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Nutr Res Pract. 2014 Apr;8(2):132-7. doi: 10.4162/nrp.2014.8.2.132. Epub 2014 Mar 28.

Induction of apoptosis by a hexane extract of aged black garlic in the human leukemic U937 cells.

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Department of Molecular Biology, College of Natural Sciences, Dongeui University, Busan 614-714, Korea.
Department of Horticultural Bioscience, College of Natural Resource and Life Sciences, Pusan National University, Gyeongnam 627-706, Korea.
Duksan B&F Co. LTD., Busan 614-853, Korea.
Laboratory of Immunobiology, Department of Marine Life Sciences, Jeju National University, Jeju 690-756, Korea.
Anti-Aging Research Center & Blue-Bio Industry RIC, Dongeui University, Busan 614-714, Korea. ; Department of Life Science and Biotechnology, College of Natural Science, Dongeui University, Busan 614-714, Korea.
Anti-Aging Research Center & Blue-Bio Industry RIC, Dongeui University, Busan 614-714, Korea. ; Department of Biochemistry, Dongeui University College of Oriental Medicine 52-57, Yangjeong-ro, Busanjin, Busan 614-052, Korea.



In this study, the apoptogenic activity and mechanisms of cell death induced by hexane extract of aged black garlic (HEABG) were investigated in human leukemic U937 cells.


Cytotoxicity was evaluated by MTT (3-(4, 5-dimethyl-thiazol-2-yl)-2, 5-diphenyl tetrazoliumbromide) assay. Apoptosis was detected using 4,6-diamidino-2-phenyllindile (DAPI) staining, agarose gel electrophoresis and flow cytometry. The protein levels were determined by Western blot analysis. Caspase activity was measured using a colorimetric assay.


Exposure to HEABG was found to result in a concentration- and time-dependent growth inhibition by induction of apoptosis, which was associated with an up-regulation of death receptor 4 and Fas legend, and an increase in the ratio of Bax/Bcl-2 protein expression. Apoptosis-inducing concentrations of HEABG induced the activation of caspase-9, an initiator caspase of the mitochodrial mediated intrinsic pathway, and caspase-3, accompanied by proteolytic degradation of poly(ADP-ribose)-polymerase. HEABG also induced apoptosis via a death receptor mediated extrinsic pathway by caspase-8 activation, resulting in the truncation of Bid, and suggesting the existence of cross-talk between the extrinsic and intrinsic pathways. However, pre-treatment of U937 cells with the caspase-3 inhibitor, z-DEVD-fmk, significantly blocked the HEABG-induced apoptosis of these cells, and increased the survival rate of HEABG-treated cells, confirming that HEABG-induced apoptosis is mediated through activation of caspase cascade.


Based on the overall results, we suggest that HEABG reduces leukemic cell growth by inducing caspase-dependent apoptosis through both intrinsic and extrinsic pathways, implying its potential therapeutic value in the treatment of leukemia.


Aged black garlic; U937 cells; apoptosis; caspase

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