Supplementary methods and figures
Supplementary methods and figures
Files in this Data Supplement:
- Figure S1 - Protocol for in vivo NA model. (A) Illustration of the time points for treatment with NA, anti-NGF and control IgG antibodies (anti-NGF, IgG), BrdU and days of analyses (d0-d29). (B) Representation of different groups of mice, their annotation in the paper, treatment and day of sacrifice and analyses. n=6 mice were sacrificed per group per analyses.
- Figure S2 - Inhibition of NGF and TrkA. (A) Inhibition of mRNA and (B, C) protein of NGF and TrkA respectively in proliferating LA4 cells after incubation with their respective siRNA�s - siRNA NGF and siRNA TrkA for 24 hours compared to cells grown in medium containing only transfection reagent (medium). (D) Inhibition of NGF protein after incubation of LA4 cells with anti-NGF (625ng/ml, black bar) compared to medium (white bar) or medium containing IgG (625ng/ml, grey bar) for 24 hours. Inhibition by anti-NGF was analysed in medium containing 1% or 10% FCS. n=6 samples were analysed per group. All numerical data are expressed as mean ± SEM. ns = non significant, * = p< 0.05, ** = p< 0.01, *** = p< 0.001.
- Figure S3 - Time and Dose response of LA4 and AECII cell proliferation. (A) Proliferation of LA4 cells in 0.1% (white bar), 1% (grey bar) and 10% (black bar) FCS containing medium, analysed at 24h, 48h and 72h. (B) FCS Concentration dependent effect of NGF on proliferation of LA4 cells. LA4 cell proliferation is analysed at 24 hours after incubation with medium (white bar), medium containing NGF (light grey bar), medium containing anti-NGF (black bar) or control IgG (dark grey bar). (C) Proliferation of AECII in 0.1% (white bar), 1% (grey bar) and 10% (black bar) FCS containing medium, analysed at 24h, 48h and 72h. Figure represents n=6 samples ± SEM for each group. * = p< 0.05, **=p <_0.01 _="p<" _0.001.--="_0.001.--" end="end" desc="desc" figure_s3.jpg="figure_s3.jpg" _--="_--">
- Figure S4 - Exfoliated Clara cells at day 1 after NA induced injury. (A) Clara cells in WT and NGFtg control animals (left panels) and in NA treated WT (WT-NA) and NGFtg (NGFtg-NA) animals after 24 hours (right panels). (B) Exfoliated cell numbers of WT (white bars) and NGFtg (black bars) untreated control animals (control) or treated with NA (NA) were counted and compared in the BALF. WT and NGFtg animals treated with NA reveal similar denudation at day 1. Bars represent n= 6 for each group. n.s = non significant.
- Figure S5 - Inhibition of NGF in vivo. 6-8 weeks old WT C57BL/6 female mice treated with or without i.p NA were applied with 24 �g intranasal IgG (IgG, NA+IgG,) or anti-NGF antibody (anti-NGF, NA+anti-NGF). NGF levels were measured in bronchoalveolar lavage fluid (BALF) at day 1, 2 and 3 by ELISA. n=9 animals, *=p <_0.05 _="p<0.01.<!--" end="end" desc="desc" figure_s5.jpg="figure_s5.jpg" _--="_--">
- Supplementary methods