The overexpression of uc.4 caused abnormal development of zebrafish. (a) The overexpression of uc.4 increased the death rates of zebrafish in a dose-dependent manner. (b) The death rates of the WT, NC and uc.4 groups (200 pg). (c) The expression of uc.4 was upregulated in the uc.4 overexpression groups. ***P<0.001 versus NC groups, Student’s t-test. (d) The overexpression of uc.4 was determined by in situ hybridization. uc.4 is primarily distributed in the atria of zebrafish. (e) The overexpression of uc.4 leads to heart looping defects and pericardial edema in zebrafish. (f) qRT-PCR determination of the mRNA expression of cardiac-specific markers (Nkx2.5, Vmhc, GATA4, hand2, Mef2c, Cx43). A minor decrease was observed in the expression of these cardiac markers. *P<0.05, **P<0.01 versus NC groups, Student’s t-test. Each experiment was repeated three times. Data are presented as the means±s.d. NC, negative control; qRT-PCR, quantitative real-time polymerase chain reaction.

Cardiac function and histological changes. (a) The overexpression of uc.4 decreased the heart rates of zebrafish embryos. (b) Histological sections of the heart showed a thinner ventricular wall and smaller valve structure compared to control embryos. Red arrows: ventricle. Blue arrows: atrium. (c) SV–BA distance, which is normalized to total body length, resulting in a ratio. The SV–BA was increased in the overexpression group compared with the WT group. (d) The edema index was increased in the overexpression group compared with the WT group. **P<0.01, ***P<0.001 versus WT groups, Student’s t-test. Each experiment was repeated three times. Data are presented as the means±s.d. SV–BA, distance between the sinus venous and the bulbus arteriosus; WT, wild type.