Establishment of an optimized murine asthma model forHumulus pollen allergy
Forty mice were randomly divided into four groups. Two different routes
and doses were used for sensitization, based on studies by Ya-li et
al.[10] and Conejero et
al.[15] (Fig. 1 A). Briefly, mice in
the SC sensitization group (SC.HSE, n=10) were sensitized via SC
injection
on the back of the neck with 25 μg
of HSE adsorbed to alum adjuvant (Imject Alum; Pierce, Rockford, IL,USA)
in a volume of 200 μL. Mice in the SC control group (SC.PBS, n=10)
received SC injections of PBS adsorbed to alum adjuvant (200 μL). Mice
in the IP sensitization group (IP.HSE, n=10) were sensitized via IP
injection—at approximately 1 cm
from the intersection point of a line running between the thighs and the
midline—with 300 μg of HSE adsorbed to alum adjuvant in a volume of
200 μL. Mice in the IP control group (IP.PBS, n=10) received IP
injections of PBS adsorbed to alum adjuvant (200 μL). Sensitization
injections were administered at weekly intervals for 3 weeks. To induce
asthma, beginning on day 21, sensitized and control mice were placed in
a plastic box and challenged for 30 min daily via the airways with
aerosolized 1% HSE or an equal volume of saline (control group) using a
jet nebulizer (BARI Co., Ltd., Germany), for 7 consecutive days. Airway
hyperresponsiveness (AHR) to
methacholine
(Mch, Sigma-Aldrich, St. Louis,
MO, USA) was measured 24 h after the last challenge in all animals. On
day 29, mice were anesthetized with 2% sodium
pentobarbital;
sera were then obtained for measurement of specific IgE levels and of
IgE profiles binding to HSE. Additionally, bronchoalveolar lavage fluid
(BALF) was prepared from the right lung for inflammatory cell
enumeration and cytokine quantitation. Paraffin sections from the left
lung were used for histological assessment. The spleen was homogenized
for cytokine measurements.