Greenwood S K, Armstrong M J, Hill R B, Bradt C I, Johnson T E, Hilliard C A, Galloway S M
Merck Research Laboratories, WP45-305, West Point, PA 19486, USA.
Mutat Res. 1998 Jun 5;401(1-2):39-53. doi: 10.1016/s0027-5107(97)00272-8.
The human lymphoblastoid cell lines TK6 (normal p53) and WI-L2-NS or WTK1 (mutant p53) differ in sensitivity to killing and induction of gene mutations and chromosome aberrations by ionizing radiation. This may be related to decreased apoptosis in the cells with mutated p53, such that more damaged cells survive. We compared the response of the two cell types to various chemicals. First, to ensure that the thymidine kinase deficiency does not increase the sensitivity of TK6 tk+/- cells to mutagens, we demonstrated that they were not hypersensitive to aberration induction by altered DNA precursor pools or DNA synthesis inhibition, by aphidicolin (APC), methotrexate, hydroxyurea (HU), cytosine arabinoside and thymidine. TK6 cells were then compared with WI-L2-NS or WTK1 cells. With APC, HU, methyl methanesulfonate (MMS), ethyl nitrosourea (ENU) and etoposide (etop), TK6 cells had more apoptosis in the first two days after treatment. Fewer aberrations were seen in normal p53 TK6 cells than the mutant p53 WI-L2-NS cells, ranging from very little difference between the two cell types with MMS to very large differences with ENU and etop. For MMS and ENU we followed cultures for several days, and found that WI-L2-NS cells underwent delayed apoptosis 3 to 5 days after treatment, in parallel with published observations with ionizing radiation. WI-L2-NS cells also had a delayed increase in aberrations (up to 5 days post-treatment) when no aberrations remained in TK6 cells. Colony forming efficiency was measured for APC, MMS and ENU, and was greater in the p53 mutant cells. Our results show that normal p53 function is required for rapid and efficient apoptosis in these lymphoblastoid cells with DNA synthesis inhibitors, alkylating agents and a topoisomerase II inhibitor, and support the hypothesis that induced levels of aberrations are higher in p53 mutant cells because of a failure to remove damaged cells by apoptosis.
人淋巴母细胞系TK6（野生型p53）和WI-L2-NS或WTK1（突变型p53）对电离辐射诱导的杀伤、基因突变和染色体畸变的敏感性不同。这可能与p53突变细胞中凋亡减少有关，从而使更多受损细胞存活下来。我们比较了这两种细胞类型对各种化学物质的反应。首先，为确保胸苷激酶缺陷不会增加TK6 tk+/-细胞对诱变剂的敏感性，我们证明它们对通过改变DNA前体库或抑制DNA合成（如阿非迪霉素（APC）、甲氨蝶呤、羟基脲（HU）、阿糖胞苷和胸苷）诱导的畸变不敏感。然后将TK6细胞与WI-L2-NS或WTK1细胞进行比较。用APC、HU、甲基磺酸甲酯（MMS）、乙基亚硝基脲（ENU）和依托泊苷（etop）处理后，TK6细胞在处理后的前两天凋亡更多。野生型p53的TK6细胞中观察到的畸变比突变型p53的WI-L2-NS细胞少，从两种细胞类型对MMS的差异很小到对ENU和etop的差异很大。对于MMS和ENU，我们对培养物进行了数天跟踪，发现WI-L2-NS细胞在处理后3至5天经历延迟凋亡，这与电离辐射的已发表观察结果一致。当TK6细胞中不再有畸变时，WI-L2-NS细胞的畸变也有延迟增加（处理后长达5天）。测量了APC、MMS和ENU的集落形成效率，发现p53突变细胞中的效率更高。我们的结果表明，在这些淋巴母细胞中，对于DNA合成抑制剂、烷化剂和拓扑异构酶II抑制剂，野生型p53功能是快速有效凋亡所必需的，并支持以下假设：由于未能通过凋亡清除受损细胞，p53突变细胞中诱导的畸变水平更高。
