Food Engineering Progress (산업식품공학)

Korean Society for Industrial Food Engineering (한국산업식품공학회)
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Effect of Aralia elata on Apoptosis in MDA-MB-231 Human Breast Cancer Cells

두릅의 인체 유방암세포 MDA-MB-231 사멸 효과

  • Ryu, Min Ju (Department of Food and Nutrition, College of Natural Sciences, Duksung Women's University) ;
  • Chung, Ha Sook (Department of Food and Nutrition, College of Natural Sciences, Duksung Women's University)
  • 류민주 (덕성여자대학교 자연과학대학 식품영양학과) ;
  • 정하숙 (덕성여자대학교 자연과학대학 식품영양학과)
  • Received : 2014.12.20
  • Accepted : 2015.06.08
  • Published : 2015.08.30
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Abstract

The cytotoxic effect and mechanism of Aralia elata were investigated in MDA-MB-231 human breast cancer cells. In this study, Aralia elata inhibited significantly the proliferation of MDA-MB-231 cells, and some typical apoptotic characteristics, such as nuclear fragmentation and chromatin condensation, were observed. In addition, flow cytometry analysis showed that Aralia elata increased the sub-G1 (apoptosis) population and apoptosis further confirmed by Annexin V-FITC and PI double staining. With respect to the mechanism underlying the induction of apoptosis, apoptosis-related mRNA and proteins were measured using a reverse transcription-polymerase chain reaction and western blot analysis. Aralia elata reduced anti-apoptotic Bcl-2 mRNA and protein levels, but pro-apoptotic Bax mRNA and protein expression were increased compared with the controls. Aralia elata also induced the cleavage of caspase-9 followed by the activation of caspase-3, resulting in the activation of poly-ADP-ribose polymerase. These results suggest that apoptotic activity of Aralia elata is probably modulated by a caspase-dependent cascade via the activation of intrinsic pathway. This is the first report to demonstrate the cytotoxic effect of Aralia elata on human breast cancer cells and to provide a possible mechanism for this activity.

본 연구는 인체 유래 유방암세포인 MDA-MB-231 세포증식을 억제하고 세포사멸을 유도하는 천연소재 발굴을 목적으로, 북한산 두릅 추출물을 MDA-MB-231 세포에 처리하여 세포사멸 및 작용기전을 규명하였다. 실험 결과, 두릅추출물 처리 농도가 증가할수록 세포증식이 감소하였고, 세포질의 응축과 핵이 분절되는 등 apoptotic bodies를 형성하였다. 또한 유세포 분석기를 사용하여 MDA-MB-231의 세포주기가 억제되었고, 세포사멸의 특징인 sub-G1 수치가 증가되는 것이 관찰되었고, 두릅 추출물 농도가 증가함에 따라 apoptotic 세포가 유의적으로 증가하였으며, 특히 early apoptosis 보다 late apoptosis가 더 많이 진행됨을 확인할수 있었다. 이를 바탕으로 MDA-MB-231 세포사멸과 관련된 유전자를 확인하고자 RT-PCR과 western blotting을 수행한 결과, 세포사멸의 주요한 조절인자인 anti-apoptotic인 bcl-2 발현이 두릅 추출물 처리 시 농도 의존적으로 감소되었고, 반대로 Bax의 발현은 증가됨을 확인하였다. 이를 통해 불활성화 형태로 존재한 pro-caspase-9과 pro-caspase-3의 발현이 시료 농도가 증가할수록 감소되었고, 활성화된 형태인 cleaved caspase-9과 cleaved caspase-3의 발현은 두릅 추출물 처리 농도에 의존적으로 증가하였다. 뿐만 아니라, 세포사멸 과정의 주요 인자인 PARP 또한 시료 농도가 증가할수록 절단 현상이 비례적으로 증가하였다. 이상의 실험 결과를 근거로, 북한산 두릅 지상부의 70% ethyl alcohol 추출물이 MDA-MB-231 인체 유방암 세포의 증식을 억제하는 효과가 있음을 확인하였고, 세포사멸과 관련된 활성기전을 규명하였다. 추후 암 예방/치료제로서 두릅을 활용하기 위해서는 활성 본체와 안정성 규명 및 임상실험 등 추가 연구가 필요할 것으로 사료된다.

Keywords

Acknowledgement

Supported by : 덕성여자대학교

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