Beas 2b human bronchial epithelial cell line

The BEAS-2B human bronchial epithelial cell line was purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). BEAS-2B cells were maintained at 37 °C in a humidified 5% CO₂ atmosphere in a serum-free bronchial epithelial cell growth basal medium (BEGM^®; Lonza, Walkersville, MD, USA) supplemented with a Bullet Kit (Cambrex Bio Science, Walkersville, MD, USA). The BEGM medium was replaced every second day, and the cells were passaged when they reached 70–80% confluency via incubation with 0.25% trypsin. Poly(I:C) (high molecular weight; Cat# tlrl-pic) was purchased from InvivoGen (San Diego, CA, USA). Poly(I:C) was freshly prepared just prior to each experiment using sterile endotoxin-free water (0.9% NaCl). BEAS-2B cells were seeded onto 100 mm culture dishes at a density of 1 × 10⁶ cells/dish and cultured with a BEGM medium in a humidified CO₂ incubator at 37 °C. When the cells reached 70–80% confluence, the cells were treated with different concentrations of Poly(I:C) (0–100 μg/mL) for up to 24 h.

Normal human bronchial epithelial cells (HBECs) were purchased from Lonza (Walkersville, MD, USA). Normal human mesothelial cells (HMCs) were purchased from Zen-Bio, Inc. (Research Triangle Park, NC, USA). The BEAS-2B human bronchial epithelial cell line was purchased from American Type Culture Collection (Manassas, VA, USA). The ACC-MESO-1 human malignant pleural mesothelioma cell line [34 (link)] was purchased from RIKEN (Ibaraki, Japan). HBECs were cultured in bronchial/tracheal epithelial cell serum-free growth medium kit with 0.1 μg/mL retinoic acid (Cell Application, San Diego, CA, USA) and passaged every 4 d, with the medium exchanged every alternate day. HMCs were cultured in mesothelial cell growth medium (Zen-Bio, Inc.) and passaged twice a week. Both types of normal cell were used within 5 passages. BEAS-2B cells were cultured in Ham's nutrient mixture F-12 (Nacalai Tesque) with 10% FBS and passaged twice a week. MESO-1 cells were cultured in RPMI 1640 (Nacalai Tesque) with 10% FBS and passaged twice a week. For each experiment, the cells were seeded at a density of 2 × 10⁵ cells/cm² and allowed to adhere for 24 h.