Tuberculosis-infected Macrophages Research Articles

The Role of Hypoxia-inducible Factor in Mycobacterium tuberculosis-infected Macrophages

Tuberculosis is caused by Mycobacterium tuberculosis infection. During M. tuberculosis infection, there is a decrease in the partial pressure of oxygen in the granuloma microenvironment, which causes the hypoxia-inducible factor (HIF) to become stable. HIF functions as a transcription factor that regulates the expression of genes crucial for metabolic adaptation in hypoxic conditions. Recent research suggests that HIF plays a vital role in infectious and inflammatory conditions. Several studies have demonstrated that HIF signaling can enhance macrophages antimicrobial activity and bactericidal effect against M. tuberculosis, such as increasing macrophage autophagy, enhancing the effects of rifampicin, inhibiting p38 MAPK signaling, enhancing the regulation of effector antimicrobial pathways mediated by human β defensin 2 (hBD2) and vitamin D receptor (VDR), redirecting energy metabolism to glycolysis, and producing various cytokines. All these responses ultimately result in the inhibition of intracellular M. tuberculosis growth. HIF has therapeutic implications, potentially being a new candidate for host-directed therapy as a complement to existing antituberculosis drugs. Understanding the role of HIF in macrophages during M. tuberculosis infection and comprehending the host-pathogen relationship with M. tuberculosis is advantageous for developing future therapies.Keywords: Mycobacterium tuberculosis, macrophages, hypoxia-inducible factor

YY1 Contributes to the Inflammatory Responses of Mycobacterium tuberculosis-Infected Macrophages Through Transcription Activation-Mediated Upregulation TLR4.

Tuberculosis (TB) is a chronic respiratory infectious disease and is induced by Mycobacterium tuberculosis (M.tb) infection. Macrophages serve as the cellular home in immunoreaction against M.tb infection, which is tightly regulated through Toll-like receptor 4 (TLR4) expression. Therefore, this study is designed to explore the role and mechanism of TLR4 in mycobacterial injury in human macrophages (THP-1 cells) after M.tb infection. Cell proliferation and apoptosis were assessed using MTT, EdU, and flow cytometry assays. ELISA kits were utilized to assess the levels of Interleukin-6 (IL-6), IL-1β, and tumor necrosis factor α (TNF-α). The binding between Yin-Yang-1 (YY1) and TLR4 promoter was predicted by JASPAR and verified using Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays. M.tbinfection might repress THP-1 cell proliferation, and induce cell apoptosis and inflammatory response in a multiplicity of infection (MOI)-dependent manner. Moreover, M.tbinfection increasedtheexpressionofTLR4 inHTP-1cellsinanMOI-dependent way, and its downregulation might overturn M.tbinfection-mediated HTP-1cell damage and inflammatory response. At the molecular level, YY1 was a transcription factor of TLR4 and promoted TLR4 transcription via binding to its promoter region. Besides, YY1 might activate the NF-kB signaling pathway via regulating TLR4. Meanwhile, TLR4 inhibitor BAY11-7082 might overturn the repression effect of TLR4 on M.tb-infected HTP-1cell damage. YY1-activated TLR4 might aggravate mycobacterial injury in human macrophages after M.tb infection by the NF-kB pathway, providing a promising therapeutic target for TB treatment.

Discovery of 2,9-diaryl-6-carbamoylpurines as a novel class of antitubercular agents

A series of novel 9-alkyl/aryl-2-aryl-6-carbamoylpurines were synthesized, and their activity against Mycobacterium tuberculosis strain H37Rv was assessed. The SAR analysis on the first set of derivatives, with an alkyl or aryl unit at N-9 and a phenolic unit at C-2, showed that the activity depends on the purine ring substituents at N-9 and C-2. A phenyl group at N-9 combined with a 3-hydroxyphenyl or 4-hydroxyphenyl at C-2 improve the activity. The most active compound of this set has a phenyl group at N-9 and a 4-hydroxyphenyl group at C-2, displaying an IC90 = 1.2 μg/mL and a selectivity index higher than 25.5. This compound served as a Hit to design the second set of derivatives. A phenyl group at N-9 was maintained, and the group at C-2 was diversified. The SAR analysis showed that the aryl unit at C-2 must have an oxygen or nitrogen atom bonded in the para position. A proton, a small alkyl or a substituted aryl group may also be bonded to the oxygen. The compound with the 4-methoxyphenyl group at C-2, 1Bd, exhibits the highest activity with an IC90 < 0.19 μg/mL. This compound is highly potent against M. tuberculosis strain H37Rv and non-toxic for VERO mammalian cells with an SI > 153.8. Compound 1Bd was also non-cytotoxic against primary macrophage cultures at IC90, 2xIC90, and 10xIC90 and significantly reduced the bacterial load in M. tuberculosis-infected macrophages at the same concentrations. Compound 1Bd showed a favorable pharmacokinetic profile when administered orally, with major lung and liver accumulation. In vivo antimycobacterial efficacy of 1Bd was tested at 25 mg/kg. At the tested regimen, a decrease in bacterial burden was observed in the liver. Optimization of the treatment regimen should be performed to fully potentiate the in vivo efficacy of our lead molecule, particularly in the lung, the main target organ of M. tuberculosis.

Triggering Receptor Expressed on Myeloid Cells 2 Mediates the Involvement of M2-Type Macrophages in Pulmonary Tuberculosis Infection.

Macrophage play a significant work in the development of tuberculosis. This study aims to investigate the relationship between TREM2 and macrophage polarization, as well as the related cytokines. This study involved 43 pulmonary tuberculosis patients and 37 healthy controls. Enzyme-linked immunosorbent assay (ELISA) was used to detect the expression levels of M1/M2 macrophage-related cytokines IL-10 and IL-12 in the peripheral blood of pulmonary tuberculosis patients. The relative mRNA expression levels of TREM2, IL-10 and IL-12 were detected using quantitative real-time PCR (qRT-PCR). Additionally, Spearman rank correlation analysis was used to preliminarily assess the correlation between TREM2 and M1 / M2 macrophages. Hematoxylin-eosin (HE) staining was performed to observe the pathological manifestations of pulmonary tuberculosis lesions. Immunohistochemical (IHC) staining was used to observe the localization of the macrophage-specific molecule CD68, the M1 specific molecule iNOS, the M2 specific molecule CD163, and TREM2. The lesions of pulmonary tuberculosis patients showed Langhans multinucleated macrophages and tuberculous granulomas. The ELISA results indicated that the expression levels of IL-10 and IL-12 were significantly increased in peripheral blood of pulmonary tuberculosis patients. Additionally, the relative mRNA expression levels of TREM2, IL-10 and IL-12 were also significantly higher in the pulmonary tuberculosis group. Furthermore, a positive correlation was observed between TREM2 and IL-10, which are secreted by M2 macrophages. IHC revealed significant positivity of TREM2 and macrophage-related markers in tuberculous granuloma. Specifically, TREM2 and M2 macrophage marker CD163 were significantly expressed in the cytoplasm and membrane of Langhans multinucleated macrophages. The role of macrophage polarization in pulmonary tuberculosis is significant, and further investigation is needed to understand relationship between TREM2 and M2 macrophages.

ZNFX1 promotes AMPK-mediated autophagy against Mycobacterium tuberculosis by stabilizing Prkaa2 mRNA.

Tuberculosis has the highest mortality rate worldwide for a chronic infectious disease caused by a single pathogen. RNA-binding proteins (RBPs) are involved in autophagy - a key defense mechanism against Mycobacterium tuberculosis (M. tuberculosis) infection - by modulating RNA stability and forming intricate regulatory networks. However, the functions of host RBPs during M. tuberculosis infection remain relatively unexplored. Zinc finger NFX1-type containing 1 (ZNFX1), a conserved RBP critically involved in immune deficiency diseases and mycobacterial infections, is significantly upregulated in M. tuberculosis-infected macrophages. Here, we aimed to explore the immunoregulatory functions of ZNFX1 during M. tuberculosis infection. We observed that Znfx1 knockout markedly compromised the multifaceted immune responses mediated by macrophages. This compromise resulted in reduced phagocytosis, suppressed macrophage activation, increased M. tuberculosis burden, progressive lung tissue injury, and chronic inflammation in M. tuberculosis-infected mice. Mechanistic investigations revealed that the absence of ZNFX1 inhibited autophagy, consequently mediating immune suppression. ZNFX1 critically maintained AMPK-regulated autophagic flux by stabilizing protein kinase AMP-activated catalytic subunit alpha 2 mRNA, which encodes a key catalytic α subunit of AMPK, through its zinc finger region. This process contributed to M. tuberculosis growth suppression. These findings reveal a function of ZNFX1 in establishing anti-M. tuberculosis immune responses, enhancing our understanding of the roles of RBPs in tuberculosis immunity and providing a promising approach to bolster antituberculosis immunotherapy.

Metabolic flux reprogramming in Mycobacterium tuberculosis-infected human macrophages.

Metabolic fluxes are at the heart of metabolism and growth in any living system. During tuberculosis (TB) infection, the pathogenic Mycobacterium tuberculosis (Mtb) adapts its nutritional behaviour and metabolic fluxes to survive in human macrophages and cause infection. The infected host cells also undergo metabolic changes. However, our knowledge of the infected host metabolism and identification of the reprogrammed metabolic flux nodes remains limited. In this study, we applied systems-based 13C-metabolic flux analysis (MFA) to measure intracellular carbon metabolic fluxes in Mtb-infected human THP-1 macrophages. We provide a flux map for infected macrophages that quantified significantly increased fluxes through glycolytic fluxes towards pyruvate synthesis and reduced pentose phosphate pathway fluxes when compared to uninfected macrophages. The tri carboxylic acid (TCA) cycle fluxes were relatively low, and amino acid fluxes were reprogrammed upon Mtb infection. The knowledge of host metabolic flux profiles derived from our work expands on how the host cell adapts its carbon metabolism in response to Mtb infection and highlights important nodes that may provide targets for developing new therapeutics to improve TB treatment.

Herp regulates intracellular survival of Mycobacterium tuberculosis H37Ra in macrophages by regulating reactive oxygen species-mediated autophagy.

Several studies have suggested that endoplasmic reticulum (ER) stress is important in the pathogenesis of infectious diseases; however, the precise function of ER stress regulation and the role of Herp as a regulator in Mtb H37Ra-induced ER stress remain elusive. Therefore, our study investigated ER stress and autophagy associated with Herp expression in Mycobacterium tuberculosis-infected macrophages to determine the role of Herp in the pathogenesis of tuberculosis.

Ursolic Acid Promotes Autophagy by Inhibiting Akt/mTOR and TNF-α/TNFR1 Signaling Pathways to Alleviate Pyroptosis and Necroptosis in Mycobacterium tuberculosis-Infected Macrophages.

As a lethal infectious disease, tuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb). Its complex pathophysiological process limits the effectiveness of many clinical treatments. By regulating host cell death, Mtb manipulates macrophages, the first line of defense against invading pathogens, to evade host immunity and promote the spread of bacteria and intracellular inflammatory substances to neighboring cells, resulting in widespread chronic inflammation and persistent lung damage. Autophagy, a metabolic pathway by which cells protect themselves, has been shown to fight intracellular microorganisms, such as Mtb, and they also play a crucial role in regulating cell survival and death. Therefore, host-directed therapy (HDT) based on antimicrobial and anti-inflammatory interventions is a pivotal adjunct to current TB treatment, enhancing anti-TB efficacy. In the present study, we showed that a secondary plant metabolite, ursolic acid (UA), inhibited Mtb-induced pyroptosis and necroptosis of macrophages. In addition, UA induced macrophage autophagy and enhanced intracellular killing of Mtb. To investigate the underlying molecular mechanisms, we explored the signaling pathways associated with autophagy as well as cell death. The results showed that UA could synergistically inhibit the Akt/mTOR and TNF-α/TNFR1 signaling pathways and promote autophagy, thus achieving its regulatory effects on pyroptosis and necroptosis of macrophages. Collectively, UA could be a potential adjuvant drug for host-targeted anti-TB therapy, as it could effectively inhibit pyroptosis and necroptosis of macrophages and counteract the excessive inflammatory response caused by Mtb-infected macrophages via modulating the host immune response, potentially improving clinical outcomes.

Cisatracurium besylate rescues Mycobacterium Tuberculosis-infected macrophages from necroptosis and enhances the bactericidal effect of isoniazid

ObjectiveTuberculosis is the leading killer among the chronic single-source infectious diseases. Mycobacterium tuberculosis can induce necrotic-dominant multiple modes of cell death in macrophages, which accelerates bacterium dissemination and expands tissue injury in host lungs. Mining drugs to counteract Mycobacterium tuberculosis-induced cell death would be beneficial to tuberculosis patients. MethodsIn this study, the protective drug was screened out from the FDA-approved drug library in Mycobacterium tuberculosis-infected macrophages with CCK-8 assay. The death mode regulated by the drug was identified using transcriptomic sequencing, cytomorphological observation, and in the experimental mouse Mycobacterium tuberculosis-infection model. The functional mechanism was explored using western blot, co-immunoprecipitation, and DARTS assay. The intracellular bacterial survival was detected using colony forming unit assays. ResultsCisatracurium besylate was identified to be highly protective for the viability of macrophages during Mycobacterium tuberculosis infection via inhibiting necroptosis. Cisatracurium besylate prevented RIPK3 to be associated with the executive molecule MLKL for forming the necroptotic complex, resulting in the inhibition of MLKL phosphorylation and pore formation on cell membrane. However, Cisatracurium besylate did not interfere with the association between RIPK3 with its upstream kinase RIPK1 or ZBP1 but regulated RIPK3 autophosphorylation. Moreover, Cisatracurium besylate significantly inhibited the expansion of intracellular Mycobacterium tuberculosis both in vitro and in vivo, which also displayed a strong auxiliary bacteriostatic effect to support the therapeutic efficacy of isoniazid and rifampicin, the first-line anti-tubercular drugs. ConclusionCisatracurium besylate performs anti-Mycobacterium tuberculosis and anti-necroptotic roles, which potentiates its application to be an adjuvant drug for antituberculosis therapy to assist the battle against drug-resistant tuberculosis.

Baicalein Suppresses NLRP3 and AIM2 Inflammasome-Mediated Pyroptosis in Macrophages Infected by Mycobacterium tuberculosis via Induced Autophagy.

Mycobacterium tuberculosis (Mtb) continues to pose a significant threat to global health because it causes granulomas and systemic inflammatory responses during active tuberculosis (TB). Mtb can induce macrophage pyroptosis, which results in the release of IL-1β and causes tissue damage, thereby promoting its spread. In the absence of anti-TB drugs, host-directed therapy (HDT) has been demonstrated to be an effective strategy against TB. In this study, we used an in vitro Mtb-infected macrophage model to assess the effect of baicalein, derived from Scutellariae radix, on pyroptosis induced in Mtb-infected macrophages. Further, we investigated the molecular mechanisms underlying the actions of baicalein. The results of the study suggest that baicalein inhibits pyroptosis in Mtb-infected macrophages by downregulating the assembly of AIM2 and NLRP3 inflammasome and promoting autophagy. Further research has also shown that the mechanism by which baicalein promotes autophagy may involve the inhibition of the activation of the Akt/mTOR pathway and the inhibition of the AIM2 protein, which affects the levels of CHMP2A protein required to promote autophagy. Thus, our data show that baicalein can inhibit Mtb infection-induced macrophage pyroptosis and has the potential to be a new adjunctive HDT drug. IMPORTANCE Current strategies for treating drug-resistant tuberculosis have limited efficacy and undesirable side effects; hence, research on new treatments, including innovative medications, is required. Host-directed therapy (HDT) has emerged as a viable strategy for modulating host cell responses in order to enhance protective immunity against infections. Baicalein, extracted from Scutellariae radix, was shown to inhibit pyroptosis caused by Mycobacterium tuberculosis-infected macrophages and was associated with autophagy. Our findings reveal that baicalein can be used as an adjunctive treatment for tuberculosis or other inflammatory diseases by regulating immune function and enhancing the antibacterial ability of the host. It also provides a new idea for exploring the anti-inflammatory mechanism of baicalein.

High Bacillary Burden and the ESX-1 Type VII Secretion System Promote MHC Class I Presentation by Mycobacterium tuberculosis-Infected Macrophages to CD8 T Cells.

We used a mouse model to study how Mycobacterium tuberculosis subverts host defenses to persist in macrophages despite immune pressure. CD4 T cells can recognize macrophages infected with a single bacillus invitro. Under identical conditions, CD8 T cells inefficiently recognize infected macrophages and fail to restrict M. tuberculosis growth, although they can inhibit M. tuberculosis growth during high-burden intracellular infection. We show that high intracellular M. tuberculosis numbers cause macrophage death, leading other macrophages to scavenge cellular debris and cross-present the TB10.4 Ag to CD8 T cells. Presentation by infected macrophages requires M. tuberculosis to have a functional ESX-1 type VII secretion system. These data indicate that phagosomal membrane damage and cell death promote MHC class I presentation of the immunodominant Ag TB10.4 by macrophages. Although this mode of Ag presentation stimulates cytokine production that we presume would be host beneficial, killing of uninfected cells could worsen immunopathology. We suggest that shifting the focus of CD8 T cell recognition to uninfected macrophages would limit the interaction of CD8 T cells with infected macrophages and impair CD8 T cell-mediated resolution of tuberculosis.

Involvement of 2′-5′ oligoadenylate synthetase-like protein in the survival of Mycobacterium tuberculosis avirulent strain in macrophages

Mycobacterium tuberculosis (M. tuberculosis) can replicate in the macrophage by interfering with many host protein functions. While it is far from known these host proteins for controlling M. tuberculosis infection. Herein, we infected macrophages including THP-1 and Raw264.7 cells with M. tuberculosis and identified the differentially expressed genes (DEGs) in the interferon signaling pathway. Among them, 2′-5′ oligoadenylate synthetase-like (OASL) underwent the greatest upregulation in M. tuberculosis-infected macrophages. Knockdown of the expression of OASL attenuated M. tuberculosis survival in macrophages. Further, bioinformatics analysis revealed the potential interaction axis of OASL-TAB3- Rv0127, which was further validated by the yeast-two-hybrid (Y2H) assay and Co-IP. This interaction axis might regulate the M. tuberculosis survival and proliferation in macrophages. The study reveals a possible role of OASL during M. tuberculosis infection as a target to control its propagation.

Intracellular Activity of Poly (DL-Lactide-co-Glycolide) Nanoparticles Encapsulated with Prothionamide, Pyrazinamide, Levofloxacin, Linezolid, or Ethambutol on Multidrug-Resistant Mycobacterium tuberculosis.

Multidrug-resistant Mycobacterium tuberculosis (MDR-TB) is a major cause of death amongst tuberculosis patients. Nanomedicine avoids some limitations of conventional drug treatment and increases therapeutic efficacy against bacterial infections. However, the effect of anti-TB drug nanoparticle (NP) compounds in anti-TB regimens against MDR-TB remains unclear. The objective of this article is to prepare levofloxacin, linezolid, ethambutol, prothionamide, and pyrazinamide encapsulated NPs and to evaluate their therapeutic efficacy against MDR-TB in macrophages. Drug-loaded PLGA NPs were prepared by the multiple emulsion method. The colocalization, intracellular release, and anti-TB activity of these NPs were investigated on cultured macrophages. The immune phenotype of the macrophages, including their mitochondrial membrane potential, reactive oxygen species (ROS), and nitric oxide (NO) production, was evaluated following treatment with NPs or free drug compounds. All drug-loaded PLGA NPs were spherical in shape, 150 to 210 nm in size, and showed 14.22% to 43.51% encapsulation efficiencies and long-duration release. Drug-loaded PLGA NPs were mainly distributed in the cytoplasm of macrophages, showed high cellular compatibility, and maintained their concentration for at least 13 days. Compared with the free drug compounds, the number of colonies after exposure to PLGA NP compounds was significantly less. The enhanced antibacterial activity of the NP compounds may be due to the enhanced levels of ROS and NO and the increased early apoptosis stress within M. tuberculosis-infected macrophages additionally. The application of PLGA NP compounds not only enhances drug efficacy but also induces innate bactericidal events in macrophages, confirming this as a promising approach for MDR-TB therapy.

IL-27 inhibits anti- Mycobacterium tuberculosis innate immune activity of primary human macrophages

Mycobacterium tuberculosis (M. tuberculosis) is an intracellular pathogen that primarily infects macrophages. Despite a robust anti-mycobacterial response, many times macrophages are unable to control M. tuberculosis. The purpose of this study was to investigate the mechanism by which the immunoregulatory cytokine IL-27 inhibits the anti-mycobacterial activity of primary human macrophages. We found concerted production of IL-27 and anti-mycobacterial cytokines by M. tuberculosis-infected macrophages in a toll-like receptor (TLR) dependent manner. Notably, IL-27 suppressed the production of anti-mycobacterial cytokines TNFα, IL-6, IL-1β, and IL-15 by M. tuberculosis-infected macrophages. IL-27 limits the anti-mycobacterial activity of macrophages by reducing Cyp27B, cathelicidin (LL-37), LC3B lipidation, and increasing IL-10 production. Furthermore, neutralizing both IL-27 and IL-10 increased the expression of proteins involved in LC3-associated phagocytosis (LAP) pathway for bacterial clearance, namely vacuolar-ATPase, NOX2, and RUN-domain containing protein RUBCN. These results implicate IL-27 is a prominent cytokine that impedes M. tuberculosis clearance.

Artemisinin promotes apoptosis of spinal tuberculosis macrophages by inhibiting NF-κB

Artemisinin has been proved to have a wide range of pharmacological effects, mainly used in anti-malaria, anti-inflammatory, antiviral, etc., but the effect and mechanism of artemisinin on tuberculosis-infected macrophages have not yet been elucidated. THP-1 cell line was cultured and randomly divided into the following 4 groups; control group, which was infected with Mtb standard human virulence strain H37Rv; low-dose, medium-dose and high-dose artemisinin groups, which were treated with artemisinin at 50 μM, 150 μM and 300 μM, respectively. The effect of artemisinin on cell proliferation was determined by MTT assay. MMP-2 and MMP-9 were detected by PCR. The inflammatory cytokines IL-6 and TNF-α were detected by ELISA. The above tests were used to understand apoptosis, inflammation and proliferation. The expressions of Bcl-2, osteoprotectin (OPG), nuclear factor κB receptor activator (RANK) and NF-κB were detected by Western blot. Acting on macrophages infected with mycobacterium tuberculosis, artemisinin can significantly inhibit cell proliferation, promote cell apoptosis, inhibit the expression of MMP-2 and MMP-9, and reduce the secretion of IL-6 and TNF-α. Artemisinin also decreased the expression of Bcl-2, OPG, RANK and NF-κB (p &lt; 0.05), and the change was more significant with the increase of dose. Artemisinin could down-regulate the expression of OPG, RANK and NF-κB, and inhibiting the expression of inflammatory factors and Matrix metalloproteinases (MMPs), and then promoting apoptosis of spinal tuberculosis macrophages.

Pathogenesis of Human Immunodeficiency Virus and Mycobacterium tuberculosis Infection as Revealed by Transcriptome and Interactome Data.

Tuberculosis (TB) among patients with human immunodeficiency virus (HIV) is a major global health burden and contributes to a high mortality rate due to HIV-mediated immunosuppression and subsequent susceptibility to TB. It is imperative to understand the pathogenesis of the association between HIV and TB for therapeutic innovation and preventive medicine. In the present study, we employed transcriptomic and bioinformatic analyses of differential gene expression data obtained from Gene Expression Omnibus (GEO) of the National Center for Biotechnology Information. The expression data of Mycobacterium tuberculosis-infected macrophages and blood samples from TB patients (GSE54992, GSE52819, and GSE19435) and blood samples from HIV patients (GSE30310) were accessed for identification of differentially expressed genes (DEGs). Data from 20 healthy subjects and 19 patients with TB and 16 healthy subjects and 16 patients with HIV were analyzed. We report here the DEGs shared by HIV and TB infection. Moreover, HIV and TB host-pathogen interaction data were collected from BIOGRID, v 4.4.210, for identifying significantly modulated genes' targets and their interactions with the host. Host targets, including PLSCR1 (phospholipid scramblase 1), STAT1 (signal transducer and activator of transcription-1 alpha/beta), FBXO6 (F-box only protein 6), ITGAL (integrin alpha-L), and APP (amyloid beta precursor protein), are commonly modulated in both diseases. The function of these targets was screened from and reconciled with the literature to understand their role in the pathogenesis of HIV and TB. Overall, the study results suggest that these targets may potentially be important contributors to the pathogenesis of this comorbidity. Further experimental work is needed for evaluating these new observations, with a view to future therapeutic innovation for patients with HIV and TB.

Transcription factor MAFB controls type I and II interferon response-mediated host immunity in Mycobacterium tuberculosis-infected macrophages.

MAFB, v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B, has been identified as a candidate gene for early tuberculosis (TB) onset in Thai and Japanese populations. Here, we investigated the genome-wide transcriptional profiles of MAFB-knockdown (KD) macrophages infected with Mycobacterium tuberculosis (Mtb) to highlight the potential role of MAFB in host immunity against TB. Gene expression analysis revealed impaired type I and type II interferon (IFN) responses and enhanced oxidative phosphorylation in MAFB-KD macrophages infected with Mtb. The expression of inflammatory chemokines, including IFN-γ-inducible genes, was confirmed to be significantly reduced by knockdown of MAFB during Mtb infection. A similar effect of MAFB knockdown on type I and type II IFN responses and oxidative phosphorylation was also observed when Mtb-infected macrophages were activated by IFN-γ. Taken together, our results demonstrate that MAFB is involved in the immune response and metabolism in Mtb-infected macrophages, providing new insight into MAFB as a candidate gene to guide further study to control TB.

Lyl1-deficiency promotes inflammatory responses and increases mycobacterial burden in response to Mycobacterium tuberculosis infection in mice.

Lymphoblastic leukemia 1 (Lyl1) is a well-studied transcription factor known to exhibit oncogenic potential in various forms of leukemia with pivotal roles in hematopoietic stem cell biology. While its role in early hematopoiesis is well established, its function in mature innate cells is less explored. Here, we identified Lyl1 as a drastically perturbed gene in the Mycobacterium tuberculosis (Mtb) infected mouse macrophage transcriptome. We report that Lyl1 downregulation upon immune stimulation is a host-driven process regulated by NFκB and MAP kinase pathways. Interestingly, Lyl1-deficient macrophages have decreased bacterial killing potential with reduced nitric oxide (NO) levels while expressing increased levels of pro-inflammatory interleukin-1 and CXCL1. Lyl1-deficient mice show reduced survival to Mtb HN878 infection with increased bacterial burden and exacerbated inflammatory responses in chronic stages. We observed that increased susceptibility to infection was accompanied by increased neutrophil recruitment and IL-1, CXCL1, and CXCL5 levels in the lung homogenates. Collectively, these results suggest that Lyl1 controls Mtb growth, reduces neutrophilic inflammation and reveals an underappreciated role for Lyl1 in innate immune responses.

Long non-coding RNA SNHG16 silencing inhibits proliferation and inflammation in Mycobacterium tuberculosis-infected macrophages by targeting miR-140-5p expression.

The study investigated the clinical diagnostic value of long non-coding RNA (LncRNA) small nucleolar RNA host gene 16 (SNHG16) and explored its underlying molecular mechanism through Mycobacterium tuberculosis (M. tuberculosiinfection of macrophages. RT-qPCR analysis of the serum SNHG16 levels of the 66 healthy individuals, 67 latent TB (LTB) patients, and 67 active TB (ATB) patients. The receiver-operating characteristic (ROC) curve to detect the clinical diagnostic value of SNHG16 in TB patients. In vitro, M. tuberculosis-infected macrophages, CCK-8 and ELISA to detect cell proliferation and inflammatory factor levels. Luciferase reported assay was performed to analyze the targeting relationship between SNHG16 and miR-140-5p. SNHG16 was significantly elevated in TB patients, and among them, ATB patients were higher than LTB patients. ROC confirmed that SNHG16 could distinguish LTB patients from healthy controls, and ATB patients from LTB patients, and can be used as a good diagnostic biomarker for TB. M. tuberculosis infection increased SNHG16 levels and promoted the proliferation and inflammation in macrophages. However, SNHG16 silencing significantly reversed the effect of infection. miR-140-5p, a direct target miRNA of SNHG16, was down-regulated in TB patients and was negatively correlated with SNHG16. When miR-140-5p was inhibited, the alleviating effect of SNHG16 silencing on M. tuberculosis infection proliferation and inflammation was significantly reversed. The present results suggested that SNHG16 may be a new diagnostic biomarker for TB patients and SNHG16 silencing may alleviate TB by inhibiting the proliferation of macrophages in TB by regulation miR-140-5p.

CD44 receptor targeted nanoparticles augment immunity against tuberculosis in mice.

We describe a role of CD44-mediated signaling during host-defense against tuberculosis (TB) using a mouse model of TB and studies in M. tuberculosis (Mtb) infected human macrophage (MФ). Liposomes targeting CD44 using thioaptamers (CD44TA-LIP) were designed and tested as new vaccines to boost host immunity in TB. CD44TA-LIP enhanced killing of Mtb in human MФ, which correlated with an increased production of pro-inflammatory cytokines IL-1β, TNF-α and IL-12. CD44TA-LIP activated MФ showed an enhanced MHC-II dependent antigen presentation to CD4 T-cells. Inhibition of cellular proliferation and cytoskeleton rearrangement pathways downstream of CD44 signaling abrogated CD44TA-LIP-induced antimicrobial effects. Blockade of inflammatory pathways also reduced antigen presentation by MФ and activation of CD4 T cells. Mtb infected MФ treated with CD44TA-LIP exhibited increased nitric oxide and HβD2 defensin peptide production. Among Mtb infected mice with increased lung and spleen loads of organisms, intranasal administration of CD44TA-LIP led to a ten-fold reduction of colony forming units of Mtb and elevated IFN-γ+CD4, effector, central and resident memory T cells. Biodistribution studies demonstrated that CD44TA-LIP preferentially accumulated in the lungs and were associated with CD11b+cells. CD44TA-LIP treated mice showed no weight loss or increased liver LDH levels. This study highlights the importance of CD44-mediated signaling in host-defense during TB and the therapeutic potential of CD44TA-LIP.