicas de Amplificação de Ácido Nucleico , Escarro , Tuberculose , Ouro/química , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Escarro/microbiologia , Humanos , Nanopartículas Metálicas/química , Técnicas de Amplificação de Ácido Nucleico/métodos , Tuberculose/diagnóstico , Tuberculose/microbiologia , Técnicas de Diagnóstico Molecular/métodos , Colorimetria/métodos , Sensibilidade e Especificidade , DNA Bacteriano/genética , DNA Bacteriano/análiseRESUMO
Animals adapt to environmental challenges with long-term changes at the behavioral, circuit, cellular, and synaptic levels which often require new protein synthesis. The discovery of reversible N6-methyladenosine (m6A) modifications of mRNA has revealed an important layer of post-transcriptional regulation which affects almost every phase of mRNA metabolism and therefore translational control. Many in vitro and in vivo studies have demonstrated the significant role of m6A in cell differentiation and survival, but its role in adult neurons is understudied. We used cell-type specific gene deletion of Mettl14, which encodes one of the subunits of the m6A methyltransferase, and Ythdf1, which encodes one of the cytoplasmic m6A reader proteins, in dopamine D1 receptor expressing or D2 receptor expressing neurons. Mettl14 or Ythdf1 deficiency blunted responses to environmental challenges at the behavioral, cellular, and molecular levels. In three different behavioral paradigms, gene deletion of either Mettl14 or Ythdf1 in D1 neurons impaired D1-dependent learning, whereas gene deletion of either Mettl14 or Ythdf1 in D2 neurons impaired D2-dependent learning. At the cellular level, modulation of D1 and D2 neuron firing in response to changes in environments was blunted in all three behavioral paradigms in mutant mice. Ythdf1 deletion resembled impairment caused by Mettl14 deletion in a cell type-specific manner, suggesting YTHDF1 is the main mediator of the functional consequences of m6A mRNA methylation in the striatum. At the molecular level, while striatal neurons in control mice responded to elevated cAMP by increasing de novo protein synthesis, striatal neurons in Ythdf1 knockout mice didn't. Finally, boosting dopamine release by cocaine drastically increased YTHDF1 binding to many mRNA targets in the striatum, especially those that encode structural proteins, suggesting the initiation of long-term neuronal and/or synaptic structural changes. While the m6A-YTHDF1 pathway has similar functional significance at cellular level, its cell type specific deficiency in D1 and D2 neurons often resulted in contrasting behavioral phenotypes, allowing us to cleanly dissociate the opposing yet cooperative roles of D1 and D2 neurons.
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Background: Excessive sodium and low potassium intake are involved in the development of hypertension. Growing evidence showed that the sodium-to-potassium ratio (Na/K) was significantly associated with blood pressure (BP). However, studies on the dose-response relationship of spot urinary Na/K ratio with hypertension and BP in the general population are scarce, especially in the Chinese population. Materials and methods: Data from the post-intervention survey of the Shandong Ministry of Health Action on Salt and Hypertension (SMASH) project was analyzed. Associations between Na/K molar ratio and hypertension prevalence and between Na/K molar ratio and BP indices were analyzed using multivariable logistic and linear regression, respectively, followed by subgroup analysis and interaction analysis. The restricted cubic spline model was used to explore the dose-response relationship. Informed by existing literature, we adjusted for potential confounding factors, including temperature and renal function, to assess the association and dose-response relationship. Results: There was a non-linear positive association between Na/K and hypertension (OR:1.09, 95%CI: 1.08-1.11) and a linear positive association between Na/K and systolic BP, diastolic BP, and mean arterial pressure (ß 0.53, 95%CI: 0.45-0.60; ß 0.36, 95%CI: 0.31-0.41; and ß 0.42, 95%CI: 0.36-0.47, respectively). The association was stronger in individuals with hypertension, female patients, those in the 50-59-year age group, and those who were obese. Environmental temperatures had little impact on associations. Conclusion: Our findings provide further evidence that the spot urinary Na/K ratio is a simple, useful, and convenient indicator for monitoring salt reduction and potassium increase, which could be used in clinical and public health practices.
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Sanitation workers' workload increases quickly with rapid urbanization, but there is almost no evidence or policy recommendations for their management in developing countries. This study describes the health status and occupational protection of sanitation workers; it also explores risk factors related to their health status in Wuhan City, China. Three hundred and eighty-five sanitation workers from 54 streets of Wuhan were surveyed. Their prevalence of 2-week illness and arthritis was relatively higher than in the general population in China. Findings related to occupational protection showed that both sanitation workers (users) and their managers (providers) neglected the role of low-cost protection measures, especially masks, soap/hand sanitizer and prejob training (use rate of 7.27%, 26.75% and 43.64%, respectively). High-intensity workload was an important risk factor for 2-week illness, and prejob training was an important protective factor against arthritis.
Assuntos
Artrite/epidemiologia , Nível de Saúde , Saúde Ocupacional , Eliminação de Resíduos , Adulto , China/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Prevalência , População Urbana , Carga de TrabalhoRESUMO
RNA-binding proteins (RBPs) are essential for gene regulation, but it remains a challenge to identify their RNA targets across cell types. Here we present PIE-Seq to investigate Protein-RNA Interaction with dual-deaminase Editing and Sequencing by conjugating C-to-U and A-to-I base editors to RBPs. We benchmark PIE-Seq and demonstrate its sensitivity in single cells, its application in the developing brain, and its scalability with 25 human RBPs. Bulk PIE-Seq identifies canonical binding features for RBPs such as PUM2 and NOVA1, and nominates additional target genes for most tested RBPs such as SRSF1 and TDP-43/TARDBP. Homologous RBPs frequently edit similar sequences and gene sets in PIE-Seq while different RBP families show distinct targets. Single-cell PIE-PUM2 uncovers comparable targets to bulk samples and applying PIE-PUM2 to the developing mouse neocortex identifies neural-progenitor- and neuron-specific target genes such as App. In summary, PIE-Seq provides an orthogonal approach and resource to uncover RBP targets in mice and human cells.