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An intranasal selective antisense oligonucleotide impairs lung cyclooxygenase-2 production and improves inflammation, but worsens airway function, in house dust mite sensitive mice.

Torres R, Herrerias A, Serra-Pagès M, Roca-Ferrer J, Pujols L, Marco A, Picado C, de Mora F - Respir. Res. (2008)

Bottom Line: Finally, mRNA levels of hPGD synthase remained unchanged.Intranasal antisense therapy against COX-2 in vivo mimicked the reported impairment of COX-2 regulation in the airway cells of asthmatic patients.This strategy revealed an unexpected novel dual effect: inflammation was improved but AHR worsened.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pneumology and Respiratory Allergy, Hospital Clínic, IDIBAPS, Universitat de Barcelona, Barcelona, Spain. rosa.torres@uab.cat

ABSTRACT

Background: Despite its reported pro-inflammatory activity, cyclooxygenase (COX)-2 has been proposed to play a protective role in asthma. Accordingly, COX-2 might be down-regulated in the airway cells of asthmatics. This, together with results of experiments to assess the impact of COX-2 blockade in ovalbumin (OVA)-sensitized mice in vivo, led us to propose a novel experimental approach using house dust mite (HDM)-sensitized mice in which we mimicked altered regulation of COX-2.

Methods: Allergic inflammation was induced in BALBc mice by intranasal exposure to HDM for 10 consecutive days. This model reproduces spontaneous exposure to aeroallergens by asthmatic patients. In order to impair, but not fully block, COX-2 production in the airways, some of the animals received an intranasal antisense oligonucleotide. Lung COX-2 expression and activity were measured along with bronchovascular inflammation, airway reactivity, and prostaglandin production.

Results: We observed impaired COX-2 mRNA and protein expression in the lung tissue of selective oligonucleotide-treated sensitized mice. This was accompanied by diminished production of mPGE synthase and PGE2 in the airways. In sensitized mice, the oligonucleotide induced increased airway hyperreactivity (AHR) to methacholine, but a substantially reduced bronchovascular inflammation. Finally, mRNA levels of hPGD synthase remained unchanged.

Conclusion: Intranasal antisense therapy against COX-2 in vivo mimicked the reported impairment of COX-2 regulation in the airway cells of asthmatic patients. This strategy revealed an unexpected novel dual effect: inflammation was improved but AHR worsened. This approach will provide insights into the differential regulation of inflammation and lung function in asthma, and will help identify pharmacological targets within the COX-2/PG system.

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(a). Relative expression of COX-2 mRNA in lung tissue from different treatment groups assayed by real-time PCR. The mRNA expression ratio in the non-sensitized mice was established as 1.0. The level of COX-2 mRNA was significantly diminished in the lungs of sensitized mice treated with the selective antisense oligonucleotide (ASO) when compared with both untreated and control oligonucleotide-treated sensitized mice (n = 12). 2 (b). Levels of COX-2 protein in the lung tissue of non-sensitized mice (n = 3) and in untreated and COX-2 ASO-treated HDM-sensitized mice (n = 6). 2 (c) PGE2 content in the bronchoalveolar lavage (BAL) of non-sensitized (n = 1) and HDM-sensitized treated (n = 4) and untreated (n = 3) mice (* p < 0.05). MM, mismatched oligonucleotide.
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Figure 2: (a). Relative expression of COX-2 mRNA in lung tissue from different treatment groups assayed by real-time PCR. The mRNA expression ratio in the non-sensitized mice was established as 1.0. The level of COX-2 mRNA was significantly diminished in the lungs of sensitized mice treated with the selective antisense oligonucleotide (ASO) when compared with both untreated and control oligonucleotide-treated sensitized mice (n = 12). 2 (b). Levels of COX-2 protein in the lung tissue of non-sensitized mice (n = 3) and in untreated and COX-2 ASO-treated HDM-sensitized mice (n = 6). 2 (c) PGE2 content in the bronchoalveolar lavage (BAL) of non-sensitized (n = 1) and HDM-sensitized treated (n = 4) and untreated (n = 3) mice (* p < 0.05). MM, mismatched oligonucleotide.

Mentions: To support lung COX-2 mRNA assessment, the enzyme's airway protein expression and activity were determined in some of the mice from each experimental group (from 3 to 6 mice see legend of Figure 2b and 2c). Briefly, proteins were extracted from the right lung lobe of each animal using a lysis buffer containing protease inhibitors (Mini complete tablet, Roche Diagnostics, Mannheim, Germany). The concentration of COX-2 protein was determined by ELISA (IBL, Hamburg, Germany).


An intranasal selective antisense oligonucleotide impairs lung cyclooxygenase-2 production and improves inflammation, but worsens airway function, in house dust mite sensitive mice.

Torres R, Herrerias A, Serra-Pagès M, Roca-Ferrer J, Pujols L, Marco A, Picado C, de Mora F - Respir. Res. (2008)

(a). Relative expression of COX-2 mRNA in lung tissue from different treatment groups assayed by real-time PCR. The mRNA expression ratio in the non-sensitized mice was established as 1.0. The level of COX-2 mRNA was significantly diminished in the lungs of sensitized mice treated with the selective antisense oligonucleotide (ASO) when compared with both untreated and control oligonucleotide-treated sensitized mice (n = 12). 2 (b). Levels of COX-2 protein in the lung tissue of non-sensitized mice (n = 3) and in untreated and COX-2 ASO-treated HDM-sensitized mice (n = 6). 2 (c) PGE2 content in the bronchoalveolar lavage (BAL) of non-sensitized (n = 1) and HDM-sensitized treated (n = 4) and untreated (n = 3) mice (* p < 0.05). MM, mismatched oligonucleotide.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2600823&req=5

Figure 2: (a). Relative expression of COX-2 mRNA in lung tissue from different treatment groups assayed by real-time PCR. The mRNA expression ratio in the non-sensitized mice was established as 1.0. The level of COX-2 mRNA was significantly diminished in the lungs of sensitized mice treated with the selective antisense oligonucleotide (ASO) when compared with both untreated and control oligonucleotide-treated sensitized mice (n = 12). 2 (b). Levels of COX-2 protein in the lung tissue of non-sensitized mice (n = 3) and in untreated and COX-2 ASO-treated HDM-sensitized mice (n = 6). 2 (c) PGE2 content in the bronchoalveolar lavage (BAL) of non-sensitized (n = 1) and HDM-sensitized treated (n = 4) and untreated (n = 3) mice (* p < 0.05). MM, mismatched oligonucleotide.
Mentions: To support lung COX-2 mRNA assessment, the enzyme's airway protein expression and activity were determined in some of the mice from each experimental group (from 3 to 6 mice see legend of Figure 2b and 2c). Briefly, proteins were extracted from the right lung lobe of each animal using a lysis buffer containing protease inhibitors (Mini complete tablet, Roche Diagnostics, Mannheim, Germany). The concentration of COX-2 protein was determined by ELISA (IBL, Hamburg, Germany).

Bottom Line: Finally, mRNA levels of hPGD synthase remained unchanged.Intranasal antisense therapy against COX-2 in vivo mimicked the reported impairment of COX-2 regulation in the airway cells of asthmatic patients.This strategy revealed an unexpected novel dual effect: inflammation was improved but AHR worsened.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pneumology and Respiratory Allergy, Hospital Clínic, IDIBAPS, Universitat de Barcelona, Barcelona, Spain. rosa.torres@uab.cat

ABSTRACT

Background: Despite its reported pro-inflammatory activity, cyclooxygenase (COX)-2 has been proposed to play a protective role in asthma. Accordingly, COX-2 might be down-regulated in the airway cells of asthmatics. This, together with results of experiments to assess the impact of COX-2 blockade in ovalbumin (OVA)-sensitized mice in vivo, led us to propose a novel experimental approach using house dust mite (HDM)-sensitized mice in which we mimicked altered regulation of COX-2.

Methods: Allergic inflammation was induced in BALBc mice by intranasal exposure to HDM for 10 consecutive days. This model reproduces spontaneous exposure to aeroallergens by asthmatic patients. In order to impair, but not fully block, COX-2 production in the airways, some of the animals received an intranasal antisense oligonucleotide. Lung COX-2 expression and activity were measured along with bronchovascular inflammation, airway reactivity, and prostaglandin production.

Results: We observed impaired COX-2 mRNA and protein expression in the lung tissue of selective oligonucleotide-treated sensitized mice. This was accompanied by diminished production of mPGE synthase and PGE2 in the airways. In sensitized mice, the oligonucleotide induced increased airway hyperreactivity (AHR) to methacholine, but a substantially reduced bronchovascular inflammation. Finally, mRNA levels of hPGD synthase remained unchanged.

Conclusion: Intranasal antisense therapy against COX-2 in vivo mimicked the reported impairment of COX-2 regulation in the airway cells of asthmatic patients. This strategy revealed an unexpected novel dual effect: inflammation was improved but AHR worsened. This approach will provide insights into the differential regulation of inflammation and lung function in asthma, and will help identify pharmacological targets within the COX-2/PG system.

Show MeSH
Related in: MedlinePlus