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Anti-inflammatory effects of levalbuterol-induced 11β-hydroxysteroid dehydrogenase type 1 activity in airway epithelial cells.

Randall MJ, Kostin SF, Burgess EJ, Hoyt LR, Ather JL, Lundblad LK, Poynter ME - Front Endocrinol (Lausanne) (2015)

Bottom Line: Since β2-agonists can induce expression of 11β-HSD1, which converts inactive 11-keto corticosteroids into active 11-hydroxy corticosteroids, thereby potentiating the effects of endogenous glucocorticoids, we examined whether this mechanism is involved in the inhibition of NF-κB activation induced by the β-agonist albuterol in airway epithelial cells.LPS- and TNFα-induced NF-κB/luc activity was diminished in MTCC cells treated with a combination of cortisone and (R)-albuterol, an effect that was inhibited by 18β-GA.These results demonstrate that levalbuterol augments expression of 11β-HSD1 in airway epithelial cells, reducing LPS-induced NF-κB transcriptional activity and pro-inflammatory cytokine production through the conversion of inactive 11-keto corticosteroids into the active 11-hydroxy form in this cell type.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary Disease and Critical Care Medicine, Department of Medicine, College of Medicine, The University of Vermont , Burlington, VT , USA.

ABSTRACT
Airway epithelial NF-κB activation is observed in asthmatic subjects and is a cause of airway inflammation in mouse models of allergic asthma. Combination therapy with inhaled short-acting β2-agonists and corticosteroids significantly improves lung function and reduces inflammation in asthmatic subjects. Corticosteroids operate through a number of mechanisms to potently inhibit NF-κB activity. Since β2-agonists can induce expression of 11β-HSD1, which converts inactive 11-keto corticosteroids into active 11-hydroxy corticosteroids, thereby potentiating the effects of endogenous glucocorticoids, we examined whether this mechanism is involved in the inhibition of NF-κB activation induced by the β-agonist albuterol in airway epithelial cells. Treatment of transformed murine Club cells (MTCC) with (R)-albuterol (levalbuterol), but not with (S)- or a mixture of (R + S)- (racemic) albuterol, augmented mRNA expression of 11β-HSD1. MTCC were stably transfected with luciferase (luc) reporter constructs under transcriptional regulation by NF-κB (NF-κB/luc) or glucocorticoid response element (GRE/luc) consensus motifs. Stimulation of NF-κB/luc MTCC with lipopolysaccharide (LPS) or tumor necrosis factor-α (TNFα) induced luc activity, which was inhibited by pretreatment with (R)-, but not (S)- or racemic albuterol. Furthermore, pretreatment of GRE/luc MTCC with (R)-, but not with (S)- or racemic albuterol, augmented 11-keto corticosteroid (cortisone) induced luc activity, which was diminished by the 11β-HSD inhibitor glycyrrhetinic acid (18β-GA), indicating that there was a conversion of inactive 11-keto to active 11-hydroxy corticosteroids. LPS- and TNFα-induced NF-κB/luc activity was diminished in MTCC cells treated with a combination of cortisone and (R)-albuterol, an effect that was inhibited by 18β-GA. Finally, pretreatment of MTCC cells with the combination of cortisone and (R)-albuterol diminished LPS- and TNFα-induced pro-inflammatory cytokine production to an extent similar to that of dexamethasone. These results demonstrate that levalbuterol augments expression of 11β-HSD1 in airway epithelial cells, reducing LPS-induced NF-κB transcriptional activity and pro-inflammatory cytokine production through the conversion of inactive 11-keto corticosteroids into the active 11-hydroxy form in this cell type.

No MeSH data available.


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Pre-exposure of MTCC to (R)-albuterol and cortisone diminishes LPS- and TNFα-induced pro-inflammatory cytokine production. MTCC were exposed to 10−6M (R)-albuterol, with or without 10−6M cortisone or 10−6M glycyrrhetinic acid, or to 10−8M dexamethasone. Twenty-four hours later, 100 ng/ml LPS (A) or 10 ng/ml TNFα (B) were added to the cell culture medium. Cell-free conditioned media were then collected 16 h later and pro-inflammatory cytokine levels were measured. n = 3 samples/group and the experiment was repeated twice; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 compared to LPS (A) or TNFα (B).
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Figure 5: Pre-exposure of MTCC to (R)-albuterol and cortisone diminishes LPS- and TNFα-induced pro-inflammatory cytokine production. MTCC were exposed to 10−6M (R)-albuterol, with or without 10−6M cortisone or 10−6M glycyrrhetinic acid, or to 10−8M dexamethasone. Twenty-four hours later, 100 ng/ml LPS (A) or 10 ng/ml TNFα (B) were added to the cell culture medium. Cell-free conditioned media were then collected 16 h later and pro-inflammatory cytokine levels were measured. n = 3 samples/group and the experiment was repeated twice; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 compared to LPS (A) or TNFα (B).

Mentions: We have previously reported the critical contribution that NF-κB activation in non-ciliated airway epithelial cells (Club cells) has in mouse models of acute lung injury (44) and allergic airway disease (13–17), wherein these cells secrete cytokines that orchestrate innate and adaptive immune responses. Using MTCC cells as an in vitro model, we left the cells untreated or pretreated the cells for 24 h with 10−8M dexamethasone, 10−6M cortisone, 10−6M (R)-albuterol, or a combination of 10−6M (R)-albuterol and 10−6M cortisone. Cells were then stimulated with 100 ng/ml LPS or 10 ng/ml TNFα for 16 h and cytokines were measured from cell-free conditioned medium. As is shown in Figure 5A, only the combined pretreatment of (R)-albuterol and cortisone significantly decreased LPS-induced production of IL-6, GM-CSF, G-CSF, MCP-1, and MIP-1α. Similarly, the combined pretreatment with (R)-albuterol and cortisone significantly decreased TNFα-induced production of IL-6, GM-CSF, KC, MCP-1, MIP-1α, and RANTES (Figure 5B). Notably, pretreatment with cortisone alone had no effect on LPS- or TNFα-induced cytokine production in the absence of (R)-albuterol. These experiments implicate (R)-albuterol-regulated 11β-HSD1 activity as a potentially important modulator of pro-inflammatory signaling and cytokine production in airway epithelial cells.


Anti-inflammatory effects of levalbuterol-induced 11β-hydroxysteroid dehydrogenase type 1 activity in airway epithelial cells.

Randall MJ, Kostin SF, Burgess EJ, Hoyt LR, Ather JL, Lundblad LK, Poynter ME - Front Endocrinol (Lausanne) (2015)

Pre-exposure of MTCC to (R)-albuterol and cortisone diminishes LPS- and TNFα-induced pro-inflammatory cytokine production. MTCC were exposed to 10−6M (R)-albuterol, with or without 10−6M cortisone or 10−6M glycyrrhetinic acid, or to 10−8M dexamethasone. Twenty-four hours later, 100 ng/ml LPS (A) or 10 ng/ml TNFα (B) were added to the cell culture medium. Cell-free conditioned media were then collected 16 h later and pro-inflammatory cytokine levels were measured. n = 3 samples/group and the experiment was repeated twice; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 compared to LPS (A) or TNFα (B).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4290686&req=5

Figure 5: Pre-exposure of MTCC to (R)-albuterol and cortisone diminishes LPS- and TNFα-induced pro-inflammatory cytokine production. MTCC were exposed to 10−6M (R)-albuterol, with or without 10−6M cortisone or 10−6M glycyrrhetinic acid, or to 10−8M dexamethasone. Twenty-four hours later, 100 ng/ml LPS (A) or 10 ng/ml TNFα (B) were added to the cell culture medium. Cell-free conditioned media were then collected 16 h later and pro-inflammatory cytokine levels were measured. n = 3 samples/group and the experiment was repeated twice; *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 compared to LPS (A) or TNFα (B).
Mentions: We have previously reported the critical contribution that NF-κB activation in non-ciliated airway epithelial cells (Club cells) has in mouse models of acute lung injury (44) and allergic airway disease (13–17), wherein these cells secrete cytokines that orchestrate innate and adaptive immune responses. Using MTCC cells as an in vitro model, we left the cells untreated or pretreated the cells for 24 h with 10−8M dexamethasone, 10−6M cortisone, 10−6M (R)-albuterol, or a combination of 10−6M (R)-albuterol and 10−6M cortisone. Cells were then stimulated with 100 ng/ml LPS or 10 ng/ml TNFα for 16 h and cytokines were measured from cell-free conditioned medium. As is shown in Figure 5A, only the combined pretreatment of (R)-albuterol and cortisone significantly decreased LPS-induced production of IL-6, GM-CSF, G-CSF, MCP-1, and MIP-1α. Similarly, the combined pretreatment with (R)-albuterol and cortisone significantly decreased TNFα-induced production of IL-6, GM-CSF, KC, MCP-1, MIP-1α, and RANTES (Figure 5B). Notably, pretreatment with cortisone alone had no effect on LPS- or TNFα-induced cytokine production in the absence of (R)-albuterol. These experiments implicate (R)-albuterol-regulated 11β-HSD1 activity as a potentially important modulator of pro-inflammatory signaling and cytokine production in airway epithelial cells.

Bottom Line: Since β2-agonists can induce expression of 11β-HSD1, which converts inactive 11-keto corticosteroids into active 11-hydroxy corticosteroids, thereby potentiating the effects of endogenous glucocorticoids, we examined whether this mechanism is involved in the inhibition of NF-κB activation induced by the β-agonist albuterol in airway epithelial cells.LPS- and TNFα-induced NF-κB/luc activity was diminished in MTCC cells treated with a combination of cortisone and (R)-albuterol, an effect that was inhibited by 18β-GA.These results demonstrate that levalbuterol augments expression of 11β-HSD1 in airway epithelial cells, reducing LPS-induced NF-κB transcriptional activity and pro-inflammatory cytokine production through the conversion of inactive 11-keto corticosteroids into the active 11-hydroxy form in this cell type.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary Disease and Critical Care Medicine, Department of Medicine, College of Medicine, The University of Vermont , Burlington, VT , USA.

ABSTRACT
Airway epithelial NF-κB activation is observed in asthmatic subjects and is a cause of airway inflammation in mouse models of allergic asthma. Combination therapy with inhaled short-acting β2-agonists and corticosteroids significantly improves lung function and reduces inflammation in asthmatic subjects. Corticosteroids operate through a number of mechanisms to potently inhibit NF-κB activity. Since β2-agonists can induce expression of 11β-HSD1, which converts inactive 11-keto corticosteroids into active 11-hydroxy corticosteroids, thereby potentiating the effects of endogenous glucocorticoids, we examined whether this mechanism is involved in the inhibition of NF-κB activation induced by the β-agonist albuterol in airway epithelial cells. Treatment of transformed murine Club cells (MTCC) with (R)-albuterol (levalbuterol), but not with (S)- or a mixture of (R + S)- (racemic) albuterol, augmented mRNA expression of 11β-HSD1. MTCC were stably transfected with luciferase (luc) reporter constructs under transcriptional regulation by NF-κB (NF-κB/luc) or glucocorticoid response element (GRE/luc) consensus motifs. Stimulation of NF-κB/luc MTCC with lipopolysaccharide (LPS) or tumor necrosis factor-α (TNFα) induced luc activity, which was inhibited by pretreatment with (R)-, but not (S)- or racemic albuterol. Furthermore, pretreatment of GRE/luc MTCC with (R)-, but not with (S)- or racemic albuterol, augmented 11-keto corticosteroid (cortisone) induced luc activity, which was diminished by the 11β-HSD inhibitor glycyrrhetinic acid (18β-GA), indicating that there was a conversion of inactive 11-keto to active 11-hydroxy corticosteroids. LPS- and TNFα-induced NF-κB/luc activity was diminished in MTCC cells treated with a combination of cortisone and (R)-albuterol, an effect that was inhibited by 18β-GA. Finally, pretreatment of MTCC cells with the combination of cortisone and (R)-albuterol diminished LPS- and TNFα-induced pro-inflammatory cytokine production to an extent similar to that of dexamethasone. These results demonstrate that levalbuterol augments expression of 11β-HSD1 in airway epithelial cells, reducing LPS-induced NF-κB transcriptional activity and pro-inflammatory cytokine production through the conversion of inactive 11-keto corticosteroids into the active 11-hydroxy form in this cell type.

No MeSH data available.


Related in: MedlinePlus