高温对人肺腺癌耐药细胞株A549／DDP放射增敏作用研究

The experimental study of radiosensitizing effect of hyperthermia on multidrug-reaiatant human lung adenocarcinoma cell line A549/DDP

目的观察高温对人肺腺癌耐药细胞株A549／DDP的放射增敏作用并探讨其机制，为临床上联合应用热放疗治疗非小细胞肺癌提供理论依据。方法流式细胞仪检测热疗或热放疗作用后A549／DDP细胞周期的改变，原位末端转移酶标记术（TUNEL）法和流式细胞仪检测热放疗对A549／DDP细胞凋亡的作用；应用蛋白芯片结合表面增强激光解吸电离飞行时间质谱（SEL-DI—TOF—MS）技术分析各组A549／DDP细胞蛋白质图谱的差异。结果流式细胞仪显示热疗后G2～M期细胞明显增多，Go～G2期细胞明显减少；TUNEL法和流式细胞仪观察到热放疗有诱导A549／DDP细胞凋亡作用；SELDI—TOF-MS技术分析热放疗组中质荷比分别为3974Da,4288Da，4680Da的3种蛋白质表达增高，质荷比为3270Da的蛋白质表达降低；热疗组质荷比分别为4288Da，4680Da的蛋白质表达增高，而质荷比为8941Da的蛋白质表达降低；放疗组只有质荷比4680Da的蛋白质表达增高。结论热疗可能通过影响细胞蛋白质表达，诱导细胞停滞于对放射线敏感的G2～M期，从而诱导细胞凋亡，增加放射敏感性。

Objective To observe the initial mechanism of radiosensitizing effect of hyperthermia on multidrug-resistant human lung adenocarcinoma cell line A549/DDP, it will provide theory evidence in thermoradiotherapy on non-small-cell lung cancer for clinic. Methods Flow cytometry assayed cell cycle variation after hyperthermia and thermoradiotherapy. TUNEL and flow cytometry assayed the effect of thermoradiotherapy on the A549/DDP apoptosis phenomenon. The difference of protein spectrum of a graph of each group A549/DDP was analyzed with Surface- enhanced laser desorption ionization mass spectrometry （SELDI-TOF-MS）. Results Flow cytometry showed hyperthermia had significant effected on cell cycle with increase of G2-M phase population and reduction of G0-G1 phase population （P〈0.01）.TUNEL assay and flow cytometry observed thermoradiatherapy could induce A549/DDP apoptosis. Three protein peaks, with mass-to-charge ratio （m/z） 3974Da,4288Da,4680Da were increased and （m/z） 3 270Da was decreased in thermoradiotherapy group compared to control group. Protein with （m/z） 4 288Da,4 680Da was increased in hyperthermia group, while （m/z） 8 941Da was decreased. Only protein （m/z） 4 680Da was increased in radiotherapy group. Conclusion Hyperthermia induced A549/DDP G2-M arrest and then induced cell apoptosis, increased radio sensitivity by affecting cell protein expression.