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[目的]研究新型光敏剂Ⅰ诱导Hep-2细胞的光氧化行为。[方法]利用MTT法检测光敏剂Ⅰ对Hep-2细胞的细胞毒性;采用活性氧特异性探针H2DCFDA通过激光共聚焦成像观察Hep-2细胞中活性氧的生成;通过测定超氧化物歧化酶(SOD)、谷胱甘肽(GSH)和丙二醛(MDA)水平及乳酸脱氢酶(LDH)渗漏检测观察Hep-2细胞的氧化应激反应。[结果]无光照时,光敏剂Ⅰ对Hep-2细胞的毒性为零,但光照后可明显抑制该细胞的生长,且其光毒性随光照剂量的增加而加强(r=-0.962,P=0.001)。光动力治疗后,细胞内DCFDA的荧光强度逐渐增强,在4h时达到高峰,随后又逐渐降低;细胞内SOD和GSH水平逐渐降低,3h后分别降低42.5%(P<0.01)和35.0%(P<0.01),而MDA含量却随时间延长逐渐增加,3h后增加54%(P<0.01)。LDH的渗出与光照剂量呈正相关(r=0.966,P=0.007)。[结论]新型光敏剂Ⅰ可有效光诱导Hep-2细胞死亡,而细胞内氧化应激反应可能是其光诱导Hep-2细胞死亡的重要作用机制。
[Objective] The research aimed to study the photooxidation of Hep-2 cells induced by a novel photosensitizer Ⅰ. [Method] The cytotoxicity of photosensitizer Ⅰ to Hep-2 cells was detected by MTT method. The generation of reactive oxygen species in Hep-2 cells was observed by laser scanning confocal microscope using reactive oxygen species specific probe H2DCFDA. The activity of superoxide dismutase (SOD), glutathione (GSH) and malondialdehyde (MDA) levels and lactate dehydrogenase (LDH) leakage were measured to observe the oxidative stress response in Hep-2 cells. [Result] When no light was applied, the photosensitizer Ⅰ had zero toxicity to Hep-2 cells, but the growth of the cells was obviously inhibited after light irradiation. The phototoxicity of photosensitizer Ⅰ was enhanced with the increase of light dose (r = -0.962, P = 0.001). After photodynamic therapy, the fluorescence intensity of intracellular DCFDA gradually increased and peaked at 4h, and then gradually decreased. The levels of SOD and GSH in cells decreased gradually after 42h (P <0.01) and 35.0% (P <0.01), while the content of MDA gradually increased with time and increased 54% after 3h (P <0.01). LDH exudation and light dose was positively correlated (r = 0.966, P = 0.007). [Conclusion] The new photosensitizer Ⅰ can effectively induce the death of Hep-2 cells, and the intracellular oxidative stress may be an important mechanism of light-induced Hep-2 cell death.