Characterization of the Troglitazone Induced Apoptosis in a Human Colon Cancer Cell Line

Abstract

Troglitazone (TRO), a peroxisome proliferator‐activated receptor (PPAR) agonist, was a drug used as an anti‐inflammatory and anti‐diabetic agent. PPARɣ specifically is a known member of the steroid receptor superfamily and is a ligand‐activated nuclear transcription factor that regulates lipid and glucose homeostasis. As a PPARɣ activator, TRO, a thiazolidinedione, has been shown to have the effect of inducing cell differentiation but also inhibiting cell growth, proliferation, and angiogenesis thereby facilitating apoptosis, making it a potential candidate to study as a cancer chemotherapeutic agent.Through the use of flow cytometry, TRO‐induced apoptosis of tissue culture grown colorectal cancer cells was examined. Apoptosis was quantified by analyzing cells that contain less than the 2N DNA content (one hallmark of apoptosis). A dose response curve was determined for increasing concentrations of TRO vs sub2N DNA content. These studies confirm that TRO was able to induce apoptosis in our cell lines at concentrations greater than 30mM.Next we examined which PPAR receptors subtypes are being activated by TRO. HCT116 cells were exposed to various concentrations of TRO for 24hr then RNA was isolated from these cells. Specific gene targets to each PPAR receptor subtypes were identified and using RT‐PCR, changes in mRNAs expression was examined. Targets for PPARdelta and PPAR gamma were consistently upregulated. We are working to refine these experiments to get better determinations on which receptors are activated by TRO.These experiments confirm that TRO induces apoptosis in HCT116 cells. Additionally, we have identified changes in at least one target of PPAR receptors in response to TRO exposure. Further experiments are needed to exactly determine which PPAR receptors are being activated by TRO, and whether the apoptosis medicated by TRO is a result of PPAR receptor activation.