Rosiglitazone And Retinoic Acid Inhibit Proliferation And Induce Apoptosis In Human Colorectal Cancer Cell Line HCT-15

Background and purpose:Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor (nuclear hormone receptor) superfamily, first discovered by Issemann and Green in 1990 in mouse cells. PPARs play important roles in the regulation of cell growth, differentiation and apoptosis. There are three known subtypes, namely PPARa, PPARP and PPARγ. They are encoded by separate genes, highly similar in amino acid sequences, particularly in the DNA binding domain and ligand binding domains. Among the three, PPARγis the subtype most studied. PPARy possesses complex and diverse biological functions. In recent years PPARγagonists has been gaining academic attentions for its anti-tumor effect. Mueller et al. confirmed that PPARγligands inhibit the growth of prostate cancer, suggesting that activation of PPARγis a potential cancer therapeutic method. Nagamine found that rosiglitazone treatment increased the number of apoptotic cells in human gastric cancer cell lines MKN-28,-45 and -74. There is growing evidence that PPARγagonists have significant anti-tumor effect.All-trans retinoic acid (ATRA) is one of the more mature types of retinoid drugs used in clinical chemotherapy and prevention of cancer. ATRA inhibits the growth of various malignant tumor cells and is a commonly used differentiation-inducing reagent. PPARs are known to form heterodimers with retinoid X receptor (RXR), bind to a specific DNA sequence-peroxisome proliferation response element (PPRE), and regulate target gene transcriptions. Elstner found in vitro and in transplanted breast tumors in nude mice that ATRA assisted PPARγligands in inhibiting tumor cell growth and decreased bcl-2 levels, suggesting that the use of PPARγagonists and retinoic acid can activate the PPARγ/RXR heterodimer and may be an effective way to treat a variety of tumors. In this study, we have investigated the effect of combined use of highly selective ATRA and PPARγagonist rosiglitazone (ROS) on proliferation and apoptosis of human colorectal cancer cell line HCT-15, and further explored the molecular mechanism involved.Methods:HCT-15 cells were divided into blank control group, vehicle control group and experimental group (ROS only or ATRA only or ROS plus ATRA). Growth inhibition was examined using MTT assay. Apoptosis and cell cycle were examined by flow cytometry (FCM). Expression of COX-2, MMP-7 and TIMP-1 was examined by immunocytochemistry.Results:ROS alone concentration-and time-dependently inhibited HCT-15 cell proliferation (P< 0.05), ROS combined with ATRA showed significant synergy (q> 1.15). ROS or ATRA alone increased the proportion of cells in G0/G1 phase, and decreased cells in S phase effectively, inducing apoptosis (P<0.05). Combination of ROS and ATRA resulted in even stronger G1 cell cycle arrest (P<0.05). HCT-15 cells expressed COX-2, MMP-7 and TIMP-1, and the positive expression rates in the control group were 66.79%,73.21%, and 64.08% respectively. After combined application of ROS and ATRA, the positive rates significantly dropped to only 19.33%,20.58% and 13.13% respectively (P< 0.01).Conclusion:ROS and ATRA reduced the expression of COX-2, MMP-7 and TIMP-1, caused cell cycle arrested at G1 phase, and induced apoptosis, which resulted in cell proliferation inhibition of human colorectal cancer cell line HCT-15.