Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
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Induction of Apoptosis in HT-29 Human Colorectal Cancer by Aloin

인간 대장암 세포 HT-29에서 Aloin에 의한 Apoptosis 유도

  • Yoo, Eun-Seon (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Woo, Joong-Seok (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Kim, Sung-Hyun (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Lee, Jae-Han (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Han, So-Hee (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Jung, Soo-Hyun (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Park, Young-Seok (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Kim, Byeong-Soo (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Kim, Sang-Ki (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Park, Byung-Kwon (Department of Companion and Laboratory Animal Science, Kongju National University) ;
  • Jung, Ji-Youn (Department of Companion and Laboratory Animal Science, Kongju National University)
  • 유은선 (공주대학교 특수동물학과) ;
  • 우중석 (공주대학교 특수동물학과) ;
  • 김성현 (공주대학교 특수동물학과) ;
  • 이재한 (공주대학교 특수동물학과) ;
  • 한소희 (공주대학교 특수동물학과) ;
  • 정수현 (공주대학교 특수동물학과) ;
  • 박영석 (공주대학교 특수동물학과) ;
  • 김병수 (공주대학교 특수동물학과) ;
  • 김상기 (공주대학교 특수동물학과) ;
  • 박병권 (공주대학교 특수동물학과) ;
  • 정지윤 (공주대학교 특수동물학과)
  • Received : 2019.08.16
  • Accepted : 2019.09.05
  • Published : 2019.10.30
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Abstract

Aloin [1,8-Dihydroxy-10-(${\beta}$-D-glucopyranosyl)-3-(hydroxymethyl)-9(10H)-anthracenone], is a natural anthraquinone from aloe. It has been shown to have antioxidant and anticancer effects in various types of human cancer cells, but the anticancer effects of aloin in human colorectal cancer cells HT-29 have not been elucidated. In this study, possible mechanisms by which aloin exerts its apoptotic action in cultured human colorectal cancer HT-29 cells were investigated. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay shows that treatment with aloin (0, 100, 200, 300 and $400{\mu}M$) reduced cell viability in a concentration-dependent manner in HT-29 and showed no effects on cell proliferation in A375SM and AGS cells. In addition, it was confirmed that apoptotic body was significantly increased as shown by 4',6-diamidino-2-phenylindole (DAPI) staining, and increased apoptosis rate by flow cytometry in HT-29 cells treated with aloin (0, 200 and $400{\mu}M$). We confirmed by western blotting that aloin activated Bax (pro-apoptotic), cleaved-poly (ADP-ribose) polymerase (PARP) and caspase-3, -8 and Bcl-2 (anti-apoptotic) were not changed compared with the control. Aloin induced up-regulation of phospho-p38 and down-regulation of phospho-extracellular signal-regulated kinase (ERK)1/2. Therefore, aloin suppressed the growth inhibitory effects by the induction of apoptosis in human colorectal cancer cells and has potential as a cancer preventive medicine.

Aloin [1,8-Dihydroxy-10-(${\beta}$-D-glucopyranosyl)-3-(hydroxymethyl)-9(10H)-anthracenone]은 알로에에서 추출한 천연 안트라퀴논이다. 다양한 유형의 인간 암세포에서 항산화, 항암 효과가 있는 것으로 밝혀졌지만 인간 대장암 세포 HT-29에서 aloin의 항암 효과는 밝혀지지 않았다. 본 연구에서는 aloin이 인간 대장암 HT-29 세포에서 세포 사멸 작용을 발휘할 수 있는 메커니즘을 조사하였다. Aloin이 세포 생존율에 영향을 미치는지 알아보기 위해 대장암 세포 HT-29, 흑색종 세포 A375SM, 위암 세포 AGS를 aloin(0, 100, 200, 300 및 $400{\mu}M$)으로 처리하였을 때, HT29에서는 농도 의존적으로 세포 생존율을 감소시켰고, A375SM과 AGS 세포에서는 암세포 생존율의 감소가 보이지 않았다. 이러한 HT-29에서의 세포 생존율 감소가 세포자멸사로 인한 감소인지 확인하기 위해 DAPI stain과 flow cytometry를 실시한 결과 apoptotic body가 유의적으로 증가하고 세포 자멸사가 증가한 것을 확인할 수 있었다. 이와 같은 결과를 바탕으로 aloin이 대장암 세포 HT-29에서 세포 사멸 관련 단백질 발현 양상에 미치는 영향을 확인하기 위해 western blotting을 실시하였다. Aloin은 Bax, PARP의 분절을 농도 의존적으로 증가시켰고, caspase3, -8을 활성화시켰지만, Bcl-2는 대조군에 비해 변화가 없었다. Aloin에 의해 유도된 세포자멸사 기전을 확인하기 위해 MAPK pathway 중 p-p38과 p-ERK의 발현을 확인한 결과, p-p38을 up-regulation시키고 p-ERK의 down-regulation을 유도했다. 따라서, aloin은 인간 대장암에서 암세포 성장 억제 효과 및 암세포 사멸 유도로 암예방 약제로서의 개발 가능성이 있다고 사료된다.

Keywords

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