Quantitative detection of 4-hydroxyequilenin-DNA adducts in mammalian cells using an immunoassay with a novel monoclonal antibody.
Quantitative detection of 4-hydroxyequilenin-DNA adducts in mammalian cells using an immunoassay with a novel monoclonal antibody.
复制标题
使用新型单克隆抗体进行免疫测定，定量检测哺乳动物细胞中的 4-羟基马萘醌-DNA 加合物。
DOI：
10.1093/nar/gkq233
复制
发表时间：
2010-07
影响因子：
14.9
通讯作者：
Mori, Toshio
中科院分区：
文献类型：
--
作者：
Okahashi, Yumiko;Iwamoto, Takaaki;Suzuki, Naomi;Shibutani, Shinya;Sugiura, Shigeki;Itoh, Shinji;Nishiwaki, Tomohisa;Ueno, Satoshi;Mori, Toshio
雌激素 - DNA加合物是评估雌激素相关癌症风险和发展的潜在生物标志物。4 - 羟基马萘雌酮（4 - OHEN）和4 - 羟基马烯雌酮（4 - OHEQ）是常见激素替代疗法（HRT）配方中含有的马雌激素代谢物，能够产生 bulky（庞大的、体积大的，此处可能指结构复杂的）4 - OHEN - DNA加合物。尽管已有关于4 - OHEN - DNA加合物形成的报道，但在哺乳动物细胞中对其进行定量检测尚未实现。为了对这类DNA加合物进行定量，我们制备了一种新型单克隆抗体（4OHEN - 1），它对4 - OHEN - DNA加合物具有特异性。其识别的主要表位是DNA中4 - OHEN - dA加合物和4 - OHEN - dC加合物的一种立体异构体。用4OHEN - 1进行的免疫测定显示，已知量的4 - OHEN - DNA加合物与抗体对这些加合物的结合之间呈线性剂量 - 反应关系，在1微克DNA样本中检测限约为每10⁸个碱基5个加合物。在人乳腺癌细胞中，定量免疫测定显示4 - OHEN产生的4 - OHEN - DNA加合物比4 - OHEQ多5倍。此外，在一个激素替代疗法的小鼠模型中，口服普力马（Premarin）以时间依赖的方式增加了包括子宫和卵巢在内的各种组织中4 - OHEN - DNA加合物的水平。因此，我们成功建立了一种用于定量检测哺乳动物细胞中4 - OHEN - DNA加合物的新型免疫测定方法。
Estrogen–DNA adducts are potential biomarkers for assessing the risk and development of estrogen-associated cancers. 4-Hydroxyequilenin (4-OHEN) and 4-hydroxyequilin (4-OHEQ), the metabolites of equine estrogens present in common hormone replacement therapy (HRT) formulations, are capable of producing bulky 4-OHEN–DNA adducts. Although the formation of 4-OHEN–DNA adducts has been reported, their quantitative detection in mammalian cells has not been done. To quantify such DNA adducts, we generated a novel monoclonal antibody (4OHEN-1) specific for 4-OHEN–DNA adducts. The primary epitope recognized is one type of stereoisomers of 4-OHEN–dA adducts and of 4-OHEN–dC adducts in DNA. An immunoassay with 4OHEN-1 revealed a linear dose–response between known amounts of 4-OHEN–DNA adducts and the antibody binding to those adducts, with a detection limit of approximately five adducts/108 bases in 1 µg DNA sample. In human breast cancer cells, the quantitative immunoassay revealed that 4-OHEN produces five times more 4-OHEN–DNA adducts than does 4-OHEQ. Moreover, in a mouse model for HRT, oral administration of Premarin increased the levels of 4-OHEN–DNA adducts in various tissues, including the uterus and ovaries, in a time-dependent manner. Thus, we succeeded in establishing a novel immunoassay for quantitative detection of 4-OHEN–DNA adducts in mammalian cells.
登录
查看更多内容
影响因子：
120.7
作者：
Chen, CL;Weiss, NS;White, E
通讯作者：
White, E
影响因子：
158.5
作者：
COLDITZ, GA;HANKINSON, SE;SPEIZER, FE
通讯作者：
SPEIZER, FE
影响因子：
56.9
作者：
LITTLEFIELD, JW
通讯作者：
LITTLEFIELD, JW
影响因子：
3.3
作者：
MORI, T;NAKANE, M;NIKAIDO, O
通讯作者：
NIKAIDO, O
影响因子：
2.3
作者：
Embrechts, J;Lemière, F;Esmans, EL
通讯作者：
Esmans, EL