The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection
| dc.contributor.advisor | Brombacher, Frank | |
| dc.contributor.advisor | Guler, Reto | |
| dc.contributor.advisor | Tamgue, Ousman | |
| dc.contributor.author | Pillay, Shandré | |
| dc.date.accessioned | 2022-08-22T18:37:03Z | |
| dc.date.available | 2022-08-22T18:37:03Z | |
| dc.date.issued | 2022 | |
| dc.date.updated | 2022-08-22T11:23:57Z | |
| dc.description.abstract | In 2020, the World Health Organization (WHO) reported 1.5 million tuberculosis (TB)- associated deaths and an incidence of 10 million new cases. The causative, Mycobacterium tuberculosis (Mtb), evades host immune responses by skewing macrophage polarization towards a less microbicidal alternative state to avoid classical effector killing functions. However, the molecular details underlying these evasion mechanisms remain incomplete and current therapy is challenged with drug resistance. Host-directed therapy (HDT) has recently gained attention, with long non-coding RNAs (lncRNAs) as potential targets due to their emerging roles in pathogenic immune responses. We previously performed cap analysis gene expression (CAGE) transcriptomics on IFN-γ stimulated (classically activated) and IL-4/IL-13 stimulated (alternatively activated) mouse macrophages, identifying 151 differentially expressed lncRNAs following Mtb infection. We validated the top 11 differentially expressed lncRNAs and two were chosen for this study, lncRNA-125, whose expression was regulated at different levels unstimulated and in response to IFN-γ and IL-4/IL-13, and lncRNA-612 whose expression was only induced by IFN-γ stimulation. Interestingly, the expression of lncRNA125 and lncRNA-612 was downregulated following Mtb infection. Therefore, this study aimed at functionally validating these lncRNAs in unstimulated, IFN-γ and IL-4/IL-13 stimulated and/or Mtb-infected mouse and human macrophages by a loss-of-function approach using chemically engineered antisense oligonucleotides (gapmeRs). Knockdown of lncRNA-125 by gapmeRs reduced Mtb growth and anti-inflammatory cytokine production mediated by increased apoptosis, nitrite and pro-inflammatory cytokine production in IL-4/IL-13 prestimulated mouse macrophages. Whereas knockdown of lncRNA-125 in IFN-γ pre-stimulated mouse macrophages favoured Mtb growth and anti-inflammatory cytokine production, with reduction of apoptosis, nitrite and pro-inflammatory cytokine production. Therefore, indicating that lncRNA-125 regulates macrophage polarization during Mtb infection. Knockdown of lncRNA-125 in human macrophages resulted in reduced Mtb growth and increased proinflammatory cytokine production in unstimulated, IFN-γ and IL-4/IL-13 pre-stimulated BMDMs infected with Mtb. Comparatively, gapmeR knockdown of lncRNA-612 reduced Mtb growth and increased pro-inflammatory cytokine production in IFN-γ pre-stimulated mouse and human macrophages. In mouse macrophages, these responses were mediated by increased apoptosis and nitrite production, with reduced anti-inflammatory cytokine production. Overall, these findings highlight lncRNAs as novel host factors to be further investigated as targets for TB diagnostics and adjunctive HDTs. | |
| dc.identifier.apacitation | Pillay, S. (2022). <i>The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection</i>. (). ,Faculty of Health Sciences ,Department of Clinical Laboratory Sciences. Retrieved from http://hdl.handle.net/11427/36719 | en_ZA |
| dc.identifier.chicagocitation | Pillay, Shandré. <i>"The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection."</i> ., ,Faculty of Health Sciences ,Department of Clinical Laboratory Sciences, 2022. http://hdl.handle.net/11427/36719 | en_ZA |
| dc.identifier.citation | Pillay, S. 2022. The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection. . ,Faculty of Health Sciences ,Department of Clinical Laboratory Sciences. http://hdl.handle.net/11427/36719 | en_ZA |
| dc.identifier.ris | TY - Doctoral Thesis AU - Pillay, Shandré AB - In 2020, the World Health Organization (WHO) reported 1.5 million tuberculosis (TB)- associated deaths and an incidence of 10 million new cases. The causative, Mycobacterium tuberculosis (Mtb), evades host immune responses by skewing macrophage polarization towards a less microbicidal alternative state to avoid classical effector killing functions. However, the molecular details underlying these evasion mechanisms remain incomplete and current therapy is challenged with drug resistance. Host-directed therapy (HDT) has recently gained attention, with long non-coding RNAs (lncRNAs) as potential targets due to their emerging roles in pathogenic immune responses. We previously performed cap analysis gene expression (CAGE) transcriptomics on IFN-γ stimulated (classically activated) and IL-4/IL-13 stimulated (alternatively activated) mouse macrophages, identifying 151 differentially expressed lncRNAs following Mtb infection. We validated the top 11 differentially expressed lncRNAs and two were chosen for this study, lncRNA-125, whose expression was regulated at different levels unstimulated and in response to IFN-γ and IL-4/IL-13, and lncRNA-612 whose expression was only induced by IFN-γ stimulation. Interestingly, the expression of lncRNA125 and lncRNA-612 was downregulated following Mtb infection. Therefore, this study aimed at functionally validating these lncRNAs in unstimulated, IFN-γ and IL-4/IL-13 stimulated and/or Mtb-infected mouse and human macrophages by a loss-of-function approach using chemically engineered antisense oligonucleotides (gapmeRs). Knockdown of lncRNA-125 by gapmeRs reduced Mtb growth and anti-inflammatory cytokine production mediated by increased apoptosis, nitrite and pro-inflammatory cytokine production in IL-4/IL-13 prestimulated mouse macrophages. Whereas knockdown of lncRNA-125 in IFN-γ pre-stimulated mouse macrophages favoured Mtb growth and anti-inflammatory cytokine production, with reduction of apoptosis, nitrite and pro-inflammatory cytokine production. Therefore, indicating that lncRNA-125 regulates macrophage polarization during Mtb infection. Knockdown of lncRNA-125 in human macrophages resulted in reduced Mtb growth and increased proinflammatory cytokine production in unstimulated, IFN-γ and IL-4/IL-13 pre-stimulated BMDMs infected with Mtb. Comparatively, gapmeR knockdown of lncRNA-612 reduced Mtb growth and increased pro-inflammatory cytokine production in IFN-γ pre-stimulated mouse and human macrophages. In mouse macrophages, these responses were mediated by increased apoptosis and nitrite production, with reduced anti-inflammatory cytokine production. Overall, these findings highlight lncRNAs as novel host factors to be further investigated as targets for TB diagnostics and adjunctive HDTs. DA - 2022 DB - OpenUCT DP - University of Cape Town KW - clinical laboratory sciences LK - https://open.uct.ac.za PY - 2022 T1 - The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection TI - The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection UR - http://hdl.handle.net/11427/36719 ER - | en_ZA |
| dc.identifier.uri | http://hdl.handle.net/11427/36719 | |
| dc.identifier.vancouvercitation | Pillay S. The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection. []. ,Faculty of Health Sciences ,Department of Clinical Laboratory Sciences, 2022 [cited yyyy month dd]. Available from: http://hdl.handle.net/11427/36719 | en_ZA |
| dc.language.rfc3066 | eng | |
| dc.publisher.department | Department of Clinical Laboratory Sciences | |
| dc.publisher.faculty | Faculty of Health Sciences | |
| dc.subject | clinical laboratory sciences | |
| dc.title | The role and host-directed targeting of long non-coding RNAs in macrophage polarization during Mycobacterium tuberculosis infection | |
| dc.type | Doctoral Thesis | |
| dc.type.qualificationlevel | Doctoral | |
| dc.type.qualificationlevel | PhD |