Immune neuroendocrine phenotypes in Coturnix coturnix: do avian species show LEWIS/FISCHER-like profiles?

Abstract

Immunoneuroendocrinology studies have identified conserved communicational paths in birds and mammals, e.g. the Hypothalamus-Pituitary-Adrenal axis with anti-inflammatory activity mediated by glucocorticoids. Immune neuroendocrine phenotypes (INPs) have been proposed for mammals implying the categorization of a population in subgroups underlying divergent immune-neuroendocrine interactions. These phenotypes were studied in the context of the LEWIS/FISCHER paradigm (rats expressing high or low pro-inflammatory profiles, respectively). Although avian species have some common immunological mechanisms with mammals, they have also evolved some distinct strategies and, until now, it has not been studied whether birds may also share with mammals similar INPs. Based on corticosterone levels we determined the existence of two divergent groups in Coturnix coturnix that also differed in other immune-neuroendocrine responses. Quail with lowest corticosterone showed higher lymphoproliferative and antibody responses, interferon-γ and interleukin-1β mRNA expression levels and lower frequencies of leukocyte subpopulations distribution and interleukin-13 levels, than their higher corticosterone counterparts. Results suggest the existence of INPs in birds, comparable to mammalian LEWIS/FISCHER profiles, where basal corticosterone also underlies responses of comparable variables associated to the phenotypes. Concluding, INP may not be a mammalian distinct feature, leading to discuss whether these profiles represent a parallel phenomenon evolved in birds and mammals, or a common feature inherited from a reptilian ancestor millions of years ago.

Fig 1. Exploration of data variability.

Principal Component Analysis Bi-plot graph. Each dot represents an animal in the study and triangles represent the explanatory variables used in the analysis. Full white dots (○) and full black dots (●) represent extreme low and high CORT birds respectively. The eigenvalues of each PC are shown in brackets next to each component.

Fig 2. Determination of divergent basal CORT level groups.

Low and High CORT animals grouped based on their CORT level. Birds belonging to the High CORT group have 2.7-fold higher CORT concentrations than their Low CORT counterparts. Data are means ± SE. Different letters indicate significant (p < 0.05) differences between groups. Number of birds in the study = 60, number of birds per group = 10.

Fig 3. Immune effector analysis in divergent Low and High CORT groups.

Correlation plots between CORT and lymphoproliferative response to PHA-P, antibody response against SRBC and FLD are presented in panels A, C and E. Effect of the aggrupation dependent on the basal levels of CORT on immune effectors is shown in panels B (lymphoproliferative response to PHA-P), D (antibody response against SRBC) and F (FLD). Data are means (number inside bars) ± SE. Number of birds per group = 10. Different letters indicate significant (p < 0.05) differences between groups. FLD number was calculated using the following formula: FLD = number of granulocytes/(number of lymphocytes + number of monocytes).

Fig 4. Analysis of molecular mediators in divergent Low and High CORT groups.

Effect of the aggrupation dependent on the basal levels of CORT on the expression of two pro-inflammatory (IFN-γ and IL-1β, panels A and B) and two anti-inflammatory mediators (IL-13 and 4, panels C and D). Data are adjusted means (number inside bars) ± SE. Number of birds per group = 10. Different letters indicate significant (p < 0.05) differences between groups. The relative expression of each target gene was calculated using the formula: Gene Level = 2^{-(Target Gene Ct – β-Actin Ct)}. The value obtained was then multiplied by 1x10⁸ in order to fit the scale of the graphs.

Fig 5. Schematic conceptual representation of INPs in Coturnix coturnix.

The variables set to determine the existence of avian INPs in the present study are represented around each bird. The size of the variable indicates if the animals show high or low response in each of the parameters in the INPs. LYMPH: lymphoproliferative response to PHA-P; Ab SRBC: antibody response against SRBC; FLD: frequency of leukocyte distribution; level of expression of mediators: IFN-γ and IL-1β (pro-inflammatory); and IL-4 and 13 (anti-inflammatory). "Fischer-like" quail with high CORT levels also manifest high FLD and IL-13, but low LYMPH, Ab SRBC, IFN-γ and IL-1β levels. "Lewis-like" counterparts have low CORT as well as low FLD and IL-13 responses, but high LYMPH, Ab SRBC, IFN-γ and IL-1β responses. These two extreme groups of birds do not differ in their IL-4 level.