Lin Jih-Shyong, Wang Chia-Jung, Li Wen-Tyng
Division of Cardiology, Department of Internal Medicine, Taoyuan General Hospital, Ministry of Health and Welfare.
Department of Biomedical Engineering.
Acta Cardiol Sin. 2019 Jan;35(1):65-74. doi: 10.6515/ACS.201901_35(1).20180731A.
Vascular smooth muscle cells play a critical role in the intimal hyperplasia of restenosis. A previous study of a rat balloon injury model demonstrated that photodynamic therapy (PDT) using indocyanine green (ICG) and near-infrared (NIR) light irradiation reduced intimal hyperplasia in carotid arteries. However, the effect of ICG-PDT on smooth muscle cells remains unclear. This study aimed to evaluate the effects of PDT with ICG and NIR irradiation on the viability of vascular smooth muscle (A-10) cells.
A-10 cells were incubated with ICG at different concentrations for different time intervals. Intracellular accumulation of ICG inside the cells was observed by light microscopy, ultraviolet-visible (UV-VIS) spectrophotometry and spectrofluorometry. Cell viability and cell death after ICG-PDT were assessed by 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide assay and lactate dehydrogenase release assay. Changes in nuclear morphology and cell cycle distribution were evaluated to determine the possible cell death mechanism mediated by ICG-PDT.
ICG uptake in A-10 cells increased with the amount of ICG in the culture media. The intracellular accumulation of ICG reached a maximum at 8 h. After ICG-PDT, cell viability decreased and cell death increased in a concentration- dependent manner. The half maximal inhibitory concentration of ICG was 8.3 μM with 4 J/cm NIR irradiation. Membrane blebbing and chromatin condensation were observed, and the percentage of cells in the sub-G phase increased after ICG-PDT. Thus, apoptosis might be responsible for decreasing the viability of A-10 cells by ICG-PDT.
This study demonstrated that ICG-PDT had an inhibitory effect on smooth muscle cells, possibly via an apoptosis pathway.
血管平滑肌细胞在再狭窄的内膜增生中起关键作用。先前一项关于大鼠球囊损伤模型的研究表明，使用吲哚菁绿（ICG）和近红外（NIR）光照射的光动力疗法（PDT）可减少颈动脉的内膜增生。然而，ICG-PDT对平滑肌细胞的影响仍不清楚。本研究旨在评估ICG和NIR照射的PDT对血管平滑肌（A-10）细胞活力的影响。
将A-10细胞与不同浓度的ICG孵育不同时间间隔。通过光学显微镜、紫外可见（UV-VIS）分光光度法和荧光分光光度法观察细胞内ICG的积累情况。通过3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐测定法和乳酸脱氢酶释放测定法评估ICG-PDT后的细胞活力和细胞死亡情况。评估核形态和细胞周期分布的变化，以确定ICG-PDT介导的可能细胞死亡机制。
A-10细胞中ICG的摄取量随培养基中ICG量的增加而增加。ICG的细胞内积累在8小时达到最大值。ICG-PDT后，细胞活力下降，细胞死亡呈浓度依赖性增加。在4 J/cm的近红外照射下，ICG的半数最大抑制浓度为8.3 μM。观察到细胞膜泡化和染色质凝聚，ICG-PDT后亚G期细胞百分比增加。因此，凋亡可能是ICG-PDT降低A-10细胞活力的原因。
本研究表明，ICG-PDT对平滑肌细胞有抑制作用，可能通过凋亡途径。
