Sokolov Mykyta V, Smilenov Lubomir B, Hall Eric J, Panyutin Igor G, Bonner William M, Sedelnikova Olga A
Department of Nuclear Medicine, Clinical Center, NIH, 9000 Rockville Pike, Bethesda, MD 20892, USA.
Oncogene. 2005 Nov 10;24(49):7257-65. doi: 10.1038/sj.onc.1208886.
That irradiated cells affect their unirradiated 'bystander' neighbors is evidenced by reports of increased clonogenic mortality, genomic instability, and expression of DNA-repair genes in the bystander cell populations. The mechanisms underlying the bystander effect are obscure, but genomic instability suggests DNA double-strand breaks (DSBs) may be involved. Formation of DSBs induces the phosphorylation of the tumor suppressor protein, histone H2AX and this phosphorylated form, named gamma-H2AX, forms foci at DSB sites. Here we report that irradiation of target cells induces gamma-H2AX focus formation in bystander cell populations. The effect is manifested by increases in the fraction of cells in a population that contains multiple gamma-H2AX foci. After 18 h coculture with cells irradiated with 20 alpha-particles, the fraction of bystander cells with multiple foci increased 3.7-fold. Similar changes occurred in bystander populations mixed and grown with cells irradiated with gamma-rays, and in cultures containing media conditioned on gamma-irradiated cells. DNA DSB repair proteins accumulated at gamma-H2AX foci, indicating that they are sites of DNA DSB repair. Lindane, which blocks gap-junctions, prevented the bystander effect in mixing but not in media transfer protocols, while c-PTIO and aminoguanidine, which lower nitric oxide levels, prevented the bystander effect in both protocols. Thus, multiple mechanisms may be involved in transmitting bystander effects. These studies show that H2AX phosphorylation is an early step in the bystander effect and that the DNA DSBs underlying gamma-H2AX focus formation may be responsible for its downstream manifestations.
有报道称，受照射细胞会影响其未受照射的“旁观者”邻居，这体现在旁观者细胞群体中克隆形成死亡率增加、基因组不稳定以及DNA修复基因表达增强。旁观者效应背后的机制尚不清楚，但基因组不稳定表明可能涉及DNA双链断裂（DSB）。DSB的形成会诱导肿瘤抑制蛋白组蛋白H2AX磷酸化，这种磷酸化形式称为γ-H2AX，会在DSB位点形成焦点。在此我们报告，对靶细胞进行照射会在旁观者细胞群体中诱导γ-H2AX焦点形成。这种效应表现为含有多个γ-H2AX焦点的细胞群体比例增加。与经20个α粒子照射的细胞共培养18小时后，具有多个焦点的旁观者细胞比例增加了3.7倍。在与经γ射线照射的细胞混合并生长的旁观者群体中，以及在含有经γ射线照射细胞的条件培养基的培养物中，也发生了类似的变化。DNA DSB修复蛋白在γ-H2AX焦点处积累，表明它们是DNA DSB修复的位点。林丹可阻断间隙连接，在混合实验中可防止旁观者效应，但在培养基转移实验中则不能，而降低一氧化氮水平的c-PTIO和氨基胍在两种实验中均可防止旁观者效应。因此，多种机制可能参与了旁观者效应的传递。这些研究表明，H2AX磷酸化是旁观者效应的早期步骤，γ-H2AX焦点形成背后的DNA DSB可能是其下游表现的原因。
