Rosiglitazone enhances fluorouracil-induced apoptosis of HT-29 cells by activating peroxisome proliferator-activated receptor gamma

Abstract

Aim: To examine whether and how rosiglitazone enhances apoptosis induced by fluorouracil in human colon cancer (HT-29) cells.

Methods: Human colon cancer HT-29 cells were cultured in vitro and treated with fluorouracil and/or rosiglitazone. Proliferation and growth of HT-29 cells were evaluated by MTT assay and trypan blue exclusion methods, respectively. The apoptosis of HT-29 cells was determined by acridine orange/ethidium bromide staining and flow cytometry using PI fluorescence staining. The expressions of peroxisome proliferator-activated receptor gamma (PPARgamma), Bcl-2 and Bax in HT-29 cells were analyzed by Western blot.

Results: Although rosiglitazone at the concentration below 30 micromol/L for 72 h exerted almost no inhibitory effect on proliferation and growth of HT-29 cells, it could significantly enhance fluorouracil-induced HT-29 cell proliferation and growth inhibition. Furthermore, 10 micromol/L rosilitazone did not induce apoptosis of HT-29 cells but dramatically enhanced fluorouracil-induced apoptosis of HT-29 cells. However, rosiglitazone did not improve apoptosis induced by fluorouracil in HT-29 cells pretreated with GW9662, a PPARgamma antagonist. Meanwhile, the expression of Bax and PPARgamma was up-regulated, while the expression of Bcl-2 was down regulated in HT-29 cells treated with rosiglitazone in a time-dependent manner. However, the effect of rosiglitazone on Bcl-2 and Bax was blocked or diminished in the presence of GW9662.

Conclusion: Rosiglitazone enhances fluorouracil-induced apoptosis of HT-29 cells by activating PPARgamma.

Figure 1

Effect of rosiglitazone and/or 5-Fu on proliferation and growth of HT-29 cells exposed to rosiglitazone or 5-Fu (A), 5-Fu with or without 10 μmol/L rosiglitazone (B), and treated with rosiglitazone and 5-Fu (C) at the indicated concentration for 72 h. ^aP < 0.05 vs control group, ^bP < 0.01 vs rosiglitazone group.

Figure 2

Apoptosis of HT-29 cells induced by AO/EB staining (A) and flow cytometrical analysis (B). HT-29 cells were treated with rosigliatzone and/or 5-Fu at the indicated concentration for 72 h, then harvested and stained with AO/EB. Results are representative of three experiments.

Figure 3

Effect of rosiglitazone on 5-Fu induced apoptosis (A), time-dependent expression of PPARγ, Bax and Bcl-2 (B), and PPARγ-dependent expression of Bax, Bcl-2 and β-actin (C). ^aP < 0.05, ^bP < 0.01.