Dissecting the inflammatory twitch in allergically inflamed mice

Abstract

We have previously advanced the hypothesis that the allergic inflammatory response in the lungs occurs as a self-limited sequence of events that begins with the onset of inflammation and then resolves back to baseline over a predetermined time course (Pothen JJ, Poynter ME, Bates JH. J Immunol 190: 3510-3516, 2013). In the present study we tested a key prediction of this hypothesis, which is that the instigation of the allergic inflammatory response should be accompanied by a later refractory period during which the response cannot be reinitiated. We challenged groups of ovalbumin-sensitized BALB/c mice for 3, 14, 21 and 31 consecutive days with aerosolized ovalbumin. We measured airways responsiveness as well as cell counts and cytokines in bronchoalveolar lavage fluid after the final challenge in subgroups from each group. In other subgroups we performed the same measurements following rest periods and after a final single recall challenge with antigen. We determined that the refractory periods for GM-CSF, KC, and IL-5 are no longer than 10 days, while those for IFNγ and IL-10 are no longer than 28 days. The refractory periods for total leukocytes and neutrophils were no greater than 28 days, while that for eosinophils was more than 28 days. The refractory period for airways resistance was less than 17, while for lung elastance it was longer than 28 days. Our results thus demonstrate that the components of the allergic inflammatory response in the lung have finite refractory periods, with the refractory period of the entire response being in the order of a month.

Keywords: bronchoalveolar lavage; cytokines; lung function; refractory period.

Fig. 1.

Closed circles show concentrations of cytokines measured in the left lobes of lungs of mice, characterized 24 h following the last day of daily antigen challenge, and on day 31 immediately preceding and following recall challenge with ovalbumin (OVA): GM-CSF (A), KC (B), IL-5 (C), IFN-γ (D), and IL-10 (E). Open circles show corresponding control measurements made in control mouse lungs 24 h after 3 days of saline and a recall challenge with saline on day 31. All points are reported as means ± SE. *Value statistically higher than control 1. **Value statistically higher than control 2. ***Value statistically higher than the other three values in OVA-challenged animals.

Fig. 2.

Closed circles show concentrations of cytokines measured in the left lobes of mouse lungs 24 h following the final daily OVA challenge, and on day 31 immediately following recall challenge: IL-4 (A), IL-17 (B), IL-3 (C), and IL-6 (D). Open circles show corresponding control measurements made 24 h after day 3 of saline challenge (control 1: left-hand point) and 1 day after saline recall challenge (control 2: right-hand point). Points represent means ± SE. *Statistically higher than control 1. **Statistically higher than control 2.

Fig. 3.

Closed circles show various cell types measured in BALF from mouse lungs 24 h after 3, 14, 21, and 31 days of daily OVA challenge: total leukocytes (A), eosinophils (B), and neutrophils (C). Open circles show corresponding control measurements made 24 h after 3 days (control 1: left-hand point) and 31 days (control 2: right-hand point) of saline challenge. Points represent means ± SE. *Statistically higher than control 1. **Statistically higher than control 2. The cell numbers on the vertical axes are in units of 1,000 cells.

Fig. 4.

Closed circles show lung mechanics parameters measured in mice measured 24 h after 3, 14, 21, and 31 days of daily OVA challenge: R_n (A) and H (B). Open circles show corresponding control measurements made 24 h after 3 days (control 1: left-hand point) and 31 days (control 2: right-hand point) of saline challenge. Points represent means ± SE. *Statistically greater than control 1. **Statistically greater than control 2. ***Statistically distinct from the other three measurements in the OVA-challenged animals.

Fig. 5.

Concentrations of cytokines in the left lobes of lungs of mice measured 24 h following 3, 14, and 21 days of OVA challenge, and both immediately preceding and following recall challenge: GM-CSF (A), KC (B), IL-5 (C), IFN-γ (D), and IL-10 (E). Control measurements were made 24 h after 3 days of saline challenge and after a recall challenge with saline on day 31. Points represent means ± SE. *Significant increase following the recall challenge. ** and ***: Significant increases following removal of an outlier from the post-recall group and pre-recall groups, respectively.

Fig. 6.

Numbers of cells measured in BALF from mouse lungs 24 h following 3, 14, and 21 days of OVA challenge, and both immediately preceding and following recall challenge: total leukocytes (A), eosinophils (B), and neutrophils (C). Control measurements were made 24 h after 3 days of saline challenge and after a recall challenge with saline on day 31. Points represent means ± SE. *Significant increase following the recall challenge.

Fig. 7.

Measurements of lung mechanics parameters in mice 24 h following 3, 14, and 21 days of OVA challenge, and both immediately preceding and following recall challenge: R_n (A) and H (B). Control measurements were made 24 h after 3 days of saline challenge and after a recall challenge with saline on day 31. Points represent means ± SE. *Significant increase following the recall challenge.