海南哥纳香醇甲GHM-10对L1210细胞DNA分子结构及拓扑异构酶Ⅱ活性的影响

EFFECT OF HOWIINOL(GHM 10) ON THE STRUCTURE OF DNA MOLECULES AND ACTIVITY OF DNA TOPOISOMERASE II IN L1210 CELLS

目的：研究海南哥纳香醇甲（ＧＨＭ１０）抑制癌细胞ＤＮＡ合成的作用机制。方法：用单细胞凝胶电泳法检测ＧＨＭ１０对Ｌ１２１０细胞ＤＮＡ分子的损伤，碱洗脱法测定ＧＨＭ１０对Ｌ１２１０细胞ＤＮＡ单链长度的影响，用ＧＨＭ１０对超螺旋ｐＵＣ１８ＤＮＡ的解旋能力测定它对ＤＮＡ拓扑异构酶ＩＩ活性的影响。结果：Ｌ１２１０细胞用ＧＨＭ１０（４～１０）μｇ·ｍｌ－１处理４５ｈ后，ＤＮＡ分子受损，表现为电泳后在荧光显微镜下可见彗星状拖尾。ＧＨＭ１０（４～２５）μｇ·ｍｌ－１处理Ｌ１２１０细胞５ｈ，可引起ＤＮＡ单链断裂。Ｌ１２１０细胞或从Ｌ１２１０细胞分离的蛋白质在用ＧＨＭ１０处理后，ＤＮＡ拓扑异构酶ＩＩ的活性均被抑制。结论：ＧＨＭ１０可引起Ｌ１２１０细胞ＤＮＡ分子损伤；无论在细胞内还是细胞外，ＧＨＭ１０可抑制拓扑异构酶ＩＩ的活性。

AIM: To study the mechanism of inhibitory action of howiinol A(GHM 10) on synthesis of DNA in cancer cells. METHODS: Single cell gel electrophoresis was used to detect the DNA molecular damage induced by GHM 10 in L1210 cells. Alkaline elution technique was used to investigate the effect of GHM 10 on the length of DNA strands of L1210 cells. Dehelix effect of GHM 10 on superhelix pUC 18 DNA was used to estimate its influence on the activity of DNA topoisomerase II. RESULTS: After L1210 cells were treated with 4 and 10 μg·ml -1 of GHM 10 for 4 5 h, comet like tails could be seen in cells under a fluorescence microscope after electrophoresis, indicating that DNA damage was induced by GHM 10. DNA single strand breaks were demonstrated when L1210 cells were treated with GHM 10 in concentrations of 4 and 25 μg·ml -1 for 5 h. When L1210 cells or protein isolated from L1210 cells were treated with GHM 10, the activity of DNA topoisomerase II was inhibited. CONCLUSION: GHM 10 induces damage to DNA molecules in L1210 cells and inhibits the activity of topoisomerase II both inside and outside the cells.