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Fig. 2 | BMC Cancer

Fig. 2

From: Rapid eradication of colon carcinoma by Clostridium perfringens Enterotoxin suicidal gene therapy

Fig. 2

Cytotoxicity of CPE gene transfer in human colon carcinoma cells. a Time-dependent cytotoxicity of CPE after transfection with the pCpG-optCPE (optCPE) and empty vector (e.v.). MTT cytotoxicity assay was performed 24, 48 and 72 h after transfection. All transfected colon cancer cells reveal strong cytotoxic effects (P < 0.0001) mediated by optCPE 48 h after gene transfer. Sk-Mel5 cells were unaffected. Measurements were done in triplicates and expressed as mean percent of untreated control. Bars: SD. Level of significance was calculated by 2way-ANOVA; * P < 0.05, ** P < 0.001, *** P < 0.0001. b Analysis of CPE release by optCPE transfected human colon carcinoma cell lines. Media of optCPE transfected cells were harvested 48 h after transfection and analyzed by western blot. Representative blot shows released CPE within the media in all transfected cells. c Analysis of biological activity of released optCPE. optCPE containing media were added once to respective non-transfected cell lines and cytotoxicity was determined by MTT 72 h after application. SW620, SW480, HCT116 and CaCo-2 cells showed strong significant decrease in cell viability, correlating with amount of released optCPE (see b). HT-29 cells released low amounts of optCPE protein (see b) after transfection and cell viability was not reduced after application of optCPE containing medium. Columns represent mean percentage of untreated control cells. Bars, SD. Measurements were performed in triplicates and levels of significance were calculated by Student’s t- test; *** P < 0.0001. d Analysis of cytotoxic effect after repeated application of CPE containing medium in HT-29 cells. optCPE containing medium of optCPE transfected cells (see b) was added twice a day at 5.4 ng CPE ml-1 (see Table 3) to non-transfected HT-29 cells. MTT assay was performed 72 h after treatment and significant decrease of cell viability (P < 0.0001) was measured. Measurements were done in triplicates and expressed as mean percent of untreated control. Bars: SD. Level of significance was calculated by Student’s t-test; *** P < 0.0001

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