二硫化二砷诱导哮喘豚鼠肺泡灌洗液中嗜酸细胞凋亡的研究

Study of As_2S_2 inducing apoptosis of eosinophils from bronchoalveolar lavage fluid in asthmatic guinea pigs

目的观察二硫化二砷(As2S2)诱发哮喘豚鼠肺泡灌洗液中嗜酸细胞凋亡的作用。方法采用卵白蛋白制备哮喘豚鼠模型并激发哮喘,提取肺泡灌洗液并分离其中的嗜酸细胞。将嗜酸细胞悬浮生长于含10%小牛血清的RPMI 1640培养液中,嗜酸细胞初始浓度均为2×109L-1。A组As2S2以1.406 3 mg/L终浓度分别加入细胞培养液中培养3,6,12,24,48 h。B组将As2S2终浓度调整为0.04,0.09,0.18,0.35,0.70,1.41,2.81 mg/L,培养6h。2组均以不加As2S2为对照。Wright’s染色、透射电子显微镜观察嗜酸细胞的形态学变化。流式细胞仪检测观察嗜酸细胞凋亡率。结果嗜酸细胞与As2S2作用6 h后,Wright’s染色、透射电子显微镜均观察到嗜酸细胞出现凋亡的形态学改变。将嗜酸细胞与终浓度为1.406 3 mg/L As2S2在培养基中分别培养3,6,12,24,48 h后,流式细胞检测可见嗜酸细胞凋亡率随着As2S2作用时间的延长而先升高,于24 h达最高,凋亡率为27.19%,而后下降。各不同时间组嗜酸细胞凋亡率与对照组比较均有显著性差异。将嗜酸细胞与不同终浓度As2S2在培养基中分别培养6 h后,流式细胞检测可见嗜酸细胞凋亡率随着As2S2浓度的增加而升高,各不同浓度组嗜酸细胞凋亡率与对照组比较亦均有显著性差异。结论As2S2可促进哮喘豚鼠肺泡灌洗液中嗜酸细胞凋亡,随着As2S2作用时间的延长和浓度增加这种作用更明显。

Objective It is to observe the inducing function of As2S2 on the apoptosis of eosinophils from bronchoalveolar lavage fluid in asthmatic guinea pigs. Methods The asthmatic guinea pig models were prepared with ovalbumin and the eosinophils were separated from bronchoalveolar lavage fluid. Eosinophils were cultured in RPMI 1640 culture .solution containing 10% calf serum. The incipient concentration of eosinophils was 2 × 10^9 L^-1 As2S2 with the final concentration of 1.406 3 mg/L was added in the cell culture fluid for 3, 6, 12, 24 and 48 hours in group A. Final concentration of As2S2 in group B was adjusted to 0.04, 0.09, 0.18, 0.35, 0.70, 1.41, 2.81 mg/L for 6 hours. Those without As2S2 were as control for the two groups. The morphology of apoptosis of eosinophils was observed by Wright＇ s staining and transmission electron microscope. The rate of apoptosis of eosinophils was assayed by the flow cytometry. Results Eosinophils apoptotic morphological changes were observed by Wright＇ s staining and transmission electron microscope 6 hours after interaction between eosinophils and As2S2. After 1. 406 3 mg/L As2S2 culturing for 3, 6, 12, 24 and 48 hours, the apoptotic rate of eosinophils increased with the prolongation of As2S2 intervention, and reached the peak with the apoptotic rate of 27.19 %, and then decreased. There were significant differences in apoptotic rate between different time groups and control group. After 6 hours treatment of As2S2 of different final concentration, the apoptotic rate of the eosinophils increased with increasing As2S2. concentration. There were significant differences in apoptotic rate between different concentration groups and control group. Conclusion As2S2 can promote the apoptosis of eosinophils from bronchoalveolar lavage fluid in asthmatic guinea pig. This effect is more obvious with the prolongation of As22 intervention and concentration increase.