Journal of the Korea Academia-Industrial cooperation Society (한국산학기술학회논문지)

The Korea Academia-Industrial cooperation Society (한국산학기술학회)
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Cell proliferation inhibition effects of epigallocatechin-3-gallate in TREK2-channel overexpressing cell line

TREK2-채널 과발현 세포주에서 에피갈로카테킨-3-갈레이트의 세포 증식 억제 효과

  • Kim, Yangmi (Dept. of Physiology, College of Medicine, Chungbuk National University) ;
  • Kim, Kyung-Ah (Dept. of Biomedical Engineering, College of Medicine, Chungbuk National University)
  • 김양미 (충북대학교 의과대학 생리학교실) ;
  • 김경아 (충북대학교 의과대학 의공학교실)
  • Received : 2016.02.12
  • Accepted : 2016.03.03
  • Published : 2016.03.31
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Abstract

Two-pore domain potassium (K2P) channels are the targets of physiological stimuli, such as intracellular pH, bioactive lipids, and neurotransmitters, and they set the resting membrane potential. Some types of K2P channels play a critical role in both apoptosis and tumoriogenesis. Among the K2P channels, no antagonists of the TREK2 channel have been reported. The aim of the present study was to determine if the TREK2 channel is blocked and whether cell proliferation is influenced by flavonoids in the TREK2 overexpressing HEK293 cells (HEKT2). The electrophysiological current was recorded using single channel patch clamp techniques and cell proliferation was measured using a XTT assay. The electrophysiological results showed that the TREK2 channel activity was reduced to $91.5{\pm}13.1%$ (n=5) and $82.2{\pm}13.7%$ (n=5) by flavonoids, such as epigallocatechin-3-gallate (EGCG) and quercetin in HEKT2 cells, respectively. In contrast, the EGCG analogue, epicatechin (EC), had no significant inhibitory effects on the TREK2 single channel activity. In addition, cell proliferation was reduced to $69.4{\pm}14.0%$ (n=4) by ECGG in the HEKT2 cells. From these results, EGCG and quercetin represent the first known TREK2 channel inhibitors and only EGCG reduced HEKT2 cell proliferation. This suggests that the flavonoids may work primarily by inhibiting the TREK2 channel, leading to a change in the resting membrane potential, and triggering the initiation of a change in intracellular signaling for cell proliferation. TREK2 channel may, at least in part, contribute to cell proliferation.

Two-pore 도메인 포타슘 채널(two-pore domain $K^+$ channel, K2P channel)은 세포내 pH, 생리 활성 지질, 신경 전달 물질과 같은 생리학적 자극의 표적이며 안정막전압(resting membrane potential)을 설정하는 것으로 알려져 있다. 일부 유형의 K2P 채널들은 세포 사멸 및 종양 형성 등에서 중요한 역할을 한다. K2P 채널 중 TREK2 채널의 길항제는 보고되지 않았다. 본 연구의 목적은 TREK2 채널을 과발현시킨 HEK293 세포(HEKT2)에서 플라보노이드에 의해 TREK2 채널이 억제되는지 그리고 HEKT2 세포의 증식이 플라보노이드에 의해 영향을 받는지 알아보고자 하였다. 전기생리학적 전류는 단일 채널 patch clamp 방법을 사용하여 기록하였고 세포 증식은 XTT 에세이방법을 이용하여 측정하였다. HEKT2 세포에서 전기생리학적 TREK2 채널 활성도는 에피갈로카테킨-3-갈레이트(EGCG) 및 케르세틴과 같은 플라보노이드에 의해 각각 $91.5{\pm}13.1%$(n=5), $82.2{\pm}13.7%$(n=5)까지 억제되었다. 반면, EGCG 유사체인 에피카테킨(EC)는 TREK2 단일 채널 활성도에 현저한 억제 효과는 없었다. 또한 HEKT2 세포에서 세포 증식이 EGCG에 의해 $69.4{\pm}14.0%$(n=4)까지 감소되었음을 확인하였다. 결과로부터 EGCG와 케르세틴이 TREK2 채널 억제제임을 처음으로 확인하였고, EGCG만 HEKT2 세포의 증식을 감소시킨다는 결론을 얻었다. 본 연구의 결과는 EGCG 및 케르세틴이 TREK2 채널을 억제함으로써 막전압의 변화 유도와 세포 증식에 필요한 세포내 신호 변화의 시작을 트리거하는데 일차적으로 작동할 수 있음을 시사한다.

Keywords

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