The Role of Epithelial Cells in Allergic Asthma

Immune Response
Emily V. Kim1, Jaclyn W. McAlees2, Ian P. Lewkowich2
1University Honors Program, University of Cincinnati, Cincinnati, OH
2Division of Immunobiology, Cincinnati Childrens Hospital Medical Center, Cincinnati, OH

Abstract
The symptoms and pathophysiology of allergic asthma
are precipitated by the action of the cytokine IL-13 on
airway structural cells. The presence of IL-17A
exacerbates IL-13-driven immune responses and is
associated with more severe allergic asthma
phenotypes. We examined the effects of IL-13 in the
presence and absence of IL-17A on mouse airway
epithelial cells mTECs, which serve as the interface
between the allergens in the environment and the
developing immune response. The mTECs are
differentiated to have an apical (air) and basal (liquid)
side as they would in the airways. Our data show that
IL-17A increases IL-13-mediated gene expression in a
concentration dependent manner and that gene
regulation differs when the cytokines are applied on the
apical or basal sides of the differentiated mTECs. These
results suggest that the airway epithelium is a potential
target for therapeutic endeavors in cases of IL-17A
associated severe allergic asthma.

Methods

Conclusion

Mouse tracheal epithelial cells (mTECs) are isolated from the

tracheas of 4-6 week old mice using enzyme digestion.
Fibroblasts are removed and the mTECs are cultured on
collagen-coated transwells under submersion conditions (media
on the basal and apical sides) until they become confluent.
Confluency is achieved when the transpeithelial resistance
reaches 1000 ohms. Once confluent, the media is removed
from the apical side of the cells to establish an air-liquid
interphase in which one side of the cells is in contact with the
collagen-coated membrane and media and the other side is
exposed to the air. The mTECs are cultured in air-liquid
interphase for a minimum of 10 days prior to treatment to allow
for differentiation3. The cells are treated either apically or
basally with IL-13 in the presence and absence of IL-17 for 1824 hours. Total RNA is isolated and gene expression is
determined by Real Time PCR.

Intl-1 and Tnfsf13 were induced to some degree by

both apical and basal IL-13 treatment.
IL-13-induced Intl-1 expression was augmented by
basal IL-17A treatment in a concentrationdependent manner, as were several other IL-13induced genes (CCL26, Intl-2, CCL24).
IL-13-induced Tnfsf13 expression was augmented
by apical IL-17A treatment in a concentrationdependent manner, as were several other IL-13induced genes. (IL-13Ra1, ALOX15, Arg1)
Figure 1. Representation of air liquid interface.
Adapted from: Role of membrane traffic in the
generation of epithelial cell asymmetry. 2012.
Nature Cell Biology 14: 1235-1243.

Results
15000

Introduction

Apical

Basal

0 .25 .5 1 2.5 10

0 .25 .5 1 2.5 10

1200

10000

5000

800
400

0 .25 .5 1 2.5 10

0 .25 .5 1 2.5 10

ng/ml of IL-13

Apical

Basal

40000

20000

12000

Intl-1:S14

Tnfsf13:S14 ratio

60000

Apical

Basal

8000

4000

Our data show that the airway epithelium is a potential

target for IL-13 and IL-17A mediated changes, which
could play a role in allergic asthma development and
progression. Because there is differential gene
regulation on the apical versus basal sides of the cells,
identifying where cytokines originate to produce these
responses could aid in creating new pharmaceutical
products for the treatment of severe allergic asthma.
Apical responses may be regulated by cytokines
produced by CD4+ T cells that have migrated to the
airspace, whereas cytokines produced by CD4+ T cells
in the parenchyma of the lung may elicit a basal
response. Future directions include defining a panel of
genes that can be screened to determine exposure to
IL-13 and IL-17A cytokines in vivo, i.e. in a mouse
model.

Acknowledgements
Jaclyn McAlees, Ph.D.
Sara Stoffers, M.S.
Alan Gordillo
Phoebe Richgels, M.S.
Ian Lewkowich, Ph.D.
NIH RO1 HL122300
University Honors Program- Megan Minton
Biomedical Research and Mentoring Program

0
0.
5n
0. g/m
5n l
g/ IL0. m 13
0. 5n l IL + 0. Me
5n g/ -1 0 5n d
0. g/mml I 3 + .25n g/mia o
5n l L- 0 g l n
g/ IL 13 .5 /m IL ly
m -1 + ng l I -1
l I 3 1 /m L- 3
L- + n l 1
13 2. g/ IL 7a
0.
5n
+ 5ng ml -17
10 /m IL- a
0. g/m
ng l 17
5n l
/m IL- a
g/ ILl I 17
0. m 13
0. 5n l IL + 0. Me L-1 a
5n g/ -1 0 5n d 7
0. g/mml I 3 + .25n g/mia o a
5n l L- 0 g l n
g/ IL 13 .5 /m IL ly
m -1 + ng l I -1
l I 3 1 /m L- 3
L- + n l 1
13 2. g/ IL 7a
+ 5ng ml -17
10 /m IL- a
ng l 17
/m IL- a
l I 17
L- a
17
a

ng/ml of IL-13

0.
5n
0. g/m
5n l
g/ IL0. m 13
0. 5n l IL + 0. Me
5n g/ -1 0 5n d
0. g/mml I 3 + .25n g/mia o
5n l L- 0 g l n
g/ IL 13 .5 /m IL ly
m -1 + ng l I -1
l I 3 1 /m L- 3
L- + n l 1
13 2. g/ IL 7a
0.
5n
+ 5ng ml -17
10 /m IL- a
0. g/m
ng l 17
5n l
/m IL- a
g/ ILl I 17
0. m 13
0. 5n l IL + 0. Me L-1 a
5n g/ -1 0 5n d 7
0. g/mml I 3 + .25n g/mia o a
5n l L- 0 g l n
g/ IL 13 .5 /m IL ly
m -1 + ng l I -1
l I 3 1 /m L- 3
L- + n l 1
13 2. g/ IL 7a
+ 5ng ml -17
10 /m IL- a
ng l 17
/m IL- a
l I 17
L- a
17
a

Allergic asthma is the result of a maladaptive immune

response and the mechanisms underlying this response
are not well defined. Airway epithelial cells are the first
cells to encounter inhaled allergens and have been
shown to shape downstream immune responses that
lead to development of allergic asthma1. A typical
allergic response involves Th2 cells, which produce the
cytokines IL-13 in addition to other factors. Patients
with Th2 immune responses are easily treated with
current therapies such as inhaled steroids. However, in
addition to Th2 cells, patients with more severe allergic
asthma have Th17 cells, which produce the cytokine IL17A. These patients disease is not well controlled by
current therapies. Data from the Lewkowich laboratory
and others suggests that IL-17A exacerbates IL-13mediated asthma symptoms to drive the development
of more severe disease2. A better understanding of
how IL-13 and IL-17 affect airway epithelial cells may
provide information about how severe disease
develops, why it is not controlled by current therapies,
and may possibly identify new therapeutic targets.
Thus, the goal of my research is to understand how IL17 affects IL-13-mediated responses in airway epithelial
cells through synergism of the two cytokines. We
hypothesize that airway epithelial cells exposed to the
cytokines IL-13 and IL-17A will exhibit exacerbated IL13-mediated responses.

1600

Basal

Intl-1:S14

Tnfsf13:S14 ratio

Apical

Discussion

Figure 2. Cultured and differentiated MTECs were treated apically or basally with (A) the indicated
amounts of IL-13 or (B) 0.5ng/ml of IL-13 and increasing concentrations of IL-17A (0.25-10ng/ml) for ~20
hours. Cells were collected, total RNA was isolated and IL-13-mediated gene expression was analyzed by
Real-Time PCR. The expression profiles of two repesentative genes are shown.

References
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D, Leigh R. Epithelial cells and airway diseases.

Immunological Reviews 2011; 242:186-204.
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activation.
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Clin
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3McAlees, JW, et al. Mucosal Immunol. 2015 July ; 8(4): 863
873. doi:10.1038/mi.2014.117.