检索历史 清除

目的:探讨5-氨基酮戊酸（5-ALA）介导的光动力疗法对宫颈癌HeLa细胞凋亡的影响。方法:分别使用紫外分光光度计和荧光光谱仪测定原卟啉Ⅸ的紫外吸收光谱和荧光光谱，采用CCK-8试剂盒检测不同质量浓度（0、10、20、40、60、80、100、120、140 μg/ml）的5-ALA对HeLa细胞的存活率的影响。使用Hoechst 33342和2,7-二氯荧光素二乙酸酯（DCFH-DA）染色法借助激光共聚焦扫描显微镜检测空白对照组和激光＋150、200 μg/ml 5-ALA组原卟啉Ⅸ和活性氧的产生。借助膜联蛋白V（Annexin V）-异硫氰酸荧光素（FITC）/碘化丙啶（PI）双染色法和活死细胞染色法检测空白对照组、激光组、5-ALA组和激光＋5-ALA组细胞的凋亡情况。结果:紫外吸收光谱显示原卟啉Ⅸ在406 nm处有吸收峰，荧光光谱显示其在635 nm处有明显的特征峰。CCK-8实验结果表明，随着5-ALA质量浓度的升高，细胞存活率呈缓慢下降趋势。激光共聚焦扫描显微镜扫描结果显示，5-ALA可在细胞内转化为原卟啉Ⅸ并发出红色荧光，激光＋5-ALA组处理后的HeLa细胞由DCFH-DA荧光探针标记后可检测到活性氧的绿色荧光。流式细胞仪检测结果表明空白对照组、激光组、5-ALA组和激光＋5-ALA组的细胞凋亡率分别为12.55%、12.41%、13.51%、28.31%，且活死细胞染色结果显示激光＋5-ALA组出现了大量细胞凋亡现象。结论:5-ALA介导的光动力疗法能产生活性氧引起HeLa细胞的凋亡。

Objective:To investigate the effects of 5-aminolevulinic acid (5-ALA)-mediated photodynamic therapy on apoptosis in cervical cancer HeLa cells.Methods:The ultraviolet absorption spectrum and fluorescence spectrum of protoporphyrin Ⅸ were measured using a UV spectrophotometer and a fluorescence spectrometer, respectively. The cell viability of HeLa cells treated with different concentrations (0, 10, 20, 40, 60, 80, 100, 120 and 140 μg/ml) of 5-ALA was assessed using the CCK-8 assay. Hoechst 33342 and dichlorofluorescein diacetate (DCFH-DA) staining methods were utilized, and the production of protoporphyrin Ⅸ and reactive oxygen species in the control group and laser + 150, 200 μg/ml 5-ALA groups were observed using a laser scanning confocal microscope. Apoptosis in the control group, laser group, 5-ALA group, and laser + 5-ALA group was detected using Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) double staining and live-dead cell staining methods.Results:The UV absorption spectrum showed an absorption peak of protoporphyrin Ⅸ at 406 nm, and the fluorescence spectrum revealed a distinct characteristic peak at 635 nm. The CCK-8 assay result indicated a gradual decrease in cell viability with increasing concentrations of 5-ALA. Laser scanning confocal microscopy results demonstrated that 5-ALA could be converted into protoporphyrin Ⅸ within cells, emitting red fluorescence. In the laser + 5-ALA group, the green fluorescence of reactive oxygen species from HeLa cells labeled with DCFH-DA fluorescent probes were detected. Flow cytometry results showed that the apoptosis rates in the control, laser, 5-ALA, and laser + 5-ALA groups were 12.55%, 12.41%, 13.51%, and 28.31%, respectively. Live-dead cell staining indicated a significant occurrence of apoptosis in laser + 5-ALA group.Conclusions:5-ALA mediated photodynamic therapy can induce apoptosis in HeLa cells through the generation of reactive oxygen species.