Function analysis of zebrafish (Danio rerio) hoxb3a gene in early heart development
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摘要: 为探究hoxb3a基因在斑马鱼（Danio rerio）心脏早期发育中的调控作用，利用基因沉默技术（Morpholino）对斑马鱼胚胎进行hoxb3a基因敲降，并结合显微镜观察分析、实时荧光定量PCR、原位杂交技术分析hoxb3a基因对斑马鱼心脏发育的影响。结果表明：hoxb3a基因敲降组（hoxb3a morphant）出现心包腔肿大、心脏环化角度异常和心房心室形态改变等现象；原位杂交试验确定了hoxb3a基因在不同时空的表达区域，其表达位置的变化与神经嵴细胞参与心脏发育的形成有关；在受精后3 d，与对照组相比，hoxb3a基因敲降组胚胎一些心脏关键基因的表达量发生变化，其中心脏祖细胞相关基因nkx2.5、gata4及房室形成关键基因nppa的表达量均具有显著升高的趋势，同时与神经嵴发育相关的fgf8a基因表达量也明显升高。研究表明，hoxb3a基因在斑马鱼胚胎的早期心脏发育中，通过调控nkx2.5、gata4、nppa与fgf8a等相关转录因子的表达，参与斑马鱼早期心脏发育的过程，本研究结果为早期心脏发育研究提供了新思路。Abstract: To investigate the regulatory role of hoxb3a gene in early development of genetically modified strains of zebrafish (Danio rerio) heart, hoxb3a gene was knocked down in the zebrafish embryos using gene silencing technology (Morpholino), and the effects of hoxb3a on zebrafish heart development were analyzed by combining microscopic observation and analysis, real-time fluorescence quantitative PCR, and in situ hybridization. The results showed that the hoxb3a knockdown group (hoxb3a morphant) showed enlarged pericardial cavity, abnormal heart looping angle and altered atrial and ventricular morphology. The in situ hybridization assay determined the expression regions of hoxb3a in different spatial and temporal regions, and changes in the location of its expression indicated that hoxb3a was related to the involvement of neural crest cells in the formation of heart development. In the hoxb3a knockdown group, the expression level of some key cardiac genes was found to be changed at 3 days after fertilization compared with the control embryos, significantly increase in the expression of cardiac progenitor genes nkx2.5 and gata4, and nppa, a key gene for atrial formation. The expression level of fgf8a, a gene related to neural crest development, was shown to be significantly increased. The findings indicate that hoxb3a plays an important role in early cardiac development in zebrafish embryos and influences cardiac development by regulating the expression level of nkx2.5, gata4, nppa and fgf8a and other related transcription factors, which may provide new ideas for early cardiovascular development research.
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Key words:
- Danio rerio /
- hoxb3a /
- cardiac development
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