siRNA targeted for NBS1 enhances heat sensitivity in human anaplastic thyroid carcinoma cells
siRNA targeted for NBS1 enhances heat sensitivity in human anaplastic thyroid carcinoma cells
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DOI：
10.3109/02656736.2010.545365
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发表时间：
2011-04
影响因子：
3.1
通讯作者：
N. Okamoto;A. Takahashi;I. Ota;K. Ohnishi;Eiichiro Mori;N. Kondo;Taichi Noda;Y. Nakagawa;H. Uemura;Katunari Yane;H. Hosoi;T. Ohnishi
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文献类型：
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作者：
N. Okamoto;A. Takahashi;I. Ota;K. Ohnishi;Eiichiro Mori;N. Kondo;Taichi Noda;Y. Nakagawa;H. Uemura;Katunari Yane;H. Hosoi;T. Ohnishi
奈梅亨断裂综合征 1 (NBS1) 作为辐射诱导的 DNA 双链断裂 (DSB) 修复中的关键蛋白发挥着重要作用，本文描述的工作旨在检查 NBS1 对人未分化甲状腺癌 8305c 细胞热敏感性的影响。通过集落形成测定评估细胞热敏感性。使用末端脱氧核苷酸转移酶介导的 dUTP 缺口末端标记 (TUNEL) 测定和 Hoechst33342 染色测定来检测和定量细胞凋亡。使用 γH2AX 抗体通过流式细胞术测量热诱导的 DSB。将NBS1-siRNA转染细胞后，特异性抑制NBS1的表达，并通过caspase途径增强热敏感性和细胞凋亡频率。此外，在用pan-caspase抑制剂热处理24小时后，在NBS1-siRNA转染的细胞中观察到比用乱序siRNA转染的对照细胞更频繁的γH2AX焦点。这些结果表明热敏化可能是由 NBS1-siRNA 介导的热诱导 DSB 修复抑制引起的，表明 NBS1-siRNA 可能作为癌症患者的热敏剂发挥作用。
Nijmegen breakage syndrome 1 (NBS1) plays an important role as a key protein in the repair of radiation-induced DNA double strand breaks (DSBs), and the work described here was designed to examine the effect of NBS1 on heat sensitivity for human anaplastic thyroid carcinoma 8305c cells. Cellular heat sensitivity was evaluated with colony formation assays. Apoptosis was detected and quantified with terminal deoxynucleotidyl transferase mediated dUTP nick end labelling (TUNEL) assay and Hoechst33342 staining assay. Heat-induced DSBs were measured with flow cytometry using γH2AX antibodies. The transfection of NBS1-siRNA into cells specifically inhibited the expression of NBS1, and enhanced heat sensitivity and the frequency of apoptosis through caspase pathway. In addition, more frequent γH2AX foci were observed in the NBS1-siRNA transfected cells than in control cells transfected with scrambled siRNA at 24 h after heat treatment with a pan-caspase inhibitor. These results suggest that heat sensitisation might result from NBS1-siRNA mediated suppression of heat-induced DSB repair, indicating that NBS1-siRNA could potentially function as a heat sensitiser for cancer patients.