Bax is Crucial for IFN-γ-Induced Resolution of Allergen- Induced Mucus Cell Metaplasia

Abstract

Allergic airway responses cause proliferation of epithelial cells and mucus cell metaplasia (MCM), and the resolution of MCM involves reduction of cell numbers. The role of inflammation and apoptosis on this process was investigated in P-selectin +/+ and −/− mice sensitized and challenged with OVA by analyzing the expression and the role of regulators of apoptosis in metaplastic mucus cells. No differences were observed in MCM at 5 days of allergen exposure between +/+ and −/− mice, despite reduced IL-13 levels in −/− mice. Although IL-4 levels were similar in both −/− and +/+ mice, IL-13 and IL-5 levels had decreased and IFN-γ levels were increased earlier in −/− compared with +/+ mice. MCM levels were decreased 4-fold at 7 days of allergen exposure in −/− mice and at 15 days in +/+ mice. The percentage of Bax-expressing mucus cells increased significantly at 7 days in −/− mice and at 10 days in +/+ mice. The Bax-positive mucus cells exhibited caspase-specific cleavage of cytokeratin 18. IFN-γ caused Bax expression in IL-13-induced MCM in microdissected airway cultures. MCM remained significantly elevated in Bax −/− mice following 15 days of allergen exposure compared with +/+ mice, while the number of eosinophils was reduced in both Bax +/+ and −/− mice at 15 days. Together, these data demonstrate that reduced IL-13 levels were sufficient to elicit maximum MCM, that IFN-γ induces Bax in metaplastic mucus cells, and that Bax plays a critical role in the resolution of MCM, but not in the resolution of eosinophils.

STAT1 Activation Causes Translocation of Bax to the Endoplasmic Reticulum during the Resolution of Airway Mucous Cell Hyperplasia by IFN-γ

“…Our previous findings showed that Bax is important for the resolution of MCH during prolonged exposure to allergen (11). However, Bax mRNA and protein levels were not increased in IFN-␥-treated human and murine AECs and our cell fractionation studies with human AECs did not detect any noticeable movement of Bax from the cytosol to the mitochondria in response to IFN-␥.…”

“…These events lead to mitochondrial membrane permeabilization (8), or calcium release from the endoplasmic reticulum (ER) (9,10) to initiate activation of caspases and DNases and the appearance of apoptotic morphology. The resolution of allergen-induced MCH by IFN-␥ is not mediated by the death signal Fas (4), but we have shown that Bax knockout (bax Ϫ/Ϫ ) mice exhibit delayed resolution of MCH compared with their wild-type littermates, suggesting that Bax is involved in the resolution of allergen-induced MCH (11). However, the path linking IFN-␥ and STAT1 to cellular apoptosis in hyperplastic mucous cells is unclear.…”

“…Bax protein detection by immunohistochemistry was performed as previously described (11). Unless otherwise specified, all fluorescent reagents used in immunofluorescent assays were procured from Molecular Probes.…”

Cigarette Smoke Suppresses Bik To Cause Epithelial Cell Hyperplasia and Mucous Cell Metaplasia

“…Tissue sections were stained with Alcian blue (AB) and periodic acid Schiff or hematoxylin and eosin as described previously (20). The number of AB-positive cells per millimeter of basal lamina and per total cell count were quantified using a light microscope (BH-2; Olympus, Melville, NY) equipped with the Image analysis system (National Institutes of Health) as described previously (26).…”

“…The cell death of airway epithelial cells (AECs) during the resolution of metaplastic mucous cells is regulated by the Bcl-2 family of proteins (20,21), and involves the intrinsic apoptotic pathway (22). During the resolution of allergen-induced epithelial and mucous cell hyperplasia that is mediated by IFN-g, we showed that STAT1 and Bik, a proapoptotic Bcl-2 family member, are crucial factors, because unlike wild-type mice, STAT1-and Bik-deficient mice fail to resolve mucous cell metaplasia (MCM).…”