Xue Qin, Wang Xiaobing, Wang Pan, Zhang Kun, Liu Quanhong
Key Laboratory of Medicinal Resources and Natural Pharmaceutical Chemistry, Ministry of Education, National Engineering Laboratory for Resource Developing of Endangered Chinese Crude Drugs in Northwest of China, College of Life Sciences, Shaanxi Normal University, Xi'an 710062, Shaanxi, China; Department of Urology, Xijing Hospital, Fourth Military, Medical University, Xi'an, China.
Key Laboratory of Medicinal Resources and Natural Pharmaceutical Chemistry, Ministry of Education, National Engineering Laboratory for Resource Developing of Endangered Chinese Crude Drugs in Northwest of China, College of Life Sciences, Shaanxi Normal University, Xi'an 710062, Shaanxi, China.
Photodiagnosis Photodyn Ther. 2015 Mar;12(1):84-91. doi: 10.1016/j.pdpdt.2014.12.001. Epub 2014 Dec 17.
Photodynamic therapy (PDT) has been undergoing clinical evaluation for the treatment of colorectal cancer. But the molecular mechanism of photodynamic injury in human colorectal cancer cells still remains unclear.
Chlorin e6 (Ce6) was used to photosensitize SW620 cells. The inhibitory effect of PDT was evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltertrazolium bromide tetrazolium) assay and colony forming assay. Apoptosis was determined by nuclear DAPI (4'-6-diamidino-2-phenylindole) staining and Annexin V-PE/7-AAD assay. Monodansylcadaverine (MDC) staining was used to evaluate the abundance of autophagic vacuoles in PDT treated cells. The apoptosis and autophagy associated proteins were analyzed by western blotting. Moreover, we applied siRNA p38MAPK and p38MAPK inhibitor SB203580 to dissect its effect on cellular response to PDT in SW620 cells.
Ce6 mediated PDT (Ce6-PDT) induced apparent autophagy and apoptosis with dependent on ROS (reactive oxygen species) generation. When p38MAPK was inhibited by siRNA or inhibitor SB203580, a marked enhancement of apoptosis and autophagy in SW620 cells was detected after PDT. Moreover, autophagy inhibitor 3-methyladenine/Bafilomycin A1 greatly aggravated PDT induced photodamage in SW620 cells.
Ce6-PDT induced ROS production to activate p38MAPK probably to prevent SW620 cells from photodamage. Inhibition of p38MAPK activation accelerated cell apoptosis, meanwhile enhanced autophagy may act as a cytoprotective process in SW620 cells.
光动力疗法（PDT）一直在接受用于治疗结直肠癌的临床评估。但人类结肠癌细胞光动力损伤的分子机制仍不清楚。
用二氢卟吩e6（Ce6）对SW620细胞进行光敏化处理。通过MTT（3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐）法和集落形成试验评估PDT的抑制作用。通过细胞核DAPI（4′-6-二脒基-2-苯基吲哚）染色和膜联蛋白V-PE/7-AAD试验确定细胞凋亡情况。用单丹磺酰尸胺（MDC）染色评估PDT处理细胞中自噬空泡的丰度。通过蛋白质免疫印迹法分析凋亡和自噬相关蛋白。此外，我们应用小干扰RNA p38丝裂原活化蛋白激酶（siRNA p38MAPK）和p38MAPK抑制剂SB203580来剖析其对SW620细胞中PDT细胞反应的影响。
Ce6介导的PDT（Ce6-PDT）诱导明显的自噬和凋亡，且依赖于活性氧（ROS）的产生。当p38MAPK被siRNA或抑制剂SB203580抑制时，PDT后SW620细胞中的凋亡和自噬显著增强。此外，自噬抑制剂3-甲基腺嘌呤/巴弗洛霉素A1极大地加重了PDT对SW620细胞的光损伤。
Ce6-PDT诱导ROS产生以激活p38MAPK，可能是为了防止SW620细胞受到光损伤。抑制p38MAPK激活加速细胞凋亡，同时增强的自噬可能在SW620细胞中起到细胞保护作用。
