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Mycobacterium tuberculosis-infected human monocytes down-regulate microglial MMP-2 secretion in CNS tuberculosis via TNFα, NFκB, p38 and caspase 8 dependent pathways.

Green JA, Dholakia S, Janczar K, Ong CW, Moores R, Fry J, Elkington PT, Roncaroli F, Friedland JS - J Neuroinflammation (2011)

Bottom Line: Inhibition of the p38 MAP kinase pathway resulted in a 228% increase in MMP-2 secretion (P < 0.01).In contrast ERK MAP kinase inhibition further decreased MMP-2 secretion by 76.6% (P < 0.05).Inhibition of the NFκB pathway resulted in 301% higher MMP-2 secretion than CoMTb alone (P < 0.01).

View Article: PubMed Central - HTML - PubMed

Affiliation: Section of Infectious Diseases and Immunity and the Imperial College Wellcome Trust Centre for Clinical Tropical Medicine, Hammersmith Campus, Imperial College London, London, W12 0NN, UK. justin.green@imperial.ac.uk

ABSTRACT
Tuberculosis (TB) of the central nervous system (CNS) is a deadly disease characterized by extensive tissue destruction, driven by molecules such as Matrix Metalloproteinase-2 (MMP-2) which targets CNS-specific substrates. In a simplified cellular model of CNS TB, we demonstrated that conditioned medium from Mycobacterium tuberculosis-infected primary human monocytes (CoMTb), but not direct infection, unexpectedly down-regulates constitutive microglial MMP-2 gene expression and secretion by 72.8% at 24 hours, sustained up to 96 hours (P < 0.01), dependent upon TNF-α. In human CNS TB brain biopsies but not controls the p38 pathway was activated in microglia/macrophages. Inhibition of the p38 MAP kinase pathway resulted in a 228% increase in MMP-2 secretion (P < 0.01). In contrast ERK MAP kinase inhibition further decreased MMP-2 secretion by 76.6% (P < 0.05). Inhibition of the NFκB pathway resulted in 301% higher MMP-2 secretion than CoMTb alone (P < 0.01). Caspase 8 restored MMP-2 secretion to basal levels. However, this caspase-dependent regulation of MMP-2 was independent of p38 and NFκB pathways; p38 phosphorylation was increased and p50/p65 NFκB nuclear trafficking unaffected by caspase 8 inhibition. In summary, suppression of microglial MMP-2 secretion by M.tb-infected monocyte-dependent networks paradoxically involves the pro-inflammatory mediators TNF-α, p38 MAP kinase and NFκB in addition to a novel caspase 8-dependent pathway.

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p38 phosphorylation acts as the break for CoMTb suppression of MMP-2 secretion. Microglial cells were incubated with SB203580 or PD98059 for 2 h, and then stimulated with CoMTb. MMP-2 secretion was analyzed at 72 h by Luminex and gelatin zymography (representative gel shown). p38 inhibition reverses CoMTb suppression and leads to supra-maximal MMP-2 secretion but ERK inhibition further suppresses secretion. Bars represent mean values ± SD of three samples, representative of at least duplicate experiments performed in triplicate. Data were analyzed by one-way analysis of variance, followed by Tukey's multiple comparison. **p < 0.01.
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Figure 3: p38 phosphorylation acts as the break for CoMTb suppression of MMP-2 secretion. Microglial cells were incubated with SB203580 or PD98059 for 2 h, and then stimulated with CoMTb. MMP-2 secretion was analyzed at 72 h by Luminex and gelatin zymography (representative gel shown). p38 inhibition reverses CoMTb suppression and leads to supra-maximal MMP-2 secretion but ERK inhibition further suppresses secretion. Bars represent mean values ± SD of three samples, representative of at least duplicate experiments performed in triplicate. Data were analyzed by one-way analysis of variance, followed by Tukey's multiple comparison. **p < 0.01.

Mentions: Inhibition of the p38 pathway abrogated CoMTb-mediated MMP-2 suppression (Figure 3). 1 μM SB230580 caused CoMTb-driven MMP-2 secretion to return to the same level as constitutively secreted and 10 μM SB230580 caused a 228% increase in MMP-2 secretion over CoMCont (P < 0.01). In contrast ERK inhibition at 1 μM PD98059 caused a 6.1% decrease in MMP-2 secretion over and above the 31.2% reduction due to CoMTb (P < 0.05). 10 μM PD98059 caused a significant 76.6% decrease in MMP-2 secretion compared to CoMCont (P < 0.01). JNK inhibition with 1 μM SP600125 led to a small but non-significant further decrease in MMP-2 secretion but 10 μM caused cell death (data not shown).


Mycobacterium tuberculosis-infected human monocytes down-regulate microglial MMP-2 secretion in CNS tuberculosis via TNFα, NFκB, p38 and caspase 8 dependent pathways.

Green JA, Dholakia S, Janczar K, Ong CW, Moores R, Fry J, Elkington PT, Roncaroli F, Friedland JS - J Neuroinflammation (2011)

p38 phosphorylation acts as the break for CoMTb suppression of MMP-2 secretion. Microglial cells were incubated with SB203580 or PD98059 for 2 h, and then stimulated with CoMTb. MMP-2 secretion was analyzed at 72 h by Luminex and gelatin zymography (representative gel shown). p38 inhibition reverses CoMTb suppression and leads to supra-maximal MMP-2 secretion but ERK inhibition further suppresses secretion. Bars represent mean values ± SD of three samples, representative of at least duplicate experiments performed in triplicate. Data were analyzed by one-way analysis of variance, followed by Tukey's multiple comparison. **p < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3113956&req=5

Figure 3: p38 phosphorylation acts as the break for CoMTb suppression of MMP-2 secretion. Microglial cells were incubated with SB203580 or PD98059 for 2 h, and then stimulated with CoMTb. MMP-2 secretion was analyzed at 72 h by Luminex and gelatin zymography (representative gel shown). p38 inhibition reverses CoMTb suppression and leads to supra-maximal MMP-2 secretion but ERK inhibition further suppresses secretion. Bars represent mean values ± SD of three samples, representative of at least duplicate experiments performed in triplicate. Data were analyzed by one-way analysis of variance, followed by Tukey's multiple comparison. **p < 0.01.
Mentions: Inhibition of the p38 pathway abrogated CoMTb-mediated MMP-2 suppression (Figure 3). 1 μM SB230580 caused CoMTb-driven MMP-2 secretion to return to the same level as constitutively secreted and 10 μM SB230580 caused a 228% increase in MMP-2 secretion over CoMCont (P < 0.01). In contrast ERK inhibition at 1 μM PD98059 caused a 6.1% decrease in MMP-2 secretion over and above the 31.2% reduction due to CoMTb (P < 0.05). 10 μM PD98059 caused a significant 76.6% decrease in MMP-2 secretion compared to CoMCont (P < 0.01). JNK inhibition with 1 μM SP600125 led to a small but non-significant further decrease in MMP-2 secretion but 10 μM caused cell death (data not shown).

Bottom Line: Inhibition of the p38 MAP kinase pathway resulted in a 228% increase in MMP-2 secretion (P < 0.01).In contrast ERK MAP kinase inhibition further decreased MMP-2 secretion by 76.6% (P < 0.05).Inhibition of the NFκB pathway resulted in 301% higher MMP-2 secretion than CoMTb alone (P < 0.01).

View Article: PubMed Central - HTML - PubMed

Affiliation: Section of Infectious Diseases and Immunity and the Imperial College Wellcome Trust Centre for Clinical Tropical Medicine, Hammersmith Campus, Imperial College London, London, W12 0NN, UK. justin.green@imperial.ac.uk

ABSTRACT
Tuberculosis (TB) of the central nervous system (CNS) is a deadly disease characterized by extensive tissue destruction, driven by molecules such as Matrix Metalloproteinase-2 (MMP-2) which targets CNS-specific substrates. In a simplified cellular model of CNS TB, we demonstrated that conditioned medium from Mycobacterium tuberculosis-infected primary human monocytes (CoMTb), but not direct infection, unexpectedly down-regulates constitutive microglial MMP-2 gene expression and secretion by 72.8% at 24 hours, sustained up to 96 hours (P < 0.01), dependent upon TNF-α. In human CNS TB brain biopsies but not controls the p38 pathway was activated in microglia/macrophages. Inhibition of the p38 MAP kinase pathway resulted in a 228% increase in MMP-2 secretion (P < 0.01). In contrast ERK MAP kinase inhibition further decreased MMP-2 secretion by 76.6% (P < 0.05). Inhibition of the NFκB pathway resulted in 301% higher MMP-2 secretion than CoMTb alone (P < 0.01). Caspase 8 restored MMP-2 secretion to basal levels. However, this caspase-dependent regulation of MMP-2 was independent of p38 and NFκB pathways; p38 phosphorylation was increased and p50/p65 NFκB nuclear trafficking unaffected by caspase 8 inhibition. In summary, suppression of microglial MMP-2 secretion by M.tb-infected monocyte-dependent networks paradoxically involves the pro-inflammatory mediators TNF-α, p38 MAP kinase and NFκB in addition to a novel caspase 8-dependent pathway.

Show MeSH
Related in: MedlinePlus