Effect of dsRNA on BSMC expression and production of IFN-β and IFN-λ₁.

<p>BSMCs were stimulated with dsRNA and expression of IFN-β (A, B) and IFN-λ₁ (E, F) mRNA was analysed by RT-qPCR after 3 h (A, E) or 24 h (B, F). Generation of IFN-β (C) and IFN-λ₁ protein (G) was determined after 24 h in the cell supernatants by ELISA. Chloroquine (Cq) and dexamethasone (Dex) added 1 h prior to dsRNA stimulation inhibited the early (3 h) response, but only dexamethasone inhibited the late (24 h) mRNA response. IFN-β protein was reduced with chloroquine treatment (D). Data are presented as mean with SEM and n = 8–10 (BSMCs from three individual donors). *p≤0.05, **p≤0.01 and ***p≤0.001 compared to non-stimulated cells (control). ^##p≤0.01 and ^###p≤0.001 compared to 10 µg/ml dsRNA.</p

dsRNA and dsRNA/LyoVec upregulate expression of the RLRs RIG-I and MDA5 in BSMCs.

<p>BSMC were treated with dsRNA or the RLR ligand dsRNA/LyoVec (dsRNA/LV) for 24 h. mRNA (A, C) and protein (B, D) expression of RIG-I (A, B) and MDA5 (C, D) was increased concentration-dependently. n = 9 for mRNA analysis and n = 6 for protein analysis (BSMCs from three individual donors). mRNA data are presented as mean with SEM and representative western blot images of each receptor protein are shown. *p≤0.05, **p≤0.01 and ***p≤0.001 compared to non-stimulated cells (control). ^#p≤0.05, ^##p≤0.01 and ^###p≤0.001 compared to 10 µg/ml dsRNA.</p

BSMC expression of RIG-I and MDA5 is upregulated upon RV1B infection.

<p>Infection with increasing doses (MOI) of RV1B for 24 h upregulated BSMC mRNA expression of RIG-I (A) and MDA5 (D) and the expression of both RLRs were further enhanced 48 h post-infection (B, E). Chloroquine (10 µg/ml) partly reduced the effect of RV1B on both RIG-I and MDA5 mRNA expression, especially after 48 h (C, F). RIG-I protein (G) was upregulated at 24 h and more enhanced at the infection dose of 0.1 MOI 48 h post-infection. MDA5 protein (H) was upregulated 48 h post-infection only. Chloroquine reduced RV1B-induced effects on both RIG-I (G) and MDA5 (H) protein at 48 h. Data are presented as mean with SEM and n = 6 for 24 h RV1B infection, n = 3 for 48 h infection and n = 2–3 for chloroquine experiments (BSMCs from three individual donors). Representative western blot images of each receptor protein are shown. *p≤0.05 and **p≤0.01 compared to non-infected cells (control).</p

RV1B induces BSMC mRNA expression of IFNs.

<p>BSMCs exposed to RV1B at increasing doses (MOI = multiplicity of infection) induced expression of IFN-β (A) and IFN-λ₁ (D) 24 h post-infection as determined by RT-qPCR. IFN-β and IFN-λ₁ expression was further enhanced after 48 h (B, E). Chloroquine (10 µg/ml) reduced RV1B-induced IFN-β expression after 24 and 48 h (C), whereas IFN- λ₁ expression was reduced after 48 h only (F). Data are presented as mean with SEM and n = 6 for 24 h RV1B infection, n = 3 for 48 h infection and n = 2–3 for chloroquine experiments (BSMCs from three individual donors). *p≤0.05 and **p≤0.01 compared to non-infected cells (control).</p

RLR activation by dsRNA/LyoVec evokes BSMC mRNA expression and protein production of IFN-β and IFN-λ₁.

<p>IFN-β and IFN-λ₁ mRNA expression was increased by dsRNA/LyoVec (dsRNA/LV) after 3 h (A, D) and 24 h (B, E) and so was secretion of IFN-β and IFN-λ₁ after 24 h (C, F). Data are presented as mean with SEM and n = 8–10 (BSMCs from three individual donors). *p≤0.05, **p≤0.01 and ***p≤0.001 compared to non-stimulated cells (control).</p

Remodeling of extra-bronchial lung vasculature following allergic airway inflammation-3

<p><b>Copyright information:</b></p><p>Taken from "Remodeling of extra-bronchial lung vasculature following allergic airway inflammation"</p><p>http://respiratory-research.com/content/9/1/18</p><p>Respiratory Research 2008;9(1):18-18.</p><p>Published online 8 Feb 2008</p><p>PMCID:PMC2254605.</p><p></p>le actin and procollagen I) increased significantly following allergen exposure. Occasional smooth muscle cells may produce procollagen I; however smooth muscle cells are not normally solitary. Sporadic myofibroblasts were also visible before allergen challenge, but when correlated to the length of the basement membrane (BM) the values are very small and closed in on zero. The data are given as mean ± SEM and compared against control using the Wilcoxon Signed-ranks test, * indicates p < 0.05

Remodeling of extra-bronchial lung vasculature following allergic airway inflammation-0

<p><b>Copyright information:</b></p><p>Taken from "Remodeling of extra-bronchial lung vasculature following allergic airway inflammation"</p><p>http://respiratory-research.com/content/9/1/18</p><p>Respiratory Research 2008;9(1):18-18.</p><p>Published online 8 Feb 2008</p><p>PMCID:PMC2254605.</p><p></p>reased following allergen challenge. Proliferation was detected using the proliferation-marker Ki67. A base line proliferation was present also in controls, however the number was very low and when correlated to the length of the basement membrane (BM), the values closed in on zero. The data are given as mean ± SEM and compared against control using the Wilcoxon Signed-ranks test, * indicates p < 0.05

Remodeling of extra-bronchial lung vasculature following allergic airway inflammation-2

<p><b>Copyright information:</b></p><p>Taken from "Remodeling of extra-bronchial lung vasculature following allergic airway inflammation"</p><p>http://respiratory-research.com/content/9/1/18</p><p>Respiratory Research 2008;9(1):18-18.</p><p>Published online 8 Feb 2008</p><p>PMCID:PMC2254605.</p><p></p>(procollagen I: brown) and myofibroblasts (here defined as solitary cells co-positive for α-smooth muscle actin and procollagen I: co-positive), in both small solitary (A-B) and mid-sized solitary (C-D) vessels. In comparison with controls (A and C) vessels from OVA exposed animals (B and D) show a significantly increased smooth muscle area as well as increased number of myofibroblasts (arrows) and procollagen I-producing cells. Vascular lumen is indicated by stars. Scale bar represents 50 μm

Remodeling of extra-bronchial lung vasculature following allergic airway inflammation-1

<p><b>Copyright information:</b></p><p>Taken from "Remodeling of extra-bronchial lung vasculature following allergic airway inflammation"</p><p>http://respiratory-research.com/content/9/1/18</p><p>Respiratory Research 2008;9(1):18-18.</p><p>Published online 8 Feb 2008</p><p>PMCID:PMC2254605.</p><p></p> approximately 5 times and in mid-sized approximately 3 times. Smooth muscle was detected by labelling with α-smooth muscle actin, and the positively stained area measured by digital image analysis and correlated to the length of the basement membrane (BM). The data are given as mean ± SEM and compared against control using the Wilcoxon Signed-ranks test, * indicates p < 0.05

Additional file 3: Figure S2. of IL-1β mediates lung neutrophilia and IL-33 expression in a mouse model of viral-induced asthma exacerbation

Increased immune cells and total protein in BALF in both WT and IL-1β−/− groups 24 h after the last HDM challenge alone. Total protein in BALF (a), total cells from BALF (b) and total neutrophil count in BALF (c). Data are presented as mean ± SEM, n = 4–7 mice in each group. ## = p < 0.01 compared to respective HDM/Saline control, and # = p < 0.05 compared to respective HDM/Saline control. (TIFF 941 kb