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|Title: ||Endocrine disrupting effects of waterborne PBDE on the brain-pituitary-gonad axis and reproduction of zebrafish (Danio rerio)|
|Other Titles: ||Shui xing bao lu duo xiu lian ben mi dui ban ma yu nao-chui ti-xing xian he sheng zhi de nei fen mi gan rao xiao ying|
|Authors: ||Han, Xiaobo ( 韓小波)|
|Department: ||Department of Biology and Chemistry|
|Degree: ||Doctor of Philosophy|
|Issue Date: ||2011|
|Publisher: ||City University of Hong Kong|
|Subjects: ||Zebra danio -- Generative organs -- Effect of polybrominated diphenyl ethers on.|
Pituitary gland -- Effect of polybrominated diphenyl ethers on.
Polybrominated diphenyl ethers -- Toxicology.
Endocrine disrupting chemicals
|Notes: ||CityU Call Number: QL638.C94 H36 2011|
xxiii, 250 leaves : ill. 30 cm.
Thesis (Ph.D.)--City University of Hong Kong, 2011.
Includes bibliographical references (leaves 204-250)
|Abstract: ||Polyborminated diphenyl ethers (PBDEs) are found ubiquitously in the
environment and biota. Since their molecular structure and physicochemical
properties are similar to those of polychlorinated biphenyl (PCBs) and dioxins,
there is increasing concern that PBDEs may disrupt endocrine systems, leading to
reproductive impairments in aquatic animals.
In this thesis, a commercial low brominated PBDEs mixture, DE-71 was selected as
a model chemical, and zebrafish (Danio rerio) employed as a model species, to test
the hypothesis that PBDE may affect fish reproduction by affecting multiple target
sites along the brain-pituitary-gonadal axis. Zebrafish were exposed to 5 ng/L, 1
µg/L and 50 µg/L of DE-71 for the whole life cycle (120 days, from eggs to adults),
and experiments were carried out to investigate the effects of PBDE on: (a) gonadal
development and reproductive processes; (b) level and balance of sex steroids, as
well as the various key enzymes regulating steroidogenesis and/or sex steroid
hormone receptors, and (c) the gene expression controlling the hypothalamic and
pituitary hormones and their receptors.
A significantly higher percentage of malformation was found in embryos when
exposed to the 1 µg/L (9.1%, p <0.05) and 50 µg/L (11.7%, p <0.01) of DE-71 at 96
hpf. At the same time, estradiol (E2) level in embryos was reduced at 48 hpf and
120 hpf (up to -48%, p <0.001). Effects of DE-71 on testosterone (T) level were
variable: the level of T increased at 48 hpf (+26%, p <0.05), while decreased at 120
hpf (at least -42%, p <0.05) and 12 dpf (up to -70%, p <0.01). The level of 11-ketotestosterone
(11KT) was decreased at all three time-points (at least -27% and up to -
62%, p <0.01) upon to exposure of DE-71. The expression of selected steroidogenic
genes was also significantly affected at these key time-points even at the lowest
DE-71 treatment (5 ng/L). At 48 hpf, expression of StAR was down-regulated (by
0.4-fold, p <0.05) in the 1 µg/L treatment, but up-regulated (by 3.0-fold, p <0.001) in
the 50 µg/L treatment group. Exposure to 5 ng/L and 1 µg/L DE-71 at 48 hpf upregulated
expression of CYP11a by 13.6-fold (p <0.01) and 18.2-fold (p <0.001),
respectively. Expression of CYP19a was down-regulated by 0.4-fold in the 5 ng/L (p <0.05) and 0.5-fold in the 50 µg/L (p <0.05) treatment group. Growth, however,
was not affected in any treatment.
At 60 dpf, E2 level of males significantly increased in the 1 µg/L (+73%, p <0.001)
and 50 µg/L (+58%, p <0.01) treatment groups. In females, E2 level increased by
32% in the 1 µg/L treatment group (p<0.05) and by 45% in the 50 µg/L treatment
group (p<0.05). T level of male fish was increased significantly in the 1 µg/L group
(+80%, p <0.05) and 11KT level was greatly increased by 375% in the minimum
exposure of 5 ng/L group (p <0.05). In female fish, T level increased only in the 50
µg/L (+61%, p <0.05) treatment group, and 11KT level was decreased by 36% only
in the 5 ng/L (p <0.05) treatment group.
Correspondingly, up-regulation (3-3.7 fold) of GnRH-2 was observed in both males
and females in the minimum 5 ng/L treatment (p <0.001). FSH-R in males was upregulated
(+1.9-fold) upon exposure to the 5 ng/L DE-71 (p <0.001), but downregulated
in the 1 µg/L and 50 µg/L treatments (p <0.01). Only the 50 µg/L
treatment led to 2-fold (p <0.01) of up-regulation of LH-R.
At 60dpf in testis, the lowest treatment 5 ng/L DE-71 stimulated the expression of
StAR (1.5-fold, p <0.05), while the highest 50 µg/L treatment depressed it (0.5-fold,
p <0.05). The expression of 3βHSD in male fish was down-regulated in all
treatments (0.2-0.4-fold, p <0.001). CYP19a was up-regulated by 1.7-fold (p <0.05)
in the 5 ng/L treatment group and down-regulated by 0.4-fold (p <0.05) in the 1
µg/L treatment group. There was no significant difference in expression of CYP11a
in all PBDE treatments. In females, all selected genes were down-regulated at 60
dpf. FSH-R and LH-R were down-regulated at higher concentrations of DE-71
(FSH-R: 0.4-0.5 fold, p <0.05; LH-R: 0.6-fold, p <0.05). The expression of 3βHSD
in female fish was down-regulated by 0.5-fold in both the 5 ng/L (p <0.05) and 1
µg/L (p <0.05) treatment groups. Ovarian CYP19a was down-regulated by 0.6-fold
(p <0.05) only in the 1 µg/L treatment group.
At 120 dpf, the level of E2 in both male and female fish was reduced upon exposure
to DE-71 (at least -30%, p <0.05). In male fish, both T and 11KT levels increased
significantly in the 1µg/L (at least +80%, p <0.05) and 50 µg/L treatment groups (up to +150%, p <0.001). 11KT level was increased greatly by 87% even in the
minimum exposure of 5 ng/L group (p <0.01). In female fish, T and 11KT levels
remained unchanged in all treatment groups, except a 24% decrease in 11KT level
in the 5 ng/L (p <0.05) treatment at 120dpf.
At 120 dpf, the mRNA level of GnRH-2 was up-regulated in both male and female
fish in the minimum 5 ng/L DE-71 treatment: 5.6-fold in males (p <0.001) and 4.7-
fold (p <0.001) in females. Both FSHβ and LHβ in both males and females were upregulated
in all concentrations after 120 days. In males, the 1 µg/L DE-71 exposure
led to 1.9-fold up-regulation in FSHβ (p <0.01) and LHβ was up-regulated by 2.9-
fold in the 1 µg/L (p <0.01) and 2.5-fold in the 50 µg/L (p <0.01) treatments. In
females, the 5 ng/L exposure caused 2.2-fold up-regulation in FSHβ (p <0.01), while
LHβ was up-regulated by 5-fold in the 5 ng/L (p <0.001) and 2.5-fold in the 50 µg/L
(p <0.01) treatment group.
This study provides the first evidence that whole life cycle exposure to PBDE (DE-
71) increased the transcripts of brain GnRH-2 and pituitary FSHβ and LHβ in both
male and female zebrafish. At 120 dpf the expression of TH and TPH of female
were more variable than male, they both were depressed by PBDE treatments,
especially at the highest concentration of 50 µg/L.
At 120 dpf, no significant difference was observed in StAR among all DE-71
treatments for both male and female fish. Only in males was CYP11a downregulated
by all three DE-71 treatments at 120 dpf. In male fish, the expression of
3βHSD was down-regulated by 0.3-fold in the 50 µg/L (p <0.01) treatment group at
120 dpf, but 2.4-fold up-regulation (p <0.01) was found in ovary in the same
treatment. The only up-regulated effect was found in testicular CYP19a expression
in the 50 µg/L (1.9-fold, p <0.05) treatment group. Our study demonstrated, for the
first time, the disruption of low brominated PBDEs on fish steroidogenesis in vivo
after long-term exposure.
The effects on the expression of key genes and hence sex hormone levels eventually
led to the impairment of gonadal development and reproductive processes. At 120
dpf, the observed disruption of sex hormones was associated with a reduction in spawning, fertilization and hatching success when fish were exposed to
environmentally realistic concentrations of DE-71. Exposure to 5 ng/L DE-71
reduced egg production by 60% (p <0.05), while exposure to 50 µg/L increased it by
40% (p <0.05). Significantly lower fertilization was also found in the 5 ng/L DE-71
group (-47%, p <0.05). Only 22% and 50% of fertilized eggs survived after 120 hpf
(p <0.05), following exposure to 5 ng/L and 50 µg/L respectively. Interestingly, the
percentage of males in the F1 generation significantly increased when exposed to
the 1 µg/L and 50 µg/L of DE-71 (by 68% and 69%, respectively; p <0.05).
Taken together, an increase in percentage of malformation, a lower production of
eggs, fertilization success, larval survival rate and a male-biased F1 generation,
suggested that low brominated PBDEs may pose a significant threat to natural fish
populations. Since hormonal regulation along the BPG axis is highly conserved in
vertebrates, the effects observed in zebrafish may also occur in other vertebrates.|
|Online Catalog Link: ||http://lib.cityu.edu.hk/record=b4086424|
|Appears in Collections:||BCH - Doctor of Philosophy |
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