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Asthmatic airway smooth muscle CXCL10 production: mitogen-activated protein kinase JNK involvement.

Alrashdan YA, Alkhouri H, Chen E, Lalor DJ, Poniris M, Henness S, Brightling CE, Burgess JK, Armour CL, Ammit AJ, Hughes JM - Am. J. Physiol. Lung Cell Mol. Physiol. (2012)

Bottom Line: However, in both, the JNK inhibitor SP600125 reduced JNK phosphorylation and CXCL10 mRNA levels but did not affect CXCL10 mRNA stability or Iκ-Bα degradation.We concluded that, in asthmatic compared with nonasthmatic ASM cells, JNK activation was reduced and CXCL10 gene expression was more rapid following cytomix stimulation.However, in both, JNK activation did not regulate early events leading to NF-κB activation.

View Article: PubMed Central - PubMed

Affiliation: Respiratory Research Group, Faculty of Pharmacy, University of Sydney, Sydney, NSW 2006, Australia.

ABSTRACT
CXCL10 (IP10) is involved in mast cell migration to airway smooth muscle (ASM) bundles in asthma. We aimed to investigate the role of cytokine-induced MAPK activation in CXCL10 production by ASM cells from people with and without asthma. Confluent growth-arrested ASM cells were treated with inhibitors of the MAPKs ERK, p38, and JNK and transcription factor NF-κB, or vehicle, and stimulated with IL-1β, TNF-α, or IFN-γ, alone or combined (cytomix). CXCL10 mRNA and protein, JNK, NF-κB p65 phosphorylation, and Iκ-Bα protein degradation were assessed using real-time PCR, ELISA, and immunoblotting, respectively. Cytomix, IL-1β, and TNF-α induced CXCL10 mRNA expression more rapidly in asthmatic than nonasthmatic ASM cells. IL-1β and/or TNF-α combined with IFN-γ synergistically increased asthmatic ASM cell CXCL10 release. Inhibitor effects were similar in asthmatic and nonasthmatic cells, but cytomix-induced release was least affected, with only JNK and NF-κB inhibitors halving it. Notably, JNK phosphorylation was markedly less in asthmatic compared with nonasthmatic cells. However, in both, the JNK inhibitor SP600125 reduced JNK phosphorylation and CXCL10 mRNA levels but did not affect CXCL10 mRNA stability or Iκ-Bα degradation. Together, the JNK and NF-κB inhibitors completely inhibited their CXCL10 release. We concluded that, in asthmatic compared with nonasthmatic ASM cells, JNK activation was reduced and CXCL10 gene expression was more rapid following cytomix stimulation. However, in both, JNK activation did not regulate early events leading to NF-κB activation. Thus JNK and NF-κB provide independent therapeutic targets for limiting CXCL10 production and mast cell migration to the ASM in asthma.

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Related in: MedlinePlus

Cytokine-induced CXCL10 mRNA production by asthmatic (A) and nonasthmatic (NA) airway smooth muscle (ASM) cells. Confluent, serum-deprived ASM cells were stimulated with 10 ng/ml of IL-1β, TNF-α, IFN-γ, or cytomix for up to 24 h, and CXCL10 mRNA levels were quantified using real-time PCR and expressed as fold change over 0 h. Bars, mean ± SE.
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Figure 1: Cytokine-induced CXCL10 mRNA production by asthmatic (A) and nonasthmatic (NA) airway smooth muscle (ASM) cells. Confluent, serum-deprived ASM cells were stimulated with 10 ng/ml of IL-1β, TNF-α, IFN-γ, or cytomix for up to 24 h, and CXCL10 mRNA levels were quantified using real-time PCR and expressed as fold change over 0 h. Bars, mean ± SE.

Mentions: The pattern of cytokine-induced CXCL10 mRNA production by ASM cells from asthmatics and nonasthmatics differed. In both cell types, the individual cytokines induced high CXCL10 mRNA levels rapidly, with IL-1β < TNF-α < IFN-γ (Fig. 1). Maximum CXCL10 mRNA levels induced were higher in asthmatic than nonasthmatic ASM cells following stimulation with IL-1β and TNF-α at 6 h but lower with IFN-γ at 24 h. Cytomix induced a synergistic upregulation of CXCL10 mRNA levels, and, similar to IL-1β and TNF-α, these were higher (P < 0.05) in asthmatic ASM cells (>48,000 × 0 h) than in nonasthmatic cells (<11,000 × 0 h) at 6 h (Fig. 1).


Asthmatic airway smooth muscle CXCL10 production: mitogen-activated protein kinase JNK involvement.

Alrashdan YA, Alkhouri H, Chen E, Lalor DJ, Poniris M, Henness S, Brightling CE, Burgess JK, Armour CL, Ammit AJ, Hughes JM - Am. J. Physiol. Lung Cell Mol. Physiol. (2012)

Cytokine-induced CXCL10 mRNA production by asthmatic (A) and nonasthmatic (NA) airway smooth muscle (ASM) cells. Confluent, serum-deprived ASM cells were stimulated with 10 ng/ml of IL-1β, TNF-α, IFN-γ, or cytomix for up to 24 h, and CXCL10 mRNA levels were quantified using real-time PCR and expressed as fold change over 0 h. Bars, mean ± SE.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3362261&req=5

Figure 1: Cytokine-induced CXCL10 mRNA production by asthmatic (A) and nonasthmatic (NA) airway smooth muscle (ASM) cells. Confluent, serum-deprived ASM cells were stimulated with 10 ng/ml of IL-1β, TNF-α, IFN-γ, or cytomix for up to 24 h, and CXCL10 mRNA levels were quantified using real-time PCR and expressed as fold change over 0 h. Bars, mean ± SE.
Mentions: The pattern of cytokine-induced CXCL10 mRNA production by ASM cells from asthmatics and nonasthmatics differed. In both cell types, the individual cytokines induced high CXCL10 mRNA levels rapidly, with IL-1β < TNF-α < IFN-γ (Fig. 1). Maximum CXCL10 mRNA levels induced were higher in asthmatic than nonasthmatic ASM cells following stimulation with IL-1β and TNF-α at 6 h but lower with IFN-γ at 24 h. Cytomix induced a synergistic upregulation of CXCL10 mRNA levels, and, similar to IL-1β and TNF-α, these were higher (P < 0.05) in asthmatic ASM cells (>48,000 × 0 h) than in nonasthmatic cells (<11,000 × 0 h) at 6 h (Fig. 1).

Bottom Line: However, in both, the JNK inhibitor SP600125 reduced JNK phosphorylation and CXCL10 mRNA levels but did not affect CXCL10 mRNA stability or Iκ-Bα degradation.We concluded that, in asthmatic compared with nonasthmatic ASM cells, JNK activation was reduced and CXCL10 gene expression was more rapid following cytomix stimulation.However, in both, JNK activation did not regulate early events leading to NF-κB activation.

View Article: PubMed Central - PubMed

Affiliation: Respiratory Research Group, Faculty of Pharmacy, University of Sydney, Sydney, NSW 2006, Australia.

ABSTRACT
CXCL10 (IP10) is involved in mast cell migration to airway smooth muscle (ASM) bundles in asthma. We aimed to investigate the role of cytokine-induced MAPK activation in CXCL10 production by ASM cells from people with and without asthma. Confluent growth-arrested ASM cells were treated with inhibitors of the MAPKs ERK, p38, and JNK and transcription factor NF-κB, or vehicle, and stimulated with IL-1β, TNF-α, or IFN-γ, alone or combined (cytomix). CXCL10 mRNA and protein, JNK, NF-κB p65 phosphorylation, and Iκ-Bα protein degradation were assessed using real-time PCR, ELISA, and immunoblotting, respectively. Cytomix, IL-1β, and TNF-α induced CXCL10 mRNA expression more rapidly in asthmatic than nonasthmatic ASM cells. IL-1β and/or TNF-α combined with IFN-γ synergistically increased asthmatic ASM cell CXCL10 release. Inhibitor effects were similar in asthmatic and nonasthmatic cells, but cytomix-induced release was least affected, with only JNK and NF-κB inhibitors halving it. Notably, JNK phosphorylation was markedly less in asthmatic compared with nonasthmatic cells. However, in both, the JNK inhibitor SP600125 reduced JNK phosphorylation and CXCL10 mRNA levels but did not affect CXCL10 mRNA stability or Iκ-Bα degradation. Together, the JNK and NF-κB inhibitors completely inhibited their CXCL10 release. We concluded that, in asthmatic compared with nonasthmatic ASM cells, JNK activation was reduced and CXCL10 gene expression was more rapid following cytomix stimulation. However, in both, JNK activation did not regulate early events leading to NF-κB activation. Thus JNK and NF-κB provide independent therapeutic targets for limiting CXCL10 production and mast cell migration to the ASM in asthma.

Show MeSH
Related in: MedlinePlus