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Vimentin regulates activation of the NLRP3 inflammasome.

dos Santos G, Rogel MR, Baker MA, Troken JR, Urich D, Morales-Nebreda L, Sennello JA, Kutuzov MA, Sitikov A, Davis JM, Lam AP, Cheresh P, Kamp D, Shumaker DK, Budinger GR, Ridge KM - Nat Commun (2015)

Bottom Line: Activation of the NLRP3 inflammasome and subsequent maturation of IL-1β have been implicated in acute lung injury (ALI), resulting in inflammation and fibrosis.Furthermore, decreased active caspase-1 and IL-1β levels are observed in vitro in Vim(-/-) and vimentin-knockdown macrophages.This study provides insights into lung inflammation and fibrosis and suggests that vimentin may be a key regulator of the NLRP3 inflammasome.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary and Critical Care Medicine, Northwestern University, Chicago, Illinois 60611, USA.

ABSTRACT
Activation of the NLRP3 inflammasome and subsequent maturation of IL-1β have been implicated in acute lung injury (ALI), resulting in inflammation and fibrosis. We investigated the role of vimentin, a type III intermediate filament, in this process using three well-characterized murine models of ALI known to require NLRP3 inflammasome activation. We demonstrate that central pathophysiologic events in ALI (inflammation, IL-1β levels, endothelial and alveolar epithelial barrier permeability, remodelling and fibrosis) are attenuated in the lungs of Vim(-/-) mice challenged with LPS, bleomycin and asbestos. Bone marrow chimeric mice lacking vimentin have reduced IL-1β levels and attenuated lung injury and fibrosis following bleomycin exposure. Furthermore, decreased active caspase-1 and IL-1β levels are observed in vitro in Vim(-/-) and vimentin-knockdown macrophages. Importantly, we show direct protein-protein interaction between NLRP3 and vimentin. This study provides insights into lung inflammation and fibrosis and suggests that vimentin may be a key regulator of the NLRP3 inflammasome.

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Vimentin deficiency prevents asbestos-induced lung injury and fibrosisWT and Vim-/- mice were treated with PBS containing either 200 μg of asbestos crocidolite or the control particle titanium dioxide (TiO2), intratracheally. Markers of inflammation and fibrosis were assessed 3 and 6 weeks after instillation, respectively. Total cell count (macrophages, neutrophils, eosinophils, erythrocytes, and lymphocytes )(A) and protein levels (B) was assessed in BALF collected from Vim-/- mice 3 weeks after asbestos administration. Inflammasome activation in WT and Vim-/- mice was measured by ELISA for caspase-1 (C) and IL-1β (D) levels in BALF at the same time point. Collagen deposition was evaluated in asbestos-treated Vim-/- and WT miceby Masson's trichrome and Picrosirius red staining of lung slices (E) (scale bars, 200 μm) and by measuring total collagen content by Sircol assay (F). Data are from 2 independent experiments n= 6-8 animals per group, and presented as mean ± SD. *P < 0.05, **P < 0.001, relative to WT versus Vim-/-, by one way ANOVA with a correction provided by the Bonferroni multiple comparisons test.
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Figure 2: Vimentin deficiency prevents asbestos-induced lung injury and fibrosisWT and Vim-/- mice were treated with PBS containing either 200 μg of asbestos crocidolite or the control particle titanium dioxide (TiO2), intratracheally. Markers of inflammation and fibrosis were assessed 3 and 6 weeks after instillation, respectively. Total cell count (macrophages, neutrophils, eosinophils, erythrocytes, and lymphocytes )(A) and protein levels (B) was assessed in BALF collected from Vim-/- mice 3 weeks after asbestos administration. Inflammasome activation in WT and Vim-/- mice was measured by ELISA for caspase-1 (C) and IL-1β (D) levels in BALF at the same time point. Collagen deposition was evaluated in asbestos-treated Vim-/- and WT miceby Masson's trichrome and Picrosirius red staining of lung slices (E) (scale bars, 200 μm) and by measuring total collagen content by Sircol assay (F). Data are from 2 independent experiments n= 6-8 animals per group, and presented as mean ± SD. *P < 0.05, **P < 0.001, relative to WT versus Vim-/-, by one way ANOVA with a correction provided by the Bonferroni multiple comparisons test.

Mentions: The NLRP3 inflammasome has been implicated in the pathological increase of IL-1β production in a variety of pulmonary inflammatory diseases that lead to fibrosis, including asbestosis21-23. In a model of asbestosis, we sought to determine whether Vim-/- mice showed diminished cytokine production and whether these animals were protected from lung injury and pulmonary fibrosis. Wild-type and Vim-/- mice were exposed to crocidolite asbestos or an inert particle, titanium dioxide (TiO2); markers of injury, inflammation, cytokine production, and fibrosis were assessed. Confirming previous reports22,23, asbestos-exposed mice had increased total cell numbers in BALF compared with TiO2-exposed animals. However, significantly fewer cells were recruited to the lungs of Vim-/- mice after asbestos exposure (Figure 2A). Additional analysis of immune cell recruitment into the lungs of WT and Vim-/- mice was performed by FACS analysis of whole lung lysates (Supplemental Figure 2A). Wild-type mice had profound inflammatory cell infiltrates following exposure to asbestos as observed by hematoxylin and eosin (H&E) staining; in contrast Vim-/- mice showed reduced inflammation (Supplemental Figure 2B). Wild-type mice exhibited an increase in total protein concentration in BALF, whereas Vim-/- mice failed to exhibit such an increase (Figure 2B). The reduction in inflammation in Vim-/- mice exposed to asbestos was associated with reduced levels of caspase-1 and IL-1β in BALF compared with WT mice (Figure 2C and 2D). These data suggest that vimentin is required for asbestos-mediated activation of the NLRP3 inflammasome (step 2), but not for NF-κB activation and upregulation of pro-IL-1β and NLRP3 (step 1). Furthermore, we observed that WT mice exposed to asbestos develop pulmonary fibrosis, as assessed by Masson trichrome and Picrosirius red stain (a dye that binds specifically to collagen fibrils) (Figure 2E), and Sircol assay (Figure 2F). Vimentin deficiency prevented pulmonary fibrosis, as no significant increase in collagen deposition was observed in Vim-/- mice (Figure 2E and 2F). Collectively, these results demonstrate that loss of vimentin prevents asbestos-induced collagen deposition and fibrosis.


Vimentin regulates activation of the NLRP3 inflammasome.

dos Santos G, Rogel MR, Baker MA, Troken JR, Urich D, Morales-Nebreda L, Sennello JA, Kutuzov MA, Sitikov A, Davis JM, Lam AP, Cheresh P, Kamp D, Shumaker DK, Budinger GR, Ridge KM - Nat Commun (2015)

Vimentin deficiency prevents asbestos-induced lung injury and fibrosisWT and Vim-/- mice were treated with PBS containing either 200 μg of asbestos crocidolite or the control particle titanium dioxide (TiO2), intratracheally. Markers of inflammation and fibrosis were assessed 3 and 6 weeks after instillation, respectively. Total cell count (macrophages, neutrophils, eosinophils, erythrocytes, and lymphocytes )(A) and protein levels (B) was assessed in BALF collected from Vim-/- mice 3 weeks after asbestos administration. Inflammasome activation in WT and Vim-/- mice was measured by ELISA for caspase-1 (C) and IL-1β (D) levels in BALF at the same time point. Collagen deposition was evaluated in asbestos-treated Vim-/- and WT miceby Masson's trichrome and Picrosirius red staining of lung slices (E) (scale bars, 200 μm) and by measuring total collagen content by Sircol assay (F). Data are from 2 independent experiments n= 6-8 animals per group, and presented as mean ± SD. *P < 0.05, **P < 0.001, relative to WT versus Vim-/-, by one way ANOVA with a correction provided by the Bonferroni multiple comparisons test.
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Related In: Results  -  Collection

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Figure 2: Vimentin deficiency prevents asbestos-induced lung injury and fibrosisWT and Vim-/- mice were treated with PBS containing either 200 μg of asbestos crocidolite or the control particle titanium dioxide (TiO2), intratracheally. Markers of inflammation and fibrosis were assessed 3 and 6 weeks after instillation, respectively. Total cell count (macrophages, neutrophils, eosinophils, erythrocytes, and lymphocytes )(A) and protein levels (B) was assessed in BALF collected from Vim-/- mice 3 weeks after asbestos administration. Inflammasome activation in WT and Vim-/- mice was measured by ELISA for caspase-1 (C) and IL-1β (D) levels in BALF at the same time point. Collagen deposition was evaluated in asbestos-treated Vim-/- and WT miceby Masson's trichrome and Picrosirius red staining of lung slices (E) (scale bars, 200 μm) and by measuring total collagen content by Sircol assay (F). Data are from 2 independent experiments n= 6-8 animals per group, and presented as mean ± SD. *P < 0.05, **P < 0.001, relative to WT versus Vim-/-, by one way ANOVA with a correction provided by the Bonferroni multiple comparisons test.
Mentions: The NLRP3 inflammasome has been implicated in the pathological increase of IL-1β production in a variety of pulmonary inflammatory diseases that lead to fibrosis, including asbestosis21-23. In a model of asbestosis, we sought to determine whether Vim-/- mice showed diminished cytokine production and whether these animals were protected from lung injury and pulmonary fibrosis. Wild-type and Vim-/- mice were exposed to crocidolite asbestos or an inert particle, titanium dioxide (TiO2); markers of injury, inflammation, cytokine production, and fibrosis were assessed. Confirming previous reports22,23, asbestos-exposed mice had increased total cell numbers in BALF compared with TiO2-exposed animals. However, significantly fewer cells were recruited to the lungs of Vim-/- mice after asbestos exposure (Figure 2A). Additional analysis of immune cell recruitment into the lungs of WT and Vim-/- mice was performed by FACS analysis of whole lung lysates (Supplemental Figure 2A). Wild-type mice had profound inflammatory cell infiltrates following exposure to asbestos as observed by hematoxylin and eosin (H&E) staining; in contrast Vim-/- mice showed reduced inflammation (Supplemental Figure 2B). Wild-type mice exhibited an increase in total protein concentration in BALF, whereas Vim-/- mice failed to exhibit such an increase (Figure 2B). The reduction in inflammation in Vim-/- mice exposed to asbestos was associated with reduced levels of caspase-1 and IL-1β in BALF compared with WT mice (Figure 2C and 2D). These data suggest that vimentin is required for asbestos-mediated activation of the NLRP3 inflammasome (step 2), but not for NF-κB activation and upregulation of pro-IL-1β and NLRP3 (step 1). Furthermore, we observed that WT mice exposed to asbestos develop pulmonary fibrosis, as assessed by Masson trichrome and Picrosirius red stain (a dye that binds specifically to collagen fibrils) (Figure 2E), and Sircol assay (Figure 2F). Vimentin deficiency prevented pulmonary fibrosis, as no significant increase in collagen deposition was observed in Vim-/- mice (Figure 2E and 2F). Collectively, these results demonstrate that loss of vimentin prevents asbestos-induced collagen deposition and fibrosis.

Bottom Line: Activation of the NLRP3 inflammasome and subsequent maturation of IL-1β have been implicated in acute lung injury (ALI), resulting in inflammation and fibrosis.Furthermore, decreased active caspase-1 and IL-1β levels are observed in vitro in Vim(-/-) and vimentin-knockdown macrophages.This study provides insights into lung inflammation and fibrosis and suggests that vimentin may be a key regulator of the NLRP3 inflammasome.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary and Critical Care Medicine, Northwestern University, Chicago, Illinois 60611, USA.

ABSTRACT
Activation of the NLRP3 inflammasome and subsequent maturation of IL-1β have been implicated in acute lung injury (ALI), resulting in inflammation and fibrosis. We investigated the role of vimentin, a type III intermediate filament, in this process using three well-characterized murine models of ALI known to require NLRP3 inflammasome activation. We demonstrate that central pathophysiologic events in ALI (inflammation, IL-1β levels, endothelial and alveolar epithelial barrier permeability, remodelling and fibrosis) are attenuated in the lungs of Vim(-/-) mice challenged with LPS, bleomycin and asbestos. Bone marrow chimeric mice lacking vimentin have reduced IL-1β levels and attenuated lung injury and fibrosis following bleomycin exposure. Furthermore, decreased active caspase-1 and IL-1β levels are observed in vitro in Vim(-/-) and vimentin-knockdown macrophages. Importantly, we show direct protein-protein interaction between NLRP3 and vimentin. This study provides insights into lung inflammation and fibrosis and suggests that vimentin may be a key regulator of the NLRP3 inflammasome.

Show MeSH
Related in: MedlinePlus