Diethylnitrosamine에 의한 랫드 간세포 clone 9의 in vitro transformation 특성

In vitro transformation of rat liver cell (clone 9) by diethylnitrosamine

본 실험에서는 화학적으로 유발되는 간 발암과정의 이해 및 in viro transformation 모델을 확립하기 위하여, Sprague-Dawley 랫드로부터 분리된 clone 9 간세포에 diethylnitrosamine (DEN)을 반복적으로 투여한 후 형태적 및 생화학적 변화를 조사하였다. In vitro transformation의 평가는 gap junction, DNA함량 측정, alcohol dehydrogenase의 활성 측정 및 soft agarose assay에 의한 anchorage-independent 세포군 형성등을 평가하였다. 결과로서 DEN 처치에 의한 in vitro transformation 과정에서 무처치 대조군에 비하여 intercellular communication과 DNA 함량은 크게 변화가 없었다. 또한 soft agarose assay에 의한 anchorage-independent 세포군의 출현은 관찰되지 않았다. 그러나 alcohol dehydrogenase는 DEN에 대한 농도 의존적인 상관관계를 나타내었으며, in vitro transformation에 대한 평가의 한 지표가 될 수 있을 것으로 사료된다.

To establish in vitro carcinogenesis model using clone 9 cell line from the normal liver of Sprague-Dawley rat was grown in media containing diethylnitrosamine (DEN). The cell was treated with 5 mM DEN from 1 to 6 times treatment and 10 mM DEN from 7 to 16 times, the intercellular communication using scraping dye loading technique, DNA content, the activity of alcohol dehydrogenase(ADH), and anchorage dependence test using soft agarose assay were evaluated. Results indicated that all DEN-treated cell and clone 9 cell could not form the cell colony in soft agarose gel. DNA content and intercellular communication also were not changed. But the activity of ADH of cell treated with DEN 10 mM was stronger than that of 5 mM. This result suggests that the activity of ADH was changed in the dose-dependent pattern and is possible as a marker to evaluate the preneoplastic alteration in DEN-induced in vitro hepatocarcinogenesis. But soft agarose assay could not provide any evidence of transformation of the cells.