Troglitazone induces G1 arrest by p27Kip1 induction that is mediated by inhibition of proteasome in human gastric cancer cells

Abstract

We examined in the present study whether human gastric cancer cells express peroxisome proliferator-activated receptor γ (PPARγ), the effect of PPARγ activation by troglitazone, a selective ligand, on cellular growth, and the mechanism of the growth arrest by troglitazone in gastric cancer cells. RT-PCR, northern blot and western blot analysis demonstrated that all four tested human gastric cancer cell lines, MKN-28, MKN-45, MKN-74 and KATO-III, expressed PPARγ mRNA and protein. WST-1 assay and flow cytometric analysis revealed that troglitazone inhibited the growth and induced G1 arrest in all four gastric cancer cell lines. To examine the role of p27Kipl, a cyclin-dependent kinase inhibitor, in the G1 arrest by troglitazone, we determined p27Kipl protein expression by western blot analysis in gastric cancer cells that had been treated with troglitazone. Troglitazone increased p27Kipl in all four gastric cancer cell lines. Since it has been reported that the ubiquitin-proteasome system plays a vital role in the degradation of p27Kipl protein, we evaluated the hypothesis that inhibition of proteasome mediates the troglitazone-induced p27Kipl accumulation. Lactacystin, a proteasome inhibitor, inhibited cell growth and increased p27Kipl expression in MKN-74 cells. It was further demonstrated that troglitazone inhibited proteasome activity in a dose-dependent manner in MKN-74 cells. All these results suggest that troglitazone inhibited proteasome activity, followed by induction of p27Kipl, which arrests cells at the G1 phase of the cell cycle in gastric cancer cells. The troglitazone-mediated inhibition of the proteasome suggests a novel mechanism for the anti-proliferative effect of this agent in cancer cells.