孕期咖啡因暴露影响神经嵴细胞表达致先天性心脏病的机制

The congenital heart disease study of caffeine exposure affect the CNCC expression during pregnancy

目的胚胎早期心脏发生的过程中过量咖啡因暴露,由于第二生心神经嵴细胞(CNCC)受到影响而导致心脏发育畸形。方法利用鸡胚为实验模型,在鸡胚心管发生发育的关键时期-HH9,按照正常组(PBS)、0.5μmol/egg组、1.0μmol/egg组、1.5μmol/egg组,连续3 d给予咖啡因暴露,至胚胎发育到12 d时获取胚胎用95%酒精固定2 d,对胚胎的心脏流出道(out flow tract,OFT)发育状态进行观察记录;,咖啡因暴露3 d后获取胚胎对CNCC进行检测,更深入研究其对心脏流出道发育机制的影响。结果胚胎早期心脏形成过程中,过量咖啡因暴露影响了CNCC的增殖、迁移,从而影响了心脏的正常发育,1.0μmol/egg组、1.5μmol/egg组分别出现心脏OFT的主动脉、肺动脉等发育畸形,并且有剂量依赖性。结论孕期大量咖啡因暴露通过影响第二生心区的CNCC的增殖和迁移路径导致心脏发育畸形,初步探究咖啡因暴露对胚胎早期心脏的发生发育影响的机制。

Objective Excessive caffeine exposure to the early-staged embryos led to cardiac malformations because of the negative effects on the embryonic cardiac neural crest cells （cardiac neural crest cells, CNCC ） in the second heart field. Methods Using chicken embryos as the experimental model, we exposed the embryos to caffeine in HH9, which was a critical period of cardiogenesis, with the doses of 5 μmol/egg, 10μmol/egg, 15μmol/egg group resperatively as there experiment groups, as well as the normal group （PBS） for three consecutive days. Then the embryos were obtained at 12 days and the cardiac outflow developmental status can be observed after the embryos were fixed in 95% alcohol for 2 clays. In order to study the mechanism of the cardiac outflow development further, the CNCC was defected after the three days＇ exposure of caffeine. Results An excess of caffeine exposure couhl affect the normal heart development in the early cardiogenesis procss by influencing the proliferation and the migration of the CNCC. And in 10μmol/egg group, 15μmol/egg group, malformations happened in a dose-dependant manner in the cardiac outflow of the aorta, puhnonary artery respectively in these two groups. Conclusion Excessive caffeine exposure influenced the proliferation and migration path of the CNCC, leading to cardiac malformations during the early stage of pregnancy, so preliminary explore the early embryonic developmental mechanism of the heart.