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目的建立胚胎干细胞实验模型,应用胚胎干细胞实验模型初步评价三聚氰胺的胚胎毒性。方法体外培养小鼠成纤维细胞3T3和小鼠胚胎干细胞E14TG2a,MTT法检测阳性对照5-氟尿嘧啶、阴性对照青霉素G和受试物三聚氰胺对3T3细胞和E14TG2a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IC50 3T3、IC50 E14TG2a;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ID50 E14TG2a。利用胚胎毒性预测模型预测三聚氰胺的胚胎毒性。结果三聚氰胺对3T3细胞和E14TG2a细胞的增殖均有抑制作用,其IC50 3T3和IC50 E14TG2a分别为485.6±25.05和304.1±20.50μg/ml。三聚氰胺对E14TG2a细胞的分化亦有抑制作用,其ID50 E14TG2a为152.6±12.62μg/ml。根据EST预测模型计算得出三聚氰胺为弱胚胎毒性。结论三聚氰胺为弱胚胎毒性。
Objective To establish an experimental model of embryonic stem cells and to evaluate the embryotoxicity of melamine using the experimental model of embryonic stem cells. Methods Mouse fibroblasts 3T3 and mouse embryonic stem cells E14TG2a were cultured in vitro. The cytotoxicity of 5-Fluorouracil, negative control penicillin G and test substance melamine on 3T3 cells and E14TG2a cells were detected by MTT assay. The cytotoxicity of 3 compounds against E14TG2a IC50 3T3 and IC50 E14TG2a were determined by MTT assay. The embryonic stem cells were induced to differentiate into cardiomyocytes in vitro by suspension-suspension-adherence method. The half-cell differentiation inhibition of the three compounds against E14TG2a cells was calculated according to the concentration-response curve Concentration value ID50 E14TG2a. Prediction of embryotoxicity of melamine using embryo toxicity prediction model. Results Melamine inhibited the proliferation of 3T3 cells and E14TG2a cells with IC50 3T3 and IC50 E14TG2a of 485.6 ± 25.05 and 304.1 ± 20.50μg / ml, respectively. Melamine also inhibited the differentiation of E14TG2a cells with ID50 E14TG2a of 152.6 ± 12.62μg / ml. According to the EST prediction model, melamine was calculated as weak embryo toxicity. Conclusion Melamine is weak embryo toxicity.