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[Establishment of model of congenital heart disease with Tbx2 gene knockdown: experiment with zebrafish].

OBJECTIVE
To establish a model of congenital heart disease with Tbx2 gene knockdown.


METHODS
1200 fertilized eggs of zebrafish (Danio rerio) were divided into 3 groups: wild type group (n = 100), Tbx2-morpholino (MO) group (n = 600, to be injected with Tbx2-MO so as to knock down the Tbx2 gene), and control MO group (n = 500, to be injected with control MO). 12, 24, 36, 48, 72, and 96 hours after fertilization the heart structure, heart rate, rhythm, contractibility, and blood flow of the embryos were observed under anatomic microscope and digital camera was used to record the findings. Whole-mount in situ hybridization was conducted to observe the expression of versican in the heart.


RESULTS
Eight hours after fertilization 27.2% of the fertilized eggs of the Tbx2-MO group died, and 21.3%, 32.8%, and 18.7% of them showed mild, moderate, or severe developmental abnormality of heart, including hypogenetic ventricle, dilatation of atria, abnormal atrioventricular canal, slow heartbeat, arrhythmia, and blood regurgitation. Whole-mount in situ hybridization showed that versican did not decrease with the passage of time but was expressed abnormally in the atria and ventricles, and did not disappear 72 hours after fertilization; however, it was expressed normally in the statolith.


CONCLUSION
MO antisense oligonucleotide knockdown is a good method for the study of heart development. Tbx2 gene plays an important role in the differentiation of atria and ventricles and formation of atrioventricular canal.