香烟烟雾溶液作用下大鼠淋巴细胞的氧化应激反应

Oxidative stress produced by cigarette smoke solution in rat lymphocytes

目的 以细胞内活性氧自由基 (ROS)水平、DNA损伤、脂质过氧化物 (LPO)水平以及超氧化物歧化酶 (SOD)活力为指标 ,研究细胞在香烟烟雾溶液作用下产生的氧化应激反应。方法 以PBS作吸收液 ,用大包氏管收集香烟主流烟雾 ,将香烟烟雾溶液 (CSS)分别以 0、1× 10 - 3、2× 10 - 3、4× 10 - 3、8× 10 - 3、12× 10 - 3、16× 10 - 3支 ml的浓度作用于大鼠淋巴细胞。用 2′ ,7′ 二乙酰二氯荧光素 (DCFH DA)测定细胞内ROS水平 ,用彗星试验检测细胞DNA损伤 ,同时检测细胞内SOD活力和LPO水平。结果 2× 10 - 3支 ml以上剂量组二氯荧光素 (DCF)荧光强度、彗星尾长以及LPO水平均显著高于对照组 ,且随剂量增加而增加。 16× 10 - 3支 ml组SOD活力显著低于对照组。与对照组相比 ,1× 10 - 3支 ml组LPO水平显著降低、SOD活力则显著增高。结论 CSS达到一定剂量时使细胞内ROS水平增高 ,引起细胞DNA损伤和脂质过氧化 ,高剂量时SOD活力降低 。

Objectives To learn oxidative stress in rat lymphocytes treated with cigarette smoke solution (CSS). Methods CSS was prepared by bubbling main stream cigarette smoke into PBS, then 0,1×10 -3 ,2×10 -3 ,4×10 -3 ,8×10 -3 ,12×10 -3 ,16×10 -3 cig/ml CSS were added into 7 groups of rat lymphocytes respectively. Intracellular reactive oxygen species (ROS) were detected by using 2′,7′ dichlorofluorescein diacetate (DCFH DA). DNA damage was evaluated by comet assay. Meanwhile, intracellular lipid peroxide (LPO) level and superoxide dismutase (SOD) activity were also measured. Results Fluorescent density of dichlorofluorescein (DCF), comet tail and LPO in groups 2×10 -3 ～16×10 -3 cig/ml were all higher than the control significantly, and there were dose effect relationships. SOD activity decreased only in 16×10 -3 cig/ml CSS group. However, in group 1×10 -3 cig/ml LPO level decreased and SOD activity increased markedly. Conclusions ROS in cells treated with CSS increased in a dose dependent pattern, and so did lipid peroxidation and DNA damage. SOD activity was induced by low concentration of CSS, but at high concentration it decreased.