缺氧诱导因子1α反义寡核苷酸对喉鳞状细胞癌的放疗增效作用

A study of enhancement of radiosensitivity of the allograft model of human laryngeal squamous carcinoma in nude mice by hypoxia inducible factor-1α(HIF-1α)anti-sense oligodeoxynucleotide

目的探讨缺氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)反义寡核苷酸结合放射对人喉鳞状细胞癌(简称喉癌)裸鼠移植瘤生长抑制敏感性的影响。方法对25只裸鼠建立喉癌裸鼠模型,分为5组:A组(正义放疗)、B组(反义放疗)、C组(空脂质体放疗)、D组(放疗)和E组(空白对照),治疗期间每3 d测量1次鼠重、瘤体最大长径及最大垂直径,治疗21 d后将实验鼠脱颈处死,瘤体称重后应用流式细胞仪、免疫组化观察抑瘤、凋亡情况及相关蛋白表达。结果反义放疗组瘤重(1.037±0.106)g明显低于其余4组(P<0.05),抑瘤率(50.21±8.10)%和凋亡率(34.52±4.159)%明显高于其余4组(P<0.05)。免疫组化PV法染色显示:HIF-1α正义放疗组胞浆有大量棕色颗粒沉积着色,而反义放疗组表达较少;正义放疗组p53表达于胞核呈棕色颗粒浓染,而反义放疗组表达较少;VEGF正义对照组胞浆有大量棕色颗粒沉积着色,而反义放疗组表达较少。流式细胞仪检测结果显示:反义放疗组的HIF-1α、p53、VEGF蛋白表达的荧光指数(fluorescen-ceindex,FI)分别为0.98±0.28、2.13±0.03、1.09±0.36明显低于其余4组(P<0.05)。结论 HIF-1α反义寡核苷酸结合放疗可以抑制喉癌异体移植瘤的生长,增加肿瘤细胞凋亡率,其机制之一可能是HIF-1α的下调降低了喉癌组织中p53、VEGF的蛋白表达,因此HIF-1α反义寡核苷酸增加喉癌对放疗的敏感性。

OBJECTIVE To investigate the inhibitory effect of hypoxia inducible factor-1α（HIF-1α）anti-sense oligodeoxynucleotide combined with radiation on human laryngeal squamous carcinoma in nude mice.METHODS Nude mice model bearing laryngeal carcinoma was established.Twenty-five nude mice were divided into five groups： group A（HIF-1α PS-ODN transfection for control）,group B（HIF-1α AS-ODN transfection for therapy）,group C（LP2000 control）,group D（radiotherapy）,and group E（blank control）.The weight and volume of tumors were determined every 3 days.The nude mice were killed 24h after the last injection.Flow cytometry（FCM）was used to detect cell apoptosis.The expression of HIF-1α,p53,VEGF was detected by immunocytochemical method of PV and？ow cytometry.RESULTS Tumor weight of group B was the smallest in all group.Tumor inhibition rate（50.21±8.10）%and apoptosis rate（34.522±4.159）%of group B were significantly higher than that in other 4 groups（P〈0.05）.The expression of HIF-1αprotein and VEGF protein in cytoplasm of laryngeal squamous cells,and P53 protein in nuclear of laryngeal squamous cells in group B were significantly lower than that in other 4 group.The results of flow cytometry（FCM）showed that the expression of HIF-1α,p53,VEGF protein in Hep-2 cells were restrained after transfected by HIF-1αAS-ODN,and their expression in group B was significantly lower than that in other 4 control groups（P〈0.05）.CONCLUSIONHIF-1αoligodeoxynucleotide combined with radiotherapy inhibited the tumor growth of allograft model of human CHIN ARCH OTOLARYNGOL HEAD NECK SURG/November 2010,Vol.17,No.11 587laryngeal squamous carcinoma,and increased the apoptosis rate of tumor cell.One of the mechanisms might be the decreased the expression of p53 and VEGF via the down-regulation of HIF-1αexpression.So HIF-1αantisense oligodeoxynucleotide increase the radiosensitivity of allograft model of human laryngeal squamous carcinoma.