IL-17A produced by T cells drives airway hyper-responsiveness in mice and enhances

mouse and human airway smooth muscle contraction

Makoto Kudo, Andrew C. Melton, Chun Chen, Mary B. Engler, Katherine E. Huang, Xin Ren,
Yanli Wang, Xin Bernstein, John T. Li, Kamran Atabai, Xiaozhu Huang, Dean Sheppard

Supplementary Figure 1. Itgb8f/f x CD11c-cre mice are protected from AHR after house
dust mite treatment, but development similar levels of AHR as control mice after
intranasal administration of IL-13. Pulmonary resistance measurements in control and Itgb8f/f
x CD11c-cre mice after treatment with house dust mite (HDM, a) or in response to increasing
doses of acetylcholine 48 hours after a single intranasal administration of 10 g IL-13 or saline
(b). *P < 0.05 for IL-13 treated control mice compared to saline treated mice and **P < 0.05 in
IL-13 treated control and Itgb8f/f x CD11c-cre mice compared to saline treated control and
Itgb8f/f x CD11c-cre mice.

Nature Medicine doi:10.1038/nm.2684

Supplementary Figure 2. Similar enhancement of airway smooth muscle contraction by IL17A after epithelium removal. (a) H&E stained sections from tracheal rings that underwent
brushing to remove epithelium or a sham treatment (scale bar = 100 m for 50X images and 10
m for 400X images). (b) Contractile force measurements in tracheal rings with epithelium
removed by brushing or sham treated. Rings were then treated for 12 hours with or without 100
ng ml-1 IL-17A and stimulated with MCh or KCl. **P < 0.01 and ***P < 0.001 in IL-17A
treated samples compared to control samples. Contractile force measurements from samples with
epithelium removed were not statistically significant (NS) compared to sham samples.

Nature Medicine doi:10.1038/nm.2684

Supplementary Figure 3. Pretreatment of lung slices with IL-17A does not affect calcium
oscillations in response to methacholine. Lung slices were treated for 24 hours with 100 ng
ml-1 IL-17A or vehicle (Control) and then loaded with Oregon Green 488 BAPTA-AM, a
fluorescent calcium indicator dye. Spinning-disk confocal microscopy was used to image
fluorescence in lung slices. (a) Regions of interest (5 X 5 pixel regions) were dened in single
airway smooth muscle cells and fluorescence changes in response to 10 M methacholine were
expressed as the ratio of uorescence intensity to the fluorescence intensity prior to methacholine
treatment (F/F0). (b) Calcium oscillations after stimulation with 10 M methacholine were
measured as the number of transient calcium peaks per minute.

Nature Medicine doi:10.1038/nm.2684