维生素K3通过ROS介导细胞质酸化诱导Hela细胞凋亡
发布时间：2018-08-12 07:40
【摘要】：目的利用维生素K3(VK3)复制氧化应激模型,探讨VK3诱导的ROS在Hela细胞溶酶体-线粒体细胞死亡途径过程中的作用及机制。方法以人宫颈癌Hela细胞为研究对象,实验分为:对照组、VK3组(30μmol/L)、NAC组(80μmol/L)和VK3与NAC联合作用组。MTT法检测细胞生存率,DCFH-DA染色观察细胞内活性氧簇(ROS)水平,吖啶橙(AO)染色观察溶酶体膜通透性变化,BCECF-AM染色流式细胞术检测细胞质p H变化,罗丹明123(Rhodamine 123)染色检测线粒体膜通透性变化。结果与对照组相比,单独应用NAC对Hela细胞无明显影响。VK3作用组Hela细胞生存率降低(P0.05),DCFH-DA染色荧光增强(P0.05),ROS水平增加;AO染色在VK3作用3 h后Hela细胞的细胞质绿色荧光增强(P0.05),红色颗粒状荧光减弱(P0.05),溶酶体通透性增加;BCECF-AM染色,VK3作用6 h后FL1/FL2比值降低(P0.05),细胞质呈酸性,酸性p H的细胞数为13.6%(P0.05);Rhodamin染色,VK3作用6 h后细胞质内红色荧光强度降低(P0.05),线粒体膜电势降低。与单独应用VK3组比较,VK3与NAC联合作用于Hela细胞时,细胞生存率增加(P0.05),ROS水平降低(P0.05),溶酶体通透性降低(P0.05),FL1/FL2比值增加(P0.05),酸性p H的细胞数为5.4%(P0.05),线粒体膜电势增加(P0.05)。结论在氧化应激诱导Hela细胞损伤过程中,VK3通过ROS依赖途径,诱导先后发生溶酶体及线粒体通透性增加,在这个过程中细胞质发生酸化。
[Abstract]:Objective to investigate the role and mechanism of VK3 induced ROS in the process of lysosomal mitochondrial cell death in Hela cells by using vitamin K3 (VK3) to replicate oxidative stress model. Methods Human cervical cancer Hela cells were divided into two groups: control group (30 渭 mol/L), control group (30 渭 mol/L) and VK3 combined with NAC group. Changes in membrane permeability of lysosome were observed by (AO) staining with acridine orange. The changes of cytosolic pH were detected by flow cytometry with BCECF-AM staining, and mitochondrial membrane permeability by Rhodamine 123 (Rhodamine 123) staining. Results compared with the control group, The survival rate of Hela cells in the Hela cells treated with NAC alone decreased (P0.05) the fluorescence intensity of DCFH-DA staining increased (P0.05) the cytoplasmic green fluorescence of Hela cells increased after 3 h of VK3 treatment (P0.05), and the red granular fluorescence decreased (P0.05). Weak (P0.05), the permeability of lysosome increased, the ratio of FL1/FL2 decreased after 6 h exposure to BCECF-AM staining (P0.05), and the cytoplasm was acidic. The number of acidic pH cells was 13.6% (P0.05). The red fluorescence intensity in cytoplasm was decreased (P0.05) and the mitochondrial membrane potential was decreased after VK3 was treated with Rhodamin staining for 6 h. Compared with the VK3 group, the cell survival rate increased (P0.05), the lysosomal permeability decreased (P0.05), the ratio of FL1 / FL2 increased (P0.05), the number of acidic pH cells increased 5.4% (P0.05), and the mitochondrial membrane potential increased (P0.05). Conclusion during oxidative stress induced Hela cell injury, VK3 induces the increase of lysosome and mitochondrial permeability through ROS dependent pathway, and the cytoplasm acidification occurs during this process.
【作者单位】： 北华大学基础医学院;国家食品药品监督管理总局药品审评中心;
【基金】：吉林省教育厅资助课题〔吉教科合字2011第117号;2009第156号〕 吉林省科技厅自然科学基金资助课题〔201215103〕 吉林省卫计委资助课题〔2013Z062〕 吉林市科技局项目〔201464064〕
【分类号】：R363
[Abstract]:Objective to investigate the role and mechanism of VK3 induced ROS in the process of lysosomal mitochondrial cell death in Hela cells by using vitamin K3 (VK3) to replicate oxidative stress model. Methods Human cervical cancer Hela cells were divided into two groups: control group (30 渭 mol/L), control group (30 渭 mol/L) and VK3 combined with NAC group. Changes in membrane permeability of lysosome were observed by (AO) staining with acridine orange. The changes of cytosolic pH were detected by flow cytometry with BCECF-AM staining, and mitochondrial membrane permeability by Rhodamine 123 (Rhodamine 123) staining. Results compared with the control group, The survival rate of Hela cells in the Hela cells treated with NAC alone decreased (P0.05) the fluorescence intensity of DCFH-DA staining increased (P0.05) the cytoplasmic green fluorescence of Hela cells increased after 3 h of VK3 treatment (P0.05), and the red granular fluorescence decreased (P0.05). Weak (P0.05), the permeability of lysosome increased, the ratio of FL1/FL2 decreased after 6 h exposure to BCECF-AM staining (P0.05), and the cytoplasm was acidic. The number of acidic pH cells was 13.6% (P0.05). The red fluorescence intensity in cytoplasm was decreased (P0.05) and the mitochondrial membrane potential was decreased after VK3 was treated with Rhodamin staining for 6 h. Compared with the VK3 group, the cell survival rate increased (P0.05), the lysosomal permeability decreased (P0.05), the ratio of FL1 / FL2 increased (P0.05), the number of acidic pH cells increased 5.4% (P0.05), and the mitochondrial membrane potential increased (P0.05). Conclusion during oxidative stress induced Hela cell injury, VK3 induces the increase of lysosome and mitochondrial permeability through ROS dependent pathway, and the cytoplasm acidification occurs during this process.
【作者单位】： 北华大学基础医学院;国家食品药品监督管理总局药品审评中心;
【基金】：吉林省教育厅资助课题〔吉教科合字2011第117号;2009第156号〕 吉林省科技厅自然科学基金资助课题〔201215103〕 吉林省卫计委资助课题〔2013Z062〕 吉林市科技局项目〔201464064〕
【分类号】：R363
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