氯乙烯诱导小鼠胚胎神经管发育畸形的研究

Vinyl chlorid induces embryo neural tube defect in mice

目的观察孕早期不同剂量氯乙烯(VC)染毒后胎鼠神经系统发育以及神经上皮细胞增殖和凋亡的变化情况,明确VC对神经系统发育的致畸作用。方法将孕鼠21只按完全随机设计分组法分为实验组和对照组,实验组分别用不同剂量的VC对孕6.5 d的小鼠进行腹腔染毒,对照组注射花生油,均于E12.5取胚胎。观察胎鼠的表观发育情况,计算胚胎畸形率以及神经系统异常的发生率。运用PCNA免疫组织化学技术和TUNEL染色技术检测并观察神经管上皮细胞的增殖及其凋亡的变化。结果①中剂量VC组胎鼠活胎率下降,活胎中畸胎发生率高达51%,其中神经管畸形发生率占40%,与正常对照组相比差异有统计学意义(P<0.05);高剂量VC组胎鼠活胎率仅占55%,活胎中畸胎率明显增高占71%,其中神经管畸形发生率占50%,与正常对照组相比差异有统计学意义(P<0.01)。②中剂量和高剂量VC组胚胎神经管上皮厚薄不一致;高剂量组神经管多数呈明显未闭合状,神经管周围的间充质细胞肿胀,轮廓不清。③正常小鼠神经管神经上皮内可见PCNA阳性细胞与TUNEL阳性细胞分布;实验组各型阳性细胞分布与对照组相似,但随着染毒剂量的增加,PCNA阳性细胞表达减少,而TUNEL阳性细胞表达增多。结论孕早期一定剂量氯乙烯染毒可导致胚胎畸形率升高,神经管畸形率也相应上升。氯乙烯主要可抑制神经上皮细胞的增殖,诱导细胞凋亡发生增多,导致增殖与凋亡比例失调。

Objective To explore the influence of different doses of vinyl chloride（VC） on the development of neural tube in mice by observing the development and diversity of mouse embryo＇s nervous system,the proliferation and apoptosis of the neuroepithelial cells,and to further investigate the underlying mechanism of neural tube defect（NTD）.Methods The pregnant mice were randomly divided into experimental group and control group.The mice of experimental group were intraperitoneally injected by arachis oil with VC at the doses of 200,400 and 600 mg/kg in E6.5,meanwhile,the mice of control group was injected by arachis oil.Both of the groups were dissected in E12.5,and the embryo was immobilized by 4% PFA,paraffin-embedded and cut continuously.The expression of PCNA in the development of neural tube was detected by immunohistochemical staining.The apoptosis in the development of neural tube was observed by TUNEL assay.Results The living embryo in the experimental group of middle dose was decreased,in which the incidence of abnormity was 51%,and 40% of them was NTD.In the experimental group of high dose,the living embryo was 55%,and the incidence of abnormity was 71%,of which 50% was NTD.Both of them had significant deviation compared with the control group（P0.05）.② The thickness of the neural epithelium in middle and high dose groups were of disparity.The majority of neural tube in high dose group was NTD,around which the swelling and unclear mesenchymal cells were found.③ The positive cells of PCNA and TUNEL were widely distributed in neural tube of normal mouse（control group）.The distribution of both kinds of positive cells in the experimental groups resembled with that of control group.But the higher dose of VC was injected to rats,the lesser positive cells of PCNA were seen and the more positive cells of TUNEL were observed.Conclusion During early pregnancy,exposure to certain dose of VC makes the incidence of abnormity increased in rats,so did the rate of NTD.VC inhibits the proliferation of neural cells and induces the cell apoptosis,leads to the ratio of proliferation and apoptosis imbalance,which might be one significant reason for NTD induced by VC.