scholarly journals Regulation of Mycobacterium tuberculosis whiB3 in the Mouse Lung and Macrophages

2006 ◽  
Vol 74 (11) ◽  
pp. 6449-6457 ◽  
Author(s):  
N. Banaiee ◽  
W. R. Jacobs ◽  
J. D. Ernst
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mouse Lung ◽  
Wild Type ◽  
Necrosis Factor Alpha ◽  
Environmental Signals ◽  
Immunodeficient Mice ◽  
Reverse Transcription Pcr ◽  
Factor Alpha

ABSTRACT Mycobacterium tuberculosis is a highly successful human pathogen, with ∼2 × 109 individuals infected globally. To understand the responses of M. tuberculosis to the in vivo environment, we studied the in vivo regulation of M. tuberculosis genes whose M. marinum homologs are induced in chronically infected frog tissues. The expression of 16S rRNA was shown to remain constant in M. tuberculosis under in vivo and in vitro conditions and therefore could be used for internal normalization in quantitative reverse transcription-PCR assays. We found whiB3, a putative transcriptional regulator implicated in mediating tissue damage, to be maximally induced at 2 weeks postinfection in the lungs of wild-type and immunodeficient (gamma interferon receptor−/−, Rag1−/−, and tumor necrosis factor alpha−/−) mice. At later time points in wild-type mice, whiB3 induction was decreased and gradually declined over the course of infection. In immunodeficient mice, whiB3 induction declined rapidly and was completely abolished in moribund animals. whiB3 was also found to be induced in naïve bone marrow-derived macrophages after 6 h of infection. whiB3 expression in vivo and in vitro was found to be inversely correlated with bacterial density. These results indicate that M. tuberculosis regulates the expression of whiB3 in response to environmental signals present in vivo and are consistent with a model of regulation by quorum sensing.

scholarly journals Ecto-5′-Nucleotidase (CD73) Deficiency in Mycobacterium tuberculosis-Infected Mice Enhances Neutrophil Recruitment

2015 ◽  
Vol 83 (9) ◽  
pp. 3666-3674 ◽  
Author(s):  
Laetitia Petit-Jentreau ◽  
Grégory Jouvion ◽  
Patricia Charles ◽  
Laleh Majlessi ◽  
Brigitte Gicquel ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Immune System ◽  
Proinflammatory Response ◽  
Necrosis Factor Alpha ◽  
Effective Response ◽  
Content Type ◽  
Factor Alpha ◽  
Endothelial Growth Factor

The immune system needs safeguards that prevent collateral tissue damage mediated by the immune system while enabling an effective response against a pathogen. The purinergic pathway is one such mechanism and finely modulates inflammation by sensing nucleotides in the environment. Extracellular ATP is considered to be a danger signal leading to a proinflammatory response, whereas adenosine is immunosuppressive. CD73, also called ecto-5′-nucleotidase, occupies a strategic position in this pathway, as it is the main enzyme responsible for the generation of adenosine from ATP. Here, we explore the role of CD73 during tuberculosis, a disease characterized by an immune response that is harmful to the host and unable to eradicateMycobacterium tuberculosis. Using CD73 knockout (KO) mice, we found that CD73 regulates the response toM. tuberculosisinfectionin vitroandin vivo. Mycobacterium-infected murine macrophages derived from CD73 KO mice secrete more keratinocyte chemoattractant (KC), tumor necrosis factor alpha (TNF-α), and interleukin-6 (IL-6) and release less vascular endothelial growth factor (VEGF) upon ATP stimulation than do those derived from wild-type (WT) mice.In vivo, CD73 limits the early influx of neutrophils to the lungs without affecting bacterial growth and dissemination. Collectively, our results support the view that CD73 fine-tunes antimycobacterial immune responses.

scholarly journals Sialyl Lewisx (sLex) and an sLexMimetic, CGP69669A, Disrupt E-Selectin–Dependent Leukocyte Rolling In Vivo

Blood ◽  
1998 ◽  
Vol 91 (2) ◽  
pp. 475-483 ◽  
Author(s):  
Keith E. Norman ◽  
Gary P. Anderson ◽  
Hartmut C. Kolb ◽  
Klaus Ley ◽  
Beat Ernst
Keyword(s):  
Leukocyte Rolling ◽  
Sialyl Lewisx ◽  
Necrosis Factor Alpha ◽  
Rolling Velocity ◽  
Beneficial Effects ◽  
Selectin Family ◽  
Factor Alpha ◽  
Observable Event

Abstract Leukocyte rolling is the earliest observable event in their recruitment from the circulation to inflamed tissue. This rolling is mediated largely by interaction between the selectin family of adhesion molecules and their glycosylated ligands. Although the nature of these ligands and their interaction with the selectins is not fully understood, it is accepted that expression of fucosylated sialylated glycans such as sialyl Lewisx (sLex) is required for function. Despite findings that sLex inhibits binding of leukocytes to E-selectin in vitro, and has beneficial effects in inflammatory disease models, inhibition of E-selectin–dependent leukocyte rolling in vivo has not been described. Functional overlap between the selectins has been noted and reduction of rolling by E-selectin antibodies only occurs if P-selectin is absent or blocked. We demonstrate that leukocyte rolling velocity in tumor necrosis factor alpha (TNFα)-stimulated mouse cremaster is increased following treatment with either sLex or the sLex-mimetic CGP69669A and that rolling is dramatically reduced if CGP69669A is applied in the presence of anti–P-selectin antibody. These effects are characteristic of E-selectin antagonism. In contrast, surgically stimulated (L- or P-selectin–dependent) rolling is unaffected by either sLex or CGP69669A. Our data demonstrate that CGP69669A is an effective and selective antagonist of E-selectin in vivo.

P-selectin glycoprotein ligand-1 supports rolling on E- and P-selectin in vivo

Blood ◽  
2000 ◽  
Vol 96 (10) ◽  
pp. 3585-3591 ◽  
Author(s):  
Keith E. Norman ◽  
Andreas G. Katopodis ◽  
Gebhard Thoma ◽  
Frank Kolbinger ◽  
Anne E. Hicks ◽  
...  
Keyword(s):  
Tumor Necrosis ◽  
Necrosis Factor Alpha ◽  
Rolling Velocity ◽  
Selectin Ligand ◽  
Factor Alpha ◽  
Observable Event ◽  
Slow Rolling ◽  
Necrosis Factor

Abstract Selectin-dependent rolling is the earliest observable event in the recruitment of leukocytes to inflamed tissues. Several glycoproteins decorated with sialic acid, fucose, and/or sulfate have been shown to bind the selectins. The best-characterized selectin ligand is P-selectin glycoprotein-1 (PSGL-1) that supports P-selectin– dependent rolling in vitro and in vivo. In vitro studies have suggested that PSGL-1 may also be a ligand for E- and L-selectins. To study the in vivo function of PSGL-1, without the influence of other leukocyte proteins, the authors observed the interaction of PSGL-1–coated microspheres in mouse venules stimulated to express P- and/or E-selectin. Microspheres coated with functional recombinant PSGL-1 rolled in surgically stimulated and tumor necrosis factor alpha (TNFα)-stimulated mouse venules. P-selectin deficiency or inhibition abolished microsphere rolling in surgically and TNFα-stimulated venules, whereas E-selectin deficiency or inhibition increased microsphere rolling velocity in TNFα-stimulated venules. The results suggest that P-selectin–PSGL-1 interaction alone is sufficient to mediate rolling in vivo and that E-selectin–PSGL-1 interaction supports slow rolling.

scholarly journals Pharmacokinetic andIn VivoEfficacy Studies of the Mycobactin Biosynthesis Inhibitor Salicyl-AMS in Mice

2013 ◽  
Vol 57 (10) ◽  
pp. 5138-5140 ◽  
Author(s):  
Shichun Lun ◽  
Haidan Guo ◽  
John Adamson ◽  
Justin S. Cisar ◽  
Tony D. Davis ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Single Dose ◽  
Iron Acquisition ◽  
Mouse Lung ◽  
Proof Of Concept ◽  
Content Type ◽  
Biosynthesis Inhibitor ◽  
Parameter Values

ABSTRACTMycobactin biosynthesis inMycobacterium tuberculosisfacilitates iron acquisition, which is required for growth and virulence. The mycobactin biosynthesis inhibitor salicyl-AMS [5′-O-(N-salicylsulfamoyl)adenosine] inhibitsM. tuberculosisgrowthin vitrounder iron-limited conditions. Here, we conducted a single-dose pharmacokinetic study and a monotherapy study of salicyl-AMS with mice. Intraperitoneal injection yielded much better pharmacokinetic parameter values than oral administration did. Monotherapy of salicyl-AMS at 5.6 or 16.7 mg/kg significantly inhibitedM. tuberculosisgrowth in the mouse lung, providing the firstin vivoproof of concept for this novel antibacterial strategy.

scholarly journals Absence of cytotoxicity towards microglia of iron oxide (α-Fe2O3) nanorhombohedra

2016 ◽  
Vol 5 (3) ◽  
pp. 836-847 ◽  
Author(s):  
Crystal S. Lewis ◽  
Luisa Torres ◽  
Jeremy T. Miyauchi ◽  
Cyrus Rastegar ◽  
Jonathan M. Patete ◽  
...  
Keyword(s):  
Interleukin 1 ◽  
Necrosis Factor Alpha ◽  
Cytotoxic Effects ◽  
Nanoparticle Exposure ◽  
Cellular Iron ◽  
Therapeutic Setting ◽  
The Central Nervous System ◽  
Factor Alpha

Abstract Understanding the nature of interactions between nanomaterials, such as commercially ubiquitous hematite (α-Fe2O3) nanorhombohedra (N-Rhomb) and biological systems is of critical importance for gaining insight into the practical applicability of nanomaterials. Microglia represent the first line of defense in the central nervous system (CNS) during severe injury or disease such as Parkinson's and Alzheimer's disease as illustrative examples. Hence, to analyze the potential cytotoxic effect of N-Rhomb exposure in the presence of microglia, we have synthesized Rhodamine B (RhB)-labeled α-Fe2O3 N-Rhomb, with lengths of 47 ± 10 nm and widths of 35 ± 8 nm. Internalization of RhB-labeled α-Fe2O3 N-Rhomb by microglia in the mouse brain was observed, and a dose-dependent increase in the cellular iron content as probed by cellular fluorescence was detected in cultured microglia after nanoparticle exposure. The cells maintained clear functional viability, exhibiting little to no cytotoxic effects after 24 and 48 hours at acceptable, physiological concentrations. Importantly, the nanoparticle exposure did not induce microglial cells to produce either tumor necrosis factor alpha (TNFα) or interleukin 1-beta (IL1β), two pro-inflammatory cytokines, nor did exposure stimulate the production of nitrites and reactive oxygen species (ROS), which are common indicators for the onset of inflammation. Finally, we propose that under the conditions of our experiments, i.e. in the presence of RhB labeled-α-Fe2O3 N-Rhomb maintaining concentrations of up to 100 μg mL−1 after 48 hours of incubation, the in vitro and in vivo internalization of RhB-labeled α-Fe2O3 N-Rhomb are likely to be clathrin-dependent, which represents a conventional mechanistic uptake route for most cells. Given the crucial role that microglia play in many neurological disorders, understanding the potential cytotoxic effects of these nanostructures is of fundamental importance if they are to be used in a therapeutic setting.

scholarly journals A Thiolase of Mycobacterium tuberculosis Is Required for Virulence and Production of Androstenedione and Androstadienedione from Cholesterol

2009 ◽  
Vol 78 (1) ◽  
pp. 275-282 ◽  
Author(s):  
Natasha M. Nesbitt ◽  
Xinxin Yang ◽  
Patricia Fontán ◽  
Irina Kolesnikova ◽  
Issar Smith ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mutant Strain ◽  
Wild Type Strain ◽  
Cholesterol Metabolism ◽  
Transcriptional Profiling ◽  
Mouse Lung ◽  
Intracellular Pathogen ◽  
Wild Type ◽  
Strain H37rv

ABSTRACT Mycobacterium tuberculosis, the causative agent of tuberculosis, is an intracellular pathogen that shifts to a lipid-based metabolism in the host. Moreover, metabolism of the host lipid cholesterol plays an important role in M. tuberculosis infection. We used transcriptional profiling to identify genes transcriptionally regulated by cholesterol and KstR (Rv3574), a TetR-like repressor. The fadA5 (Rv3546) gene, annotated as a lipid-metabolizing thiolase, the expression of which is upregulated by cholesterol and repressed by KstR, was deleted in M. tuberculosis H37Rv. We demonstrated that fadA5 is required for utilization of cholesterol as a sole carbon source in vitro and for full virulence of M. tuberculosis in the chronic stage of mouse lung infection. Cholesterol is not toxic to the fadA5 mutant strain, and, therefore, toxicity does not account for its attenuation. We show that the wild-type strain, H37Rv, metabolizes cholesterol to androst-4-ene-3,17-dione (AD) and androsta-1,4-diene-3,17-dione (ADD) and exports these metabolites into the medium, whereas the fadA5 mutant strain is defective for this activity. We demonstrate that FadA5 catalyzes the thiolysis of acetoacetyl-coenzyme A (CoA). This catalytic activity is consistent with a β-ketoacyl-CoA thiolase function in cholesterol β-oxidation that is required for the production of androsterones. We conclude that the attenuated phenotype of the fadA5 mutant is a consequence of disrupted cholesterol metabolism that is essential only in the persistent stage of M. tuberculosis infection and may be caused by the inability to produce AD/ADD from cholesterol.

scholarly journals Deletion of the Mycobacterium tuberculosis α-Crystallin-Like hspX Gene Causes Increased Bacterial Growth In Vivo

2006 ◽  
Vol 74 (2) ◽  
pp. 861-868 ◽  
Author(s):  
Yanmin Hu ◽  
Farahnaz Movahedzadeh ◽  
Neil G. Stoker ◽  
Anthony R. M. Coates
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Constitutive Expression ◽  
Active Role ◽  
Structural Genes ◽  
Wild Type ◽  
Major Antigen ◽  
Oxide Stress ◽  
Unusual Phenotype

ABSTRACT Hypervirulent mutants of Mycobacterium tuberculosis, whose growth rates are higher in vivo, have now been reported to have mutations in both regulatory and structural genes, but the basis for this unusual phenotype is not understood. One hypervirulence gene, dosR (devR, Rv2031c), activates transcription of approximately 50 genes in this pathogen in response to hypoxia and nitric oxide stress. The most dramatic activation (∼80-fold) is activation of the hspX (acr, Rv2031c) gene, which encodes a 16-kDa α-crystallin-like protein that is a major antigen. In this study we found that a Δacr mutant exhibited increased growth following infection of BALB/c mice in vivo and in both resting and activated macrophages in vitro (as measured by the number of CFU). The increased growth in macrophages was equal to that of a ΔdosR mutant, while introduction of a constitutively expressed hspX gene reduced the ΔdosR virulence to wild-type levels. These results suggest that the increased number of CFU of the ΔdosR mutant was largely due to loss of hspX expression. We also confirmed that constitutive expression of hspX slows growth in vitro, and we propose that hspX plays an active role in slowing the growth of M. tuberculosis in vivo immediately following infection.

scholarly journals Construction and Phenotypic Characterization of an Auxotrophic Mutant of Mycobacterium tuberculosis Defective in l-Arginine Biosynthesis

2002 ◽  
Vol 70 (6) ◽  
pp. 3080-3084 ◽  
Author(s):  
Bhavna G. Gordhan ◽  
Debbie A. Smith ◽  
Heidi Alderton ◽  
Ruth A. McAdam ◽  
Gregory J. Bancroft ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mutant Strain ◽  
Auxotrophic Mutant ◽  
Phenotypic Characterization ◽  
Wild Type ◽  
Arginine Biosynthesis ◽  
High Concentrations

ABSTRACT A mutant of Mycobacterium tuberculosis defective in the metabolism of l-arginine was constructed by allelic exchange mutagenesis. The argF mutant strain required exogenous l-arginine for growth in vitro, and in the presence of 0.96 mM l-arginine, it achieved a growth rate and cell density in stationary phase comparable to those of the wild type. The mutant strain was also able to grow in the presence of high concentrations of argininosuccinate, but its auxotrophic phenotype could not be rescued by l-citrulline, suggesting that the ΔargF::hyg mutation exerted a polar effect on the downstream argG gene but not on argH. The mutant strain displayed reduced virulence in immunodeficient SCID mice and was highly attenuated in immunocompetent DBA/2 mice, suggesting that l-arginine availability is restricted in vivo.

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