Unit of Radiation Biology and Human Health, ENEA CR Casaccia, Via Anguillarese 301, 00123 Rome, Italy.
Mutat Res. 2011 Sep 1;714(1-2):44-52. doi: 10.1016/j.mrfmmm.2011.06.010. Epub 2011 Jul 5.
The early pronucleus stage of the mouse zygote has been characterised in vitro as radiosensitive, due to a high rate of induction of chromosome-type chromosome abnormalities (CA). We have investigated the repair of irradiation induced double strand DNA breaks in vivo by γH2AX foci and first cleavage metaphase analysis. Breaks were induced in sperm and in the early zygote stages comprising sperm chromatin remodelling and early pronucleus expansion. Moreover, the role of PARP1 in the formation and repair of spontaneous and radiation-induced double strand breaks in the zygote was evaluated by comparing observations in C57BL/6J and PARP1 genetically ablated females. The results confirmed in vivo that the rate of chromosome aberration induction by X-rays was approximately 3-fold higher in the zygote than in mouse lymphocytes. This finding was related to a diminished efficiency of double strand break signalling, as shown by a lower rate of γH2AX radiation-induced foci compared to that measured in most other somatic cell types. The spontaneous frequency of CA in PARP1 depleted zygotes was slightly but significantly higher than in wild type zygotes. Also, these zygotes showed some impairment of the radiation-induced DNA Damage Response when exposed closer to the start of S-phase, revealed by a higher number of γH2AX foci and a longer cell cycle delay. The rate of chromosome aberrations, however, was not elevated over that of wild type zygotes, possibly thanks to backup repair pathways and/or selection mechanisms against damaged cells. When comparing with the literature data on irradiation induced CA in mouse zygotes in vitro, the levels of induction were strikingly similar as was the frequency of misrepair of double strand breaks (γH2AX foci). This result can be reassuring for in vitro human gamete and embryo handling, because it shows that culture conditions do not significantly affect double strand DNA break repair.
小鼠受精卵的早期原核阶段在体外被描述为对辐射敏感，因为它具有诱导染色体型染色体异常 (CA) 的高频率。我们通过 γH2AX 焦点和第一次卵裂中期分析研究了体内照射诱导的双链 DNA 断裂的修复。在精子和早期受精卵阶段诱导了断裂，包括精子染色质重塑和早期原核扩张。此外，通过比较 C57BL/6J 和 PARP1 基因缺失雌性的观察结果，评估了 PARP1 在受精卵中自发和辐射诱导的双链断裂的形成和修复中的作用。结果证实，与小鼠淋巴细胞相比，X 射线诱导的染色体异常发生率在受精卵中约高 3 倍。这一发现与双链断裂信号的效率降低有关，因为与大多数其他体细胞类型相比，γH2AX 辐射诱导焦点的速率较低。PARP1 耗尽的受精卵中自发 CA 的频率略高，但明显高于野生型受精卵。此外，当接近 S 期开始时，这些受精卵的辐射诱导 DNA 损伤反应显示出一些缺陷，表现为更多的 γH2AX 焦点和更长的细胞周期延迟。然而，染色体异常的发生率并没有超过野生型受精卵，这可能要归功于备用修复途径和/或对受损细胞的选择机制。与体外小鼠受精卵中照射诱导的 CA 的文献数据进行比较时，诱导水平惊人地相似，双链断裂的错误修复频率（γH2AX 焦点）也是如此。这一结果令人放心，因为它表明培养条件不会显著影响双链 DNA 断裂的修复，这对体外人类配子和胚胎处理是有利的。
