Project description:The peritoneal macrophages were infected with Mtb H37Rv for 4 hours, and the miRNA expression profile were analyzed with deep sequencing.
Project description:The goal of this study is to perform transcriptome profiling of both infected and uninfected WT and HIF-1ɑ-/- peritoneal macrophages using RNA sequencing techniques. For that, WT and HIF-1ɑ-/- peritoneal macrophages were infeced for 6 hours with L. donovani promastigotes (or left uninfected as a control) and RNA samples were stored in TRI reagent (Sigma), for further analysis.
Project description:Noncoding RNAs regulate the process of Mycobacterium tuberculosis (M. tb) infecting the host, but there is no simultaneous transcriptional information of long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) and the global regulatory networks of non-coding RNA. Rv1759c, a virulence factor, is a member of protein family containing the proline-glutamic acid (PE) in M. tb, which can increase M. tb survival. To reveal the noncoding RNA regulatory networks and the effect of Rv1759c on non-coding RNA expression during M. tb infection, we collected samples of H37Rv- and H37Rv△1759c-infected macrophages and explored the full transcriptome expression profile. We found 356 mRNAs, 433 lncRNAs, 168 circRNAs, and 12 miRNAs differentially expressed during H37Rv infection, 356 mRNAs, 433 lncRNAs, 168 circRNAs, and 12 miRNAs differentially expressed during H37Rv△1759c infection. We constructed lncRNA/circRNA-miRNA-mRNA regulatory networks during H37Rv and H37Rv△1759c infection. We demonstrated the role of one of the hubs of the networks, hsa-miR-181b-3p, for H37Rv survival in macrophages. We discovered that the expression changes of 68 mRNAs, 92 lncRNAs, 26 circRNAs, and 3 miRNAs were only related to the deletion of Rv1759c by comparing the transcription profiles of H37Rv and H37Rv△1759c. Here, our study comprehensively characterizes the transcriptional profiles in THP1-derived-macrophages infected with H37Rv and H37Rv△1759c, which provides support and new directions for in-depth exploration of noncoding RNA and PE/PPE family functions during the infection process.
Project description:To explore the regulatory network of noncoding RNAs after M.tb infection and the role of Rv1759c in the infection process, we collected samples of H37Rv- and H37Rv△1759c-infected macrophages and explored the full transcriptome expression profile. We constructed DE-lncRNA/DE-circRNA-DE-miRNA-DE-mRNA regulatory networks during H37Rv and H37Rv△1759c infection. In addition, we first discovered the close relationship between Rv1759c and chemokines during M.tb infection by comparing the transcription profiles of H37Rv and H37Rv△1759c and bioinformatics analysis. Here, our study comprehensively characterizes the ncRNA and mRNA profiles in macrophages infected with H37Rv and H37Rv△1759c, which provides support and new directions for in-depth exploration of ncRNA and PE/PPE family functions during the infection process.
Project description:The C57B/6 mice were infected with Mtb H37Rv (CFU=200) for 28 days, and the miRNA expression profile from lung tissues were analyzed with deep sequencing.
Project description:Tuberculosis (TB) remains a worldwide healthcare concern, and the exploration of the host-pathogen interaction is essential to develop therapeutic modalities and strategies to control Mycobacterium tuberculosis (M.tb). In this study, RNA sequencing (transcriptome sequencing) was employed to investigate the global transcriptome changes in the macrophages during the different strains of M.tb infection. THP-1 cells derived from macrophages were exposed to the virulent M.tb strain H37Rv (Rv) or the avirulent M.tb strain H37Ra (Ra), and the M.tb BCG vaccine strain was used as a control. The cDNA libraries were prepared from M.tb-infected macrophages and then sequenced. To assess the transcriptional differences between the expressed genes, the bioinformatics analysis was performed using a standard pipeline of quality control, reference mapping, differential expression analysis, protein-protein interaction (PPI) networks, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Q-PCR and Western blot assays were also performed to validate the data. Our findings indicated that, when compared to BCG or M.tb H37Ra infection, the transcriptome analysis identified 66 differentially expressed genes in the M.tb H37Rv-infected macrophages, out of which 36 genes were up-regulated, and 30 genes were down-regulated. The up-regulated genes were associated with immune response regulation, chemokine secretion, and leucocyte chemotaxis. In contrast, the down-regulated genes were associated with amino acid biosynthetic and energy metabolism, connective tissue development and extracellular matrix organization. The Q-PCR and Western blot assays confirmed increased expression of pro-inflammatory factors, altered energy metabolic processes, enhanced activation of pro-inflammatory signalling pathways and increased pyroptosis in H37Rv-infected macrophage. Overall, our RNA sequencing-based transcriptome study successfully identified a comprehensive, in-depth gene expression/regulation profile in M.tb-infected macrophages. The results demonstrated that virulent M.tb strain H37Rv infection triggers a more severe inflammatory immune response associated with increased tissue damage, which helps in understanding the host-pathogen interaction dynamics and pathogenesis features in different strains of M.tb infection.
