Survivin signaling is regulated through nuclear factor-kappa B pathway during glycochenodeoxycholate-induced hepatocyte apoptosis.

Kewei Wang, John J Brems, Richard L Gamelli, Ai-Xuan Holterman

Department of Pediatrics and Surgery/Section of Pediatric Surgery, Rush, University Medical Center, Chicago, IL 60612, USA. kewei_wang@rush.edu

Biochimica et biophysica acta 2010 Dec

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Hepatocytes in primary culture undergo apoptosis upon exposure to glycochenodeoxycholate (GCDC). The signaling mechanisms of GCDC-induced apoptosis remain unclear. To investigate the role of antiapoptotic genes, we compared apoptotic response in primary hepatocytes following GCDC treatment. The hepatocytes from adult Sprague-Dawley rats were cultured in collagen-coated dishes and treated with GCDC in varying concentrations, or the same concentration at different time intervals. Apoptosis was detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay, DNA fragmentation assay, and caspase assays. Expression of apoptosis-related genes and proteins was evaluated by RT-PCR, quantitative real-time PCR (qRT-PCR), and Western blotting, respectively. The DNA-binding property of a nuclear protein was assessed by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay. An interesting result was that GCDC caused hepatocyte apoptosis to display a biphasic phenomenon at a dosage of 50μM, whereas it was not found at higher dosages such as 200μM. GCDC stimulated the expression of antiapoptotic Survivin, which also presented a biphasic response. The activation of nuclear factor-kappaB (NF-κB) corresponded with the up-regulation of Survivin. The inhibitor of NF-κB, BAY 11-7082, suppressed the expression of Survivin and simultaneously eliminated the biphasic response. The expression of Survivin was transcriptionally mediated by the activation of NF-κB, as shown by EMSA and ChIP assay. Conclusions: These results demonstrated that a low dosage of GCDC induced the hepatocyte apoptosis to exhibit the biphasic response, which was regulated by the expression of Survivin through NF-κB signaling pathway.

Citation

Kewei Wang, John J Brems, Richard L Gamelli, Ai-Xuan Holterman. Survivin signaling is regulated through nuclear factor-kappa B pathway during glycochenodeoxycholate-induced hepatocyte apoptosis. Biochimica et biophysica acta. 2010 Dec;1803(12):1368-75