RESUMO
BACKGROUND: Cell-free DNA (cfDNA) is a noninvasive marker of cellular injury. Its significance in pulmonary arterial hypertension (PAH) is unknown. METHODS: Plasma cfDNA was measured in 2 PAH cohorts (A, n=48; B, n=161) and controls (n=48). Data were collected for REVEAL 2.0 (Registry to Evaluate Early and Long-Term PAH Disease Management) scores and outcome determinations. Patients were divided into the following REVEAL risk groups: low (≤6), medium (7-8), and high (≥9). Total cfDNA concentrations were compared among controls and PAH risk groups by 1-way analysis of variance. Log-rank tests compared survival between cfDNA tertiles and REVEAL risk groups. Areas under the receiver operating characteristic curve were estimated from logistic regression models. A sample subset from cohort B (n=96) and controls (n=16) underwent bisulfite sequencing followed by a deconvolution algorithm to map cell-specific cfDNA methylation patterns, with concentrations compared using t tests. RESULTS: In cohort A, median (interquartile range) age was 62 years (47-71), with 75% female, and median (interquartile range) REVEAL 2.0 was 6 (4-9). In cohort B, median (interquartile range) age was 59 years (49-71), with 69% female, and median (interquartile range) REVEAL 2.0 was 7 (6-9). In both cohorts, cfDNA concentrations differed among patients with PAH of varying REVEAL risk and controls (analysis of variance P≤0.002) and were greater in the high-risk compared with the low-risk category (P≤0.002). In cohort B, death or lung transplant occurred in 14 of 54, 23 of 53, and 35 of 54 patients in the lowest, middle, and highest cfDNA tertiles, respectively. cfDNA levels stratified as tertiles (log-rank: P=0.0001) and REVEAL risk groups (log-rank: P<0.0001) each predicted transplant-free survival. The addition of cfDNA to REVEAL improved discrimination (area under the receiver operating characteristic curve, 0.72-0.78; P=0.02). Compared with controls, methylation analysis in patients with PAH revealed increased cfDNA originating from erythrocyte progenitors, neutrophils, monocytes, adipocytes, natural killer cells, vascular endothelium, and cardiac myocytes (Bonferroni adjusted P<0.05). cfDNA concentrations derived from erythrocyte progenitor cells, cardiac myocytes, and vascular endothelium were greater in patients with PAH with high-risk versus low-risk REVEAL scores (P≤0.02). CONCLUSIONS: Circulating cfDNA is elevated in patients with PAH, correlates with disease severity, and predicts worse survival. Results from cfDNA methylation analyses in patients with PAH are consistent with prevailing paradigms of disease pathogenesis.
Assuntos
Ácidos Nucleicos Livres , Hipertensão Arterial Pulmonar , Idoso , Biomarcadores , Ácidos Nucleicos Livres/genética , Hipertensão Pulmonar Primária Familiar , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Hipertensão Arterial Pulmonar/diagnóstico , Hipertensão Arterial Pulmonar/genética , Curva ROCRESUMO
The pathophysiology of sickle cell disease (SCD) is driven by chronic inflammation fueled by damage associated molecular patterns (DAMPs). We show that elevated cell-free DNA (cfDNA) in patients with SCD is not just a prognostic biomarker, it also contributes to the pathological inflammation. Within the elevated cfDNA, patients with SCD had a significantly higher ratio of cell-free mitochondrial DNA (cf-mtDNA)/cell-free nuclear DNA compared with healthy controls. Additionally, mitochondrial DNA in patient samples showed significantly disproportionately increased hypomethylation compared with healthy controls, and it was increased further in crises compared with steady-state. Using flow cytometry, structured illumination microscopy, and electron microscopy, we showed that circulating SCD red blood cells abnormally retained their mitochondria and, thus, are likely to be the source of the elevated cf-mtDNA in patients with SCD. Patient plasma containing high levels of cf-mtDNA triggered the formation of neutrophil extracellular traps (NETs) that was substantially reduced by inhibition of TANK-binding kinase 1, implicating activation of the cGAS-STING pathway. cf-mtDNA is an erythrocytic DAMP, highlighting an underappreciated role for mitochondria in sickle pathology. These trials were registered at www.clinicaltrials.gov as #NCT00081523, #NCT03049475, and #NCT00047996.
Assuntos
Anemia Falciforme/sangue , Ácidos Nucleicos Livres/sangue , Metilação de DNA , DNA Mitocondrial/sangue , Adulto , Idoso , Biomarcadores/sangue , Armadilhas Extracelulares/metabolismo , Feminino , Humanos , Inflamação/sangue , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Nucleotidiltransferases/metabolismo , Transdução de SinaisRESUMO
The metabolite acetyl-coenzyme A (acetyl-CoA) serves as an essential element for a wide range of cellular functions including adenosine triphosphate (ATP) production, lipid synthesis, and protein acetylation. Intracellular acetyl-CoA concentrations are associated with nutrient availability, but the mechanisms by which a cell responds to fluctuations in acetyl-CoA levels remain elusive. Here, we generate a cell system to selectively manipulate the nucleo-cytoplasmic levels of acetyl-CoA using clustered regularly interspaced short palindromic repeat (CRISPR)-mediated gene editing and acetate supplementation of the culture media. Using this system and quantitative omics analyses, we demonstrate that acetyl-CoA depletion alters the integrity of the nucleolus, impairing ribosomal RNA synthesis and evoking the ribosomal protein-dependent activation of p53. This nucleolar remodeling appears to be mediated through the class IIa histone deacetylases (HDACs). Our findings highlight acetylation-mediated control of the nucleolus as an important hub linking acetyl-CoA fluctuations to cellular stress responses.
Assuntos
Acetilcoenzima A/biossíntese , Nucléolo Celular/metabolismo , ATP Citrato (pro-S)-Liase/deficiência , ATP Citrato (pro-S)-Liase/genética , ATP Citrato (pro-S)-Liase/metabolismo , Acetatos/metabolismo , Acetilação , Linhagem Celular , Nucléolo Celular/ultraestrutura , Expressão Gênica , Técnicas de Inativação de Genes , Células HCT116 , Histona Desacetilases/metabolismo , Humanos , Modelos Biológicos , Proteínas Nucleares/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas Ribossômicas/metabolismo , Proteína Supressora de Tumor p53/metabolismoRESUMO
Allostatic load (AL) refers to the cumulative "wear and tear" on an organism throughout its lifetime. One of the primary contributing factors to AL is prolonged exposure to stress or its primary catabolic agent cortisol. Chronic exposure to stress or cortisol is associated with numerous diseases, including cardiovascular disease, metabolic disorders, and psychiatric disorders. Therefore, a molecular marker capable of integrating a past history of cortisol exposure would be of great utility for assessing disease risk. To this end, we recruited 87 healthy males and females of European ancestry between 18 and 60 years old, extracted genomic DNA and RNA from leukocytes, and implemented a gene-centric DNA enrichment method coupled with bisulfite sequencing and RNA-Seq of total RNA for the determination of genome-wide methylation and gene transcription, respectively. Sequencing data were analyzed against awakening and bedtime cortisol data to identify differentially methylated regions (DMRs) and CpGs (DMCs) and differentially expressed genes (DEGs). Six candidate DMCs (punadjusted < 0.005) and nine DEGs (punadjusted < 0.0005) were used to construct a prediction model that could capture past 30+ days of both bedtime and awakening cortisol levels. Utilizing a cross-validation approach, we obtained a regression coefficient of R2 = 0.308 for predicting continuous awakening cortisol and an area under the curve (AUC) = 0.753 for dichotomous (high vs. low tertile) awakening cortisol, and R2 = 0.224 and AUC = 0.723 for continuous and dichotomous bedtime cortisol levels, respectively. To our knowledge, the current study represents the first attempt to identify genome-wide predictors of cortisol exposure that utilizes both methylation and transcription targets. The utility of our approach needs to be replicated in an independent cohort of samples for which similar cortisol metrics are available.
Assuntos
Alostase , Hidrocortisona , Adolescente , Adulto , Metilação de DNA , Feminino , Humanos , Hidrocortisona/metabolismo , Masculino , Pessoa de Meia-Idade , Saliva/metabolismo , Estresse Psicológico/metabolismo , Transcriptoma , Adulto JovemRESUMO
The primary function of APOE (apolipoprotein E) is to mediate the transport of cholesterol- and lipid-containing lipoprotein particles into cells by receptor-mediated endocytosis. APOE also has pro- and antiinflammatory effects, which are both context and concentration dependent. For example, Apoe-/- mice exhibit enhanced airway remodeling and hyperreactivity in experimental asthma, whereas increased APOE levels in lung epithelial lining fluid induce IL-1ß secretion from human asthmatic alveolar macrophages. However, APOE-mediated airway epithelial cell inflammatory responses and signaling pathways have not been defined. Here, RNA sequencing of human asthmatic bronchial brushing cells stimulated with APOE identified increased expression of mRNA transcripts encoding multiple proinflammatory genes, including CXCL5 (C-X-C motif chemokine ligand 5), an epithelial-derived chemokine that promotes neutrophil activation and chemotaxis. We subsequently characterized the APOE signaling pathway that induces CXCL5 secretion by human asthmatic small airway epithelial cells (SAECs). Neutralizing antibodies directed against TLR4 (Toll-like receptor 4), but not TLR2, attenuated APOE-mediated CXCL5 secretion by human asthmatic SAECs. Inhibition of TAK1 (transforming growth factor-ß-activated kinase 1), IκKß (inhibitor of nuclear factor κ B kinase subunit ß), TPL2 (tumor progression locus 2), and JNK (c-Jun N-terminal kinase), but not p38 MAPK (mitogen-activated protein kinase) or MEK1/2 (MAPK kinase 1/2), attenuated APOE-mediated CXCL5 secretion. The roles of TAK1, IκKß, TPL2, and JNK in APOE-mediated CXCL5 secretion were verified by RNA interference. Furthermore, RNA interference showed that after APOE stimulation, both NF-κB p65 and TPL2 were downstream of TAK1 and IκKß, whereas JNK was downstream of TPL2. In summary, elevated levels of APOE in the airway may activate a TLR4/TAK1/IκKß/NF-κB/TPL2/JNK signaling pathway that induces CXCL5 secretion by human asthmatic SAECs. These findings identify new roles for TLR4 and TPL2 in APOE-mediated proinflammatory responses in asthma.
Assuntos
Apolipoproteínas E/metabolismo , Asma/metabolismo , Quimiocina CXCL5/metabolismo , Células Epiteliais/metabolismo , Sistema Respiratório/metabolismo , Transdução de Sinais/fisiologia , Receptor 4 Toll-Like/metabolismo , Quimiocinas/metabolismo , Humanos , Inflamação/metabolismo , Neutrófilos/metabolismo , RNA Mensageiro/metabolismoRESUMO
The addition of a methyl group to, typically, a cytosine-guanine dinucleotide (CpG) creates distinct DNA methylation patterns across the genome that can regulate gene expression. Aberrant DNA methylation of CpG sites has been associated with many psychiatric disorders including bipolar disorder (BD) and suicide. Using the SureSelectXT system, Methyl-Seq, we investigated the DNA methylation status of CpG sites throughout the genome in 50 BD individuals (23 subjects who died by suicide and 27 subjects who died from other causes) and 31 nonpsychiatric controls. We identified differentially methylated regions (DMRs) from three analyses: (a) BD subjects compared to nonpsychiatric controls (BD-NC), (b) BD subjects who died by suicide compared to nonpsychiatric controls (BDS-NC), and (c) BDS subjects compared to BD subjects who died from other causes (BDS-BDNS). One DMR from the BDS-NC analysis, located in ARHGEF38, was significantly hypomethylated (23.4%) in BDS subjects. This finding remained significant after multiple testing (PBootstrapped = 9.0 × 10-3 ), was validated using pyrosequencing, and was more significant in males. A secondary analysis utilized Ingenuity Pathway Analysis to identify enrichment in nominally significant DMRs. This identified an association with several pathways including axonal guidance signaling, calcium signaling, ß-adrenergic signaling, and opioid signaling. Our comprehensive study provides further support that DNA methylation alterations influence the risk for BD and suicide. However, further investigation is required to confirm these associations and identify their functional consequences.
Assuntos
Transtorno Bipolar/genética , Metilação de DNA/genética , Suicídio/psicologia , Ilhas de CpG/genética , Epigênese Genética/genética , Feminino , Genoma/genética , Estudo de Associação Genômica Ampla , Humanos , Masculino , Regiões Promotoras Genéticas/genética , Transdução de Sinais/genéticaRESUMO
BACKGROUND: Numerous genome-wide gene expression studies of bipolar disorder (BP) have been carried out. These studies are heterogeneous, underpowered and use overlapping samples. We conducted a systematic review of these studies to synthesize the current findings. METHODS: We identified all genome-wide gene expression studies on BP in humans. We then carried out a quantitative mega-analysis of studies done with post-mortem brain tissue. We obtained raw data from each study and used standardized procedures to process and analyze the data. We then combined the data and conducted three separate mega-analyses on samples from 1) any region of the brain (9 studies); 2) the prefrontal cortex (PFC) (6 studies); and 3) the hippocampus (2 studies). To minimize heterogeneity across studies, we focused primarily on the most numerous, recent and comprehensive studies. RESULTS: A total of 30 genome-wide gene expression studies of BP done with blood or brain tissue were identified. We included 10 studies with data on 211 microarrays on 57 unique BP cases and 229 microarrays on 60 unique controls in the quantitative mega-analysis. A total of 382 genes were identified as significantly differentially expressed by the three analyses. Eleven genes survived correction for multiple testing with a q-value < 0.05 in the PFC. Among these were FKBP5 and WFS1, which have been previously implicated in mood disorders. Pathway analyses suggested a role for metallothionein proteins, MAP Kinase phosphotases, and neuropeptides. CONCLUSION: We provided an up-to-date summary of results from gene expression studies of the brain in BP. Our analyses focused on the highest quality data available and provided results by brain region so that similarities and differences can be examined relative to disease status. The results are available for closer inspection on-line at Metamoodics [http://metamoodics.igm.jhmi.edu/], where investigators can look up any genes of interest and view the current results in their genomic context and in relation to leading findings from other genomic experiments in bipolar disorder.
Assuntos
Transtorno Bipolar/genética , Transtorno Bipolar/metabolismo , Encéfalo/metabolismo , Expressão Gênica , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Genótipo , HumanosRESUMO
Glucocorticoid resistance is commonly observed in depression, and has been linked to reduced expression and/or function of the glucocorticoid receptor (NR3C1 in human, hereafter referred to as GR). Previous studies have shown that GR-mutant zebrafish exhibit behavioural abnormalities that are indicative of an affective disorder, suggesting that GR plays a role in brain function. We compared the brain methylomes and brain transcriptomes of adult wild-type and GR-mutant zebrafish, and identified 249 differentially methylated regions (DMRs) that are regulated by GR. These include a cluster of CpG sites within the first intron of fkbp5, the gene encoding the glucocorticoid-inducible heat shock protein co-chaperone Fkbp5. RNA-sequencing analysis revealed that genes associated with chaperone-mediated protein folding, the regulation of circadian rhythm and the regulation of metabolism are particularly sensitive to loss of GR function. In addition, we identified subsets of genes exhibiting GR-regulated transcription that are known to regulate behaviour, and are linked to unipolar depression and anxiety. Taken together, our results identify key biological processes and novel molecular mechanisms through which the GR is likely to mediate responses to stress in the adult zebrafish brain, and they provide further support for the zebrafish GR mutant as a model for the study of affective disorders.
Assuntos
Relógios Circadianos , Receptores de Glucocorticoides , Animais , Adulto , Humanos , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Relógios Circadianos/genética , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Encéfalo/metabolismo , Transtornos do Humor/metabolismoRESUMO
Numerous candidate gene association studies of bipolar disorder (BP) have been carried out, but the results have been inconsistent. Individual studies are typically underpowered to detect associations with genes of small effect sizes. We conducted a meta-analysis of published candidate gene studies to evaluate the cumulative evidence. We systematically searched for all published candidate gene association studies of BP. We then carried out a random-effects meta-analysis on all polymorphisms that were reported on by three or more case-control studies. The results from meta-analyses of these genes were compared with the findings from a recent mega-analysis of eleven genome-wide association studies (GWAS) in BP performed by the Psychiatric GWAS Consortium (PGC). A total of 487 articles were included in our review. Among these, 33 polymorphisms in 18 genes were reported on by three or more case-control studies and included in the random-effects meta-analysis. Polymorphisms in BDNF, DRD4, DAOA, and TPH1, were found to be nominally significant with a P-value < 0.05. However, none of the findings were significant after correction for multiple testing. Moreover, none of these polymorphisms were nominally significant in the PGC-BP GWAS. A number of plausible candidate genes have been previously associated with BP. However, the lack of robust findings in our review of these candidate genes highlights the need for more atheoretical approaches to study the genetics of BP afforded by GWAS. The results of this meta-analysis and from other on-going genomic experiments in BP are available online at Metamoodics (http://metamoodics.igm.jhmi.edu).
Assuntos
Transtorno Bipolar/genética , Estudos de Associação Genética , Predisposição Genética para Doença , Estudos de Casos e Controles , Estudo de Associação Genômica Ampla , Humanos , Polimorfismo GenéticoRESUMO
Epidemiological studies, such as family, twin, and adoption studies, demonstrate the presence of a heritable component to both attempted and completed suicide. Some of this heritability is accounted for by the presence of comorbid psychiatric disorders, but the evidence also indicates that a portion of this heritability is specific to suicidality. The serotonergic system has been studied extensively in this phenotype, but findings have been inconsistent, possibly due to the presence of multiple susceptibility variants and/or gene-gene interactions. In this study, we genotyped 174 tag and coding single nucleotide polymorphisms (SNPs) from 17 genes within the serotonin pathway on 516 subjects with a major mood disorder and a history of a suicide attempt (cases) and 515 healthy controls, with the goal of capturing the common genetic variation across each of these candidate genes. We tested the 174 markers in single-SNP, haplotype, gene-based, and epistasis analyses. While these association analyses identified multiple marginally significant SNPs, haplotypes, genes, and interactions, none of them survived correction for multiple testing. Additional studies, including assessment in larger sample sets and deep resequencing to identify rare causal variants, may be required to fully understand the role that the serotonin pathway plays in suicidal behavior.
Assuntos
Estudos de Associação Genética , Serotonina/genética , Transdução de Sinais/genética , Tentativa de Suicídio , Humanos , Polimorfismo de Nucleotídeo Único/genética , Tentativa de Suicídio/psicologia , Transmissão Sináptica/genéticaRESUMO
Acute pain, the most prominent complication of sickle cell disease (SCD), results from vaso-occlusion triggered by sickling of deoxygenated red blood cells (RBCs). Concentration of 2,3-diphosphoglycerate (2,3-DPG) in RBCs promotes deoxygenation by preferentially binding to the low-affinity T conformation of HbS. 2,3-DPG is an intermediate substrate in the glycolytic pathway in which pyruvate kinase (gene PKLR, protein PKR) is a rate-limiting enzyme; variants in PKLR may affect PKR activity, 2,3-DPG levels in RBCs, RBC sickling, and acute pain episodes (APEs). We performed a candidate gene association study using 2 cohorts: 242 adult SCD-HbSS patients and 977 children with SCD-HbSS or SCD-HbSß0 thalassemia. Seven of 47 PKLR variants evaluated in the adult cohort were associated with hospitalization: intron 4, rs2071053; intron 2, rs8177970, rs116244351, rs114455416, rs12741350, rs3020781, and rs8177964. All 7 variants showed consistent effect directions in both cohorts and remained significant in weighted Fisher's meta-analyses of the adult and pediatric cohorts using P < .0071 as threshold to correct for multiple testing. Allele-specific expression analyses in an independent cohort of 52 SCD adults showed that the intronic variants are likely to influence APE by affecting expression of PKLR, although the causal variant and mechanism are not defined.
Assuntos
Dor Aguda , Anemia Falciforme , Piruvato Quinase , 2,3-Difosfoglicerato/metabolismo , Dor Aguda/genética , Dor Aguda/metabolismo , Adulto , Anemia Falciforme/complicações , Anemia Falciforme/genética , Criança , Eritrócitos Anormais/metabolismo , Hemoglobina Falciforme/metabolismo , Humanos , Piruvato Quinase/genética , Piruvato Quinase/metabolismoRESUMO
COVID-19 pathogenesis is associated with an exuberant inflammatory response. However, the tissue injury pattern and immune response in solid-organ transplant recipients (SOTRs) taking immunosuppressive therapy have not been well characterized. Here, we perform both cfDNA and cytokine profiling on plasma samples to map tissue damage, including allograft injury and delineate underlying immunopathology. We identified injuries from multiple-tissue types, including hematopoietic cells, vascular endothelium, hepatocyte, adipocyte, pancreas, kidney, heart, and lung in SOTRs with COVID-19 that correlates with disease severity. SOTRs with COVID-19 have higher plasma levels of cytokines such as IFN-λ1, IFN-γ, IL-15, IL-18 IL-1RA, IL-6, MCP-2, and TNF-α as compared to healthy controls, and the levels of GM-CSF, IL-15, IL-6, IL-8, and IL-10 were associated with disease severity in SOTRs. Strikingly, IFN-λ and IP-10 were markedly increased in SOTRs compared to immunocompetent patients with COVID-19. Correlation analyses showed a strong association between monocyte-derived cfDNA and inflammatory cytokines/chemokines in SOTRs with COVID-19. Moreover, compared to other respiratory viral infections, COVID-19 induced pronounced injury in hematopoietic, vascular endothelial and endocrine tissues. Allograft injury, measured as donor-derived cfDNA was elevated in SOTRs with COVID-19, including allografts distant from the primary site of infection. Allograft injury correlated with inflammatory cytokines and cfDNA from immune cells. Furthermore, longitudinal analysis identified a gradual decrease of cfDNA and inflammatory cytokine levels in patients with a favorable outcome. Our findings highlight distinct tissue injury and cytokine features in SOTRs with COVID-19 that correlate with disease severity.
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Recent genetic studies have identified variants associated with bipolar disorder (BD), but it remains unclear how brain gene expression is altered in BD and how genetic risk for BD may contribute to these alterations. Here, we obtained transcriptomes from subgenual anterior cingulate cortex and amygdala samples from post-mortem brains of individuals with BD and neurotypical controls, including 511 total samples from 295 unique donors. We examined differential gene expression between cases and controls and the transcriptional effects of BD-associated genetic variants. We found two coexpressed modules that were associated with transcriptional changes in BD: one enriched for immune and inflammatory genes and the other with genes related to the postsynaptic membrane. Over 50% of BD genome-wide significant loci contained significant expression quantitative trait loci (QTL) (eQTL), and these data converged on several individual genes, including SCN2A and GRIN2A. Thus, these data implicate specific genes and pathways that may contribute to the pathology of BP.
Assuntos
Transtorno Bipolar , Giro do Cíngulo , Tonsila do Cerebelo/metabolismo , Transtorno Bipolar/genética , Transtorno Bipolar/metabolismo , Encéfalo/metabolismo , Giro do Cíngulo/metabolismo , Humanos , TranscriptomaRESUMO
Genome-wide association studies (GWAS) have identified several susceptibility loci for bipolar disorder (BP), most notably ANK3. However, most of the inherited risk for BP remains unexplained. One reason for the limited success may be the genetic heterogeneity of BP. Clinical sub-phenotypes of BP may identify more etiologically homogeneous subsets of patients, which can be studied with increased power to detect genetic variation. Here, we report on a mega-analysis of two widely studied sub-phenotypes of BP, age at onset and psychotic symptoms, which are familial and clinically significant. We combined data from three GWAS: NIMH Bipolar Disorder Genetic Association Information Network (GAIN-BP), NIMH Bipolar Disorder Genome Study (BiGS), and a German sample. The combined sample consisted of 2,836 BP cases with information on sub-phenotypes and 2,744 controls. Imputation was performed, resulting in 2.3 million SNPs available for analysis. No SNP reached genome-wide significance for either sub-phenotype. In addition, no SNP reached genome-wide significance in a meta-analysis with an independent replication sample. We had 80% power to detect associations with a common SNP at an OR of 1.6 for psychotic symptoms and a mean difference of 1.8 years in age at onset. Age at onset and psychotic symptoms in BP may be influenced by many genes of smaller effect sizes or other variants not measured well by SNP arrays, such as rare alleles.
Assuntos
Transtorno Bipolar/epidemiologia , Transtorno Bipolar/genética , Estudo de Associação Genômica Ampla , Transtornos Psicóticos/epidemiologia , Transtornos Psicóticos/genética , Idade de Início , Transtorno Bipolar/complicações , Demografia , Feminino , Alemanha/epidemiologia , Humanos , Masculino , Transtornos Psicóticos/complicações , Adulto JovemRESUMO
There is accumulating evidence that long noncoding RNAs (lncRNAs) play crucial roles in biological processes and diseases. In recent years, computational models have been widely used to predict potential lncRNA-disease relations. In this chapter, we systematically describe various computational algorithms and prediction tools that have been developed to elucidate the roles of lncRNAs in diseases, coding potential/functional characterization, or ascertaining their involvement in critical biological processes as well as provide a comprehensive summary of these applications.
Assuntos
Biologia Computacional/métodos , RNA Longo não Codificante/genética , Algoritmos , Doença/genética , Humanos , Proteínas/genética , RNA Longo não Codificante/metabolismoRESUMO
Analyzing host cells' transcriptional response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection will help delineate biological processes underlying viral pathogenesis. First, analysis of expression profiles of lung cell lines A549 and Calu3 revealed upregulation of antiviral interferon signaling genes in response to all three SARS-CoV-2, MERS-CoV, or influenza A virus (IAV) infections. However, perturbations in expression of genes involved in inflammatory, mitochondrial, and autophagy processes were specifically observed in SARS-CoV-2-infected cells. Next, a validation study in infected human nasopharyngeal samples also revealed perturbations in autophagy and mitochondrial processes. Specifically, mTOR expression, mitochondrial ribosomal, mitochondrial complex I, lysosome acidification, and mitochondrial fission promoting genes were concurrently downregulated in both infected cell lines and human samples. SARS-CoV-2 infection impeded autophagic flux either by upregulating GSK3B in lung cell lines or by downregulating autophagy genes, SNAP29, and lysosome acidification genes in human samples, contributing to increased viral replication. Therefore, drugs targeting lysosome acidification or autophagic flux could be tested as intervention strategies. Finally, age-stratified SARS-CoV-2-positive human data revealed impaired upregulation of chemokines, interferon-stimulated genes, and tripartite motif genes that are critical for antiviral signaling. Together, this analysis has revealed specific aspects of autophagic and mitochondrial function that are uniquely perturbed in SARS-CoV-2-infected host cells.
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Haploidentical hematopoietic stem cell transplantation (haplo-HSCT) is a widely available curative option for patients with sickle cell disease (SCD). Our original non-myeloablative haplo-HSCT trial employing post-transplant (PT) cyclophosphamide had a low incidence of GVHD but had high rejection rates. Here, we aimed to evaluate immune reconstitution following haplo-HSCT and identify cytokines and cells associated with graft rejection/engraftment. 50 cytokines and 10 immune cell subsets were screened using multiplex-ELISA and flow cytometry, respectively, at baseline and PT-Days 30, 60, 100, and 180. We observed the most significant differences in cytokine levels between the engrafted and rejected groups at PT-Day 60, corresponding with clinical findings of secondary graft rejection. Of the 44 cytokines evaluated, plasma concentrations of 19 cytokines were different between the two groups at PT-Day 60. Factor analysis suggested two independent factors. The first factor (IL-17A, IL-10, IL-7, G-CSF, IL-2, MIP-1a, VEGF, and TGFb1 contributed significantly) was strongly associated with engraftment with OR = 2.7 (95%CI of 1.4 to 5.4), whereas the second factor (GROa and IL-18 contributed significantly) was not significantly associated with engraftment. Sufficient donor myeloid chimerism (DMC) is critical for the success of HSCT; here, we evaluated immune cells among high (H) DMC (DMC≥20%) and low (L) DMC (DMC<20%) groups along with engrafted and rejected groups. We found that early myeloid-derived suppressor cell (eMDSC) frequencies were elevated in engrafted patients and patients with HDMC at PT-Day 30 (P< 0.04 & P< 0.003, respectively). 9 of 20 patients were evaluated for the source of eMDSCs. The HDMC group had high mixed chimeric eMDSCs as compared to the LDMC group (P< 0.00001). We found a positive correlation between the frequencies of eMDSCs and Tregs at PT-Day 100 (r=0.72, P <0.0007); eMDSCs at BSL and Tregs at PT-Day 100 (r=0.63, P <0.004). Of 10 immune regulatory cells and 50 cytokines, we observed mixed chimeric eMDSCs and IL-17A, IL-10, IL-7, G-CSF, IL-2, MIP-1a, VEGF, TGFb1 as potential hits which could serve as prognostic markers in predicting allograft outcome towards engraftment following haploidentical HSCT employing post-transplant cyclophosphamide. The current findings need to be replicated and further explored in a larger cohort.
Assuntos
Anemia Falciforme/terapia , Transplante de Células-Tronco Hematopoéticas , Reconstituição Imune/imunologia , Quimeras de Transplante , Adulto , Anemia Falciforme/imunologia , Quimerismo , Ciclofosfamida/uso terapêutico , Citocinas/imunologia , Rejeição de Enxerto/imunologia , Sobrevivência de Enxerto/imunologia , Humanos , Imunossupressores/uso terapêutico , Células Supressoras Mieloides , Prognóstico , Condicionamento Pré-Transplante , Transplante Haploidêntico , Resultado do TratamentoRESUMO
Activation of NOTCH signaling in human hematopoietic stem/progenitor cells (HSPCs) by treatment with an engineered Delta-like ligand (DELTA1ext-IgG [DXI]) has enabled ex vivo expansion of short-term HSPCs, but the effect on long-term repopulating hematopoietic stem cells (LTR-HSCs) remains uncertain. Here, we demonstrate that ex vivo culture of human adult HSPCs with DXI under low oxygen tension limits ER stress in LTR-HSCs and lineage-committed progenitors compared with normoxic cultures. A distinct HSC gene signature was upregulated in cells cultured with DXI in hypoxia and, after 21 days of culture, the frequency of LTR-HSCs increased 4.9-fold relative to uncultured cells and 4.2-fold compared with the normoxia + DXI group. NOTCH and hypoxia pathways intersected to maintain undifferentiated phenotypes in cultured HSPCs. Our work underscores the importance of mitigating ER stress perturbations to preserve functional LTR-HSCs in extended cultures and offers a clinically feasible platform for the expansion of human HSPCs.
Assuntos
Hipóxia Celular , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Receptores Notch/metabolismo , Antígenos CD34/metabolismo , Biomarcadores , Técnicas de Cultura de Células , Diferenciação Celular , Proliferação de Células , Biologia Computacional/métodos , Humanos , Anotação de Sequência Molecular , Receptores Notch/genética , Transdução de Sinais , TranscriptomaRESUMO
A growing number of long noncoding RNAs (lncRNAs) have emerged as vital metabolic regulators. However, most human lncRNAs are nonconserved and highly tissue specific, vastly limiting our ability to identify human lncRNA metabolic regulators (hLMRs). In this study, we established a pipeline to identify putative hLMRs that are metabolically sensitive, disease relevant, and population applicable. We first progressively processed multilevel human transcriptome data to select liver lncRNAs that exhibit highly dynamic expression in the general population, show differential expression in a nonalcoholic fatty liver disease (NAFLD) population, and respond to dietary intervention in a small NAFLD cohort. We then experimentally demonstrated the responsiveness of selected hepatic lncRNAs to defined metabolic milieus in a liver-specific humanized mouse model. Furthermore, by extracting a concise list of protein-coding genes that are persistently correlated with lncRNAs in general and NAFLD populations, we predicted the specific function for each hLMR. Using gain- and loss-of-function approaches in humanized mice as well as ectopic expression in conventional mice, we validated the regulatory role of one nonconserved hLMR in cholesterol metabolism by coordinating with an RNA-binding protein, PTBP1, to modulate the transcription of cholesterol synthesis genes. Our work overcame the heterogeneity intrinsic to human data to enable the efficient identification and functional definition of disease-relevant human lncRNAs in metabolic homeostasis.
Assuntos
Bases de Dados de Ácidos Nucleicos , Homeostase/genética , Hepatopatia Gordurosa não Alcoólica , RNA Longo não Codificante , Animais , Humanos , Camundongos , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismoRESUMO
Intermittent fasting blunts inflammation in asthma1 and rheumatoid arthritis2, suggesting that fasting may be exploited as an immune-modulatory intervention. However, the mechanisms underpinning the anti-inflammatory effects of fasting are poorly characterized3-5. Here, we show that fasting in humans is sufficient to blunt CD4+ T helper cell responsiveness. RNA sequencing and flow cytometry immunophenotyping of peripheral blood mononuclear cells from volunteers subjected to overnight or 24-h fasting and 3 h of refeeding suggest that fasting blunts CD4+ T helper cell activation and differentiation. Transcriptomic analysis reveals that longer fasting has a more robust effect on CD4+ T-cell biology. Through bioinformatics analyses, we identify the transcription factor FOXO4 and its canonical target FK506-binding protein 5 (FKBP5) as a potential fasting-responsive regulatory axis. Genetic gain- or loss-of-function of FOXO4 and FKBP5 is sufficient to modulate TH1 and TH17 cytokine production. Moreover, we find that fasting-induced or genetic overexpression of FOXO4 and FKBP5 is sufficient to downregulate mammalian target of rapamycin complex 1 signalling and suppress signal transducer and activator of transcription 1/3 activation. Our results identify FOXO4-FKBP5 as a new fasting-induced, signal transducer and activator of transcription-mediated regulatory pathway to blunt human CD4+ T helper cell responsiveness.