Western Blot Analysis of UVB-Induced Stress

Cells were incubated with CAE (5–50 μg/mL) for 24 h after UVB irradiation. Cells were collected and homogenized with lysis buffer as described in previous studies [53 (link),54 (link)]. Equal protein amounts (30 μg) were loaded and separated on sodium dodecyl sulfate polyacrylamide gels and then transferred to a PVDF membrane. The membrane was incubated with specific antibodies, which comprised antibodies against IκB, p-IκB, and COX-2 (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA). The blots were then incubated with antiimmunoglobulin G-horseradish peroxidase, and the ECL™ detection reagent (Amersham Biosciences, Buckinghamshire, UK) was added. Immunoreactive proteins were detected using an ECL Western blotting detection system (LAS-4000, Fujifilm, Tokyo, Japan), and density of bands was determined using a densitometric program (Multi Gauge V2.2, Fujifilm, Tokyo, Japan).