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Sudden unexpected death in epilepsy (SUDEP) is the leading cause of death among patients with epilepsy, causing a global public health burden. The underlying mechanisms of SUDEP remain elusive, and effective prevention or treatment strategies require further investigation. A major challenge in current SUDEP research is the lack of an ideal model that maximally mimics the human condition. Animal models are important for revealing the potential pathogenesis of SUDEP and preventing its occurrence; however, they have potential limitations due to species differences that prevent them from precisely replicating the intricate physiological and pathological processes of human disease. This Review provides a comprehensive overview of several available SUDEP animal models, highlighting their pros and cons. More importantly, we further propose the establishment of an ideal model based on brain-computer interfaces and artificial intelligence, hoping to offer new insights into potential advancements in SUDEP research. In doing so, we hope to provide valuable information for SUDEP researchers, offer new insights into the pathogenesis of SUDEP and open new avenues for the development of strategies to prevent SUDEP.
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BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is a globally increasing health epidemic. Lifestyle intervention is recommended as the main therapy for NAFLD. However, the optimal approach is still unclear. This study aimed to evaluate the effects of a comprehensive approach of intensive lifestyle intervention (ILI) concerning enhanced control of calorie-restricted diet (CRD), exercise, and personalized nutrition counseling on liver steatosis and extrahepatic metabolic status in Chinese overweight and obese patients with NAFLD. METHODS: This study was a multicenter randomized controlled trial (RCT) conducted across seven hospitals in China. It involved 226 participants with a body mass index (BMI) above 25. These participants were randomly assigned to two groups: the ILI group, which followed a low carbohydrate, high protein CRD combined with exercise and intensive counseling from a dietitian, and a control group, which adhered to a balanced CRD along with exercise and standard counseling. The main measure of the study was the change in the fat attenuation parameter (FAP) from the start of the study to week 12, analyzed within the per-protocol set. Secondary measures included changes in BMI, liver stiffness measurement (LSM), and the improvement of various metabolic indexes. Additionally, predetermined subgroup analyses of the FAP were conducted based on variables like gender, age, BMI, ethnicity, hyperlipidemia, and hypertension. RESULTS: A total of 167 participants completed the whole study. Compared to the control group, ILI participants achieved a significant reduction in FAP (LS mean difference, 16.07 [95% CI: 8.90-23.25] dB/m) and BMI (LS mean difference, 1.46 [95% CI: 1.09-1.82] kg/m2) but not in LSM improvement (LS mean difference, 0.20 [95% CI: -0.19-0.59] kPa). The ILI also substantially improved other secondary outcomes (including ALT, AST, GGT, body fat mass, muscle mass and skeletal muscle mass, triglyceride, fasting blood glucose, fasting insulin, HbA1c, HOMA-IR, HOMA-ß, blood pressure, and homocysteine). Further subgroup analyses showed that ILI, rather than control intervention, led to more significant FAP reduction, especially in patients with concurrent hypertension (p < 0.001). CONCLUSION: In this RCT, a 12-week intensive lifestyle intervention program led to significant improvements in liver steatosis and other metabolic indicators in overweight and obese Chinese patients suffering from nonalcoholic fatty liver disease. Further research is required to confirm the long-term advantages and practicality of this approach. TRIAL REGISTRATION: This clinical trial was registered on ClinicalTrials.gov (registration number: NCT03972631) in June 2019.
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Restrição Calórica , Estilo de Vida , Hepatopatia Gordurosa não Alcoólica , Obesidade , Sobrepeso , Humanos , Masculino , Feminino , Restrição Calórica/métodos , China , Hepatopatia Gordurosa não Alcoólica/dietoterapia , Hepatopatia Gordurosa não Alcoólica/terapia , Hepatopatia Gordurosa não Alcoólica/complicações , Pessoa de Meia-Idade , Obesidade/dietoterapia , Obesidade/terapia , Obesidade/complicações , Sobrepeso/terapia , Sobrepeso/complicações , Sobrepeso/dietoterapia , Adulto , Fígado/metabolismo , Índice de Massa Corporal , Exercício Físico/fisiologia , Aconselhamento/métodosRESUMO
Methylation-based liquid biopsies show promises in detecting cancer using circulating cell-free DNA; however, current limitations impede clinical application. Most assays necessitate substantial DNA inputs, posing challenges. Additionally, underrepresented tumor DNA fragments may go undetected during exponential amplification steps of traditional sequencing methods. Here, we report linear amplification-based bisulfite sequencing (LABS), enabling linear amplification of bisulfite-treated DNA fragments in a genome-wide, unbiased fashion, detecting cancer abnormalities with sub-nanogram inputs. Applying LABS to 100 patient samples revealed cancer-specific patterns, copy number alterations, and enhanced cancer detection accuracy by identifying tissue-of-origin and immune cell composition.
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Metilação de DNA , Neoplasias , Análise de Sequência de DNA , Sulfitos , Humanos , Neoplasias/genética , Análise de Sequência de DNA/métodos , Ácidos Nucleicos Livres , Técnicas de Amplificação de Ácido Nucleico/métodos , Variações do Número de Cópias de DNA , DNA de Neoplasias/genética , DNA Tumoral Circulante/genéticaRESUMO
The endoperoxide group of artemisinins is universally accepted an essential group for their anti-cancer effects. In this study, a series of D-ring-contracted artemisinin derivatives were constructed by combining ring-contracted artemisinin core with fragments of functional heterocyclic molecules or classical CDK4/6 inhibitors to identify more efficacious breast cancer treatment agents. Twenty-six novel hybridized molecules were synthesized and characterized by HRMS, IR, 1H-NMR and 13Câ NMR. In antiproliferative activities and kinase inhibitory effects assays, we found that the antiproliferative effects of B01 were close to those of the positive control Palbociclib, with GI50 values of 4.87±0.23â µM and 9.97±1.44â µM towards T47D cells and MDA-MB-436 cells respectively. In addition, the results showed that B01 was the most potent compound against CDK6/cyclin D3 kinase, with an IC50 value of 0.135±0.041â µM, and its activity was approximately 1/3 of the positive control Palbociclib.
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Antineoplásicos , Artemisininas , Neoplasias da Mama , Proliferação de Células , Quinase 4 Dependente de Ciclina , Quinase 6 Dependente de Ciclina , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores de Proteínas Quinases , Humanos , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/metabolismo , Artemisininas/farmacologia , Artemisininas/química , Artemisininas/síntese química , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 4 Dependente de Ciclina/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proliferação de Células/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Relação Estrutura-Atividade , Linhagem Celular Tumoral , Estrutura Molecular , Feminino , Relação Dose-Resposta a Droga , Simulação de Acoplamento MolecularRESUMO
Widespread alterations in RNA alternative splicing (AS) have been identified in adult gliomas. However, their regulatory mechanism, biological significance, and therapeutic potential remain largely elusive. Here, using a computational approach with both bulk and single-cell RNA-Seq, we uncover a prognostic AS signature linked with neural developmental hierarchies. Using advanced iPSC glioma models driven by glioma driver mutations, we show that this AS signature could be enhanced by EGFRvIII and inhibited by in situ IDH1 mutation. Functional validations of 2 isoform switching events in CERS5 and MPZL1 show regulations of sphingolipid metabolism and SHP2 signaling, respectively. Analysis of upstream RNA binding proteins reveals PTBP1 as a key regulator of the AS signature where targeting of PTBP1 suppresses tumor growth and promotes the expression of a neuron marker TUJ1 in glioma stem-like cells. Overall, our data highlights the role of AS in affecting glioma malignancy and heterogeneity and its potential as a therapeutic vulnerability for treating adult gliomas.
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Processamento Alternativo , Glioma , Proteína de Ligação a Regiões Ricas em Polipirimidinas , Glioma/genética , Glioma/patologia , Glioma/metabolismo , Glioma/terapia , Humanos , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Proteína de Ligação a Regiões Ricas em Polipirimidinas/metabolismo , Animais , Camundongos , Ribonucleoproteínas Nucleares Heterogêneas/genética , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/terapia , Adulto , Células-Tronco Pluripotentes Induzidas/metabolismo , Linhagem Celular Tumoral , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismoRESUMO
IL-17C is an epithelial cell-derived proinflammatory cytokine whose transcriptional regulation remains unclear. Analysis of the IL17C promoter region identified TCF4 as putative regulator, and siRNA knockdown of TCF4 in human keratinocytes (KCs) increased IL17C. IL-17C stimulation of KCs (along with IL-17A and TNF-α stimulation) decreased TCF4 and increased NFKBIZ and ZC3H12A expression in an IL-17RA/RE-dependent manner, thus creating a feedback loop. ZC3H12A (MCPIP1/Regnase-1), a transcriptional immune-response regulator, also increased following TCF4 siRNA knockdown, and siRNA knockdown of ZC3H12A decreased NFKBIZ, IL1B, IL36G, CCL20, and CXCL1, revealing a proinflammatory role for ZC3H12A. Examination of lesional skin from the KC-Tie2 inflammatory dermatitis mouse model identified decreases in TCF4 protein concomitant with increases in IL-17C and Zc3h12a that reversed following the genetic elimination of Il17c, Il17ra, and Il17re and improvement in the skin phenotype. Conversely, interference with Tcf4 in KC-Tie2 mouse skin increased Il17c and exacerbated the inflammatory skin phenotype. Together, these findings identify a role for TCF4 in the negative regulation of IL-17C, which, alone and with TNF-α and IL-17A, feed back to decrease TCF4 in an IL-17RA/RE-dependent manner. This loop is further amplified by IL-17C-TCF4 autocrine regulation of ZC3H12A and IL-17C regulation of NFKBIZ to promote self-sustaining skin inflammation.
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Proteínas Adaptadoras de Transdução de Sinal , Interleucina-17 , Queratinócitos , Receptores de Interleucina-17 , Ribonucleases , Transdução de Sinais , Fator de Transcrição 4 , Animais , Fator de Transcrição 4/metabolismo , Fator de Transcrição 4/genética , Humanos , Interleucina-17/metabolismo , Interleucina-17/genética , Camundongos , Queratinócitos/metabolismo , Ribonucleases/metabolismo , Ribonucleases/genética , Receptores de Interleucina-17/metabolismo , Receptores de Interleucina-17/genética , Inflamação/metabolismo , Inflamação/genética , Modelos Animais de Doenças , Epiderme/metabolismo , Dermatite/metabolismo , Dermatite/genética , Dermatite/imunologia , Dermatite/patologia , Retroalimentação Fisiológica , Regulação da Expressão GênicaRESUMO
OBJECTIVE: To evaluate the bacteria debridement efficacy of two generations of sonic root canal irrigant activation systems: EndoActivator (Dentsply Sirona), the first generation, and SmartLite Pro EndoActivator, the second generation. METHODS: Instrumented, autoclaved, single-rooted human premolars were inoculated with Enterococcus faecalis (ATCC-29212) for 21 days. The bacteria biofilm-containing teeth were randomly divided into 5 groups (N=8): Group 1: Syringe-side-vented needle (S-N) delivery of saline for 1 min; Group 2: S-N delivery of 2% NaOCl for 1 min; Group 3: S-N delivery of 2% NaOCl for 5 min; Group 4: EndoActivator activation of 2% NaOCl for 1 min; Group 5: SmartLite Pro EndoActivator activation of 2% NaOCl for 1 min. The teeth were evaluated for bacterial reduction using CFU counts, and the percentages of dead bacteria within the dentinal tubules using confocal laser scanning microscopy. RESULTS: Activation of NaOCl with EndoActivator or SmartLite Pro EndoActivator significantly reduced the overall intracanal bacterial load, compared with S-N irrigant delivery (P<0.05), with no significant difference between the two agitation devices (P>0.05). Nevertheless, S-N delivery of 2% NaOCl for 5 min produced better bacteria debridement than either sonic agitation system. Different degrees of bacteria kill were identified in the coronal-middle portions and apical portion of the canal space. CONCLUSION: Delivery time of NaOCl affects the efficacy of bacteria disinfection. Activation for 1 min with the EndoActivator or SmartLite Pro EndoActivator demonstrated comparable canal wall biofilm and intracanal bacteria reduction efficacy when 2% NaOCl was used as irrigant for disinfecting E. faecalis in single-rooted teeth. CLINICAL SIGNIFICANCE: Although the sonic root canal irrigant activation devices investigated do not completely eliminate live bacteria biofilms from the canal space, they help reduce bacteria load during irrigant activation.
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Cavidade Pulpar , Irrigantes do Canal Radicular , Humanos , Cavidade Pulpar/microbiologia , Irrigantes do Canal Radicular/farmacologia , Irrigantes do Canal Radicular/uso terapêutico , Hipoclorito de Sódio/farmacologia , Hipoclorito de Sódio/uso terapêutico , Desbridamento , Enterococcus faecalis , Preparo de Canal RadicularRESUMO
With the acceleration of the pace of society, the problem of scarcity of time resources is becoming more prominent, which may lead to short-sighted behavior. This study aimed to examine the impact of time scarcity on intertemporal choice and its underlying mechanisms through three experiments. Study 1 aimed to investigate the causal relationship between time scarcity and intertemporal choice and the mediating role of anxiety between time scarcity and intertemporal choice. The results showed that participants with a stronger perception of time scarcity were more likely to choose smaller and sooner gains, and that the mediating role of anxiety was significant. In Study 2, the control of time was introduced to explore whether the sense of control could alleviate the negative emotion caused by time scarcity. The results showed that individuals with high levels of control had significantly less anxiety under higher time-scarcity conditions than those with low levels of control. In Study 3, we manipulated time scarcity and found that the sense of control moderated the mediating effect of anxiety in time scarcity and intertemporal choice. Overall, these findings suggest that the sense of control over time is associated with less anxiety from time scarcity, which in turn fosters preferences for delayed gains.
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Ansiedade , Humanos , Feminino , Masculino , Ansiedade/psicologia , Adulto , Tomada de Decisões , Desvalorização pelo Atraso , Comportamento de Escolha , Percepção do Tempo , Adulto Jovem , Fatores de TempoRESUMO
BACKGROUND: Grade 4 glioma is the most aggressive and currently incurable brain tumor with a median survival of one year in adult patients. Elucidating novel transcriptomic and epigenetic contributors to the molecular heterogeneity underlying its aggressiveness may lead to improved clinical outcomes. METHODS: To identify grade 4 glioma -associated 5-hydroxymethylcytosine (5hmC) and transcriptomic features as well as their cross-talks, genome-wide 5hmC and transcriptomic profiles of tissue samples from 61 patients with grade 4 gliomas and 9 normal controls were obtained for differential and co-regulation/co-modification analyses. Prognostic models on overall survival based on transcriptomic features and the 5hmC modifications summarized over genic regions (promoters, gene bodies) and brain-derived histone marks were developed using machine learning algorithms. RESULTS: Despite global reduction, the majority of differential 5hmC features showed higher modification levels in grade 4 gliomas as compared to normal controls. In addition, the bi-directional correlations between 5hmC modifications over promoter regions or gene bodies and gene expression were greatly disturbed in grade 4 gliomas regardless of IDH1 mutation status. Phenotype-associated co-regulated 5hmC-5hmC modules and 5hmC-mRNA modules not only are enriched with different molecular pathways that are indicative of the pathogenesis of grade 4 gliomas, but also are of prognostic significance comparable to IDH1 mutation status. Lastly, the best-performing 5hmC model can predict patient survival at a much higher accuracy (c-index = 74%) when compared to conventional prognostic factor IDH1 (c-index = 57%), capturing the molecular characteristics of tumors that are independent of IDH1 mutation status and gene expression-based molecular subtypes. CONCLUSIONS: The 5hmC-based prognostic model could offer a robust tool to predict survival in patients with grade 4 gliomas, potentially outperforming existing prognostic factors such as IDH1 mutations. The crosstalk between 5hmC and gene expression revealed another layer of complexity underlying the molecular heterogeneity in grade 4 gliomas, offering opportunities for identifying novel therapeutic targets.
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Neoplasias Encefálicas , Glioma , Humanos , Transcriptoma , Glioma/patologia , Prognóstico , Neoplasias Encefálicas/patologia , Mutação , Epigênese Genética , Isocitrato Desidrogenase/genéticaRESUMO
BACKGROUND: Basal zone hyperplasia (BZH) and dilated intercellular spaces (DISs) are thought to contribute to the clinical manifestations of eosinophilic esophagitis (EoE); however, the molecular pathways that drive BZH remain largely unexplored. OBJECTIVE: We sought to define the role of IL-13-induced transcriptional programs in esophageal epithelial proliferation in EoE. METHODS: We performed RNA sequencing, bioinformatics, Western blot, reverse transcriptase quantitative PCR, and histologic analyses on esophageal biopsies from healthy control and patients with EoE, primary esophageal cells derived from patients with EoE, and IL-13-stimulated esophageal epithelial keratinocytes grown at the air-liquid interface (EPC2-ALI). Genetic (shRNA) and pharmacologic (proteolysis-targeting chimera degrader) approaches and in vivo model of IL-13-induced esophageal epithelial remodeling (Krt5-rtTA x tetO-IL-13Tg) were used to define the role of signal transducer and activator of transcription 3 (STAT3) and STAT6 and secreted frizzled-related protein 1 (SFRP1) in esophageal epithelial proliferation. RESULTS: RNA-sequencing analysis of esophageal biopsies (healthy control vs EoE) and EPC2-ALI revealed 82 common differentially expressed genes that were enriched for putative STAT3 target genes. In vitro and in vivo analyses revealed a link between IL-13-induced STAT3 and STAT6 phosphorylation, SFRP1 mRNA expression, and esophageal epithelial proliferation. In vitro studies showed that IL-13-induced esophageal epithelial proliferation was STAT3-dependent and regulated by the STAT3 target SFRP1. SFRP1 mRNA is increased in esophageal biopsies from patients with active EoE compared with healthy controls or patients in remission and identifies an esophageal suprabasal epithelial cell subpopulation that uniquely expressed the core EoE proinflammatory transcriptome genes (CCL26, ALOX15, CAPN14, ANO1, and TNFAIP6). CONCLUSIONS: These studies identify SFRP1 as a key regulator of IL-13-induced and STAT3-dependent esophageal proliferation and BZH in EoE and link SFRP1+ esophageal epithelial cells with the proinflammatory and epithelial remodeling response in EoE.
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Esofagite Eosinofílica , Humanos , Esofagite Eosinofílica/patologia , Interleucina-13/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Células Epiteliais/metabolismo , RNA Mensageiro/metabolismo , Proliferação de CélulasRESUMO
Objective: To investigate the relationship between single nucleotide polymorphisms (SNPs) related to vitamin D (VitD) metabolism and post-stroke depression (PSD) in patients with ischemic stroke. Methods: A total of 210 patients with ischemic stroke were enrolled at the Department of Neurology in Xiangya Hospital, Central South University, from July 2019 to August 2021. SNPs in the VitD metabolic pathway (VDR, CYP2R1, CYP24A1, and CYP27B1) were genotyped using the SNPscan™ multiplex SNP typing kit. Demographic and clinical data were collected using a standardized questionnaire. Multiple genetic models including dominant, recessive, and over-dominant models were utilized to analyze the associations between SNPs and PSD. Results: In the dominant, recessive, and over-dominant models, no significant association was observed between the selected SNPs in the CYP24A1 and CYP2R1 genes and PSD. However, univariate and multivariate logistic regression analysis revealed that the CYP27B1 rs10877012 G/G genotype was associated with a decreased risk of PSD (OR: 0.41, 95% CI: 0.18-0.92, p = 0.030 and OR: 0.42, 95% CI: 0.18-0.98, p = 0.040, respectively). Furthermore, haplotype association analysis indicated that rs11568820-rs1544410-rs2228570-rs7975232-rs731236 CCGAA haplotype in the VDR gene was associated with a reduced risk of PSD (OR: 0.14, 95% CI: 0.03-0.65, p = 0.010), whereas no significant association was observed between haplotypes in the CYP2R1 and CYP24A1 genes and PSD. Conclusion: Our findings suggest that the polymorphisms of VitD metabolic pathway genes VDR and CYP27B1 may be associated with PSD in patients with ischemic stroke.
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Epigenetic modifications play critical roles in gene regulation and disease pathobiology. Highly sensitive enabling technologies, including microarray- and sequencing-based approaches have allowed genome-wide profiling of cytosine modifications in DNAs in clinical samples to facilitate discovery of epigenetic biomarkers for disease diagnosis and prognosis. Historically, many previous studies, however, did not distinguish the most investigated 5-methylcytosines (5mC) from other modified cytosines, especially the biochemically stable 5-hydroxymethylcytosines (5hmC), which have been shown to have a distinct genomic distribution and regulatory role from 5mC. Notably, during the past several years, the 5hmC-Seal, a highly sensitive chemical labeling technique, has been demonstrated to be a powerful tool for genome-wide profiling of 5hmC in clinically feasible biospecimens (e.g. a few milliliter of plasma or serum). The 5hmC-Seal technique has been utilized by our team in biomarker discovery for human cancers and other complex diseases using circulating cell-free DNA (cfDNA), as well as the characterization of the first 5hmC Human Tissue Map. Convenient access to the accumulating 5hmC-Seal data will allow the research community to validate and re-use these results, potentially providing novel insights into epigenetic contribution to a range of human diseases. Here we introduce the PETCH-DB, an integrated database that was implemented to provide 5hmC-related results generated using the 5hmC-Seal technique. We aim the PETCH-DB to be a central portal, which will be available to the scientific community with regularly updated 5hmC data in clinical samples to reflect current advances in this field. Database URL http://petch-db.org/.
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5-Metilcitosina , Pesquisa Biomédica , Humanos , Citosina , Bases de Dados FactuaisRESUMO
Epigenetic modifications play critical roles in gene regulation and disease pathobiology. Highly sensitive enabling technologies, including microarray- and sequencing-based approaches have allowed genome-wide profiling of cytosine modifications in DNAs in clinical samples to facilitate discovery of epigenetic biomarkers for disease diagnosis and prognosis. Historically, many previous studies, however, did not distinguish the most investigated 5-methylcytosines (5mC) from other modified cytosines, especially the biochemically stable 5-hydroxymethylcytosines (5hmC), which have been shown to have a distinct genomic distribution and regulatory role from 5mC. Notably, during the past several years, the 5hmC-Seal, a highly sensitive chemical labeling technique, has been demonstrated to be a powerful tool for genome-wide profiling of 5hmC in clinically feasible biospecimens (e.g. a few milliliter of plasma or serum). The 5hmC-Seal technique has been utilized by our team in biomarker discovery for human cancers and other complex diseases using circulating cell-free DNA (cfDNA), as well as the characterization of the first 5hmC Human Tissue Map. Convenient access to the accumulating 5hmC-Seal data will allow the research community to validate and re-use these results, potentially providing novel insights into epigenetic contribution to a range of human diseases. Here we introduce the PETCH-DB, an integrated database that was implemented to provide 5hmC-related results generated using the 5hmC-Seal technique. We aim the PETCH-DB to be a central portal, which will be available to the scientific community with regularly updated 5hmC data in clinical samples to reflect current advances in this field. Database URL http://petch-db.org/.
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5-Metilcitosina , Pesquisa Biomédica , Humanos , Citosina , Bases de Dados FactuaisRESUMO
PURPOSE: Establishing a cross-species animal model of human immunodeficiency virus (HIV) infection is crucial for the study of HIV/acquired immunodeficiency syndrome (AIDS). However, due to the species-specific characteristics of HIV-1, the virus can only infect directly humans and a small number of non-human primates. It cannot directly infect mouse cells across species. METHODS: A mouse leukemia cell line with high CD4 (clusters of differentiation 4)/CCR5 (CC-chemokine receptor 5)/CyclinT1 expression was constructed in this study. First, CD4/CCR5/CyclinT1 lentiviral vector was used to infect a murine leukemia cell line L1210 to express the receptor CD4, co-receptor CCR5 and tat protein driving factor CyclinT1, which are required to infect L1210 cells with HIV-1. RESULTS: The results of sequencing identification and fluorescence expression showed that the plasmid expressing CD4, CCR5, and CyclinT1 vector was successfully constructed and wrapped as the lentiviral vector. Moreover, it was observed that CD4, CCR5, and CyclinT1 proteins were highly expressed in mouse leukemia cells L1210 compared to empty lentiviral vector-transfected cells. Next, viral entry and replication were demonstrated when HIV-1 RNA was detected in body cells and cultured supernatants. Transgenic mice cells L1210 showed significantly greater content of HIV-1 RNA compared to control L1210 cells. Finally, CEMx174 was infected with cell culture supernatants to clarify that the progeny virus is an active virus with infection ability. HIV-1 RNA was highly expressed in CEMx174 cells. CONCLUSIONS: This study made the foundation for future studies evaluating HIV-1 cross-species infection in a murine animal model. The results provided new direction for studies investigating the development of vaccines, antiviral drugs screening, and HIV/AIDS pathogenesis.
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Síndrome da Imunodeficiência Adquirida , Infecções por HIV , HIV-1 , Animais , Humanos , Camundongos , Linhagem Celular , Infecções por HIV/genética , HIV-1/genética , HIV-1/metabolismo , Camundongos Transgênicos , Receptores CCR5/genética , Receptores CCR5/metabolismoRESUMO
Background: The immune-inflammatory response has been widely considered to be involved in the pathogenesis of post-stroke depression (PSD), but there is ambiguity about the mechanism underlying such association. Methods: According to Diagnostic and Statistical Manual of Mental Disorders (5th edition), depressive symptoms were assessed at 2 weeks after stroke onset. 15 single nucleotide polymorphisms (SNPs) in genes of indoleamine 2,3-dioxygenase (IDO, including IDO1 and IDO2) and its inducers (including pro-inflammatory cytokines interferon [IFN]-γ, tumor necrosis factor [TNF]-α, interleukin [IL]-1ß, IL-2 and IL-6) were genotyped using SNPscan™ technology, and serum IDO1 levels were detected by double-antibody sandwich enzyme-linked immune-sorbent assay. Results: Fifty-nine patients (31.72%) were diagnosed with depression at 2 weeks after stroke onset (early-onset PSD). The IDO1 rs9657182 T/T genotype was independently associated with early-onset PSD (adjusted odds ratio [OR] = 3.008, 95% confidence interval [CI] 1.157-7.822, p = 0.024) and the frequency of rs9657182 T allele was significantly higher in patients with PSD than that in patients with non-PSD (χ2 = 4.355, p = 0.037), but these results did not reach the Bonferroni significance threshold (p > 0.003). Serum IDO1 levels were also independently linked to early-onset PSD (adjusted OR = 1.071, 95% CI 1.002-1.145, p = 0.044) and patients with PSD had higher serum IDO1 levels than patients with non-PSD in the presence of the rs9657182 T allele but not homozygous C allele (t = -2.046, p = 0.043). Stroke patients with the TNF-α rs361525 G/G genotype had higher serum IDO1 levels compared to those with the G/A genotype (Z = -2.451, p = 0.014). Conclusions: Our findings provided evidence that IDO1 gene polymorphisms and protein levels were involved in the development of early-onset PSD and TNF-α polymorphism was associated with IDO1 levels, supporting that IDO1 which underlie strongly regulation by cytokines may be a specific pathway for the involvement of immune-inflammatory mechanism in the pathophysiology of PSD.
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Indolamina-Pirrol 2,3,-Dioxigenase , Acidente Vascular Cerebral , Humanos , Citocinas/genética , Depressão/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Polimorfismo de Nucleotídeo Único , Acidente Vascular Cerebral/genética , Fator de Necrose Tumoral alfa/genéticaRESUMO
CRISPR/Cas9 has been proposed as a treatment for genetically inherited skin disorders. Here we report that CRISPR transfection activates STING-dependent antiviral responses in keratinocytes, resulting in heightened endogenous interferon (IFN) responses through induction of IFN-κ, leading to decreased plasmid stability secondary to induction of the cytidine deaminase gene APOBEC3G. Notably, CRISPR-generated KO keratinocytes had permanent suppression of IFN-κ and IFN-stimulated gene (ISG) expression, secondary to hypermethylation of the IFNK promoter region by the DNA methyltransferase DNMT3B. JAK inhibition via baricitinib prior to CRISPR transfection increased transfection efficiency, prevented IFNK promoter hypermethylation, and restored normal IFN-κ activity and ISG responses. This work shows that CRISPR-mediated gene correction alters antiviral responses in keratinocytes, has implications for future gene therapies for inherited skin diseases using CRISPR technology, and suggests pharmacologic JAK inhibition as a tool for facilitating and attenuating inadvertent selection effects in CRISPR/Cas9 therapeutic approaches.
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Interferon Tipo I , Antivirais , DNA/metabolismo , Interferon Tipo I/metabolismo , Queratinócitos/metabolismo , HumanosRESUMO
BACKGROUND: Long noncoding RNAs (lncRNAs) regulate the etiology of complex diseases and cancers, including glioblastoma (GBM). However, lncRNA-based therapies are limited because the mechanisms of action of many lncRNAs with their binding partners are not completely understood. METHODS: We used transcriptomic and genomic data to analyze correlations between LINC02283 and PDGFRA (platelet-derived growth factor receptor A). The biological functions of the novel lncRNA were assessed in vivo using patient-derived glioma stem-like cells (GSCs), and orthotopic GBM xenografts. Immunoblotting, qRT-PCR, RNA pull down, crosslinked RNA immunoprecipitation, fluorescence in situ hybridization, and antisense oligo-mediated knockdown were performed to explore the regulation of LINC02283 on PDGFRA signaling. Expression of LINC02283 in clinical samples was assessed using pathologically diagnosed GBM patient samples. RESULTS: We identified a novel oncogenic lncRNA, LINC02283, that is highly expressed in the PDGFRA mutation-driven cohort of glioma patients and associated with worse prognosis. LINC02283 gene co-amplifies with the PDGFRA locus and shows high correlation with PDGFRA expression. Deprivation of LINC02283 in GSCs with PDGFRA amplification mutation, attenuated tumorigenicity and enhanced survival in orthotopic GBM xenograft models, while overexpression of LINC02283 in GSCs with wild-type PDGFRA, enhances PDGFRA signaling, and decreases survival. Further, LINC02283 interacts with PDGFRA to enhance its signaling and that of its downstream targets AKT and ERK, thus promoting oncogenesis in GBM. CONCLUSIONS: Our results provide strong evidence of LINC02283 as a regulator of PDGFRA oncogenic activity and GBM malignancy and support the potential of lncRNAs as possible therapeutic targets.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Glioma , RNA Longo não Codificante , Humanos , Glioblastoma/patologia , RNA Longo não Codificante/genética , Hibridização in Situ Fluorescente , Glioma/genética , Transformação Celular Neoplásica/genética , Receptores do Fator de Crescimento Derivado de Plaquetas/genética , Receptores do Fator de Crescimento Derivado de Plaquetas/metabolismo , Regulação Neoplásica da Expressão Gênica , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias Encefálicas/patologiaRESUMO
Background: Homocysteine (Hcy) has been indicated to be involved in pathophysiology of post stroke depression (PSD). There is a lack of research to study the relationship between Hcy metabolism genes and PSD. Our study aims to investigate the relationship among Hcy metabolism genes, Hcy, and early-onset PSD. Materials and methods: We recruited 212 patients with stroke and collected their peripheral blood sample, clinical data, and laboratory test on admission. 12 single nucleotide polymorphisms (SNPs) in methylenetetrahydrofolate reductase (MTHFR), methionine synthase reductase (MTRR), and methionine synthase (MTR) genes were genotyped by high-resolution melt analysis. PSD was diagnosed by DSM-V at 2 weeks after stroke. Binary logistic regression and haplotype analysis were used to examine the association between Hcy metabolism genes and PSD. Mediation analysis was performed to clarify whether the SNPs exerted their effect on PSD by affecting the Hcy level. Results: 81 patients were diagnosed with PSD, and the incidence rate was 38.2%. Hcy level in PSD group was significantly higher than it in non-PSD group (p = 0.019). MTHFR rs1801133 AA genotype an A allele were associated with an elevated risk of PSD after adjustment for some confounding factors (OR = 4.021, 95% CI: 1.459â¼11.080, p = 0.007 for AA genotype; OR = 1.808, 95% CI: 1.172â¼2.788, p = 0.007 for A allele). Furthermore, the effect of MTHFR rs1801133 AA genotype on PSD was mediated by Hcy (OR = 1.569, 95% CI: 0.013â¼3.350, p < 0.05). Conclusion: MTHFR rs1801133 and Hcy were associated with PSD, and MTHFR rs1801133 may exert an effect on PSD via mediating Hcy level. This offers a new perspective for treating PSD and understanding the mechanism of PSD.
RESUMO
The Illumina EPIC array is widely used for high-throughput profiling of DNA cytosine modifications in human samples, covering more than 850,000 modification sites across various genomic features. The application of this platform is expected to provide novel insights into the epigenetic contribution to human complex traits and diseases. Considering the diverse inter-population genetic and epigenetic variation, it will benefit the research community with a comprehensive characterization of this platform for its applicability to major global populations. Specifically, we mapped 866,836 CpG probes from the EPIC array to the human genome reference. We detected 91,034 CpG probes that did not align reliably to the human genome reference. In addition, 21,256 CpG probes were found to ambiguously map to multiple loci in the human genome, and 448 probes showing inaccurate genomic information from the original Illumina annotations. We further characterized those uniquely mapped CpG probes in terms of whether they contained common genetic variants, i.e., single nucleotide polymorphisms (SNPs), in major global populations, by utilizing the 1000 Genomes Project data. A list of optimal CpG probes on the EPIC array was generated for major global populations, with the aim of providing a resource to facilitate future studies of diverse human populations. In conclusion, our analysis indicated that studies of diverse human populations using the EPIC array would be benefited by taking into account of the technical features of this platform.
RESUMO
Sudden unexpected death in epilepsy (SUDEP) is the leading cause of death among patients with epilepsy. However, the underlying mechanism of SUDEP remains elusive. Previous studies showed seizure-induced respiratory arrest (S-IRA) is the main factor in SUDEP, and that enhancement of serotonin (5-HT) function in the dorsal raphe nucleus (DR) can significantly reduce the incidence of S-IRA in the DBA/1 mouse model of SUDEP. The pre-Bötzinger complex (PBC), known for its role in regulating respiratory rhythm, can express the 5-HT2A receptor (5-HT2AR). Here, using the pharmacological and optogenetic methods, respectively, we observed that the serotonergic neural circuit between DR and PBC was involved in S-IRA evoked by either acoustic stimulation or pentylenetetrazole (PTZ) injection in the DBA/1 mice, and found 5-HT2AR located in PBC plays an important role in it. Our findings will further significantly improve our understanding of SUDEP and provide a promising therapeutic target for SUDEP prevention.