Table 1.
Gene-specific primers for quantitative real-time PCR.
Fig 1.
Cortisol upregulates SOCS mRNA abundance.
The effect of cortisol on the temporal profiles of SOCS-1 (A) and SOCS-2 (B) mRNA abundance in rainbow trout liver. Liver slices were incubated with either control media or media containing cortisol (100 ng/ml) for 1, 4, 6, 8 and 24 h. Values are plotted as % control and show mean ± S.E.M (n = 6 fish livers); different lower case letters denote significant treatment effects within each timepoint; (two way ANOVA, p < 0.05).
Fig 2.
Glucocorticoid receptor signaling is involved in SOCS upregulation.
The effect of cortisol and mifepristone either alone or in combination on SOCS-1 (A) and SOCS-2 (B) mRNA abundance in rainbow trout liver. Liver slices were incubated with control media or media containing cortisol (100 ng/ml), mifepristone (MP; 1000 ng/mL; Sigma) or a combination of mifepristone and cortisol for 24 h. Values are plotted as % control and show mean ± S.E.M (n = 6 fish livers); different upper case letters denote significant treatment effects (One way repeated measures ANOVA, p < 0.05).
Fig 3.
Pre-exposure to cortisol suppresses acute GH signaling.
The effect of cortisol and GH on IGF-1 mRNA abundance (A), STAT5 phosphorylation (B) and total JAK2 protein expression (C) in rainbow trout liver. Liver slices were pre-incubated with cortisol (100ng/ml; Sigma) or control media for 24 h and then stimulated with GH (100ng/ml or 1000ng/ml) for either 10 min (JAK/STAT) or 6 h (IGF-1). Values are plotted as % no cortisol control and shown as mean ± S.E.M (n = 6 fish livers); different lower case letters denote significant treatment effects and interactions; different upper case letters denote overall GH effects between the control, 100 GH and 1000 GH groups; the inset shows overall cortisol effects (two way repeated measures ANOVA, p < 0.05).
Fig 4.
Pre-exposure to cortisol suppresses LPS signaling.
Cortisol modulates LPS-induced IL-6 (A) but not IL-8 (B) mRNA abundance in rainbow trout liver. This is paralleled by an increase in SOCS-2 expression with cortisol exposure (C). Liver slices were pre-incubated with control media or media containing cortisol (100ng/ml) for 24 h after which they were incubated with control media or media containing LPS (30μg/ml) for 6 h. Values are plotted as % control and shown as mean ± S.E.M (n = 7 fish livers); different lower case letters denote significant treatment effects and interactions; * denotes overall cortisol effects (two way repeated measures ANOVA, p < 0.05).
Fig 5.
Pre-exposure to cortisol and LPS affect GH signaling.
The graphs show the effects of cortisol and LPS either singly or in combination in modulating GH stimulation of IGF-1 mRNA abundance (A), STAT5 phosphorylation (B) and total JAK2 protein expression (C) in rainbow trout liver. Rainbow trout liver slices were pre-incubated with control media or media containing cortisol (100 ng/ml; Sigma), LPS (30 μg/ml) or a combination of cortisol and LPS for 24 h, after which they were incubated with or without GH (500 ng/ml) for either 10 min (JAK/STAT) or 6 h (IGF-1). Values are plotted as % no GH control and show mean ± S.E.M (n = 6 fish livers); different lower case letters denote significant treatment effects and interations; * denotes overall GH effects; the inset shows overall cortisol effects (two way repeated measures ANOVA, p < 0.05).
Fig 6.
Effect of cortisol and LPS on GH receptors.
The effect of cortisol and LPS on GHR1 (A) and GHR2 (B) mRNA abundance in rainbow trout liver. Rainbow trout liver slices were pre-incubated with control media or media containing cortisol (100 ng/ml; Sigma), LPS (30 μg/ml) or a combination of cortisol and LPS for 24 h. Values are plotted as % control and show mean ± S.E.M (n = 6 fish livers); different lower case letters denote significant treatment effects; * denotes overall cortisol effects (two way repeated measures ANOVA, p < 0.05).
Fig 7.
A proposed model for cortisol effect on growth and immune suppression in trout liver.
Stressed levels of cortisol elevate SOCS transcript levels and reduce GH signaling and the corresponding IGF-1 expression in rainbow trout liver by preventing STAT5 phosphorylation and decreasing total JAK2 protein expression. The cortisol-induced upregulation of SOCS may be playing a role in the suppression of LPS-induced IL-6 expression (a cytokine signaling through the JAK/STAT pathway). Immune challenge with LPS may indirectly inhibit GH signaling by elevating plasma cortisol levels or directly inhibit GH signaling and the corresponding IGF-1 expression by downregulating growth hormone receptors 1 and 2 and by preventing STAT5 phosphorylation.