5-氨基酮戊酸介导的光动力疗法对宫颈癌HeLa细胞凋亡的影响  

Effects of 5-aminolevulinic acid-mediated photodynamic therapy on apoptosis in cervical cancer HeLa cells

作　　者：熊加宝 朱丽君 闫婷 古丽妮尕尔·阿里木 努尔尼沙·阿力甫 Xiong Jiabao;Zhu Lijun;Yan Ting;Gulinigaer Alimu;Nuernisha Alifu(The Second Affiliated Hospital of Xinjiang Medical University,Urumqi 830028,China;College of Public Health,Xinjiang Medical University,Urumqi 830011,China;College of Medical Engineering and Technology,Xinjiang Medical University,Urumqi 830011,China)

机构地区：[1]新疆医科大学第二附属医院,乌鲁木齐830028 [2]新疆医科大学公共卫生学院,乌鲁木齐830011 [3]新疆医科大学医学工程技术学院,乌鲁木齐830011

出　　处：《国际生物医学工程杂志》2024年第3期213-218,共6页International Journal of Biomedical Engineering

基　　金：自治区区域协同创新专项(科技援疆计划)(2022E02130)。

摘　　要：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。ObjectiveTo investigate the effects of 5-aminolevulinic acid(5-ALA)-mediated photodynamic therapy on apoptosis in cervical cancer HeLa cells.MethodsThe ultraviolet absorption spectrum and fluorescence spectrum of protoporphyrinⅨwere 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 140μg/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 protoporphyrinⅨand reactive oxygen species in the control group and laser+150,200μg/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.ResultsThe UV absorption spectrum showed an absorption peak of protoporphyrinⅨat 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 protoporphyrinⅨwithin 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.Conclusions5-ALA mediated photodynamic therapy can induce apoptosis in HeLa cells through the generation of reactive oxygen species.

关 键 词：光动力疗法 宫颈癌 5-氨基酮戊酸 活性氧 凋亡 

分 类 号：R737.33[医药卫生—肿瘤]