应用胚胎干细胞实验模型评价塑化剂邻苯二甲酸二丁酯的胚胎毒性

Embryonic stem cell test for the study of the embryotoxicity of dibutyl phthalate

目的应用胚胎干细胞实验模型初步评价塑化剂邻苯二甲酸二丁酯的胚胎毒性。方法体外培养小鼠胚胎干细胞E14TG2a和小鼠成纤维细胞3T3,MTT法检测阳性对照5-氟尿嘧啶、阴性对照青霉素G和受试物邻苯二甲酸二丁酯对E14TG2a细胞和3T3细胞的细胞毒性,计算3种化合物对E14TG2a细胞和3T3细胞的半数生长抑制浓度值IC503T3、IC50E14TG2a;采用悬滴-悬浮-贴壁法体外诱导胚胎干细胞向心肌细胞分化,根据浓度-反应曲线计算3种化合物对E14TG2a细胞的半数分化抑制浓度值ID50E14TG2a。利用胚胎毒性预测模型预测邻苯二甲酸二丁酯的胚胎毒性。结果邻苯二甲酸二丁酯对E14TG2a细胞和3T3细胞的增殖均有抑制作用,其IC503T3和IC50E14TG2a分别为(321.20±8.60)μg/ml和(180.60±4.84)μg/ml。邻苯二甲酸二丁酯对E14TG2a细胞的分化亦有抑制作用,其ID50E14TG2a为(34.40±3.91)μg/ml。根据EST预测模型计算得出邻苯二甲酸二丁酯为弱胚胎毒性。结论邻苯二甲酸二丁酯为弱胚胎毒性。

Objective To evaluate the embryotoxicity of dibutyl phthalate by the embryonic stem cell test（ EST）. Methods Mouse ES cells（ E14TG2a） and mouse fibroblasts（ 3T3） were cultured in vitro,and MTT test was performed to detect the cytotoxicity of positive control 5- fluorouracil,negative control penicillin G and tested compound dibutyl phthalate by using 3T3 cells and E14TG2 a cells. The concentration of the test compounds that inhibited 50% viability of ES cells（ IC50E14TG2a） and 3T3 fibroblasts（ IC503T3） was calculated. The hanging- suspension- adherent culture system was employed to induce mouse embryonic stem cells E14TG2 a into cardiomyocytes,and the concentration of test compounds that caused 50% inhibition of differentiation of E14TG2 a cells into cardiomyocytes（ ID50E14TG2a） was calculated. The embryotoxic potential of dibutyl phthalate was classified by prediction model（ PM） of the EST. Results The proliferation of E14TG2 a and 3T3 cells were inhibited by dibutyl phthalate,of which the IC503T3 and IC50E14TG2 a values were（ 321. 20 ± 8. 60） μg / ml and（ 180. 60 ± 4. 84） μg / ml. The differentiation of E14TG2 a was also inhibited by dibutyl phthalate,of which the ID50E14TG2 a was（ 34. 40 ± 3. 91） μg / ml. Dibutyl phthalate was evaluated as a weak embryotoxicity by EST. Conclusion Dibutyl phthalate exhibited a weak embryotoxicity.