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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.

Author information

1
Department of Molecular Biology, College of Natural Sciences, Dongeui University, Busan 614-714, Korea.
2
Department of Horticultural Bioscience, College of Natural Resource and Life Sciences, Pusan National University, Gyeongnam 627-706, Korea.
3
Duksan B&F Co. LTD., Busan 614-853, Korea.
4
Laboratory of Immunobiology, Department of Marine Life Sciences, Jeju National University, Jeju 690-756, Korea.
5
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.
6
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.

Abstract

BACKGROUND/OBJECTIVES:

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.

MATERIALS/METHODS:

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.

RESULTS:

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.

CONCLUSIONS:

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.

KEYWORDS:

Aged black garlic; U937 cells; apoptosis; caspase

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