Autonomous inhibition of apoptosis correlates with responsiveness of colon carcinoma cell lines to ciglitazone.

Colorectal cancer is a leading cause of mortality worldwide. Resistance to therapy is common and often results in patients succumbing to the disease. The mechanisms of resistance are poorly understood. Cells basically have two possibilities to survive a treatment with potentially apoptosis-inducing substances. They can make use of their existing proteins to counteract the induced reactions or quickly upregulate protective factors to evade the apoptotic signal. To identify protein patterns involved in resistance to apoptosis, we studied two colorectal adenocarcinoma cell lines with different growth responses to low-molar concentrations of the thiazolidinedione Ciglitazone: HT29 cells underwent apoptosis, whereas SW480 cells increased cell number. Fluorescence detection and autoradiography scans of 2D-PAGE gels were performed in both cell lines to assess protein synthesis and turnover, respectively. To verify the data we performed shotgun analysis using the same treatment procedure as in 2D-experiments. Biological functions of the identified proteins were mainly associated with apoptosis regulation, chaperoning, intrinsic inflammation, and DNA repair. The present study suggests that different growth response of two colorectal carcinoma cell lines after treatment with Ciglitazone results from cell-specific protein synthesis and differences in protein regulation

Cell viability and mitochondrial membrane potential after treatment with thiazolidinediones.

<p>Cell viability was assessed by neutral red uptake in SW480 and HT29 cells treated with increasing concentrations of (A) Ciglitazone or (B) Troglitazone for 6 and 24 hours. *p<0.05, values of HT29 cells differ from those of SW480 cells. Mitochondrial membrane potential (Δψ_m) was assessed by JC-1 FACS analysis in (C) HT29 and (D) SW480 cells treated with increasing concentrations of Ciglitazone for 24 and 48 hours. *p<0.05, values at 48 hours differ from those at 24 hours. #p<0.05, values at indicated concentrations differ from baseline.</p

Cell responsiveness of colorectal adenocarcinoma cell lines after treatment with Ciglitazone.

<p>2D-PAGE gels were performed in (A) HT29 and (B) SW480 cells after treatment with 5 µM Ciglitazone. (1) Fluorescence scans of untreated cells, (2) fluorescence scans of cells treated with Ciglitazone, (3) autoradiography scans of untreated cells, (4) autoradiography scans of cells treated with Ciglitazone. Cell cycle distribution of (C) HT29 and (D) SW480 cells treated with increasing concentrations of Ciglitazone.</p

Protein synthesis in untreated colorectal adenocarcinoma cell lines.

<p>Representative 2D-PAGE gels of untreated (A) HT29 and (B) SW480 cells. Proteins synthesized to a greater extent in HT29 cells are indicated by hexagons, those in SW480 cells by circles. Accession numbers of proteins listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0114158#pone-0114158-t001" target="_blank">Table 1</a> are annotated according to the UniProtKB/Swiss-Prot database. (C) Total proteins identified by shotgun analysis in HT29 and SW480 cells.</p

Autonomous Inhibition of Apoptosis Correlates with Responsiveness of Colon Carcinoma Cell Lines to Ciglitazone

Colorectal cancer is a leading cause of mortality worldwide. Resistance to therapy is common and often results in patients succumbing to the disease. The mechanisms of resistance are poorly understood. Cells basically have two possibilities to survive a treatment with potentially apoptosis-inducing substances. They can make use of their existing proteins to counteract the induced reactions or quickly upregulate protective factors to evade the apoptotic signal. To identify protein patterns involved in resistance to apoptosis, we studied two colorectal adenocarcinoma cell lines with different growth responses to low-molar concentrations of the thiazolidinedione Ciglitazone: HT29 cells underwent apoptosis, whereas SW480 cells increased cell number. Fluorescence detection and autoradiography scans of 2D-PAGE gels were performed in both cell lines to assess protein synthesis and turnover, respectively. To verify the data we performed shotgun analysis using the same treatment procedure as in 2D-experiments. Biological functions of the identified proteins were mainly associated with apoptosis regulation, chaperoning, intrinsic inflammation, and DNA repair. The present study suggests that different growth response of two colorectal carcinoma cell lines after treatment with Ciglitazone results from cell-specific protein synthesis and differences in protein regulation