孟熙, 李媛媛, 蒋孑庆, 陈越, 黎小梅, 贺前勇, 张蓬新, 赵朝芬, 唐雅雪, 甘加应, 周建奖, 周定安, 金风. 时钟基因Bmal1对鼻咽癌移植瘤放疗敏感性的影响[J]. 中国肿瘤临床, 2020, 47(1): 12-17. DOI: 10.3969/j.issn.1000-8179.2020.01.327
引用本文: 孟熙, 李媛媛, 蒋孑庆, 陈越, 黎小梅, 贺前勇, 张蓬新, 赵朝芬, 唐雅雪, 甘加应, 周建奖, 周定安, 金风. 时钟基因Bmal1对鼻咽癌移植瘤放疗敏感性的影响[J]. 中国肿瘤临床, 2020, 47(1): 12-17. DOI: 10.3969/j.issn.1000-8179.2020.01.327
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Xi Meng, Yuanyuan Li, Jieqing Jiang, Yue Chen, Xiaomei Li, Qianyong He, Pengxin Zhang, Chaofen Zhao, Yaxue Tang, Jiaying Gan, Jianjiang Zhou, Ding'an Zhou, Feng Jin. Effect of Bmal1 expression on radiotherapy sensitivity of nasopharyngeal carcinoma xenograft[J]. CHINESE JOURNAL OF CLINICAL ONCOLOGY, 2020, 47(1): 12-17. DOI: 10.3969/j.issn.1000-8179.2020.01.327
Citation: Xi Meng, Yuanyuan Li, Jieqing Jiang, Yue Chen, Xiaomei Li, Qianyong He, Pengxin Zhang, Chaofen Zhao, Yaxue Tang, Jiaying Gan, Jianjiang Zhou, Ding'an Zhou, Feng Jin. Effect of Bmal1 expression on radiotherapy sensitivity of nasopharyngeal carcinoma xenograft[J]. CHINESE JOURNAL OF CLINICAL ONCOLOGY, 2020, 47(1): 12-17. DOI: 10.3969/j.issn.1000-8179.2020.01.327
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时钟基因Bmal1对鼻咽癌移植瘤放疗敏感性的影响

Effect of Bmal1 expression on radiotherapy sensitivity of nasopharyngeal carcinoma xenograft

    摘要
  • 摘要:
      目的  探讨双向调控时钟基因Bmal1（brain and muscle arnt-like1）对鼻咽癌CNE1裸鼠移植瘤生长的影响及与放疗敏感性的关系。
      方法  采用慢病毒感染方法，构建Bmal1基因CNE1OE（过表达组）、CNE1OENC（过表达对照组）、CNE1sh3（低表达组）、CNE1shNC（低表达对照组）4株细胞，Western blot验证各组细胞Bmal1蛋白表达情况；4株细胞分别皮下注射相应的4组裸鼠，移植瘤生长后测量其体积；给予6-MeV电子线15 Gy照射裸鼠移植瘤，观察放疗后各组移植瘤体积变化情况。剥离移植瘤，RTPCR及Western blot分别检测Bmal1、P53、P21 mRNA及蛋白表达情况。
      结果  Western blot结果提示，与各自对照组对比，CNE1OE组Bmal1蛋白过表达，CNE1sh3组Bmal1蛋白低表达，表明细胞转染成功。成功构建裸鼠移植瘤模型，CNE1OE组裸鼠移植瘤体积明显小于CNE1OENC组，CNE1sh3组体积明显大于CNE1shNC组（P < 0.05）。放疗后，CNE1OE、CNE1OENC、CNE1shNC组裸鼠移植瘤体积均缩小（P < 0.05），其中CNE1OE组缩小最明显。CNE1sh3组放疗前后自身体积变化差异无统计学意义。Bmal1基因、P53及P21的mRNA及蛋白相对表达量在CNE1OE组明显高于CNE1OENC组（P < 0.05），CNE1sh3组则明显低于CNE1shNC组（P < 0.05）。
      结论  Bmal1基因过表达能抑制鼻咽癌CNE1裸鼠移植瘤生长，增强其放疗敏感性，可能与P53、P21蛋白上调相关；敲低则促进移植瘤生长，导致其辐射抗拒，可能与P53、P21蛋白下调相关。

     

    Abstract:
      Objective  To explore the effects of up- and down-regulation of circadian clock gene Bmal1 on the growth and radiation sensitivity of nasopharyngeal carcinoma after CNE1 xenograft in nude mice.
      Methods  We produced four groups of cells using lentiviral transfection:cells overexpressing CNE1 (CNE1OE), negative control in which there was no overexpression of CNE1 (OENC), cells with short hairpin RNAi (CNE1sh3), and RNAi negative cells (CNE1shNC). We investigated the expression of Bmal1 protein in the aforementioned groups with Western blot. After subcutaneously injecting the four groups of cells in nude mice, the size of the xenograft was measured. Subsequently, the xenografts were irradiated with 15 Gy at 6 MeV, and variation in the xenograft volume was recorded. mRNA and protein expression levels of Bmal1, p53, and p21 in the xenograft were measured with RT-PCR and Western blot, respectively.
      Results  The CNE1OE group highly expressed Bmal1 protein whereas the CNE1sh3 group was silenced by RNAi as shown with the Western blot, indicating successful transfection. The xenograft in the nude mice developed well. The CNE1OE xenograft volume was lower than that of the CNE1OENC xenograft, whereas the CNE1sh3 xenograft was larger than the CNE1shNC xenograft (P < 0.05). CNE1OE, CNE1OENC, and CNE1shNC xenograft volumes shrank after being irradiated (t=4.32, 5.38, 5.16, respectively; P < 0.05) and the effect was the highest in the CNE1OE group. However, there was almost no variation in xenograft volume in the CNE1sh3 group. The relative amounts of mRNA and protein of Bmal1, P53, and P21 were higher in the CNE1OE group than in the CNE1OENC group, while they were lower in the CNE1sh3 group compared to the CNE1shNC group (P < 0.05).
      Conclusions  Overexpression of Bmal1 inhibited the growth of the CNE1 xenograft in nude mice and enhanced its radiation sensitivity whereas silencing the Bmal1 gene by RNAi promoted the growth of the xenograft and led to radiation resistance. We believe that Bmal1 overexpression leads to P53 and P21 overexpression, thereby inhibiting the growth of the xenograft.

     

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