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Small interfering RNA against CD86 during allergen challenge blocks experimental allergic asthma.

Asai-Tajiri Y, Matsumoto K, Fukuyama S, Kan-O K, Nakano T, Tonai K, Ohno T, Azuma M, Inoue H, Nakanishi Y - Respir. Res. (2014)

Bottom Line: However, it remains uncertain whether this pathway also has an essential role in the effector phase.These results suggest that local administration of CD86 siRNA during the effector phase ameliorates lines of asthma phenotypes.Targeting airway dendritic cells with siRNA suppresses airway inflammation and hyperresponsiveness in an experimental model of allergic asthma.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan. yukari-a@kokyu.med.kyushu-u.ac.jp.

ABSTRACT

Background: CD86-CD28 interaction has been suggested as the principal costimulatory pathway for the activation and differentiation of naïve T cells in allergic inflammation. However, it remains uncertain whether this pathway also has an essential role in the effector phase. We sought to determine the contribution of CD86 on dendritic cells in the reactivation of allergen-specific Th2 cells.

Methods: We investigated the effects of the downregulation of CD86 by short interfering RNAs (siRNAs) on Th2 cytokine production in the effector phase in vitro and on asthma phenotypes in ovalbumin (OVA)-sensitized and -challenged mice.

Results: Treatment of bone marrow-derived dendritic cells (BMDCs) with CD86 siRNA attenuated LPS-induced upregulation of CD86. CD86 siRNA treatment impaired BMDCs' ability to activate OVA-specific Th2 cells. Intratracheal administration of CD86 siRNA during OVA challenge downregulated CD86 expression in the airway mucosa. CD86 siRNA treatment ameliorated OVA-induced airway eosinophilia, airway hyperresponsiveness, and the elevations of OVA-specific IgE in the sera and IL-5, IL-13, and CCL17 in the bronchoalveolar lavage fluid, but not the goblet cell hyperplasia.

Conclusion: These results suggest that local administration of CD86 siRNA during the effector phase ameliorates lines of asthma phenotypes. Targeting airway dendritic cells with siRNA suppresses airway inflammation and hyperresponsiveness in an experimental model of allergic asthma.

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Related in: MedlinePlus

Silencing CD86 expression by transfection with CD86 small interfering RNA (siRNA) on BMDCs. BMDCs were cultured in the presence of GM-CSF for 7 days. The DCs were transfected with 100nM CD86 siRNA and non-specific (control) siRNA by GeneSilencer®. Twenty-four hours after gene silencing, DCs were activated with LPS for 18 hours. Cells were stained with FITC-conjugated anti-CD11c mAb and biotinylated anti-CD86 mAb followed by phycoerithrin (PE)-conjugated streptavidin. CD86 expression on CD11c-positive cells was assessed by flow cytometry. Histograms represent the number of cells at various fluorescent intensities. MFI data was expressed as the mean ± SEM of 4 mice. * indicates statistically significant (P <0.05) difference between the indicated two groups.
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Fig1: Silencing CD86 expression by transfection with CD86 small interfering RNA (siRNA) on BMDCs. BMDCs were cultured in the presence of GM-CSF for 7 days. The DCs were transfected with 100nM CD86 siRNA and non-specific (control) siRNA by GeneSilencer®. Twenty-four hours after gene silencing, DCs were activated with LPS for 18 hours. Cells were stained with FITC-conjugated anti-CD11c mAb and biotinylated anti-CD86 mAb followed by phycoerithrin (PE)-conjugated streptavidin. CD86 expression on CD11c-positive cells was assessed by flow cytometry. Histograms represent the number of cells at various fluorescent intensities. MFI data was expressed as the mean ± SEM of 4 mice. * indicates statistically significant (P <0.05) difference between the indicated two groups.

Mentions: We first investigated the efficacy of CD86 siRNA in BMDCs. Immature BMDCs showed moderate expression of CD86, which was upregulated by maturation-inducing stimulation with LPS and pOVA for 18 hrs. We compared the expression level of CD86 between non-targeting control siRNA-treated and –untreated BMDCs that were stimulated with LPS and pOVA. There was no significant difference in CD86 expression (control siRNA-treated BMDCs, MFI 2593 ± 193; untreated BMDCs, MFI 3065 ± 191; P =0.106 ). We considered that the transfection protocol per se was neutral in the expression status of CD86. The expression level of CD86 on CD86 siRNA-treated BMDCs was significantly lower than that on non-targeting control siRNA-treated BMDCs [P =0.0014] (Figure 1). The expression of CD80 was unaffected by treatment with CD86 siRNA in our preliminary study [9]. These results suggest that treatment with CD86 siRNA specifically suppresses the activation-induced upregulation of CD86 on BMDCs in vitro.Figure 1


Small interfering RNA against CD86 during allergen challenge blocks experimental allergic asthma.

Asai-Tajiri Y, Matsumoto K, Fukuyama S, Kan-O K, Nakano T, Tonai K, Ohno T, Azuma M, Inoue H, Nakanishi Y - Respir. Res. (2014)

Silencing CD86 expression by transfection with CD86 small interfering RNA (siRNA) on BMDCs. BMDCs were cultured in the presence of GM-CSF for 7 days. The DCs were transfected with 100nM CD86 siRNA and non-specific (control) siRNA by GeneSilencer®. Twenty-four hours after gene silencing, DCs were activated with LPS for 18 hours. Cells were stained with FITC-conjugated anti-CD11c mAb and biotinylated anti-CD86 mAb followed by phycoerithrin (PE)-conjugated streptavidin. CD86 expression on CD11c-positive cells was assessed by flow cytometry. Histograms represent the number of cells at various fluorescent intensities. MFI data was expressed as the mean ± SEM of 4 mice. * indicates statistically significant (P <0.05) difference between the indicated two groups.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4216659&req=5

Fig1: Silencing CD86 expression by transfection with CD86 small interfering RNA (siRNA) on BMDCs. BMDCs were cultured in the presence of GM-CSF for 7 days. The DCs were transfected with 100nM CD86 siRNA and non-specific (control) siRNA by GeneSilencer®. Twenty-four hours after gene silencing, DCs were activated with LPS for 18 hours. Cells were stained with FITC-conjugated anti-CD11c mAb and biotinylated anti-CD86 mAb followed by phycoerithrin (PE)-conjugated streptavidin. CD86 expression on CD11c-positive cells was assessed by flow cytometry. Histograms represent the number of cells at various fluorescent intensities. MFI data was expressed as the mean ± SEM of 4 mice. * indicates statistically significant (P <0.05) difference between the indicated two groups.
Mentions: We first investigated the efficacy of CD86 siRNA in BMDCs. Immature BMDCs showed moderate expression of CD86, which was upregulated by maturation-inducing stimulation with LPS and pOVA for 18 hrs. We compared the expression level of CD86 between non-targeting control siRNA-treated and –untreated BMDCs that were stimulated with LPS and pOVA. There was no significant difference in CD86 expression (control siRNA-treated BMDCs, MFI 2593 ± 193; untreated BMDCs, MFI 3065 ± 191; P =0.106 ). We considered that the transfection protocol per se was neutral in the expression status of CD86. The expression level of CD86 on CD86 siRNA-treated BMDCs was significantly lower than that on non-targeting control siRNA-treated BMDCs [P =0.0014] (Figure 1). The expression of CD80 was unaffected by treatment with CD86 siRNA in our preliminary study [9]. These results suggest that treatment with CD86 siRNA specifically suppresses the activation-induced upregulation of CD86 on BMDCs in vitro.Figure 1

Bottom Line: However, it remains uncertain whether this pathway also has an essential role in the effector phase.These results suggest that local administration of CD86 siRNA during the effector phase ameliorates lines of asthma phenotypes.Targeting airway dendritic cells with siRNA suppresses airway inflammation and hyperresponsiveness in an experimental model of allergic asthma.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan. yukari-a@kokyu.med.kyushu-u.ac.jp.

ABSTRACT

Background: CD86-CD28 interaction has been suggested as the principal costimulatory pathway for the activation and differentiation of naïve T cells in allergic inflammation. However, it remains uncertain whether this pathway also has an essential role in the effector phase. We sought to determine the contribution of CD86 on dendritic cells in the reactivation of allergen-specific Th2 cells.

Methods: We investigated the effects of the downregulation of CD86 by short interfering RNAs (siRNAs) on Th2 cytokine production in the effector phase in vitro and on asthma phenotypes in ovalbumin (OVA)-sensitized and -challenged mice.

Results: Treatment of bone marrow-derived dendritic cells (BMDCs) with CD86 siRNA attenuated LPS-induced upregulation of CD86. CD86 siRNA treatment impaired BMDCs' ability to activate OVA-specific Th2 cells. Intratracheal administration of CD86 siRNA during OVA challenge downregulated CD86 expression in the airway mucosa. CD86 siRNA treatment ameliorated OVA-induced airway eosinophilia, airway hyperresponsiveness, and the elevations of OVA-specific IgE in the sera and IL-5, IL-13, and CCL17 in the bronchoalveolar lavage fluid, but not the goblet cell hyperplasia.

Conclusion: These results suggest that local administration of CD86 siRNA during the effector phase ameliorates lines of asthma phenotypes. Targeting airway dendritic cells with siRNA suppresses airway inflammation and hyperresponsiveness in an experimental model of allergic asthma.

Show MeSH
Related in: MedlinePlus