Additional file 1 of Feasibility and acceptability of remotely monitoring spirometry and pulse oximetry as part of interstitial lung disease clinical care: a single arm observational study

Supplementary Material 1: Additional Figure 1: Heatmap demonstrating frequency of spirometry attempts throughout the 91 day study period. Each horizontal bar represents a participant within the study. Each vertical line within a bar represents a day on which home spirometry was performed

Exogenous TGF-β₂ suppresses IFN-β release from virally infected (A) or poly IC exposed (B) PBEC cultures from non-asthmatic donors.

<p>PBEC cultures were infected with RV1B (5000 TCID₅₀ units/10⁵ cells) (n = 10) or treated with poly IC (n = 5) in the presence or absence of TGF-β₂ which was used at 1 (black bars in B) or 10 ng/ml (panel A and grey bars in B). Culture supernatants were harvested 48 hours p.i (A) or 8 h post stimulation (B) and IFN-β protein levels were measured by ELISA. In B, the data are expressed as a % of control cultures treated with poly IC in the absence of TGF-β₂ (median (IQR) IFN-β release  = 346 (1135) and 369 (1390) pg/ml for cells treated with 1 or 10 µg/ml Poly IC, respectively. The data were analyzed using Wilcoxon’s rank sum test (A) or using a paired t-test for normally distributed data (B).</p

The effect of anti-TGF-β antibodies on release of IFN-β and IFNλ1/IL-29 protein in response to poly IC.

<p>PBECs from 6 asthmatic donors were treated with 0.1–10 µg/ml poly IC in the presence of neutralizing anti-TGF-β antibodies (black bars) or an IgG isotype control (open bars) and incubated for 24 h. Supernatants were removed and IFN-β (A) or IFN-λ1/IL-29 protein levels (B) were measured by ELISA. Graphs show (mean±SEM) IFN produced in pg/ml the presence of the control or anti TGF-β antibodies.</p

The effect of neutralizing endogenous TGF-β on RV replication.

<p>PBECs from 8 asthmatic donors or 6 non-asthmatic control subjects were pretreated for 24 h in the presence of a neutralizing anti pan TGF-β antibody or isotype control antibody before infection with RV1B (1000 units/10⁵ cells) for 48 h. The fold-decrease in viral replication by the neutralizing antibody was plotted as a ratio of the TCID₅₀/ml of antibody-treated versus isotype controls. The figure shows median and interquartile range, with individual data points superimposed. Data were analysed using a Mann Whitney U test.</p

The effect of TGF-β neutralization on basal SMAD2 activation.

<p>PBECs from asthmatic donors were treated with RV and anti TGF-β antibody, as indicated, as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044580#pone-0044580-g005" target="_blank">Figure 5</a>. Cell lysates were harvested at 1, 4, and 6 hours post-virus infection and Smad-2 phosphorylation was analysed by Western blotting. A representative Western blot is shown in (A) and densitometric quantification of the experiment repeated using PBECs from 3 different asthmatic subjects is shown in (B).</p

The effect of exogenous TGF-β₂ on RV replication.

<p>PBECs from 3 non-asthmatic volunteers were pre-incubated with 0, 1, 10, and 25 ng/ml of TGF- β₂ for 24 h, followed by infection with RV1B at 5000 TCID₅₀ units/10⁵ cells. Cells were then further incubated for 48 h in the presence or absence of TGF-β₂, as indicated. Viral replication at 24 h was measured as vRNA by RT-qPCR (A) and at 48 h by release of infectious virions into culture supernatants by TCID₅₀ assays (B). The graph (C) shows data for infectious virus release from PBECs from 10 non-asthmatic donors treated without or with 10 ng/ml TGF-β₂, followed by infection with RV1B at 5000 TCID₅₀ units/10⁵ cells for 48 hours. Statistical comparison was made using a Wilcoxon rank sum test. The # mark in C indicates where 2 data points overlap (1.8e⁶→3.1e⁶ TCID₅₀ units/ml).</p

Suppression of viral replication in PBECs from asthmatic donors by neutralization of endogenous TGF-β.

<p>PBECs from asthmatic donors were pretreated for 24 h in the presence of a neutralizing anti pan TGF-β antibody or isotype control antibody before infection with RV1B (1000 units/10⁵ cells) for 48 h. In A, viral titre was determined as TCID₅₀/ml using culture supernatants obtained 48 h p.i. In B, IFN-β protein was measured at 48 h and was expressed as a ratio of the viral load measured as TCID₅₀ units. The data were analyzed using Wilcoxon’s rank sum test.</p

Neutralizing endogenous TGF-β suppresses RV1B mediated SOCS-1 and SOCS-3 gene expression in asthmatic PBECs.

<p>Samples were treated as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044580#pone-0044580-g005" target="_blank">Figure 5</a>. SOCS-1 (A) and SOCS-3 (B) gene expression were measured in 7 asthmatic subjects at 48 h p.i. by RT-qPCR and normalized to housekeeping genes. Results were plotted as relative fold-induction using the ΔΔCt method. The Wilcoxon rank sum test was used to analyse statistical significance.</p

Exogenous TGF-β₂ suppresses IFN-λ1/IL-29 release from virally infected (A) (n = 10) or poly IC (n = 4) exposed (B) PBEC cultures from non-asthmatic donors.

<p>IFNλ1/IL-29 protein levels were measured by ELISA from RV-infected or poly IC exposed PBECs treated with TGF-β₂ as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044580#pone-0044580-g002" target="_blank">Figure 2</a>. Median (IQR) IFN-λ₁ release  = 3896 (2766) and 4932 (4941) pg/ml for cells treated with 1 or 10 µg/ml Poly IC, respectively.</p

Rheumatoid arthritis and idiopathic pulmonary fibrosis: a bidirectional Mendelian randomisation study

BackgroundA usual interstitial pneumonia (UIP) pattern of lung injury is a key feature of idiopathic pulmonary fibrosis (IPF) and is also observed in up to 40% of individuals with rheumatoid arthritis (RA)-associated interstitial lung disease (RA-ILD). The RA-UIP phenotype could result from either a causal relationship of RA on UIP or vice versa, or from a simple co-occurrence of RA and IPF due to shared demographic, genetic or environmental risk factors.MethodsWe used two-sample bidirectional Mendelian randomisation (MR) to test the hypothesis of a causal effect of RA on UIP and of UIP on RA, using variants from genome-wide association studies (GWAS) of RA (separately for seropositive (18 019 cases and 991 604 controls) and seronegative (8515 cases and 1 015 471 controls) RA) and of IPF (4125 cases and 20 464 controls) as genetic instruments. Sensitivity analyses were conducted to assess the robustness of the results to violations of the MR assumptions.FindingsIPF showed a significant causal effect on seropositive RA, with developing IPF increasing the risk of seropositive RA (OR=1.06, 95% CI: 1.04 to 1.08, p</p