Role of caspase activation in butyrate-induced apoptosis of HT-29 colon carcinoma cells

AIM:Sodium butyrate has antitumor effects on colon cancer cells such as inhibiting cell growth and promoting differentiation and apoptosis.The aim of this study is to investigate whether sodium butyrate induces apoptosis in human colon cancer cell line HT-29 and to examine the intracellular mechanisms involved,especially the role of caspase activation in the process.METHODS:HT-29 cells were cultured to logarithmic phase before treatment with sodium butyrate at concentration of 5.0 mmol/L and caspase inhibitors at the concentration of 20 μmol/L.The latter were added in the medium ahead of sodium butyrate for 1 h.Then,the staining of Annexin V-FITC and PI were used to analyze HT-29 apoptosis and the dye JC-1 was applied to detect mitochondrial membrane potential by flow cytometry.Caspase activity within the cells was measured respectively using a specific caspase activity assay kit and a microplate reader.RESULTS:Preincubation of HT-29 cells with sodium butyrate significantly increased apoptosis [(35.40±0.70)%] and decreased mitochondrial membrane potential(5.53±0.91).This effect was blocked when pretreatments were enforced with z-VAD-fmk,z-DEVD-fmk and z-LEHD-fmk.The apoptosis percentages were(1.33±0.59)%,(1.40±0.53)% and(1.27±0.91)%,respectively and mitochondrial membrane potentials were 9.80±1.15,10.23±0.50 and 10.33±1.02,respectively.However,the role of reduction by z-IETD-fmk,which presented the apoptosis percentage of(32.10±2.33)% and mitochondrial membrane potential of 5.93±1.31,was not observed.An enhancement of caspase-3 and-9 activities(2-3-fold)but no change of caspase-8 activity was confirmed.CONCLUSION:Apoptosis of HT-29 colon carcinoma cells induced by sodium butyrate is tightly linked to caspase activation via mitochondrial pathway other than tumor necrosis factor-alpha and has the potential to inhibit proliferation and thereby may contribute to the progression of colon cancer.