Tian Si, Yong Min, Zhu Jiang, Zhang Li, Pan Li, Chen Qing, Li Kai-Ting, Kong Yu-Han, Jiang Yuan, Yu Ting-He, Yu Le-Hua, Bai Ding-Qun
Department of Rehabilitation Medicine, the First Affiliated Hospital of Chongqing Medical University, Chongqing. China.
Department of Obstetrics and Gynecology, the Second Affiliated Hospital of Chongqing Medical University, Chongqing. China.
Anticancer Agents Med Chem. 2017;17(13):1824-1836. doi: 10.2174/1871520617666170327145857.
Emerging evidence indicates that the transcription factor nuclear factor-E2-related factor 2 (Nrf2) plays an essential role in cellular defense against oxidative stress; its activation has been related to cytoprotection.
Here, we investigated the role of Nrf2 in improving the efficacy of methyl pyropheophorbide-amediated photodynamic therapy (Mppa-PDT) via the downregulation of Nrf2.
Human ovarian cancer A2780 cells and SKOV3 cells were treated with Mppa-PDT and siRNA transfection was performed to inhibit Nrf2. After treated with siRNA and Mppa-PDT, the cell viability was examined with CCK-8 assay; cell apoptosis was detected tested by flow cytometry with Annexin V-FITC/PI; the celluar reactive oxygen species (ROS) and mitochondrial membrane potential were measured with DCFHDA and JC-1 staining; expression of protein was assessed by western blot analysis.
We found that Nrf2 translocated from the cytoplasm to the nucleus in vitro and in vivo, and the expression of Nrf2 and P-Nrf2 increased through a possible mechanism regulated by mitogen-activated protein kinase (MAPK) after Mppa-PDT treatment. Furthermore, cytotoxicity and apoptosis induced by Mppa-PDT increased after Nrf2down-regulation. Nrf2 down -regulation increased reactive oxygen species (ROS) levels by attenuating antioxidants or pumping Mppa out of cells,which resulted from the inhibition of Nrf2-HO-1 or Nrf2- ABCG2 signaling. In addition, SKOV3 cells exhibited increased resistance to Mppa-PDT, and the expression levels of P-Nrf2 and ABCG2 were higher in SKOV3 cells than in A2780 cells, suggesting that Nrf2-ABCG2 signaling might be involved in the intrinsic resistanceto Mppa-PDT.
These results provided evidence that Nrf2 down-regulation can enhance the effect of Mppa-PDT.
新出现的证据表明，转录因子核因子E2相关因子2（Nrf2）在细胞抵御氧化应激中起重要作用；其激活与细胞保护有关。
在此，我们通过下调Nrf2来研究其在提高焦脱镁叶绿酸-a甲酯介导的光动力疗法（Mppa-PDT）疗效中的作用。
用人卵巢癌A2780细胞和SKOV3细胞进行Mppa-PDT处理，并进行小干扰RNA（siRNA）转染以抑制Nrf2。用siRNA和Mppa-PDT处理后，采用CCK-8法检测细胞活力；用Annexin V-FITC/PI通过流式细胞术检测细胞凋亡；用DCFHDA和JC-1染色检测细胞活性氧（ROS）和线粒体膜电位；通过蛋白质印迹分析评估蛋白质表达。
我们发现Nrf2在体外和体内从细胞质转移到细胞核，并且在Mppa-PDT处理后，Nrf2和磷酸化Nrf2（P-Nrf2）的表达通过丝裂原活化蛋白激酶（MAPK）调控的可能机制增加。此外，Nrf2下调后，Mppa-PDT诱导的细胞毒性和凋亡增加。Nrf2下调通过减弱抗氧化剂或促使Mppa排出细胞来增加活性氧（ROS）水平，这是由Nrf2-血红素加氧酶-1（HO-1）或Nrf2-三磷酸腺苷结合盒转运蛋白G2（ABCG2）信号传导的抑制导致的。此外，SKOV3细胞对Mppa-PDT的抗性增加，并且SKOV3细胞中P-Nrf2和ABCG2的表达水平高于A2780细胞，表明Nrf2-ABCG2信号传导可能参与对Mppa-PDT的内在抗性。
这些结果证明Nrf2下调可增强Mppa-PDT的效果。
