Olive P L, Durand R E, Banáth J P, Evans H H
B.C. Cancer Research Centre, Vancouver, Canada.
Int J Radiat Biol. 1991 Sep;60(3):453-66. doi: 10.1080/09553009114552311.
Chinese hamster V79 cells grown in suspension culture as spheroids are more resistant than monolayers to killing and mutation by ionizing radiation. A change in DNA conformation appears to accompany this increase in radiation resistance. We were therefore interested in whether the activity of topoisomerase II, a nuclear enzyme involved in DNA conformational changes and possibly in DNA repair, might differ in monolayers and small spheroids. One-day-old spheroid cells were more resistant than monolayer cells to the toxic effects of etoposide, a topoisomerase II inhibitor. Fewer strand breaks were induced by etoposide in spheroid DNA than monolayer DNA, as measured by the DNA precipitation and alkali unwinding assays, although identical amounts of damage were produced in monolayers and spheroids by the topoisomerase I inhibitor camptothecin, and the cell cycle specific agent, 5-fluorouracil. There was no evidence of a subpopulation of spheroid cells which were more resistant to etoposide, and no change in the rate of incorporation or DNA chain elongation in spheroids compared to monolayers. Topoisomerase II activity in 1-day-old spheroids, measured by decatenation of trypanosome kinetoplast DNA, was reduced to 68% of the monolayer value; in 3-4-day-old spheroids the level was 32.5%. These results indicate that topoisomerase II activity and sensitivity to a topoisomerase II inhibitor are reduced in 1-day-old spheroid cells. We suggest that the decrease in the activity of this enzyme may be linked to the change in DNA conformation in spheroids and the decrease in their radiation sensitivity.
在悬浮培养中生长成球体的中国仓鼠V79细胞比单层细胞对电离辐射的杀伤和诱变更具抗性。DNA构象的变化似乎伴随着这种辐射抗性的增加。因此，我们感兴趣的是，拓扑异构酶II（一种参与DNA构象变化并可能参与DNA修复的核酶）的活性在单层细胞和小球体中是否会有所不同。1日龄的球体细胞比单层细胞对拓扑异构酶II抑制剂依托泊苷的毒性作用更具抗性。通过DNA沉淀和碱解链试验测量，依托泊苷在球体DNA中诱导的链断裂比在单层DNA中少，尽管拓扑异构酶I抑制剂喜树碱以及细胞周期特异性药物5-氟尿嘧啶在单层细胞和球体中产生的损伤量相同。没有证据表明存在对依托泊苷更具抗性的球体细胞亚群，并且与单层细胞相比，球体中掺入率或DNA链延伸率没有变化。通过锥虫动质体DNA解连环测量，1日龄球体中的拓扑异构酶II活性降至单层细胞活性值的68%；在3 - 4日龄的球体中，该水平为32.5%。这些结果表明，1日龄球体细胞中拓扑异构酶II的活性以及对拓扑异构酶II抑制剂的敏感性降低。我们认为这种酶活性的降低可能与球体中DNA构象的变化及其辐射敏感性的降低有关。
