Background
5-ethynyl-2′-deoxyuridine (EdU), as a thymidine analoghas, a terminal alkyne group replacing a methyl group at the 5 position of the pyrimidine ring and can be readily incorporated into DNA during synthesis[1]. EdU is a inhibitor of the cell growth of human breast cancer cells (MCF-7 and MDA-MP-231) with the IC50 of 0.4 μM for MCF-7 cells and 4.4 μM for MDA-MB-231 cells[2].
In vitro, 10 μMEdU induced gamma-H2AX foci for 51D1 and KO40 in CHO cells for DNA damage responses[3]. In vitro experiment it shown that A549 cells were treated with 20 μM EdU for 6 h increased the level of expression of γH2AX and ATM-S1981P by 88% and 116%, respectively[4].
In vivo efficacy test it shown that femalJKe mice were administrated 10, 20, 50, 100 or 200 mg/kg EdU intraperitoneally, there is an increase in the number of EdU positive cells in the DG slightlyin a dose-dependent manner[5]. In vivo, few EdU-positive cells were observed in Wistar and GK rats at a dose of 5 mg/kg and 25 mg/kg EdU intraperitoneally. Prominent EdU-positive cells were observed in Wistar and GK rats at a dose of 100 mg/kg and 200 mg/kg, respectively[6]. In vivo, use of 5-Ethynyl-2'-deoxyuridine (EdU) incorporation to in vivo monitor T lymphocyte proliferation by flow cytometry with an adoptive transfer model. The result shown that the percentage of EdU-positive cells increased in a dose-dependent manner, and the saturated dose of EdU was 20mg/kg. Intraperitoneal and intravenous injection had no differences in lymphocyte proliferation detection with EdU in vivo[7].
5-乙炔基-2′-脱氧尿苷（EdU），作为胸苷类似物，是取代嘧啶环5位甲基的末端炔基，在合成过程中可以很容易地结合到DNA中[1]。EdU是人类乳腺癌症细胞（MCF-7和MDA-MP-231）的细胞生长抑制剂，MCF-7细胞的IC50为0.4 μM，MDA-MB-231细胞为4.4μM[2]。
在体外，10 μM EdU诱导CHO细胞中51D1和KO40的γ-H2AX焦点产生DNA损伤反应[3]。体外实验表明，用20 μM EdU处理A549细胞6小时，γH2AX和ATM-S1981P的表达水平分别提高了88%和116%[4]。
体内疗效测试表明，雌性JKe小鼠腹膜内给予10、20、50、100或200 mg/kg EdU，DG中EdU阳性细胞的数量略有增加，呈剂量依赖性[5]。在体内，在Wistar和GK大鼠中观察到腹膜内剂量为5 mg/kg和25 mg/kg EdU的EdU阳性细胞很少。在Wistar和GK大鼠中分别观察到剂量为100 mg/kg和200 mg/kg的显著EdU阳性细胞[6]。在体内，使用5-乙炔基-2'-脱氧尿苷（EdU）掺入通过过继转移模型的流式细胞术在体内监测T淋巴细胞增殖。结果表明，EdU阳性细胞的百分比呈剂量依赖性增加，EdU的饱和剂量为20 mg/kg。腹膜内注射和静脉注射在体内用EdU检测淋巴细胞增殖方面没有差异[7]。
参考文献:
[1] Kaiser CL, et al. 5-Ethynyl-2'-deoxyuridine labeling detects proliferating cells in the regenerating avian cochlea. Laryngoscope. 2009 Sep;119(9):1770-5.
[2] Meneni S, et al. (2007) 5-Alkynyl-2′-deoxyuridines: chromatography-free synthesis and cytotoxicity evaluation against human breast cancer cells. Bioorg Med Chem 15: 3082–3088.
[3] Haskins JS, et al. Evaluating the Genotoxic and Cytotoxic Effects of Thymidine Analogs, 5-Ethynyl-2'-Deoxyuridine and 5-Bromo-2'-Deoxyurdine to Mammalian Cells. Int J Mol Sci. 2020 Sep 10;21(18):6631.
[4] Zhao H, et al. DNA damage signaling, impairment of cell cycle progression, and apoptosis triggered by 5-ethynyl-2'-deoxyuridine incorporated into DNA. Cytometry A. 2013 Nov;83(11):979-88.
[5] Zeng C, et al. Evaluation of 5-ethynyl-2'-deoxyuridine staining as a sensitive and reliable method for studying cell proliferation in the adult nervous system. Brain Res. 2010 Mar 10;1319:21-32.
[6] Guo J, Li D, et al. Detecting DNA synthesis of neointimal formation after catheter balloon injury in GK and in Wistar rats: using 5-ethynyl-2'-deoxyuridine. Cardiovasc Diabetol. 2012 Dec 13;11:150.
[7] Sun X, Zhang C, et al. Flow cytometric analysis of T lymphocyte proliferation in vivo by EdU incorporation. Int Immunopharmacol. 2016 Dec;41:56-65.
Protocol
| Cell experiment [1]: |
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Cell lines
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HeLa cells
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Preparation Method
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HeLa cells were incubated with either 10 μM EdU or 10 μM EdC for 4 h. Then, the detection of EdU and DNA using DAPI was performed.
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Reaction Conditions
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10 μM; 4h
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Applications
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The result shown approximately 75% of the cells contained a signal corresponding to the localization of incorporated EdU.
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| Animal experiment [2]: |
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Animal models
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bird
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Preparation Method
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In order to label mitotically active supporting cells, a single, subcutaneous injection of EdU (50 mg/kg) in sterile, phosphate buffered saline (PBS, pH 7.4) was administered to each bird 72h after the gentamicin injection.
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Dosage form
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50 mg/kg; s.c.
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Applications
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When EdU is injected at 72h and allowed to incorporate for 4h, some of the supporting cells that first entered S phase around 65h are labeled with EdU at the end of their S phase. They are able to reach the mitotic phase and some even divide during this time.
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参考文献:
Ligasová A, et al. Dr Jekyll and Mr Hyde: a strange case of 5-ethynyl-2'-deoxyuridine and 5-ethynyl-2'-deoxycytidine. Open Biol. 2016 Jan;6(1):150172.
Kaiser CL, et al. 5-Ethynyl-2'-deoxyuridine labeling detects proliferating cells in the regenerating avian cochlea. Laryngoscope. 2009 Sep;119(9):1770-5.
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