Radiation Oncology Journal

The Korean Society for Radiation Oncology (대한방사선종양학회)
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The Use of MTT Assay, In Vitro and Ex Vivo, to Predict the Radiosensitivity of Colorectal Cancer

In-vitro와 Ex-vivo MTT Assay를 통한 직장암의 방사선치료 감수성 예측 가능성 검증

  • Kim, Ji-Eun (Laboratory of Radiation Treatment Research, Korea Institute of radiological and Medical Sciences) ;
  • Kim, Mi-Sook (Department of Radiation Oncology, Korea Institute of radiological and Medical Sciences) ;
  • Kang, Chang-Mo (Laboratory of Cytogenetics and Tissue Regeneration, Korea Institute of radiological and Medical Sciences) ;
  • Kim, Jong-Il (Department of Food and Microbial Technology, Seoul Women's University) ;
  • Shin, Hye-Kyung (Laboratory of Radiation Treatment Research, Korea Institute of radiological and Medical Sciences) ;
  • Choi, Chul-Won (Department of Radiation Oncology, Korea Institute of radiological and Medical Sciences) ;
  • Seo, Young-Seok (Department of Radiation Oncology, Korea Institute of radiological and Medical Sciences) ;
  • Ji, Young-Hoon (Department of Radiation Oncology, Korea Institute of radiological and Medical Sciences)
  • 김지은 (원자력의학원 방사선치료기술연구팀) ;
  • 김미숙 (원자력의학원 방사선종양학과) ;
  • 강창모 (원자력의학원 세포유전조직재생연구팀) ;
  • 김종일 (서울여자대학교 식품.미생물공학과) ;
  • 신혜경 (원자력의학원 방사선치료기술연구팀) ;
  • 최철원 (원자력의학원 방사선종양학과) ;
  • 서영석 (원자력의학원 방사선종양학과) ;
  • 지영훈 (원자력의학원 방사선종양학과)
  • Published : 2008.09.30
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Abstract

Purpose: The measurement of radiosensitivity of individuals is useful in radiation therapy. Unfortunately, the measurement of radiation survival using a clonogenic assay, which is the established standard, can be difficult and time consuming. The aim of this study is to compare radiosensitivity results obtained from the MTT and clonogenic assays, and to evaluate whether the MTT assay can be used on clinical specimens. Materials and Methods: HCT-8, LoVo, CT-26, and WiDr were the colon cancer cell lines used for this study. The clonogenic assay was performed to obtain the cell survival curves and surviving fractions at a dose of 2 Gy ($SF_2$) as the standard technique for radiosensitivity. Also, the MTT assay was performed for each of the cell lines (in vitro). To simulate clinical specimens, the cell lines were inoculated into nude mice, removed when the tumors reached 1 cm in diameter, and chopped. Next, the tumors were subjected to the same process involved with the MTT assay in vitro. The inhibition rates (IR) of 10 Gy or 20 Gy of irradiation for in vitro and ex vivo were calculated based on the optical density of the MTT assay, respectively. Results: According to $SF_2$ and the cell survival curve, the HCT-8 and WiDr cell lines were more resistant to radiation than LoVo and CT-26 (p<0.05). The IR was measured by in vitro. The MTT assay IR was 17.3%, 21%, 30% and 56.5% for the WiDr, HCT-8, LoVo and CT-26 cell lines, respectively. In addition, the IR measured ex vivo by the MTT assay was 23.5%, 26%, 38% and 53% in the HCT-8, WiDr, LoVo and CT-26 tumors, respectively. Conclusion: The radiosensitivity measured by the MTT assay was correlated with the measures obtained from the clonogenic assay. This result highlights the possibility that the MTT assay could be used in clinical specimens for individual radiosensitivity assays.

목 적: 암환자의 방사선 치료 전 방사선에 대한 감수성을 미리 측정할 수 있다면 임상적으로 많은 도움이 될 것이다. 본 연구는 전 임상 실험을 통하여 MTT assay가 세포집락 측정기법과 비교해서 방사선 감수성을 예측할 수 있고, 직장암 환자의 조직에 사용할 수 있는지 가능성을 확인하고자 하였다. 대상 및 방법: 대장암 세포 주인 HCT-8, LoVo, CT-26, WiDr을 이용하여 세포집락 측정기법을 통해 세포생존곡선 및 2 Gy에서의 세포생존확률(SF2)을 구하였다. 세포 주 자체를 대상으로 MTT assay를 시행하는 실험(in vitro) 및 환자의 암 조직과 같은 상태를 만들기 위하여, 누드 마우스에 세포 주를 주입하여 암 조직을 형성한 후 in vitro와 같은 방식으로 MTT assay를 시행(ex vivo)하였다. 이 두 실험에 대한 흡광도 값에 따른 저해율(inhibition rate, %)을 구하였다. 결 과: $SF_2$ 및 세포생존곡선에 따르면 CT-26 및 LoVo가 HCT-8, WiDr에 비해 방사선에 민감하였다(p<0.05). In vitro MTT assay 결과 WiDr, HCT-8, LoVo와 CT-26의 방사선 저해율이 각각 17.3%, 21%, 30%, 56.5%를 나타내었다. 또한 ex vivo MTT assay의 저해율은 HCT-8, WiDr, LoVo와 CT-26에서 각각 23.5%, 26%, 38%, 53%를 나타내었다. 통계적인 차이를 감안하였을 때 세포생존곡선을 통해 얻은 방사선 감수성의 결과와 동일한 순서를 가졌다. 결 론: 4개의 세포 주의 방사선의 감수성의 순서가 세포집락 측정기법 및 in vitro와 ex vivo MTT assay 결과에서 거의 일치함을 보였다. 이는 직장암 환자에서 MTT assay를 통해 방사선 감수성을 예측할 수 있는 가능성을 제시하였다.

Keywords

References

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