Peroxisome proliferator‐activated receptor γ ligand troglitazone induces cell cycle arrest and apoptosis of hepatocellular carcinoma cell lines

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Abstract
BACKGROUND Ligand activation of peroxisome proliferator‐activated receptor γ (PPARγ) results in the inhibition of proliferation of various cancer cells. The aim of this study is to investigate the mechanisms of cell growth inhibition of hepatocellular carcinoma (HCC) cell lines by the PPARγ ligand, troglitazone. METHODS Six HCC cell lines were used to study the effects of troglitazone on cell growth by 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyltetrazolium bromide (MTT) assay, on cell cycle by flow cytometry, and on the cell cycle‐regulating factors of late G1 phase by Western blotting. Apoptosis assays were performed by flow cytometry using membrane, nuclear, cytoplasmic, and mitochondrial markers. Caspase inhibitors were used to analyze the mechanisms of apoptosis induced by troglitazone. RESULTS Troglitazone showed a potent dose‐dependent effect on the growth inhibition of all six HCC cell lines, which were suppressed to under 50% of control at the concentration of 10 μmol/L. The growth inhibition was linked to the G1 phase cell cycle arrest through the up‐expression of the cyclin‐dependent kinase inhibitors, p21 and p27 proteins, and the hypophosphorylation of retinoblastoma protein. Troglitazone induced apoptosis by caspase‐dependent (mitchondrial transmembrane potential decrease, cleavage of poly [adenosine diphosphate ribose] polymerase, 7A6 antigen exposure, Bcl‐2 decrease, and activation of caspase 3) and caspase‐independent (phosphatidylserine externalization) mechanisms. CONCLUSIONS Our data suggest that ligand activation of PPARγ by troglitazone or modified analogs of the thiazolidinedione class of drugs is a novel target for effective therapy against HCC, because of the significant antiproliferative and programmed cell death induction capabilities demonstrated by troglitazone. Cancer 2002;95:2243–51. © 2002 American Cancer Society. DOI 10.1002/cncr.10906