Transcriptomic analysis reveals significant changes in gene expression and a potential mitochondria-mediated apoptotic pathway in murine RAW 264.7 macrophages exposed to tert-butylquinone

Tert-butylquinone (TBBQ), a major oxidation product of the tert-butylhydroquinone,showed higher cytotoxicity. Cell death induced by TBBQ is related with apoptosis, however detailed mechanism was not well understood. TBBQ exposure resulted in dose dependently increase in level of ROS in RAW 264.7 cells as well as decrease in level of intracellular GSH, suggesting that oxidation stress maybe responsible for the cytotoxicity of TBBQ. Transcriptome analysis of RAW 264.7 cells exposed to TBBQ at concentration of 1 mu g/mL for 12 h finding that 442 genes were up-regulated and 316 genes were down-regulated. Among them, 15 differentially expressed genes were found to be associated with apoptosis, respectively fall into mitochondria (CTS L, BCL-2, BAX, CYTOc, CASPASE-9,-3), endoplasmic reticulum stress (CAPN2, CASPASE-12) and death receptors pathway(FAS, FADD, DAXX, JNK, JUN, ASK1, CASPASE-8). Results of qRT-PCR showed that all the gene express were unregulated, except for BCL-2 with negative regulation. Western blot results showed the expression of key target proteins (Bcl2, Fas, Cytochrome c, Caspase-3, -8, and -12) in exogenous and endogenous apoptosis pathways tracked the qRTPCR gene results well, indicated that ROS- Bcl-2-mitochondria mediated apoptosis pathway accounted for the cytotoxicity of TBBQ. In addition, death receptor/Fas, endoplasmic reticulum stress were partially involved into the TBBQ-induced apoptosis in RAW 264.7 cells. The finding on cytotoxic mechanism of TBBQ would guide the exploration on attenuation measure of TBBQ bio-toxicity.