Fenretinide-induced apoptosis of Huh-7 hepatocellular carcinoma is retinoic acid receptor beta dependent.
BACKGROUND
Retinoids are used to treat several types of cancer; however, their effects on liver cancer have not been fully characterized. To investigate the therapeutic potential of retinoids on hepatocellular carcinoma (HCC), the present study evaluates the apoptotic effect of a panel of natural and synthetic retinoids in three human HCC cell lines as well as explores the underlying mechanisms.


METHODS
Apoptosis was determined by caspase-3 cleavage using western blot, DNA double-strand breaks using TUNEL assay, and phosphatidylserine translocation using flow cytometry analysis. Gene expression of nuclear receptors was assessed by real-time PCR. Transactivation assay and chromatin immunoprecipitation (ChIP) were conducted to evaluate the activation of RXRalpha/RARbeta pathway by fenretinide. Knockdown of RARbeta mRNA expression was achieved by siRNA transfection.


RESULTS
Our data revealed that fenretinide effectively induces apoptosis in Huh-7 and Hep3B cells. Gene expression analysis of nuclear receptors revealed that the basal and inducibility of retinoic acid receptor beta (RARbeta) expression positively correlate with the susceptibility of HCC cells to fenretinide treatment. Furthermore, fenretinide transactivates the RXRalpha/RARbeta-mediated pathway and directly increases the transcriptional activity of RARbeta. Knockdown of RARbeta mRNA expression significantly impairs fenretinide-induced apoptosis in Huh-7 cells.


CONCLUSIONS
Our findings reveal that endogenous expression of retinoids receptor RARbeta gene determines the susceptibility of HCC cells to fenretinide-induced apoptosis. Our results also demonstrate fenretinide directly activates RARbeta and induces apoptosis in Huh-7 cells in a RARbeta-dependent manner. These findings suggest a novel role of RARbeta as a tumor suppressor by mediating the signals of certain chemotherapeutic agents.
Bu
Pengli
P
Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS 66160, USA. pbu@kumc.edu
Wan
Yu-Jui Yvonne
YJ
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D017313:Fenretinide / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D020287:In Situ Nick-End Labeling; D008113:Liver Neoplasms / Q000473:pathology*; D010718:Phosphatidylserines / Q000378:metabolism; D018168:Receptors, Retinoic Acid / Q000378:metabolism*; D015533:Transcriptional Activation
D000970:Antineoplastic Agents; D010718:Phosphatidylserines; D018168:Receptors, Retinoic Acid; C082925:retinoic acid receptor beta; D017313:Fenretinide; D053148:Caspase 3