He Chongxin, Xia Jie, Gao Yinghua, Chen Zhilong, Wan Xinjian
Shanghai Key Laboratory of Pancreatic Diseases and Department of Gastroenterology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine Shanghai, China.
Department of Gastroenterology, The Second Hospital of Changzhou Affiliated to Nanjing Medical University Changzhou, China.
Am J Transl Res. 2020 Sep 15;12(9):5080-5094. eCollection 2020.
Photodynamic therapy (PDT) is a promising strategy for multiple cancers. Chlorin e6 and its derivative 13-[2'-(2-pyridyl)ethylamine] Chlorin e6 (Chlorin A) are effective photosensitizers, although their cytotoxic mechanisms have not yet been fully characterized.
Cell viability and apoptosis were evaluated by CCK8 assay, TUNEL assay, and Annexin V/PI staining. The expression levels of different proteins were analyzed by Western blot analysis and immunofluorescence. The crosstalk between autophagy, endoplasmic reticulum stress (ERS), and mitochondrial dysfunction was investigated using reactive oxygen species (ROS) scavenger N-acetyl cysteine (NAC), PERK inhibitor GSK2606414, autophagy inhibitor 3-MA, and mitochondrial stabilizer elamipretide. Furthermore, the extent of ROS production, lysosomal damage, autophagy flux, and mitochondrial membrane potential (MMP) were tracked using established probes. An xenograft model of cholangiocarcinoma (CCA) was established in BALB/c-nude mice by inoculation with EGI-1 cells, and Chlorin A was administered topically or intravenously, followed by light irradiation.
Chlorin A-PDT decreased the viability of CCA cells and induced apoptosis. Intriguingly, Chlorin A-PDT promoted autophagy via activation of ROS-induced ERS-related PERK/p-eif2α/CHOP axis, and blocked the ensuing autophagy flux by lysosomal damage. The PERK inhibitor GSK2606414 and NAC alleviated apoptosis and autophagy induced by Chlorin A-PDT. Furthermore, mitochondrial dysfunction aggravated ERS, and stabilizing the mitochondria reduced both apoptosis and autophagy. Finally, Chlorin A-PDT significantly reduced tumor growth .
Chlorin A-PDT induced apoptosis in CCA cells by initiating autophagy and impaired the autophagy flux via ROS-mediated ERS and lysosomal damage.
光动力疗法（PDT）是一种针对多种癌症的有前景的治疗策略。二氢卟吩e6及其衍生物13-[2'-(2-吡啶基)乙胺]二氢卟吩e6（二氢卟吩A）是有效的光敏剂，尽管它们的细胞毒性机制尚未完全明确。
通过CCK8法、TUNEL法和Annexin V/PI染色评估细胞活力和凋亡情况。采用蛋白质免疫印迹分析和免疫荧光分析不同蛋白质的表达水平。使用活性氧（ROS）清除剂N-乙酰半胱氨酸（NAC）、PERK抑制剂GSK2606414、自噬抑制剂3-MA和线粒体稳定剂依拉米肽研究自噬、内质网应激（ERS）和线粒体功能障碍之间的相互作用。此外，使用已建立的探针追踪ROS产生的程度、溶酶体损伤、自噬通量和线粒体膜电位（MMP）。通过接种EGI-1细胞在BALB/c裸鼠中建立胆管癌（CCA）异种移植模型，局部或静脉给予二氢卟吩A，随后进行光照。
二氢卟吩A-PDT降低了CCA细胞的活力并诱导凋亡。有趣的是，二氢卟吩A-PDT通过激活ROS诱导的与ERS相关的PERK/p-eif2α/CHOP轴促进自噬，并通过溶酶体损伤阻断随后的自噬通量。PERK抑制剂GSK2606414和NAC减轻了二氢卟吩A-PDT诱导的凋亡和自噬。此外，线粒体功能障碍加重了ERS，而稳定线粒体则减少了凋亡和自噬。最后，二氢卟吩A-PDT显著降低了肿瘤生长。
二氢卟吩A-PDT通过启动自噬诱导CCA细胞凋亡，并通过ROS介导的ERS和溶酶体损伤损害自噬通量。
