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目的探讨1,2-二氯乙烷(1,2-DCE)及其体内代谢产物对人星形胶质细胞(HAs)的毒性。方法取对数生长期的HAs建立离体细胞培养实验模型,分别采用不同剂量的1,2-DCE(5.00、10.00、25.00、50.00、100.00mmol/L)、2-氯乙醇(5.00、25.00、50.00、100.00、200.00 mmol/L)、2-氯乙醛(1.00、5.00、10.00、20.00、50.00mmol/L)和氯乙酸(0.01、0.05、0.10、0.50、1.00 mmol/L)对HAs进行染毒,培养24 h后,采用荧光倒置相差显微镜观察HAs形态,采用CCK-8比色法检测HAs的存活率和抑制率,估算24 h半抑制浓度(24 h-IC50),采用磷脂结合蛋白Ⅴ-异硫氰酸荧光素/碘化丙啶双标记流式细胞技术检测细胞的凋亡情况。结果染毒24 h后,1,2-DCE、2-氯乙醇、2-氯乙醛和氯乙酸均可使HAs发生不同程度的改变,表现为胞体变小、伪足变尖、胞内颗粒增多,细胞透明度下降,圆形悬浮细胞增多。随着1,2-DCE、2-氯乙醇、2-氯乙醛和氯乙酸染毒剂量的增加,HAs的细胞存活率均下降(P<0.01),细胞抑制率均增加(P<0.01),均呈剂量-效应关系;上述4种受试物对HAs的24 h-IC50分别为56.25、235.00、26.43和1.38 mmol/L。1,2-DCE、2-氯乙醇和氯乙酸均可诱导HAs凋亡,HAs的凋亡率与上述3种受试物的染毒剂量均呈正相关(P<0.01)。结论 1,2-DCE及其代谢产物2-氯乙醇、2-氯乙醛和氯乙酸均可对HAs造成毒性损伤,并可诱导HAs发生凋亡,以氯乙酸毒性作用最强。
Objective To investigate the toxicity of 1,2-dichloroethane (1,2-DCE) and its metabolites in vivo to human astrocytes (HAs). Methods HAS in logarithmic growth phase was used to establish an in vitro cell culture model. The effects of 1,2-DCE (5.00,10.00,25.00,50.00,100.00mmol / L), 2-chloroethanol (5.00,25.00, 50.00, 100.00, 200.00 mmol / L), 2-chloroacetaldehyde (1.00, 5.00, 10.00, 20.00 and 50.00 mmol / L) and chloroacetic acid (0.01,0.05,0.10,0.50,1.00 mmol / L) 24 hours after culture, the morphology of HAs was observed by fluorescence inverted phase contrast microscope. The survival rate and inhibition rate of HAs were measured by CCK-8 colorimetric assay, and the 24 h half-inhibitory concentration (24 h-IC50) was estimated by phospholipid binding protein V Dual - labeled Flow Cytometry Using Fluorescein Isothiocyanate / Propidium Iodide for Detection of Cell Apoptosis. Results After 24 h of exposure, 1,2-DCE, 2-chloroethanol, 2-chloroacetaldehyde and chloroacetic acid all changed HAs in varying degrees. The results showed that the cell bodies became smaller and the pseudopods became sharp. Increased, cell transparency decreased, rounded cells increased. With the increase of 1,2-DCE, 2-chloroethanol, 2-chloroacetaldehyde and chloroacetic acid, the cell viability of HAs decreased (P <0.01) and the cell inhibitory rate increased , All showed a dose-effect relationship. The 24 h-IC50 of HAs in the four tested samples were 56.25, 235.00, 26.43 and 1.38 mmol / L, respectively. 1,2-DCE, 2-chloroethanol and chloroacetic acid could induce the apoptosis of HAs, and the apoptosis rate of HAs was positively correlated with the doses of the above three test substances (P <0.01). CONCLUSION 1,2-DCE and its metabolites 2-chloroethanol, 2-chloroacetaldehyde and chloroacetic acid can cause toxic damage to HAs and induce apoptosis of HAs. Chloroacetic acid has the strongest toxicity.