Zhang Xiaowei, Wu Rudolf S S, Fu Wenyu, Xu Lihong, Lam Paul K S
Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, Kowloon, Hong Kong, People's Republic of China.
Chemosphere. 2004 Jun;55(10):1303-8. doi: 10.1016/j.chemosphere.2003.12.004.
Previous studies have shown that ultraviolet (UV) A light and the polycyclic aromatic hydrocarbon benzo[a]pyrene (BaP) can synergistically enhance the formation of 8-hydroxy-2'deoxyguanosine (8-OHdG) in living cells. It has been postulated that the underlying mechanism is production of reactive oxygen species (ROS) via photosensitization, but direct evidence supporting this hypothesis has been lacking. This study examined intracellular ROS production in living cells co-exposed to UV-A and BaP as well as the relationship between intracellular production of ROS and formation of 8-OHdG. KB cells were exposed to BaP for 24 h, followed by exposure to UV-A (365 nm) or UV-B (312 nm). The levels of intracellular ROS were directly measured by use of the fluorescent probe dihydrorhodamine 123 (DHR-123) in flow cytometry. Levels of 8-OHdG were measured by high performance liquid chromatography coupled with electrochemical detection (HPLC-ECD). The results demonstrated that UV-B itself induced a much greater level of intracellular ROS than did UV-A alone under the same dose of energy (0.10 mW/cm(2), 20 min). The presence of BaP (13.3 microM) substantially increased ROS production in UV-A-treated cells (2.9-fold), but only slightly enhanced ROS production in UV-B-treated cells (1.3-fold). These results show that BaP acts mainly as a photosensitizer of UV-A, but not UV-B. Furthermore, greater intracellular ROS production was proportional to both BaP concentration and UV-A dosage. There was a linear relationship between ROS production and 8-OHdG formation in cells co-exposed to BaP and UV-A. Results of this study suggest that UV-A and BaP act synergistically to enhance ROS production and formation of 8-OHdG, resulting in increased DNA damage.
先前的研究表明，紫外线（UV）A光和多环芳烃苯并[a]芘（BaP）可协同增强活细胞中8-羟基-2'-脱氧鸟苷（8-OHdG）的形成。据推测，其潜在机制是通过光敏化产生活性氧（ROS），但一直缺乏支持这一假设的直接证据。本研究检测了同时暴露于UV-A和BaP的活细胞内ROS的产生情况，以及细胞内ROS产生与8-OHdG形成之间的关系。将KB细胞暴露于BaP 24小时，然后再暴露于UV-A（365nm）或UV-B（312nm）。通过流式细胞术中使用荧光探针二氢罗丹明123（DHR-123）直接测量细胞内ROS水平。通过高效液相色谱-电化学检测法（HPLC-ECD）测量8-OHdG水平。结果表明，在相同能量剂量（0.10 mW/cm²，20分钟）下，UV-B本身诱导产生的细胞内ROS水平比单独的UV-A高得多。BaP（13.3μM）的存在显著增加了UV-A处理细胞中的ROS产生（2.9倍），但仅略微增强了UV-B处理细胞中的ROS产生（1.3倍）。这些结果表明，BaP主要作为UV-A的光敏剂，而不是UV-B的光敏剂。此外，细胞内ROS产生增加与BaP浓度和UV-A剂量均成正比。在同时暴露于BaP和UV-A的细胞中，ROS产生与8-OHdG形成之间存在线性关系。本研究结果表明，UV-A和BaP协同作用可增强ROS产生和8-OHdG形成，从而导致DNA损伤增加。
