Figure 1.
Paradoxical Effect of Rapamycin on Bronchoalveolar Lavage Fluid Inflammatory Cells in Induction and Treatment Models of House Dust Mite-induced Asthma.
Balb/c mice received daily nasal challenges with HDM (25 µg) 5 days per week. In the induction model (Panel A), HDM challenges were initiated concurrent with rapamycin administration (3 mg/kg) by gavage 5 days per week for 3 weeks (n = 7–10 animals per group). In the treatment model (Panel B), HDM challenges were administered for 6 weeks and rapamycin administration was given during weeks 4 through 6 (n = 12–13 animals per group). * P<0.05 vs. Saline+Vehicle, ** P<0.001. Results are representative of two independent experiments.
Figure 2.
Paradoxical Effect of Rapamycin on Lung Histology in Induction and Treatment Models of House Dust Mite-induced Asthma.
Histologic sections of lung were stained with hematoxylin and eosin (H & E) or periodic acid-Schiff (PAS) stains and images obtained at 200× or 1000×. Results are representative of 2 independent experiments.
Figure 3.
Paradoxical Effect of Rapamycin on Lung Th2 and Th17 Cytokine Expression in Induction and Treatment Models of House Dust Mite-induced Asthma.
Quantification of lung mRNA levels for IL-4, IL-13, and IL-17A by qRT-PCR presented as relative mRNA expression. Results for the induction experiment are shown in Panel A (n = 6–8 animals per group, * P<0.05, HDM+Vehicle vs. HDM+Rapamycin), while results for the treatment experiment are shown in Panel B (n = 6–10 animals per group, * P<0.001). Results are representative of 2 independent experiments.
Figure 4.
Paradoxical Effect of Rapamycin on Lung C-C Chemokine Expression in Induction and Treatment Models of House Dust Mite-induced Asthma.
Quantification of lung mRNA levels for CCL11, CCL24, CCL7 and CCL17 by qRT-PCR presented as relative mRNA expression. Results for the induction experiment are shown in Panel A (n = 6 animals per group, * P<0.01), while results for the treatment experiment are shown in Panel B (n = 5–10 animals per group, * P<0.05). Results are representative of 2 independent experiments.
Figure 5.
Effects of Rapamycin on Mediastinal Lymph Node Th2 Cytokine Production.
Mediastinal lymph nodes from HDM-challenged mice that had or had not been treated with rapamycin concurrent with HDM stimulation for 3 weeks (induction model) were cultured ex vivo with or without HDM re-stimulation and/or rapamycin. The amount of IL-4, IL-5 and IL-13 released into the medium was quantified by ELISA (* P<0.05, n = 5–6). Results are representative of two independent experiments.
Figure 6.
Paradoxical Effect of Rapamycin on Plasma Immunoglobulin Levels in House Dust Mite-induced Asthma.
Plasma levels of IgE, IgG1 and IgG2a were quantified. Results for the induction experiment are shown in Panels A, C and E, while results for the treatment experiment are shown in Panels B, D and F (n = 8–20 animals per group, * P<0.05 vs. Saline+Vehicle, ** P<0.001).
Figure 7.
Paradoxical Effect of Rapamycin on Goblet Cell Hyperplasia in House Dust Mite-induced Asthma.
Quantification of lung mRNA levels for Muc5AC and Clca3 by qRT-PCR are presented as relative mRNA expression. Results for the induction experiment are shown in Panel A (n = 6 animals per group, * P<0.01), while the results for the treatment experiment are shown in Panel B (n = 5–6 animals per group, P = NS). Results are representative of 2 independent experiments. Goblet cell hyperplasia is presented as the percentage of airways containing PAS-positive cells (n = 8–10 animals per group, * P<0.001). 35.3±0.6 airways were inspected in each mouse.
Figure 8.
Paradoxical Effect of Rapamycin on Airway Hyperreactivity in House Dust Mite-induced Asthma.
Airway resistance (cm H20/ml/s) was directly measured following administration of increasing doses of nebulized methacholine. Results for the induction experiment are shown in Panel A (n = 10 animals per group, * P<0.05), while results form the treatment experiment are shown in Panel B (n = 9–10 animals per group, * P<0.05). Results are representative of 2 independent experiments.
Figure 9.
Rapamycin attenuates mTORC1 effectors.
Phosphorylation of the mTORC1 effector, S6 (phospho-S6 Ser 235/236), or the mTORC2 effector, Akt (phospho-Akt S473), was assessed by Western blot analysis. The phosphorylation of STAT6 (phospho-STAT6 Y641) was determined as a control for activation of Th2 pathways. A representative blot from 5 experiments is shown.