Assessment of estrogenic endocrine-disrupting chemical actions in the brain using in vivo somatic gene transfer.
Estrogenic endocrine-disrupting chemicals abnormally stimulate vitellogenin gene expression and production in the liver of many male aquatic vertebrates. However, very few studies demonstrate the effects of estrogenic pollutants on brain function. We have used polyethylenimine-mediated in vivo somatic gene transfer to introduce an estrogen response element-thymidine kinase-luciferase (ERE-TK-LUC) construct into the brain. To determine if waterborne estrogenic chemicals modulate gene transcription in the brain, we injected the estrogen-sensitive construct into the brains of Nieuwkoop-Faber stage 54 Xenopus laevis tadpoles. Both ethinylestradiol (EE2; p < 0.002) and bisphenol A (BPA; p < 0.03) increased luciferase activity by 1.9- and 1.5-fold, respectively. In contrast, low physiologic levels of 17ss-estradiol had no effect (p > 0.05). The mixed antagonist/agonist tamoxifen was estrogenic in vivo and increased (p < 0.003) luciferase activity in the tadpole brain by 2.3-fold. There have been no previous reports of somatic gene transfer to the fish brain; therefore, it was necessary to optimize injection and transfection conditions for the adult goldfish (Carassius auratus). Following third brain ventricle injection of cytomegalovirus (CMV)-green fluorescent protein or CMV-LUC gene constructs, we established that cells in the telencephalon and optic tectum are transfected. Optimal transfections were achieved with 1 microg DNA complexed with 18 nmol 22 kDa polyethylenimine 4 days after brain injections. Exposure to EE2 increased brain luciferase activity by 2-fold in males (p < 0.05) but not in females. Activation of an ERE-dependent luciferase reporter gene in both tadpole and fish indicates that waterborne estrogens can directly modulate transcription of estrogen-responsive genes in the brain. We provide a method adaptable to aquatic organisms to study the direct regulation of estrogen-responsive genes in vivo.
Trudeau
Vance L
VL
Centre for Advanced Research in Environmental Genomics, Department of Biology, University of Ottawa, Ottawa, Ontario, Canada. trudeauv@uottawa.ca
Turque
Nathalie
N
Le Mével
Sébastien
S
Alliot
Caroline
C
Gallant
Natacha
N
Coen
Laurent
L
Pakdel
Farzad
F
Demeneix
Barbara
B
D000818:Animals; D001681:Biological Assay / Q000379:methods; D001921:Brain / Q000187:drug effects* / Q000502:physiology*; D003587:Cytomegalovirus / Q000235:genetics; D004703:Endocrine System / Q000187:drug effects; D004967:Estrogens / Q000494:pharmacology / Q000633:toxicity*; D020869:Gene Expression Profiling; D018014:Gene Transfer Techniques; D006054:Goldfish / Q000235:genetics / Q000502:physiology; D008156:Luciferases / Q000235:genetics* / Q000494:pharmacology; D013937:Thymidine Kinase / Q000235:genetics* / Q000494:pharmacology; D014162:Transfection; D014874:Water Pollutants, Chemical / Q000494:pharmacology / Q000633:toxicity; D014982:Xenopus laevis / Q000235:genetics
D004967:Estrogens; D014874:Water Pollutants, Chemical; D008156:Luciferases; D013937:Thymidine Kinase
Environ Health Perspect. 2002 Mar;110(3):277-84
11882479
Environ Health Perspect. 1999 Feb;107 Suppl 1:89-108
10229711
Environ Sci Technol. 2002 Mar 15;36(6):1202-11
11944670
Biol Reprod. 2002 Jun;66(6):1881-92
12021076
Dev Dyn. 2002 Aug;224(4):381-90
12203730
Environ Toxicol Chem. 2002 Sep;21(9):1864-75
12206426
Chemosphere. 2002 Oct;49(1):97-103
12243336
Comp Biochem Physiol B Biochem Mol Biol. 2002 Dec;133(4):559-70
12470819
J Steroid Biochem Mol Biol. 2002 Oct;82(2-3):181-94
12477484
Environ Int. 2002 Dec;28(6):545-51
12503920
J Cell Sci. 2003 Feb 15;116(Pt 4):585-6
12538758
Ecotoxicol Environ Saf. 2003 Mar;54(3):315-22
12651187
Mol Cell Endocrinol. 2003 Jun 30;204(1-2):169-77
12850291
Chemosphere. 2003 Oct;53(3):223-35
12919782
Gen Comp Endocrinol. 2003 Sep;133(2):189-98
12928009
Environ Toxicol Chem. 2003 Oct;22(10):2466-75
14552012
J Steroid Biochem Mol Biol. 2003 Sep;86(3-5):387-91
14623535
Environ Res. 2004 Jan;94(1):102-11
14643292
Cell. 2003 Dec 12;115(6):751-63
14675539
Environ Toxicol Chem. 2003 Dec;22(12):2872-80
14713026
Endocrinology. 2004 Jul;145(7):3055-61
14976146
Reprod Biol Endocrinol. 2003 Jul 9;1:52
12904263
J Comp Neurol. 1975 Jan 1;159(1):69-101
1088950
J Comp Neurol. 1978 Dec 15;182(4):611-20
721971
Endocrinology. 1988 Apr;122(4):1349-56
3345716
Environ Health Perspect. 1995 Oct;103 Suppl 7:157-64
8593864
Environ Health Perspect. 1995 Oct;103 Suppl 7:173-8
8593867
J Comp Neurol. 1995 Dec 18;363(3):461-74
8847411
Carcinogenesis. 1996 Nov;17(11):2279-84
8968038
Rev Reprod. 1997 Jan;2(1):55-68
9414466
Neuroendocrinology. 1999 Sep;70(3):168-74
10516479
Int J Dev Biol. 1999 Nov;43(8):823-30
10707906
J AOAC Int. 2000 Mar-Apr;83(2):290-7
10772166
Chemosphere. 2001 Feb;42(4):415-8
11100793
J Steroid Biochem Mol Biol. 2000 Nov 30;74(5):255-9
11162933
J Neurochem. 2001 Feb;76(4):945-56
11181814
Hum Reprod. 2001 May;16(5):1030-6
11331656
Curr Opin Mol Ther. 2001 Apr;3(2):178-82
11338931
Environ Toxicol Chem. 2001 Feb;20(2):297-308
11351429
Endocr Rev. 2001 Jun;22(3):319-41
11399747
Mar Pollut Bull. 2001 Aug;42(8):643-55
11525282
J Steroid Biochem Mol Biol. 2001 Jul;78(1):25-32
11530281
Mol Endocrinol. 2001 Nov;15(11):1953-70
11682626
J Steroid Biochem Mol Biol. 2001 Dec;79(1-5):305-14
11850237
Environ Health Perspect. 2002 Mar;110(3):269-75
11882478
J Mol Endocrinol. 1997 Dec;19(3):321-35
9460653
Chemosphere. 1998 Apr;36(10):2149-73
9566294
Crit Rev Toxicol. 1998 Jul;28(4):319-61
9711432
FASEB J. 1998 Dec;12(15):1755-64
9837866
Int J Dev Biol. 1998 Nov;42(8):1159-64
9879714
Sci Total Environ. 1999 Jan 12;225(1-2):59-68
10028703
Sci Total Environ. 1999 Jan 12;225(1-2):81-90
10028705
Bioessays. 2002 Mar;24(3):244-54
11891761