Correction: The transcriptome of equine peripheral blood mononuclear cells.
Ontology highlight
ABSTRACT:
SUBMITTER: PLOS ONE Staff
PROVIDER: S-EPMC4410916 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Complete transcriptomic data at high resolution are available only for a few model organisms with medical importance. The gene structures of non-model organisms are mostly computationally predicted based on comparative genomics with other species. As a result, more than half of the horse gene models are known only by projection. Experimental data supporting these gene models are scarce. Moreover, most of the annotated equine genes are single-transcript genes. Utilizing RNA sequencing (RNA-seq) the experimental validation of predicted transcriptomes has become accessible at reasonable costs. To improve the horse genome annotation we performed RNA-seq on 561 samples of peripheral blood mononuclear cells (PBMCs) derived from 85 Warmblood horses. The mapped sequencing reads were used to build a new transcriptome assembly. The new assembly revealed many alternative isoforms associated to known genes or to those predicted by the Ensembl and/or Gnomon pipelines. We also identified 7,531 transcripts not associated with any horse gene annotated in public databases. Of these, 3,280 transcripts did not have a homologous match to any sequence deposited in the NCBI EST database suggesting horse specificity. The unknown transcripts were categorized as coding and noncoding based on predicted coding potential scores. Among them 230 transcripts had high coding potential score, at least 2 exons, and an open reading frame of at least 300 nt. We experimentally validated 9 new equine coding transcripts using RT-PCR and Sanger sequencing. Our results provide valuable detailed information on many transcripts yet to be annotated in the horse genome.
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Project description:Lipopolysaccharide (endotoxin, LPS) is a strong inducer of the innate immune response. It is widespread in our environment, e.g. in house dust and contributes to asthma. Compared to humans, horses are even more sensitive to LPS. However, data on LPS effects on the equine transcriptome are very limited. Using RNA-seq we analysed LPS-induced differences in the gene expression in equine peripheral blood mononuclear cells at the gene and gene-network level in two half-sib families and one group of unrelated horses.24 h-LPS challenge of equine immune cells resulted in substantial changes in the transcriptomic profile (1,265 differentially expressed genes) showing partial overlap with human data. One of the half-sib families showed a specific response different from the other two groups of horses. We also identified co-expressed gene modules that clearly differentiated 24 h-LPS- from non-stimulated samples. These modules consisted of 934 highly interconnected genes and included genes involved in the immune response (e.g. IL6, CCL22, CXCL6, CXCL2), however, none of the top ten hub genes of the modules have been annotated as responsive to LPS in gene ontology.Using weighted gene co-expression network analysis we identified ten co-expressed gene modules significantly regulated by in vitro stimulation with LPS. Apart from 47 genes (5%) all other genes highly interconnected within the most up- and down-regulated modules were also significantly differentially expressed (FDR?<?0.05). The LPS-regulated module hub genes have not yet been described as having a role in the immune response to LPS (e.g. VAT1 and TTC25).
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Project description:BackgroundSarcoidosis is a granulomatous systemic disease of unknown etiology. Mononuclear cells such as macrophages or lymphocytes in lung tissue and hilar or mediastinal lymph nodes have been recognized to play an essential role in granuloma formation in pulmonary sarcoidosis. Peripheral blood mononuclear cells (PBMCs) consist of several immunocompetent cells and have been shown to play a mechanistic role in the pathogenesis of sarcoidosis. However, the genetic modifications that occur in bulk PBMCs of sarcoidosis remain to be elucidated.PurposeThis study aimed to explore the pathobiological markers of sarcoidosis in PBMCs by comparing the transcriptional signature of PBMCs from patients with pulmonary sarcoidosis with those of healthy controls by RNA sequencing.MethodsPBMC samples were collected from subjects with pulmonary sarcoidosis with no steroid/immunosuppressant drugs (n = 8) and healthy controls (n = 11) from August 2020 to April 2021, and RNA sequencing was performed with the PBMC samples.ResultsPrincipal component analysis using RNA sequencing datasets comparing pulmonary sarcoidosis with healthy controls revealed that the two groups appeared to be differentiated, in which 270 differentially expressed genes were found in PBMCs between sarcoidosis and healthy controls. Enrichment analysis for gene ontology suggested that some biological processes related to the pathobiology of sarcoidosis, such as cellular response to interleukin (IL)-1 and IFN-γ, regulation of IL-6 production, IL-8 secretion, regulation of mononuclear cell migration, and response to lipopolysaccharide, were involved. Enrichment analysis of the KEGG pathway indicated the involvement of tumor necrosis factor (TNF), toll-like receptor signaling, IL-17 signaling pathways, phagosomes, and ribosomes. Most of the genes involved in TNF and IL-17 signaling pathways and phagosomes were upregulated, while most of the ribosome-related genes were downregulated.ConclusionThe present study demonstrated that bulk gene expression patterns in PBMCs were different between patients with pulmonary sarcoidosis and healthy controls. The changes in the gene expression pattern of PBMCs could reflect the existence of sarcoidosis lesions and influence granuloma formation in sarcoidosis. These new findings are important to strengthen our understanding of the etiology and pathobiology of sarcoidosis and indicate a potential therapeutic target for sarcoidosis.
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Project description:Equine herpesvirus 1 (EHV-1) affects horses worldwide and causes respiratory disease, abortions, and equine herpesvirus myeloencephalopathy (EHM). Following infection, a cell-associated viremia is established in the peripheral blood mononuclear cells (PBMCs). This viremia is essential for transport of EHV-1 to secondary infection sites where subsequent immunopathology results in diseases such as abortion or EHM. Because of the central role of PBMCs in EHV-1 pathogenesis, our goal was to establish a gene expression analysis of host and equine herpesvirus genes during EHV-1 viremia using RNA sequencing. When comparing transcriptomes of PBMCs during peak viremia to those prior to EHV-1 infection, we found 51 differentially expressed equine genes (48 upregulated and 3 downregulated). After gene ontology analysis, processes such as the interferon defense response, response to chemokines, the complement protein activation cascade, cell adhesion, and coagulation were overrepresented during viremia. Additionally, transcripts for EHV-1, EHV-2, and EHV-5 were identified in pre- and post-EHV-1-infection samples. Looking at micro RNAs (miRNAs), 278 known equine miRNAs and 855 potentially novel equine miRNAs were identified in addition to 57 and 41 potentially novel miRNAs that mapped to the EHV-2 and EHV-5 genomes, respectively. Of those, 1 EHV-5 and 4 equine miRNAs were differentially expressed in PBMCs during viremia. In conclusion, this work expands our current knowledge about the role of PBMCs during EHV-1 viremia and will inform the focus on future experiments to identify host and viral factors that contribute to clinical EHM.
| S-EPMC7825769 | biostudies-literature
Project description:Background: Sepsis, as a clinical emergency, usually causes multiorgan dysfunction and can lead to high mortality. Establishment of specific and sensitive biomarkers for early diagnosis is critical to identify patients who would benefit from targeted therapy. In this study, we investigated this syndrome by analyzing the transcriptome of peripheral blood mononuclear cells (PBMCs) from patients with sepsis and identified sepsis-specific biomarkers. Methods: In this study, a total of 87 patients with sepsis and 40 healthy controls from a prospective multicenter cohort were enrolled. Samples from 44 subjects (24 patients with sepsis and 20 healthy controls) were sequenced and the remaining patients were included in the validation group. Using high-throughput sequencing, a gene expression profile of PBMCs from patients with sepsis was generated to elucidate the pathophysiology of sepsis and identify sepsis-specific biomarkers. Results: Principal component analysis (PCA) and unsupervised hierarchical cluster analysis showed that patients with sepsis separated from healthy controls. A total of 1639 differentially expressed genes (DEGs) were identified (|log2 fold change|>2, adjusted P value <0.05) between these two groups, with 1278 (78.0%) upregulated and 361 (22.0%) downregulated in patients with sepsis. Gene Ontology (GO) analysis of the upregulated DEGs identified 194 GO terms that were clustered into 27 groups, and analysis of the downregulated DEGs identified 20 GO terms that were clustered into 4 groups. Four unique genes were identified that could be predictive of patients with sepsis. External validation of the four genes using quantitative real-time polymerase chain reaction (qRT-PCR) was consistent with the results of mRNA sequencing, revealing their potential in sepsis diagnosis. Conclusions: The transcriptome characteristics of PBMCs, which were significantly altered in sepsis patients, provide new insights into sepsis pathogenesis. The four identified gene expression changes differentiated patients with sepsis from healthy subjects, which could serve as a convenient tool contributing to sepsis diagnosis.
| S-EPMC7484663 | biostudies-literature
Project description:Peripheral blood mononuclear cells (PBMCs) are blood cells that are a critical part of the immune system used to fight off infection, defending our bodies from harmful pathogens. In biomedical research, PBMCs are commonly used to study global immune response to disease outbreak and progression, pathogen infections, for vaccine development and a multitude of other clinical applications. Over the past few years, the revolution in single-cell RNA sequencing (scRNA-seq) has enabled an unbiased quantification of gene expression in thousands of individual cells, which provides a more efficient tool to decipher the immune system in human diseases. In this work, we generate scRNA-seq data from human PBMCs at high sequencing depth (>100,000 reads/cell) for more than 30,000 cells, in resting, stimulated, fresh and frozen conditions. The data generated can be used for benchmarking batch correction and data integration methods, and to study the effect of freezing-thawing cycles on the quality of immune cell populations and their transcriptomic profiles.
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Project description:The elite controller (EC)-long term non-progressor (LTNP) phenotype represent a spontaneous and advantageous model of HIV-1 control in the absence of therapy. The transcriptome of peripheral blood mononuclear cells (PBMCs) collected from EC-LTNPs was sequenced by RNA-Seq and compared with the transcriptomes from other phenotypes of disease progression. The transcript abundance estimation combined with the use of supervised classification algorithms allowed the selection of 20 genes and pseudogenes, mainly involved in interferon-regulated antiviral mechanisms and cell machineries of transcription and translation, as the best predictive genes of disease progression. Differential expression analyses between phenotypes showed an altered calcium homeostasis in EC-LTNPs evidenced by the upregulation of several membrane receptors implicated in calcium-signaling cascades and intracellular calcium-mobilization and by the overrepresentation of NFAT1/Elk-1-binding sites in the promoters of the genes differentially expressed in these individuals. A coordinated upregulation of host genes associated with HIV-1 reverse transcription and viral transcription was also observed in EC-LTNPs -i.e. p21/CDKN1A, TNF, IER3 and GADD45B. We also found an upregulation of ANKRD54 in EC-LTNPs and viremic LTNPs in comparison with typical progressors and a clear alteration of type-I interferon signaling as a consequence of viremia in typical progressors before and after receiving antiretroviral therapy.
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Project description:The innate immune response to lipopolysaccharide contributes substantially to the morbidity and mortality of gram-negative sepsis. Horses and humans share an exquisite sensitivity to lipopolysaccharide and thus the horse may provide valuable comparative insights into this aspect of the inflammatory response. MicroRNAs, small non-coding RNA molecules acting as post-transcriptional regulators of gene expression, have key roles in toll-like receptor signaling regulation but have not been studied in this context in horses. The central hypothesis of this study was that lipopolysaccharide induces differential microRNA expression in equine peripheral blood mononuclear cells in a manner comparable to humans. Illumina Next Generation Sequencing was used to characterize the basal microRNA transcriptome in isolated peripheral blood mononuclear cells from healthy adult horses, and to evaluate LPS-induced changes in microRNA expression in cells cultured for up to four hours. Selected expression changes were validated using quantitative reverse-transcriptase PCR. Only miR-155 was significantly upregulated by LPS, changing in parallel with supernatant tumor necrosis factor-? concentration. Eight additional microRNAs, including miR-146a and miR-146b, showed significant expression change with time in culture without a clear LPS effect. Target predictions indicated a number of potential immunity-associated targets for miR-155 in the horse, including SOCS1, TAB2 and elements of the PI3K signaling pathway, suggesting that it is likely to influence the acute inflammatory response to LPS. Gene alignment showed extensive conservation of the miR-155 precursor gene and associated promoter regions between horses and humans. The basal and LPS-stimulated microRNA expression pattern characterized here were similar to those described in human leukocytes. As well as providing a resource for further research into the roles of microRNAs in immune responses in horses, this will facilitate inter-species comparative study of the role of microRNAs in the inflammatory cascade during endotoxemia and sepsis.
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Project description:Non-emphysematous chronic obstructive pulmonary disease (COPD), which is defined based on chest computed tomography findings, presented different transcriptome features of peripheral blood mononuclear cells (PBMCs) compared with emphysematous COPD. Enrichment analysis of transcriptomic data in COPD demonstrated that the "Hematopoietic cell lineage" pathway in Kyoto Encyclopedia of Genes and Genomes pathway analysis was highly upregulated, suggesting that cellular dynamic dysregulation in COPD lungs is affected by pathologically modified PBMCs. The differentially expressed genes (DEGs) upregulated in PBMCs reflected the disease state of non-emphysematous COPD. Upregulated DEGs such as XCL1, PRKCZ, TMEM102, CD200R1, and AQP1 activate T lymphocytes and eosinophils. Upregulating keratan sulfate biosynthesis and metabolic processes is associated with protection against the destruction of the distal airways. ITGA3 upregulation augments interactions with extracellular matrix proteins, and COL6A1 augments the profibrotic mast cell phenotype during alveolar collagen VI deposition. Upregulating HSPG2, PDGFRB, and PAK4 contributes to the thickening of the airway wall, and upregulating SERPINF1 expression explains the better-preserved vascular bed. Therefore, gene expression and pathway analysis in PBMCs in patients with non-emphysematous COPD represented type 2 immune responses and airway remodeling features. Therefore, these patients have asthmatic potential despite no clinical signs of asthma, in contrast to those with emphysematous COPD.
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Project description:The present study aimed to investigate the differential expression of long non-coding RNAs (lncRNAs) in rheumatoid arthritis (RA). High-throughput gene sequencing technology was used to detect the expression of lncRNA and mRNA in three patients with RA (RA group) and normal controls (NC group). A Bioinformatics analysis was used to assess the effects of differentially expressed mRNAs on signaling pathways and biological functions. The selected dysregulated lncRNAs were verified by reverse transcription-quantitative (RT-q)PCR in the peripheral blood mononuclear cells (PBMCs) of patients with RA and age- and sex-matched controls. A correlation analysis was used to analyze the relationship between lncRNAs and clinical indexes. From the lncRNA sequencing data, significantly differentially expressed lncRNAs between the RA and NC groups were identified by a fold change ≥2 and P<0.05. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis suggested that the differentially expressed mRNAs were mainly involved in organelle composition, intracellular regulation, signaling pathways, cancer, virus and inflammation. A total of four of these lncRNAs were confirmed by RT-qPCR to be significantly differentially expressed (LINC00304, MIR503HG, LINC01504 and FAM95B1). Through the correlation analysis, it was confirmed that there was a strong correlation between these lncRNAs and clinical laboratory indicators and indexes such as course of disease, arthrocele and joint tenderness. Overall, the present results suggested that the expression levels of LINC00304, MIR503HG, LINC01504 and FAM95B1 in PBMCs from patients with RA may serve as potential biomarkers for RA diagnosis, influencing the occurrence and progress of RA.
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