抗clusterin反义核酸治疗提高肺癌H460细胞放射敏感性的实验研究

Anti-sense oligonucleotide against clusterin sensitizes lung carcinoma H460 cells to irradiation

目的:研究抑制clusterin的表达对肺癌H460细胞放射敏感性的影响。方法:用脂质体转染的方法将抗clusterin反义核酸转染肺癌H460细胞,以137Cs作为放射源对肺癌细胞进行放射干预。用流式细胞仪检测抗clusterin反义核酸治疗和放射干预对肺癌细胞凋亡的影响,用MTT法和克隆形成分析方法检测抗clusterin反义核酸治疗和放射干预对肺癌细胞增殖的影响。结果:抗clusterin反义核酸特异性抑制了分泌型clusterin的表达,而对核型clusterin的表达无明显作用。单纯放射干预或单纯反义核酸转染将凋亡细胞数提高了10%,反义核酸治疗联合放射干预使肺癌H460细胞的凋亡细胞数增加了25%,和单纯放疗或单纯反义核酸转染组比较有显著性差异(P<0.01)。MTT和体外克隆计数分析显示抗clusterin反义核酸治疗联合放疗干预显著抑制了肺癌H460细胞的增殖(P<0.01)。结论:体外实验提示抗clusterin反义核酸治疗可以提高肺癌细胞对放射的敏感性。

Objective：To study whether the inhibition of clusterin sensitizes lung cancer H460 cells to irradiation. Methods： The H460 cells were transfected with antisense oligonucleotide（oligo） against clusterin mediated by lipofectin. Lung cancer H460 cells were radiated by 137Cs irradiator. The percentage of apoptotic cells was determined by flow-cytometry analysis of 7-AAD stained cells. Cell proliferation was measured by MTT and clonogenic assay. Results： Antisense oligonucleotides of clusterin caused specific attenuation of secreted clusterin, but exerted no effect on nuclear clusterin. Compared with control group, transfection with antisense oligonucleotide against clusterin or irradiation alone increased the apoptotic ratio from 5% to 10%, but irradiation combined with antisense oligonucleotide therapy significantly increased the apoptotic population to almost 25 % （P〈 0.01）. MTT and clonogenic assay showed that irradiation combined with antisense oligonucleotide therapy significantly reduced the proliferation of H460 cells in vitro compared with control group （P^0.01）. Conclusion.. Antisense oligonucleotide against clusterin sensitizes lung cancer H460 cells to radiotherapy in vitro.