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Systemic inhibition of NF-kappaB activation protects from silicosis.

Di Giuseppe M, Gambelli F, Hoyle GW, Lungarella G, Studer SM, Richards T, Yousem S, McCurry K, Dauber J, Kaminski N, Leikauf G, Ortiz LA - PLoS ONE (2009)

Bottom Line: At the time of lung transplantation, we found the lungs of silica-exposed subjects to contain multiple foci of inflammatory cells and silicotic nodules with proximal TNFalpha expressing macrophage and NF-kappaB activation in epithelial cells.Using a mouse experimental model in which the endotracheal instillation of silica reproduces the silica-induced lung injury observed in humans we found that systemic inhibition of NF-kappaB activation with a pharmacologic inhibitor (BAY 11-7085) of IkappaB alpha phosphorylation decreased silica-induced inflammation and collagen deposition.In contrast, transgenic mice expressing a dominant negative IkappaB alpha mutant protein under the control of epithelial cell specific promoters demonstrate enhanced apoptosis and collagen deposition in their lungs in response to silica.

View Article: PubMed Central - PubMed

Affiliation: Division of Occupational and Environmental Medicine, Department of Environmental and Occupational Health, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.

ABSTRACT

Background: Silicosis is a complex lung disease for which no successful treatment is available and therefore lung transplantation is a potential alternative. Tumor necrosis factor alpha (TNFalpha) plays a central role in the pathogenesis of silicosis. TNFalpha signaling is mediated by the transcription factor, Nuclear Factor (NF)-kappaB, which regulates genes controlling several physiological processes including the innate immune responses, cell death, and inflammation. Therefore, inhibition of NF-kappaB activation represents a potential therapeutic strategy for silicosis.

Methods/findings: In the present work we evaluated the lung transplant database (May 1986-July 2007) at the University of Pittsburgh to study the efficacy of lung transplantation in patients with silicosis (n = 11). We contrasted the overall survival and rate of graft rejection in these patients to that of patients with idiopathic pulmonary fibrosis (IPF, n = 79) that was selected as a control group because survival benefit of lung transplantation has been identified for these patients. At the time of lung transplantation, we found the lungs of silica-exposed subjects to contain multiple foci of inflammatory cells and silicotic nodules with proximal TNFalpha expressing macrophage and NF-kappaB activation in epithelial cells. Patients with silicosis had poor survival (median survival 2.4 yr; confidence interval (CI): 0.16-7.88 yr) compared to IPF patients (5.3 yr; CI: 2.8-15 yr; p = 0.07), and experienced early rejection of their lung grafts (0.9 yr; CI: 0.22-0.9 yr) following lung transplantation (2.4 yr; CI:1.5-3.6 yr; p<0.05). Using a mouse experimental model in which the endotracheal instillation of silica reproduces the silica-induced lung injury observed in humans we found that systemic inhibition of NF-kappaB activation with a pharmacologic inhibitor (BAY 11-7085) of IkappaB alpha phosphorylation decreased silica-induced inflammation and collagen deposition. In contrast, transgenic mice expressing a dominant negative IkappaB alpha mutant protein under the control of epithelial cell specific promoters demonstrate enhanced apoptosis and collagen deposition in their lungs in response to silica.

Conclusions: Although limited by its size, our data support that patients with silicosis appear to have poor outcome following lung transplantation. Experimental data indicate that while the systemic inhibition of NF-kappaB protects from silica-induced lung injury, epithelial cell specific NF-kappaB inhibition appears to aggravate the outcome of experimental silicosis.

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Systemic inhibition of NF-κB activity ameliorates silica-induced lung injury in mice.A) DNA binding activity of NF-κB in crude nuclear extracts from whole lung isolated from C57BL/6 mice 14 and 28 days after silica, or silica+BAY exposure. Excess probe represents NF-κB binding in lung nuclear extract of a silica-treated mouse, assayed in the presence of excess unlabelled oligonucleotide as a competitor. Antibody supershifts were performed using the nuclear extract of a C57BL/6 silica-treated mouse as described in Methods section. B) Northern blot analysis of TNFα, α1(I) collagen, and 18S (loading control) mRNA expression in mouse lung 14 days following the intratracheal injection of saline as control, silica alone, or silica+BAY as described in Methods section. C) The effect of BAY on metalloproteinase (MMPs) RNA expression in mouse lung following silica exposure. Total lung RNA was isolated from the lungs of C57BL/6 mice 14 days after control, silica, or silica+BAY and subjected to RPA as described in Methods section. D) The panels show low (×100) power magnification photomicrographs of the lung obtained from lung tissues of C57BL/6 mice exposed to saline as control (A), silica (B), or silica+BAY (C) as described in the Methods section. TB = terminal bronchiole. Bar = 20 µm. Gel illustrations are representative of 4 different experiments.
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pone-0005689-g005: Systemic inhibition of NF-κB activity ameliorates silica-induced lung injury in mice.A) DNA binding activity of NF-κB in crude nuclear extracts from whole lung isolated from C57BL/6 mice 14 and 28 days after silica, or silica+BAY exposure. Excess probe represents NF-κB binding in lung nuclear extract of a silica-treated mouse, assayed in the presence of excess unlabelled oligonucleotide as a competitor. Antibody supershifts were performed using the nuclear extract of a C57BL/6 silica-treated mouse as described in Methods section. B) Northern blot analysis of TNFα, α1(I) collagen, and 18S (loading control) mRNA expression in mouse lung 14 days following the intratracheal injection of saline as control, silica alone, or silica+BAY as described in Methods section. C) The effect of BAY on metalloproteinase (MMPs) RNA expression in mouse lung following silica exposure. Total lung RNA was isolated from the lungs of C57BL/6 mice 14 days after control, silica, or silica+BAY and subjected to RPA as described in Methods section. D) The panels show low (×100) power magnification photomicrographs of the lung obtained from lung tissues of C57BL/6 mice exposed to saline as control (A), silica (B), or silica+BAY (C) as described in the Methods section. TB = terminal bronchiole. Bar = 20 µm. Gel illustrations are representative of 4 different experiments.

Mentions: Following the early activation of NF-κB, lung IκB mRNA increased in silica-exposed C57BL/6 mice (not shown) and IκB protein returned to baseline (Figure 4). Enhanced NF-κB activation was appreciated in the lungs of silica-exposed C57BL/6 mice for the entire duration of the experiments, and it was readily demonstrated 14 and 28 d after silica exposure (Figure 5). Therefore, it is conceivable that inhibition of NF-κB activity could in turn modulate expression of NF-κB sensitive genes and alters the development of silica-induced lung injury in these mice.


Systemic inhibition of NF-kappaB activation protects from silicosis.

Di Giuseppe M, Gambelli F, Hoyle GW, Lungarella G, Studer SM, Richards T, Yousem S, McCurry K, Dauber J, Kaminski N, Leikauf G, Ortiz LA - PLoS ONE (2009)

Systemic inhibition of NF-κB activity ameliorates silica-induced lung injury in mice.A) DNA binding activity of NF-κB in crude nuclear extracts from whole lung isolated from C57BL/6 mice 14 and 28 days after silica, or silica+BAY exposure. Excess probe represents NF-κB binding in lung nuclear extract of a silica-treated mouse, assayed in the presence of excess unlabelled oligonucleotide as a competitor. Antibody supershifts were performed using the nuclear extract of a C57BL/6 silica-treated mouse as described in Methods section. B) Northern blot analysis of TNFα, α1(I) collagen, and 18S (loading control) mRNA expression in mouse lung 14 days following the intratracheal injection of saline as control, silica alone, or silica+BAY as described in Methods section. C) The effect of BAY on metalloproteinase (MMPs) RNA expression in mouse lung following silica exposure. Total lung RNA was isolated from the lungs of C57BL/6 mice 14 days after control, silica, or silica+BAY and subjected to RPA as described in Methods section. D) The panels show low (×100) power magnification photomicrographs of the lung obtained from lung tissues of C57BL/6 mice exposed to saline as control (A), silica (B), or silica+BAY (C) as described in the Methods section. TB = terminal bronchiole. Bar = 20 µm. Gel illustrations are representative of 4 different experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2682759&req=5

pone-0005689-g005: Systemic inhibition of NF-κB activity ameliorates silica-induced lung injury in mice.A) DNA binding activity of NF-κB in crude nuclear extracts from whole lung isolated from C57BL/6 mice 14 and 28 days after silica, or silica+BAY exposure. Excess probe represents NF-κB binding in lung nuclear extract of a silica-treated mouse, assayed in the presence of excess unlabelled oligonucleotide as a competitor. Antibody supershifts were performed using the nuclear extract of a C57BL/6 silica-treated mouse as described in Methods section. B) Northern blot analysis of TNFα, α1(I) collagen, and 18S (loading control) mRNA expression in mouse lung 14 days following the intratracheal injection of saline as control, silica alone, or silica+BAY as described in Methods section. C) The effect of BAY on metalloproteinase (MMPs) RNA expression in mouse lung following silica exposure. Total lung RNA was isolated from the lungs of C57BL/6 mice 14 days after control, silica, or silica+BAY and subjected to RPA as described in Methods section. D) The panels show low (×100) power magnification photomicrographs of the lung obtained from lung tissues of C57BL/6 mice exposed to saline as control (A), silica (B), or silica+BAY (C) as described in the Methods section. TB = terminal bronchiole. Bar = 20 µm. Gel illustrations are representative of 4 different experiments.
Mentions: Following the early activation of NF-κB, lung IκB mRNA increased in silica-exposed C57BL/6 mice (not shown) and IκB protein returned to baseline (Figure 4). Enhanced NF-κB activation was appreciated in the lungs of silica-exposed C57BL/6 mice for the entire duration of the experiments, and it was readily demonstrated 14 and 28 d after silica exposure (Figure 5). Therefore, it is conceivable that inhibition of NF-κB activity could in turn modulate expression of NF-κB sensitive genes and alters the development of silica-induced lung injury in these mice.

Bottom Line: At the time of lung transplantation, we found the lungs of silica-exposed subjects to contain multiple foci of inflammatory cells and silicotic nodules with proximal TNFalpha expressing macrophage and NF-kappaB activation in epithelial cells.Using a mouse experimental model in which the endotracheal instillation of silica reproduces the silica-induced lung injury observed in humans we found that systemic inhibition of NF-kappaB activation with a pharmacologic inhibitor (BAY 11-7085) of IkappaB alpha phosphorylation decreased silica-induced inflammation and collagen deposition.In contrast, transgenic mice expressing a dominant negative IkappaB alpha mutant protein under the control of epithelial cell specific promoters demonstrate enhanced apoptosis and collagen deposition in their lungs in response to silica.

View Article: PubMed Central - PubMed

Affiliation: Division of Occupational and Environmental Medicine, Department of Environmental and Occupational Health, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.

ABSTRACT

Background: Silicosis is a complex lung disease for which no successful treatment is available and therefore lung transplantation is a potential alternative. Tumor necrosis factor alpha (TNFalpha) plays a central role in the pathogenesis of silicosis. TNFalpha signaling is mediated by the transcription factor, Nuclear Factor (NF)-kappaB, which regulates genes controlling several physiological processes including the innate immune responses, cell death, and inflammation. Therefore, inhibition of NF-kappaB activation represents a potential therapeutic strategy for silicosis.

Methods/findings: In the present work we evaluated the lung transplant database (May 1986-July 2007) at the University of Pittsburgh to study the efficacy of lung transplantation in patients with silicosis (n = 11). We contrasted the overall survival and rate of graft rejection in these patients to that of patients with idiopathic pulmonary fibrosis (IPF, n = 79) that was selected as a control group because survival benefit of lung transplantation has been identified for these patients. At the time of lung transplantation, we found the lungs of silica-exposed subjects to contain multiple foci of inflammatory cells and silicotic nodules with proximal TNFalpha expressing macrophage and NF-kappaB activation in epithelial cells. Patients with silicosis had poor survival (median survival 2.4 yr; confidence interval (CI): 0.16-7.88 yr) compared to IPF patients (5.3 yr; CI: 2.8-15 yr; p = 0.07), and experienced early rejection of their lung grafts (0.9 yr; CI: 0.22-0.9 yr) following lung transplantation (2.4 yr; CI:1.5-3.6 yr; p<0.05). Using a mouse experimental model in which the endotracheal instillation of silica reproduces the silica-induced lung injury observed in humans we found that systemic inhibition of NF-kappaB activation with a pharmacologic inhibitor (BAY 11-7085) of IkappaB alpha phosphorylation decreased silica-induced inflammation and collagen deposition. In contrast, transgenic mice expressing a dominant negative IkappaB alpha mutant protein under the control of epithelial cell specific promoters demonstrate enhanced apoptosis and collagen deposition in their lungs in response to silica.

Conclusions: Although limited by its size, our data support that patients with silicosis appear to have poor outcome following lung transplantation. Experimental data indicate that while the systemic inhibition of NF-kappaB protects from silica-induced lung injury, epithelial cell specific NF-kappaB inhibition appears to aggravate the outcome of experimental silicosis.

Show MeSH
Related in: MedlinePlus