Influence of culture conditions on metal-induced responses in a cultured rainbow trout gill epithelium.
A primary culture technique for rainbow trout (Oncorhynchus mykiss) gill cells was optimized to better represent the intact gill in vivo in response to waterborne toxic metals. Modifications in cell seeding density and culture conditions resulted in a gill epithelial cell culture model, which displayed classic in vivo responses to toxic metals. Metallothionein-A (MTA), metallothionein-B (MTB), zinc transporter-1 (ZnT-1), glutathione-S-transferase (GST), and glucose-6-phosphate-dehydrogenase (G6PD) all showed dose-dependent increases in expression at the mRNA level in response to waterborne zinc. Of these genes, the change in zinc-induced expression relative to the control was greatest for MTA, MTB, and ZnT-1. MT expression was also induced by silver, lead, copper, and cadmium. Cells cultured with freshwater on the apical side maintained the net transepithelial influx of Ca2+ displayed by freshwater trout gills in vivo, and there was an active inward movement of Ca2+. Waterborne zinc applied to the apical compartment reduced the net uptake of Ca2+ by stimulating the efflux component. The use of endogenous metal-responsive gene expression and inhibition of ion transport in the developed cell culture system will facilitate studies of metal-gill interactions and may prove to have future practical applications within biomonitoring of natural waters.
Walker
Paul A
PA
Nutritional Sciences Research Division, King's College London, Franklin-Wilkins Building, 150 Stamford Street, London SE1 9NH, UK. Paul.Walker@man.ac.uk
Bury
Nic R
NR
Hogstrand
Christer
C
D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D003470:Culture Media / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D005880:Gills / Q000166:cytology*; D005954:Glucosephosphate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D008668:Metallothionein / Q000235:genetics / Q000378:metabolism; D008670:Metals / Q000494:pharmacology*; D017686:Oncorhynchus mykiss; D020133:Reverse Transcriptase Polymerase Chain Reaction
D002352:Carrier Proteins; D003470:Culture Media; D008670:Metals; C017800:zinc-binding protein; D008668:Metallothionein; D005954:Glucosephosphate Dehydrogenase; D005982:Glutathione Transferase