摘要
目的利用新型模式生物斑马鱼，采用外源性视黄醛脱氢酶2抑制剂——对二乙氨基苯甲醛（4-diethylaminobenzaldehyde，DEAB），建立维甲酸（retinoicacid，RA）缺乏的斑马鱼模型，探讨其对斑马鱼胚胎心脏发育的影响。方法在斑马鱼胚胎受精后5、8、10．3h，分别用1×10^-6、5×10^-6、10×10^-6、25×10^-6mol／L的DEAB处理，在解剖显微镜下实时观察胚胎发育的全过程，在受精后5h给予1×10^-9mol／L外源性RA干预，观察其对DEAB致畸的拮抗作用。通过胚胎心脏表型观察、心率和心室收缩分数比较以及心脏特异分子标记——心房利钠肽A基因整体原位杂交实验分析RA缺乏对胚胎心脏发育的影响。结果外源性DEAB处理后，胚胎生存率随着处理浓度增加而降低，随着处理时间点后移而升高。当DEAB浓度≥5×10^-6mol／L时，斑马鱼畸胎率为100％，异常表型一致，并能被1×10^-6mol／L外源性RA有效援救。RA缺乏时斑马鱼心脏表现出管状心脏、无向右环化或环化不完全、房室分化异常及房室管区血液反流。与野生型胚胎相比，DEAB处理后斑马鱼胚胎心率和心室收缩分数降低，心房利钠肽A基因表达改变，在心室部位表达清晰强烈，在心房部位表达明显减弱。结论DEAB影响胚胎发育有剂量依赖性和时效性，其致畸作用能被外源性RA有效拮抗。RA缺乏影响心脏早期发育的多个重要环节，导致心脏收缩功能受损。心脏心房利钠肽A基因表达受RA信号调控。
Objective To study the effect of retinal dehydrogenase type 2 inhibitor （4-diethylaminobenzaldehyde, DEAB）on embryonic cardiac development of zebrafish model with retinoic acid （RA）deficiency. Methods gebrafish embryos were treated with DEAB at various concentrations including 1^-6, 5^-6, 10^-6, 25^-6mol/Lat5, 8 and 10.3 hours post fertilization, respectively. The effects of DEAB on the embryonic development were assessed under microscope. 1^-9 mol/L exogenous RA was then added to detect the antagonistic effect against DEAB. The abnormal cardiac phenotype, heart rate and ventrieular systolic fraction were observed and analyzed between wild type and DEAB treated groups. The expression of specific cardiac gene, natriuretic peptide precursor A, was monitored by whole-mount in situ hybridization to demonstrate the effect of RA signaling on early cardiac development. Results The survival rate of zebrafish embryos declined with the increase of DEAB concentration at different developmental stage. The percentage of abnormal embryos reached 100% when DEAB over 5 ^-6mol/L. 1^-6 mol/L exogenous RA could eliminate the teratogenie effect of DEAB （95^-6 mol/L）. DEAB treated embryos presented abnormal cardiac phenotype, including tubular heart, incomplete D-loop,abnormal atrioventricular development, regurgitation, slow blood flow and weak heart beat. The difference of heart rate and ventricular systolic fraction between wild type and RA deficiency embryos was of statistical significance （P〈0.05）. The natriuretic peptide precursor A expression remained in the ventricle, but reduced obviously in the atrium with RA signaling deficiency. Conclusions The effects of DEAB on the embryonic development are dose-dependent and time-dependent, and could be rescued by exogenous RA. RA signaling plays a critical role in several key stages of early cardiac development and natriuretie peptide precursor A expression.
出处
《中华围产医学杂志》
CAS
2010年第2期117-122,共6页
Chinese Journal of Perinatal Medicine
基金
上海市科学技术委员会科研计划项目（054119512）
关键词
维甲酸
视黄醛脱氢酶
斑马鱼
心脏
胚胎发育
Tretinoin
Retinal dehydrogenase
Zebrafich
Heart
Embryonic development
