BACKGROUND: We previously reported that water extract of Mekabu, a kind of seaweed, induced apoptosis in a human breast cancer cell line. In the present study we investigated intracellular signaling in apoptosis, with a focus on caspases. METHODS: Mekabu extract, obtained with ultrapure water, was used to induce apoptosis in a human breast cancer cell line, MDA-MB231, and DNA fractionation was investigated by flow cytometry and electrophoresis. In addition, using the caspase detection kit Caspa Tag, activation of caspases 3, 6, 8, and 9 was observed under a fluorescence microscope. Furthermore, using antibodies to caspases 3, 8, 9, and Bid, we conducted a protein analysis by Western blotting to determine the activation of these substances. RESULTS: Obvious ladder formation demonstrating DNA fractionation was seen, confirming that Mekabu extract induced apoptosis. In the fluorescence microscope observations, activation of caspases 3, 6, and 8, but not caspase 9, was seen. Activated caspases 3 and 8 were detected in the Western blotting analysis, but no proteins of activated caspase 9 or Bid were detected. CONCLUSION: Mekabu extract activates caspases 3, 6, and 8 and contributes to intracellular signaling to induce apoptosis in a human breast cancer cell line. This signaling is not via the mitochondria.
BACKGROUND: We previously reported that water extract of Mekabu, a kind of seaweed, induced apoptosis in a humanbreast cancer cell line. In the present study we investigated intracellular signaling in apoptosis, with a focus on caspases. METHODS: Mekabu extract, obtained with ultrapure water, was used to induce apoptosis in a humanbreast cancer cell line, MDA-MB231, and DNA fractionation was investigated by flow cytometry and electrophoresis. In addition, using the caspase detection kit Caspa Tag, activation of caspases 3, 6, 8, and 9 was observed under a fluorescence microscope. Furthermore, using antibodies to caspases 3, 8, 9, and Bid, we conducted a protein analysis by Western blotting to determine the activation of these substances. RESULTS: Obvious ladder formation demonstrating DNA fractionation was seen, confirming that Mekabu extract induced apoptosis. In the fluorescence microscope observations, activation of caspases 3, 6, and 8, but not caspase 9, was seen. Activated caspases 3 and 8 were detected in the Western blotting analysis, but no proteins of activated caspase 9 or Bid were detected. CONCLUSION: Mekabu extract activates caspases 3, 6, and 8 and contributes to intracellular signaling to induce apoptosis in a humanbreast cancer cell line. This signaling is not via the mitochondria.