International Journal of Marine Science 2016, Vol.6, No.4, 1-12
5
cardiovascular phenotype by exposing them
continuously from 5 hpf to 96 hpf.
Table 2 survival rate, hatching rate and malformation rate of
zebrafish embryos exposed to different concentration of 4-t-OP
at 3 dpf.
4t-OP conc.
(
M)
Survival rate
(%)
a
Hatching
rate (%)
a
Malformation
rate (%)
a
Control
b
96±2.1%
96±2.9%
0 %
0.1
M
95±1.8%
95±1.1%
0%
0.5
M
79±1.5%
83±1.7%
28±1.6%
1.0
M
52±1.8%
71±2.8%
60±1.3%
2.5
M
26±2.8%
45±2.3%
87±4.4%
5.0
M
12±0.4%
23±1.8%
100%
Note:
a
Data was evaluated from three times experiment and
each experiment was triplicated (n=100).
b
Control indicated
the embryo medium only containing 0.2% PTU.
3.2 4-tert-octylphenol induced cardiovascular
defects
To easily observe the cardiovascular defect, Tg (fil-1:
EGFP) zebrafish embryos were used for immersion
administration. As result shown in Table 3, exposure
of Tg (fil-1: EGFP) zebrafish embryos to 4-t-OP
resulted in visible cardiovascular defects from 24 hpf.
The cardiovascular development was normal in the
control group (Fig. 1A, C and D), however the
severity of the cardiovascular defects were not
consistent among 4-t-OP treated embryos. The
majority of embryos exhibited pericardial edema (56%
at 2 dpf) and irregular shape or incomplete looping of
ventricle and atrium (28% at 2 dpf) (Fig. 1B). The
proportion of these phenotypes was increased
following 4-t-OP exposure for 3 days, and then
declined at 4 days due to the increased mortality. At
certain region in the notochord tail, the absence of
intersegmental vessel caused the change of the
distance between intersegmental vessels and it was
also exhibited in 4-t-OP treated embryos (Fig. 1C).
Furthermore, unformed parachordal vessel and kinks
in the caudal vein resulted in blockage of blood flow
were exhibited in the 4-t-OP treated embryos (Fig.
1F).These phenotypes were categorized as abnormal
blood vessel development, and the proportion was
increased accompanying with the time of 4-t-OP
treatment (Table 2).
Table 3 Cardiovascular defects induced by 4t-OP
Developmenta
l stages
Treatment
Number of embryos
examined
Embryos with cardiovascular defects
Pericardial
edema
Irregular
shape
of
atrium and ventricle
Abnormal blood vessel
development
1 dpf
Control
312
None
None
None
1
M 4t-OP
287
123 (43%) 49 (17%)
26 (9%)
2 dpf
Control
304
None
None
None
1
M 4t-OP
273
153 (56%) 76 (28%)
44 (16%)
3 dpf
Control
301
None
None
None
1
M 4t-OP
178
110 (62%) 83 (47%)
66 (37%)
4 dpf
Control
297
None
None
None
1
M 4t-OP
143
61 (43%)
31 (21%)
67 (47%)
Note: Zebrafish embryos were exposed to control (embryo medium contain PTU only) or 1
M 4t-OP and the cardiovascular
defects was examined on days 1, 2, 3 and 4. Data shown are the pooled results of triplicated experiments.
