沉默PKM2对人肺腺癌细胞的放射增敏 研究
Effect of silencing of PKM2 expression on radiosensitivity of lung adenocarcinoma cell
目的 研究沉默丙酮酸激酶M2型(PKM2)对人肺腺癌A549细胞的放射敏感性,并探讨其潜在的相关机制。方法 将PKM2基因干扰质粒pshRNA-PKM2稳定转染至A549 细胞(pshRNA-PKM2组),同时设立空载质粒转染组(pshRNA-Control组)和未转染组(Control组)。采用免疫印迹法(Western Blot)检测pshRNA-PKM2的沉默效率;台盼蓝和克隆形成实验分别检测沉默PKM2后放疗对细胞存活能力及克隆形成能力的影响;透射电镜和Western Blot法分别检测沉默PKM2的A549细胞放疗后自噬体和自噬小泡形成及微管相关蛋白1轻链3(LC3)的表达水平。结果 成功获得转染pshRNA-PKM2的A549稳定细胞株,pshRNA-PKM2组可显著下调PKM2的蛋白表达水平(p<0.05)。pshRNA-PKM2可降低放疗后细胞的活性。pshRNA-PKM2组与pshRNA-Control组的放射增敏比(SERD0值)分别为1.47与1.01。干扰PKM2可增加放疗所诱导的细胞内自噬体及自噬小泡形成,同时显著增加LC3-II/I的比值(p<0.05)。结论 沉默PKM2调节自噬提高人肺腺癌A549细胞的放射敏感性,其有望成为放疗非小细胞肺癌的有效增敏靶点。
Methods A plasmid of pshRNA-PKM2 for expressing a short hairpin RNA targeting PKM2, pshRNA-Control, and Control were simulitantly constructed and transfered into A549 cells. The protein expression of PKM2 was measured. The change in cell radiosensitivity was observed by colony-forming assay and the frequency of viable cells, and the autophagosome as well as the protein level of microtubule-associated protein 1 light chain 3 (LC3) were also measured, respectively. Results Treatment with pshRNA-PKM2 effectively inhibited PKM2 expression in A549 cells (p<0.05). The results demonstrated that pshRNA-PKM2 and ionizing radiation (IR) significantly reduced the frequency of viable A549 cells, and the sensitization enhancement ratio (SERD0 ratio) for pshRNA-PKM2 and pshRNA-Control were 1.47 and 1.01. Knockdown of PKM2 expression and IR induced A549 cell autophagy, which was associated with significantly increase levels of LC3 (p<0.05). Conclusion Silencing of PKM2 expression enhances the radiosensitivity of A549 cells by inducing autophagy, and may be used as a novel strategy for the radiotherapy of advanced lung adenocarcinoma.
袁智勇、王平、曾宪亮、孟茂斌、王欢欢
肿瘤学基础医学临床医学
放射丙酮酸激酶M2型自噬人肺腺癌
IrradiationPyruvate kinase M2Autophagylung adenocarcinom
袁智勇,王平,曾宪亮,孟茂斌,王欢欢.沉默PKM2对人肺腺癌细胞的放射增敏 研究[EB/OL].(2016-05-18)[2025-08-18].http://www.paper.edu.cn/releasepaper/content/201605-623.点此复制
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