Modulation of Tongue Cancer Cell Apoptosis

Tongue cancer cells were pretreated with or without an ERK inhibitor (U0126, Cell Signaling Technology, Danvers, MA, USA), a JNK1/2 inhibitor (JNK-IN-8, Calbiochem, San Diego, CA, USA), or a p38 inhibitor (SB203580, Calbiochem, San Diego, CA, USA) for 2 h and maintained in the presence or absence of DSK for 24 h. Total protein lysates (20 μg) were harvested and subjected to SDS-PAGE analyses [43 (link),44 (link)]. Specific antibodies targeting the following molecules were used for detection: Anti-cleaved Caspase-8 (#9496), Anti-cleaved Caspase-9 (#9505), Anti-cleaved Caspase-3 (#9664), Anti-Caspase-8 (#9746), Anti-Caspase-9 (#9502), Anti-PARP (#9542), Anti-Phospho-Erk1/2 (#4370), Anti-Erk1/2 (#9102), Anti-Phospho-JNK (#4668), Anti-JNK2 (#9258), Anti-c-IAP1 (#7065), and Anti-XIAP (#2045) antibodies from Cell Signaling Technology (Danvers, MA, USA); Anti-Caspase-3 (610323), Anti-phospho-p38 (612281), and Anti-p38 (612168) antibodies from BD biosciences (San Jose, CA, USA); Anti-β-actin (ab8226) and anti-HO-1 (ab68477) antibodies from Abcam (Cambridge, UK); Anti-Nrf2 (GTX55732) antibodies from GeneTex (Irvine, CA, USA); and HRP-conjugated secondary antibodies (Dako Corporation, Carpinteria, CA, USA). Densitometry data of immunoblots were generated and analyzed by ImageJ software.