Differential Oxidative Stress Responses in Type II Airway Epithelial Cells Impact Premature Senescence and Lung Fibrosis Susceptibility

Abstract

Radiation-Induced Pulmonary Fibrosis (RIPF) is a late toxicity characterized by premature senescence in Type II airway epithelial cells (AECII) and accumulation of alternatively activated (M2) macrophages. Differential susceptibility to RIPF is observed across mouse strains. Based on our prior study of the effects of macrophage variation on RIPF, we hypothesized that intrinsic differences in AECII oxidative stress response across mouse strains also impact susceptibility to RIPF. Ten-week-old female mice from C57L, C57BL6 and C3H/HeN strains were exposed to thoracic irradiation (5x6 Gy, n>5 per group). Fifteen weeks after radiation, lung tissue was collected and examined with Masson-Trichrome staining (histologic changes) and β-galactosidase activity assay (senescence). AECII prepared from mice of each strain were exposed to irradiation. To assess differential gene expression, total RNA was extracted and assessed with a multiplex analysis platform and quantitative PCR. Senescence was assessed by β-galactosidase activity assay in primary AECII after irradiation or after co-culture with M2 macrophages polarized with IL13. Susceptibility to radiation-induced lung injury, survival, and premature AECII senescence vary by mouse strain: C57L (fibrosis-prone), C57BL6J (-intermediate) and C3H/HeN (-resistant). Enriched AECII from each strain exhibited differential expression of genes related to inflammatory responses including SASP production after irradiation. Minimal increased expression of Il1r1 was observed in irradiated and unirradiated AECII from C3H/HeN, however Il1rn levels were markedly elevated in response to irradiation. The expression of Thioredoxin (Txn) and Thioredoxin reductase 1 (Txnrd1) in AECII from C3H/HeN was significantly higher than those observed in other strains. In Vivo, C3H/HeN mouse lungs exhibited the least premature senescence in AECII after irradiation. Premature senescence in AECII irradiated In Vitro or co-cultured with superoxide anion-producing M2 macrophages was substantially less in AECII from C3H/HeN compared to other strains. A comparison of primary AECII from three different mouse strains identified intrinsic differences in expression of major inflammatory signaling (IL1R and IL1RN) and redox homeostasis status (TXN and TXNDR1) molecules. This study is the first to demonstrate that intrinsic differences in AECII impacts susceptibility to premature senescence and lung fibrosis after irradiation.