Exposure to continuous bromodeoxyuridine (BrdU) differentially affects cell cycle progression of human breast and bladder cancer cell lines
Exposure to continuous bromodeoxyuridine (BrdU) differentially affects cell cycle progression of human breast and bladder cancer cell lines
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DOI：
10.1111/j.1365-2184.2004.00296.x
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发表时间：
2004-04-01
影响因子：
8.5
通讯作者：
Brockhoff, G
中科院分区：
文献类型：
--
作者：
Diermeier, S;Schmidt-Bruecken, E;Brockhoff, G
在DNA复制过程中掺入溴脱氧尿苷（BrdU）常用于细胞周期分析。流式细胞仪BrdU/Hoechst淬灭技术有助于高分辨率评估细胞周期动力学，但需要连续的BrdU治疗，这可能具有细胞抑制或细胞毒性作用。在这里，我们研究了BrdU对BT474和SK-BR-3乳腺癌细胞系增殖的影响，并将观察到的效果与RT4和J82膀胱癌细胞的细胞增殖进行了比较，先前分别描述为对BrdU敏感和不敏感。进行单参数和双参数DNA测量以鉴定BrdU诱导的S期分数和细胞周期进程的改变。使用膜联蛋白V/碘化丙啶（PI）测定来鉴定BrdU对细胞凋亡的潜在诱导作用。由于连续用60、5.0和3.5 μ M BrdU处理，BT474、SK-BR-3和RT4培养物中的还原活性在不同的细胞周期阶段降低。在J82培养物中未发现这种效应，其依赖于暴露时间（96小时与48小时），并且对于RT 4和SK-BR-3也具有剂量依赖性。BrdU应用不诱导细胞凋亡或坏死，如膜联蛋白V/PI测定所示。我们的结论是，连续BrdU治疗不影响细胞活力，但基本上改变了细胞周期的进展，在三个四个细胞系测试。需要并建议对强大的BrdU/Hoechst淬灭技术的可行性进行细胞类型特异性验证。
Incorporation of bromodeoxyuridine (BrdU) during DNA replication is frequently used for cell cycle analysis. The flow cytometric BrdU/Hoechst quenching technique is conducive to high-resolution assessment of cell cycle kinetics, but requires continuous BrdU treatment, which may have cytostatic or cytotoxic effects. Here, we have examined the impact of BrdU on the proliferation of BT474 and SK-BR-3 breast cancer cell lines and compared the observed effects with cell proliferation of RT4 and J82 bladder carcinoma cells, previously described to be sensitive and insensitive to BrdU, respectively. Both uni- and bi-parametric DNA measurements were performed to identify BrdU-induced alterations in the S-phase fraction and in cell cycle progression. An annexinV/propidium iodide (PI) assay was used to identify potential induction of apoptosis by BrdU. Proliferative activity in BT474, SK-BR-3, and RT4 cultures was reduced in different cell cycle phases due to continuous treatment with 60, 5.0, and 3.5 muM BrdU. This effect, which was not found in J82 cultures, was dependent on exposure time (96 versus 48 h) and was also dose-dependent for RT4 and SK-BR-3. BrdU application does not induce apoptosis or necrosis as revealed with the annexin V/PI assay. We concluded that continuous BrdU treatment did not affect cell viability, but essentially alters cell cycle progression in three out of four cell lines tested. Cell-type specific validation of the feasibility of the powerful BrdU/Hoechst quenching technique is required and recommended.