摘要
目的:探讨三氯乙烯(TCE)对人胚胎干细胞心肌发育分化的影响。方法:以人胚胎干细胞H9体外心肌定向分化为模型,分化培养液中添加不同剂量的TCE(分别为处理组100 ppb组,1 ppm组,10 ppm组)以及DMSO(对照组)。对诱导后形成的拟胚体以及不同发育阶段的细胞进行检测,通过MTT法测定细胞的生存能力,统计有自主节律跳动的拟胚体的数量;流式细胞仪计数分析细胞分化比例;荧光定量PCR检测心肌特异性基因表达;并应用基因芯片方法初步筛选检测三氯乙烯对心肌细胞钙离子通道相关的基因表达的影响。结果:TCE的三个浓度处理组均未显示细胞毒性,但高浓度组明显抑制向心肌细胞分化,能产生自主节律搏动的拟胚体比例显著降低,同时心肌特异性标记物c Tn T表达随剂量的增加而显著降低。在具有自主节律性的拟胚体中,钙离子通路的相关基因表达受到影响。结论:推测TCE通过降低成熟心肌的比例,以及影响心肌细胞中钙离子通道相关基因表达来共同导致心脏发育异常。
Objective： To explore the effects of trichloroethylene （TCE） on cardiac developmental differentiation in human embryonic stem cells. Methods： In this study, based on the human embryonic stem ceils in vitro cardiac differentiation assay, we investigated tile poten- tial effect of TCE exposure on the cardiac toxicity in embryo development. Human embryonic stem cells were treated with TCE at different con- centrations of 100 ppb, 1 ppm, and 10 ppm and dimethyl sulfoxide（DMSO）treated as control. The MTF assay was performed to examine the cytoplasmic toxicity of TCE exposure. The beating percentages were recorded and the expression of cardiac specific gene was evaluated by PCR or flow cytometry. Also, real time PCR was performed to verify the micro array analysis on the expression level changes of genes which were involved in the Ca2＋ signal pathways. Results： Compared with the control group, there was no significant difference in cell viability when cells were treated with TCE at the concentrations of 100 ppb, 1 ppm, and 10 ppm. However, TCE could inhibit the expression of cTnT protein in a concentration-dependant manner. And the most interestingly, TCE significantly inhibited the cardiac differentiation characterized by the de- crease beating percentages. Genes involved in Ca2＋ signaling pathway were severely disrupted by TCE. Conclusion： TCE inhibited the cardiac specific differentiation of human embryonic stem cell and at the meanwhile the genes responsible for Ca2 ＋ signaling pathway were severely dis- rupted, which could contribute the severe effects of TCE cardiotoxicity.
出处
《中国应用生理学杂志》
CAS
CSCD
2015年第3期216-219,224,共5页
Chinese Journal of Applied Physiology
基金
国家自然科学基金项目(81300143)
中国博士后基金(2013M541718)
关键词
三氯乙烯
人胚胎干细胞
心肌分化
基因表达
钙离子通道
trichloroethylene
human embryonic stem cells
cardiac differentiation
gene expression
calcium channel
