Deoxyribonucleoside Triphosphate Imbalance: The Mechanism of Cell Death and DNA-Double Strand Breaks.

研究了小鼠FM 3A细胞内脱氧核苷三磷酸（dNTP）库失衡导致细胞死亡和dNA双链断裂的机制。在用20 m2-氯脱氧腺苷处理的3小时内发生dNTP池的不平衡; dATP和dGTP池被耗尽，而dTTP池增加。2-加入到培养基中的氯脱氧腺苷使成熟的DNA链断裂，通过正交场交替凝胶电泳发现，产生100-200个酶对的片段。通过该技术测量的DNA链断裂在添加后约12小时在处理的细胞中观察到。细胞在约12小时也失去活力。断裂的DNA的单链和双链，作为测量的碱性和中性过滤器洗脱，成为明显的18小时后，治疗与20米2-氯脱氧腺苷，有尽可能多的OD单链断裂会造成130拉德的射线照射。双链断裂相当于由2180拉德的射线照射引起的断裂。比较2-氯脱氧腺苷与辐射后引起的单链和双链断裂的比率表明，2-氯脱氧腺苷仅断裂双链。环己酰亚胺可抑制DNA双链断裂和细胞死亡。流式细胞术研究2-氯脱氧腺苷对FM 3A细胞的细胞抑制作用，即细胞在S期的早期积累。2-氯脱氧腺苷减少DNA合成比RNA或蛋白质合成。双链DNA的断裂可能是2-氯脱氧腺苷引起细胞死亡的重要原因。细胞内dNTP失衡可能触发这些事件。

The mechanism of intracellular deoxyribonucleoside-triphosphate (dNTP)-pool imbalance induced cell death and dNA double strand breaks in mouse FM3A cells was investigated. imbalance of the dNTP pools occurred within 3 h of treatment with 20 m 2-chlorodeoxyadenosine; the dATP and dGTP pools were depleted and the dTTP pool increased. 2-Chlorodeoxyadenosine added to the culture medium broke mature DNA strands, giving fragments of 100-200 kilobase pairs as found by orthogonal-field-alternation gel electrophoresis. DNA strand breaks, measured by this technique, were observed in the treated cells about 12 h after the addition. The cells also lost viability at about 12 h. Breaks in the single and double strands of DNA, as measured by alkaline and neutral filter elution, became evident 18 h after treatment with 20 m 2-chlorodeoxyadenosine; there were as many od single-strand breaks as would be caused by 130 rads of -ray irradiation. Double-strand breaks were equivalent to those caused by 2180 rads of -ray irradiation. Comparison of the ratio of singl- and double-strand breaks caused by 2-chlorodeoxyadenosine to that following radiation suggested that 2-chlorodeoxyadenosine broke only double strands. Cycloheximide inhibited the breakage of DNA double strands and the cell death caused by this compound. Flow cytometric studies of cytostatit brought about by 2-chlorodeoxyadenosine in FM3A cells that cells accumulated in the earlier part of the s phase. 2-Chlorodeoxyadenosine decreased DNA synthesis more than RNA or protein synthesis. The breaks in double strands DNA were probably important in the cell death caused by 2-chlorodeoxyadenosine. The intracellular dNTP imbalance may trigger these events.