Primary cultured cells as sensitive in vitro model for assessment of toxicants--comparison to hepatocytes and gill epithelia.
In an effort to develop cultured cell models for toxicity screening and environmental biomonitoring, we compared primary cultured gill epithelia and hepatocytes from freshwater tilapia (Oreochromis niloticus) to assess their sensitivity to AhR agonist toxicants. Epithelia were cultured on permeable supports (terephthalate membranes, "filters") and bathed on the apical with waterborne toxicants (pseudo in vivo asymmetrical culture conditions). Hepatocytes were cultured in multi-well plates and exposed to toxicants in culture medium. Cytochrome P4501A (measured as 7-Ethoxyresorufin-O-deethylase, EROD) was selected as a biomarker. For cultured gill epithelia, the integrity of the epithelia remained unchanged on exposure to model toxicants, such as 1,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), benzo(a)pyrene B[a]P, polychlorinated biphenyl (PCB) mixture (Aroclor 1254), and polybrominated diphenyl ether (PBDE) mixture (DE71). A good concentration-dependent response of EROD activity was clearly observed in both cultured gill epithelia and hepatocytes. The time-course response of EROD was measured as early as 3h, and was maximal after 6h of exposure to TCDD, B[a]P and Aroclor 1254. The estimated 6h EC50 for TCDD, B[a]P, and Aroclor 1254 was 1.2 x 10(-9), 5.7 x 10(-8) and 6.6 x 10(-6)M. For the cultured hepatocytes, time-course study showed that a significant induction of EROD took place at 18 h, and the maximal induction of EROD was observed at 24h after exposure. The estimated 24h EC50 for TCDD, B[a]P, and Aroclor 1254 was 1.4 x 10(-9), 8.1 x 10(-8) and 7.3 x 10(-6)M. There was no induction or inhibition of EROD in DE71 exposure to both gill epithelia and hepatocytes. The results show that cultured gill epithelia more rapidly induce EROD and are slightly more sensitive than cultured hepatocytes, and could be used as a rapid and sensitive tool for screening chemicals and monitoring environmental AhR agonist toxicants.
Zhou
Bingsheng
B
State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, PR China. bszhou@ihb.ac.cn
Liu
Chunsheng
C
Wang
Jingxian
J
Lam
Paul K S
PK
Wu
Rudolf S S
RS
D000818:Animals; D001564:Benzo(a)pyrene / Q000633:toxicity; D002478:Cells, Cultured; D020111:Chlorodiphenyl (54% Chlorine) / Q000633:toxicity; D023681:Cichlids; D019363:Cytochrome P-450 CYP1A1 / Q000032:analysis / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D004784:Environmental Monitoring / Q000379:methods*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D005880:Gills / Q000166:cytology / Q000187:drug effects*; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity; D013997:Time Factors; D018675:Toxicity Tests / Q000379:methods*; D014874:Water Pollutants, Chemical / Q000633:toxicity*
D000072317:Polychlorinated Dibenzodioxins; D014874:Water Pollutants, Chemical; D020111:Chlorodiphenyl (54% Chlorine); D001564:Benzo(a)pyrene; D019363:Cytochrome P-450 CYP1A1