International Journal of Marine Science 2016, Vol.6, No.4, 1-12
9
4 Discussion
Alkylphenol polyethoxylate (APEs) as nonionic
surfactants has been widely used in a variety of
industrial and surfactant applications. However,
several investigations have reported that the unstable
property of APEs in environment cause rapid
degradation to hydrophobic and more toxic
alkylphenols including 4-nonylphenol (4-NP) and
4-t-OP. Based on hematological and biochemical
parameters examined, the study elucidated that OP
had a relatively greater effect than NP and affected
hematological enzymes leading to serious impairment
of the metabolism and physiology in African
sharptooth catfish (C. gariepinus) (Senthil et al., 2011).
Other study reported that zebrafish embryos exposed
to 1
M of 4-t-OP developed normally (Chandrasekar
et al., 2011), however our results showed that
zebrafish exposure to 1
M of 4-t-OP resulted in
cardiovascular defect. These results also suggest that
the toxicity effect of 4-t-OP was higher than 4-NP,
and affect blood circulation of fish. In the present
study the developmental toxicity of 4-t-OP on
zebrafish embryos was first demonstrated that 4-t-OP
disrupts zebrafish cardiovascular system. 4-t-OP
exposure at 1
M significantly decreased heart rate in
zebrafish
hatchlings.
4-t-OP
and
other
endocrine-disrupting compounds has been linked to
endocrine disruption mediated via interference with
the estrogen and thyroid hormone systems (Ghisari
and Bonefeld, 2009). A strong positive correlation
between levels of thyroid hormone and heart rate has
been demonstrated (Roef et al., 2013). Thus, based on
those studies, we assume that 4-t-OP may reduce heart
rate through its effects on reducing thyroid hormone.
Transgenic biosensor zebrafish embryos which
express the green florescent protein (GFP) under the
control of estrogen-inducible promoter had been
developed for studying potential health effects of
environmental estrogens (Petersen et al., 2013).
Exposure of the transgenic biosensor of zebrafish to
4-t-OP induced GFP expressed demonstrating that
4-t-OP possesses ability to act as natural estrogen
activity in zebrafish (Brion et al., 2012). Moreover,
exposure to alkylphenol induced GFP expressed in
heart of transgenic biosensor zebrafish suggesting
4-t-OP act action in cardiovascular system (Lee et al.,
2012). It is well-known that estrogen mediates
estrogen receptors (ERs) to activate transcription
factors (TFs) that modulating estrogen target gene
activity. Exposure of zebrafish embryos to 4-t-OP
caused carodio vascular defects can be done through
4-t-OP binding of ERs. In zebrafish, the three estrogen
receptors, ER, ER 1 and ER 2, had been characterized,
and three ERs with a distinct feature in gene structure
and tissue distribution pattern (Menuet et al., 2002). In
the present study, the expression of estrogen receptors
including ER, ER 1 and ER 2 were analyzed. The
presence of 4-t-OP at 0.5
M and 1
M significantly
induced ER, ER 1 and ER 2 expression in zebrafish,
and higher induction level was revealed in ER and ER
2. Reports have showed that ERs expression can be
induced by diverse estrogens or estrogen analog, and
different type of ER have a different binding affinity
for the different ligands. Using HELN assay, which
ERE-driven full- length zebrafish ER, ER 1 and ER 2
expression in HeLa cells, 4-t-OP has been
demonstrated to have greater affinity towards
zebrafish ER and ER 2 relative to ER 1 (Pinto et al.,
2014). Our study present higher expression level
induced by 4-t-OP in ER and ER 2 also potentially
suggesting that zebrafish ER and ER 2 have higher
affinity for 4-t-OP.
The heart is the first organ to form and function
during embryogenesis and its circulatory function is
critical for the viability of zebrafish embryos. The
presence of 4-t-OP in zebrafish cause cardiovascular
defects including incomplete looping of ventricle and
atrium, defects in formation of intersegmental vessels
and organization of caudal vein, and these indicators
signifying that the heart development and circulation
function were injured. Several genes encoding
transcription factors are required for normal heart and
blood vessel development. The tinmen gene encodes a
NK-class of homeobox transcription factor which
plays key roles in the establishment of myogenic
lineages. In zebrafish Nkx2.5 and Nkx2.7 are
expressed in heart field of lateral plate mesoderm and
required for cardiac morphogenesis (Stainier, 2001).
Report has showed that morpholino (MO) knockdown
Nkx2.5 in zebrafish did not affect heart development.
Furthermore Nkx2.7 has been demonstrated to play a
critical function in the lateral development of the heart
and normal cardiac looping and chamber formation
(Tu et al., 2009). The hand2 gene encodes bHLH
transcription factor that regulate differentiation and
the morphogenesis of the myocardial cells and
