Laboratorio de Citogenética y Mutagénesis, Instituto Multidisciplinario de Biología Celular (IMBICE, CCT-CONICET La Plata-CICPBA), C.C. 403, 1900 La Plata, Argentina.
Mutat Res. 2012 Jun 1;734(1-2):5-11. doi: 10.1016/j.mrfmmm.2012.04.008. Epub 2012 May 4.
We analyzed the chromosomal aberrations involving telomeres in the progeny of mammalian cells exposed to the radiomimetic compound bleomycin (BLM) in order to determine if this antineoplastic drug induces long-term telomere instability. To this end, rat cells (ADIPO-P2 cell line, derived from adipose cells from Sprague-Dawley rat) were treated with a single concentration of BLM (2.5 μg/ml), and chromosomal aberrations were analyzed 18 h and 10 days after treatment by using PNA-FISH with a pan-telomeric probe [(TTAGGG)n repeats]. Cytogenetic analysis revealed a higher frequency of aberrations at 18 h and 10 days after treatment in BLM-exposed cultures vs. untreated cultures, although the yield of BLM-induced aberrations 10 days after treatment decreased about 25% compared with the one at 18 h after treatment. Moreover, the level of telomerase activity in BLM-treated cells compared with that of untreated control cells was significantly higher at 10 days after treatment, but did not differ at 18 h after treatment. These data indicate that in terms of unstable aberrations, the in vitro clastogenic effect of BLM on ADIPO-P2 cells persists for at least 10 days after exposure. In addition, our data demonstrate, for the first time, that BLM-induced telomere instability in mammalian cells (cytogenetically detectable as incomplete chromosome elements and telomere FISH signal loss and duplication) persists for several generations after exposure. Moreover, the appearance of telomere fusions in BLM-exposed cells 10 days after treatment suggests that this compound can induce delayed telomere instability. The increase in telomerase activity in BLM-exposed cells 10 days after treatment is accompanied by the presence of aberrations directly related to telomere dysfunction. This fact suggests that telomerase is not directly involved in BLM-induced telomere instability.
我们分析了哺乳动物细胞系在受到放射模拟化合物博来霉素（BLM）处理后涉及端粒的染色体畸变，以确定这种抗肿瘤药物是否会导致端粒的长期不稳定。为此，用单一浓度 BLM（2.5μg/ml）处理大鼠细胞（ADIPO-P2 细胞系，源自 Sprague-Dawley 大鼠的脂肪细胞），并在处理后 18 小时和 10 天用 PNA-FISH 分析带有全端粒探针的染色体畸变 [(TTAGGG)n 重复]。细胞遗传学分析显示，与未处理的培养物相比，BLM 暴露培养物在处理后 18 小时和 10 天的染色体畸变频率更高，尽管与处理后 18 小时相比，处理后 10 天 BLM 诱导的畸变产率降低了约 25%。此外，与未经处理的对照细胞相比，BLM 处理细胞的端粒酶活性在处理后 10 天明显更高，但在处理后 18 小时没有差异。这些数据表明，就不稳定的畸变而言，BLM 对 ADIPO-P2 细胞的体外致裂效应在暴露后至少持续 10 天。此外，我们的数据首次表明，BLM 诱导的哺乳动物细胞端粒不稳定（在细胞遗传学上可检测为不完整的染色体元素和端粒 FISH 信号丢失和复制）在暴露后几个代际中持续存在。此外，在处理后 10 天，BLM 暴露细胞中端粒融合的出现表明该化合物可以诱导延迟的端粒不稳定。在处理后 10 天，BLM 暴露细胞中端粒酶活性的增加伴随着与端粒功能障碍直接相关的畸变的出现。这一事实表明，端粒酶不直接参与 BLM 诱导的端粒不稳定。
