Expression of phase I and phase II genes in mouse embryonic stem cells cultured in the presence of 2,3,7,8-tetrachlorodibenzo-para-dioxin.
Embryonic stem (ES) cells have features that resemble the pluripotent cells of peri-implantation embryos and have been used as an in vitro model to assess the effects of test substances on these stages of development. Here, for the first time, we report on the effects of the xenobiotic 2,3,7,8-tetrachlorodibenzo-para-dioxin (TCDD) on mouse ES cells cultured with TCDD at concentrations ranging from 0.0001 to 100 nM for 15 min to 48 h. TCDD effects were determined by analysing the induction of Cyp1A1, Cyp1A2, Cyp1B1 (phase I) and Nqo1, Gsta1, Ugt1a6 (phase II) genes. Cyp1A1 was the phase I gene most rapidly induced (4 h at 1 nM); Cyp1B1 was induced at 48 h (1 nM), whereas Cyp1A2 expression was not affected. TCDD did not alter phase II gene expression, which remained at basal levels throughout the 48 h of culture. We studied more accurately the expression of Cyp1A1, the earliest gene to respond to the presence of TCDD. We found that: 1) Cyp1A1 gene induction is dependent on the duration of exposure (precisely it is first induced after 3 h of culture at 1 nM, the minimum effective-dose); 2) Cyp1A1 induction requires the continuous presence of TCDD, being interrupted 4 h after removal of the xenobiotic; and 3) induced expression of CYP1A1 protein is dependent on TCDD concentration, the higher the concentration the earlier the production of the enzyme. Furthermore, after 48 h of treatment, TCDD did not promote either apoptosis or changes to the differentiation status of the ES cells. These results are the first important step to investigate the effects of dioxin on the very early stages of mammalian development.
Neri
Tui
T
Laboratorio di Biologia dello Sviluppo, Dipartimento di Biologia Animale, Universita' di Pavia, Pavia, 27100, Italy.
Merico
Valeria
V
Garagna
Silvia
S
Redi
Carlo Alberto
CA
Zuccotti
Maurizio
M
D000818:Animals; D000943:Antigens, Differentiation / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics / Q000378:metabolism; D019388:Cytochrome P-450 CYP1A2 / Q000235:genetics / Q000378:metabolism; D065633:Cytochrome P-450 CYP1B1; D004305:Dose-Response Relationship, Drug; D053595:Embryonic Stem Cells / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D016660:NAD(P)H Dehydrogenase (Quinone); D009252:NADPH Dehydrogenase / Q000235:genetics / Q000378:metabolism; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
D000943:Antigens, Differentiation; D007527:Isoenzymes; D000072317:Polychlorinated Dibenzodioxins; D019253:Proto-Oncogene Proteins c-bcl-2; D018336:Receptors, Aryl Hydrocarbon; D051028:bcl-2-Associated X Protein; D001189:Aryl Hydrocarbon Hydroxylases; C585498:Cyp1b1 protein, mouse; D019363:Cytochrome P-450 CYP1A1; D019388:Cytochrome P-450 CYP1A2; D065633:Cytochrome P-450 CYP1B1; D016660:NAD(P)H Dehydrogenase (Quinone); C491732:Nqo1 protein, mouse; D009252:NADPH Dehydrogenase; C110655:UDP-glucuronosyltransferase, UGT1A6; D014453:Glucuronosyltransferase; D005982:Glutathione Transferase; C119995:glutathione S-transferase alpha