Purification and isolation of corticosteroidogenic cells from head kidney of rainbow trout (Oncorhynchus mykiss) for testing cell-specific effects of a pesticide.
The teleost head kidneys contain corticosteroidogenic cells, chromaffin cells, lymphoid cells, and melanomacrophages. We have developed and validated a method using a Percoll density gradient and differential staining for 3beta-hydroxysteroid dehydrogenase (3beta-HSD) to prepare fractions enriched with specific head kidney cell types. The proportion of steroidogenic cells to other cells in the head kidney was 1:8000 in rainbow trout, Oncorhynchus mykiss. To test the hypothesis that steroidogenic cells are more vulnerable to a pesticide than other cell types in the head kidney, head kidney cells were separated by a Percoll gradient and the steroidogenic cell-enriched fractions and lymphoid cell-enriched fractions were exposed to the pesticide endosulfan in vitro, and their functional integrity and viability were assessed. The effective concentration of the pesticide (EC50, concentration that inhibits 50% of the secretory response to ACTH) in the mixed head kidney cell preparation was similar to the EC50 in the fraction enriched with steroidogenic cells, but differences in viability were detected. The Percoll method for isolation of different cell types from the head kidney facilitated a study of cell-specific effects of a pesticide.
Hontela
Alice
A
Department of Biological Sciences, University of Lethbridge, Lethbridge, Alberta, Canada T1K 3M4. alice.hontela@uleth.ca
Leblond
Vincent S
VS
Chang
John P
JP
D015096:3-Hydroxysteroid Dehydrogenases / Q000032:analysis; D000305:Adrenal Cortex Hormones / Q000096:biosynthesis*; D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D000818:Animals; D002469:Cell Separation / Q000379:methods*; D048430:Cell Shape; D002470:Cell Survival / Q000187:drug effects; D002499:Centrifugation, Density Gradient; D004305:Dose-Response Relationship, Drug; D004726:Endosulfan / Q000633:toxicity*; D029941:Fish Proteins / Q000032:analysis; D006854:Hydrocortisone / Q000378:metabolism; D007668:Kidney / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D008214:Lymphocytes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D017686:Oncorhynchus mykiss / Q000378:metabolism*; D010575:Pesticides / Q000633:toxicity*; D011205:Povidone; D015203:Reproducibility of Results; D012822:Silicon Dioxide; D013194:Staining and Labeling
D000305:Adrenal Cortex Hormones; D029941:Fish Proteins; D010575:Pesticides; C016039:Percoll; D012822:Silicon Dioxide; D000324:Adrenocorticotropic Hormone; D015096:3-Hydroxysteroid Dehydrogenases; D011205:Povidone; D004726:Endosulfan; D006854:Hydrocortisone