View Abstract

A1320 - Investigation of Bronchial Epithelial Cell Repairing and Regulation of Epithelial Cytokine Expression by Glucocorticoid
Author Block: K. Chibana, T. Shiobara, T. Watanabe, Y. Horigane, Y. Nakamura, Y. Shimizu, Y. Ishii; Department of Pulmonary Medicine and Clinical Immunology, Dokkyo University School of Medicine, Mibu Tochigi, Japan.
[Rationale]
It is recognized that pathological features of the bronchial asthma are injured or desquamated bronchial epithelial cells (BECs), increasing goblet cells, thickening of basement membrane, eosinophil infiltration and enlargement of the airway smooth muscle cell bundles. So-called epithelial cytokines, including IL-33 are released by BECs injury and that stimulate ILC2 and DC to amplify type-2 immunity. Also, comparing airway specimens from untreated asthma and treated with inhaled glucocorticoid (GC) asthma subjects, BECs were regenerated and IL-33 mRNA significantly decreased in treated asthma subjects. However, it is still unclear whether GC could act on repairing BECs directly or regulate production of epithelial cytokines.
[Methods]
First, normal human BECs cell-line, NuLi-1 was cultured in BEGM without hydrocortisone (HC) and seeded in 96-well culture plates. After BECs were attached on the plates, culture medium was exchanged with HBSS for 24 hours to synchronize cell cycles. BECs were cultured by BEGM with or without GC (HC, BUD, FP, and MF) for another 24 hours and then quantified BECs proliferations for an indexed by BrdU up-taking. Second, to examine the production and release of epithelial cytokines from injured BECs, NuLi-1 was seeded in 24-well culture plates, and cultured by BEGM without HC stimulated with several concentrations of papain. We stimulated BECs with papain at a concentration of (10-11 ~ 10-6 M) for 24 hours and assessed expression of IL-33 mRNA with qPCR and observed cell-injury using phase contrast microscope, BZX-710. Furthermore, to assess the regulation of IL-33 mRNA expression by GC, BECs was pre-treated with BUD for 24 hours before papain stimulation. After another 24 hours papain stimulation, BECs were collected and examined IL-33 mRNA expression using qPCR.
[Results]
BECs showed significant proliferation treated with BUD, FP, and MF, except HC. BECs was damaged by papain and IL-33 mRNA expression was increased up-to 10-7 M, however, it decreased with further high concentration of papain. The de novo expression of IL-33 mRNA from BECs were significantly inhibited by pre-treatment of BUD even in low concentration (10-13 M). However, IL-33 mRNA expression was decreased by BUD pre-treatment, BECs injury was still remained.
[Conclusion]
BECs were proliferated by GC directly and it was suggested that BECs were promoted regeneration. Expression of IL-33 mRNA was induced by papain and it was inhibited by GC pre-treatment. However, this inhibition occurred regardless of BECs injury, and suggested other mechanism was existed for production of IL-33.