H. S. Siegel

Stress, strains and resistance 1

1. Stress describes the birds defence mechanisms and a stressor is the situation that elicits the defence response. 2. As the environment can be viewed as a composite of interacting stressors, the birds success in coping with its environment depends on the severity of the stressors and the physiological ability to respond properly and thus maintain homeostasis. 3. The neural, endocrine and more recently immune systems are considered to be integrators of the stress response. Although stress responses may be necessary for survival in wild bird populations, they are often detrimental to efficient growth, skeletal integrity and disease resistance in domesticated fowl. 4. Stress responses are modified by the genetic components.

Effect of genetic selection and MHC haplotypes on lymphocyte proliferation and Interleukin-2 like activity in chicken lines selected for high and low antibody production against sheep red blood cells

Chickens from third generation matings of lines of chickens selected for high (HA) and low (LA) antibody production to sheep red blood cells (SRBC) and typed for MHC genotypes B13/13, B13/21, and B21/21 were used in this study. Chickens from both lines carried all the three genotypes B13/13, B13/21, and B21/21. To study T- and B-lymphocytes mitogenic activity, 12-week-old female chickens were injected intravenously with 0.2 ml of 9% SRBC and spleens were collected at 0, 6 h, and 6 day post-antigen injection (pAg). Isolated lymphocytes were incubated with either Concanavalin-A (Con-A) for T-cell activity, or Pokeweed mitogen (PWM) for B-cell activity and thymidine 3H uptakes were measured. To study the Interleukin-2 (IL-2)-like activity in the same lines and genotypes, splenic lymphocytes from 12-week-old chickens were passed through nylon wool columns to enrich the T-cell population. After a 24 h incubation with Con-A, the conditioned media (CM) were collected. The CM were tested for IL-2 like activity by determining whether they altered the proliferation of Con-A stimulated T cells. This proliferation effect was then compared to that of a reference conditioned media (RCM) prepared from K-strain birds and that were used as the standard for the assay. There was no significant difference (p > 0.05) in IL-2 like activity between HA and LA lines, however, the LA was significantly higher than HA (p < 0.05) in T- and B-cell mitogenic activity. The genotype B13/13 had significantly higher (p < 0.05) IL-2 like activity than the B21/21. The genotype B13/13 was also significantly higher (p < 0.05) in T- and B-cell mitogenic activity than the B21/21. At 0 h, pAg T- and B-mitogenic activity was significantly higher (p < 0.05) than 6 h. In summary, our results indicate that although the birds were selected for high antibody production to SRBC, their lymphocyte mitogenic activity was lower than those selected for low antibody production. Hence, humoral and cell-mediated immune responses appear to be under different genetic controls, and that selection for greater humoral response may be at the expense of cellular responses. Our results also suggest differences in IL-2 like activity production between chickens carrying different MHC B-haplotypes, and that genetic control of such activity is possibly linked to the MHC genes.

Total Cholesterol, Total Triglycerides, and Cholesterol Distribution Among Lipoproteins as Predictors of Atherosclerosis in Selected Lines of Japanese Quail

The proportions of plasma high and low density lipoprotein cholesterol have been linked to inherited tendency for atherosclerosis in humans. Studies were conducted with Japanese quail males from lines genetically selected for high and low TC and a randombred (unselected) control line that were fed 0.0 or 0.5% cholesterol for 12 weeks. Atherosclerotic plaques were more severe in the high than in the low line quail and in those fed cholesterol compared to non-cholesterol-fed quail. Serum TG, TC, VLDLC, LDLC, and HDLC were also higher in the high than in the low line quail and in cholesterol-fed vs. non-cholesterol-fed quail. Significant interactions indicated that TC and LDLC concentrations were more affected by dietary cholesterol in the high line than in the low line. The low line quail maintained higher HDLC and lower LDLC than the high line. Regression and correlation analyses revealed that although VLDLC, LDLC, and TC were significant predictors of atherosclerosis in the high line birds, the TC/HDLC ratio was a better predictor in the low line. The Japanese quail lines used herein represent useful experimental models for studies of genetic differences in atherosclerosis in humans.

Changes in the profiles of circulating white blood cells, corticosterone, T3 and T4 during the initiation of humoral immunity in immature male chickens

Eight-week-old male White Leghorn chickens were injected intravenously with either 0.1 ml of Brucella abortus (BA) or saline. Blood samples were collected before injection, 1 hr, 3 hr, 6 hr, 12 hr, 24 hr, and then daily for the following 6 days after the injection. Total white blood cells (WBC), differential counts and corticosterone (CS), T3 and T4 levels were measured in all blood samples. These data represented the results obtained from the primary response. To obtain data for the secondary response, the same birds were injected 4 weeks following the first challenge and the same procedure was followed. During primary and secondary responses, circulating lymphocytes and monocytes decreased significantly 3 hr following BA injection and then increased steadily and reached a peak approximately 5 days post-BA injection, however, circulating heterophils increased significantly 6-12 hr and returned to normal levels 2 days post-BA injection. Serum corticosterone increased significantly 3 hr following BA injection. Serum T3 decreased significantly 3 to 12 hr then increased significantly 2 days following BA injection. There were no significant changes in any of the measurements following saline injection. Correlations between circulating hormones and white blood cells were calculated and are reported. These results indicate that an early step in the initiation of humoral immunity is probably an increase in serum CS. This increase in CS is followed by a decrease in circulating lymphocyte and monocyte numbers. These decreases may be the result of lymphocyte trapping in secondary lymphoid organs. This series of events may occur to modulate and activate the immune response.

Changes in blood and spleen lymphocyte populations following antigen challenge in immature male chickens

1. The effects of antigen (Ag) injection on the distribution of lymphocyte populations of Cornell K-strain male chickens were studied. 2. Two experiments were conducted. In the first, chickens were injected with Brucella abortus (BA), a purported T-independent antigen. In the second, chickens were injected with sheep red blood cells (SRBC), a T-dependent antigen. Peripheral blood lymphocytes (PBL) and spleen lymphocytes isolated at 0, 3, 6, 9, 12 and 24 h following Ag injection were stained with monoclonal antibodies (mAb) detecting B-lymphocytes, CD4+ and CD8+ cells. 3. B-lymphocytes in the blood or spleen showed no significant changes following either BA or SRBC injection. In contrast, CD4+ cells were decreased in the blood and increased in the spleen following BA and SRBC injections. CD8+ cells were decreased in both blood and spleen following BA injection but were unchanged in either blood or the spleen following SRBC injection. 4. These results indicate that there is a change in both spleen and circulating lymphocyte populations, especially T-helper cells, following Ag injection. T-helper cells are apparently the primary population involved in the initiation of humoral immunity.

Ovine Corticotropin-Releasing Factor Increases Endocrine and Immunologic Activity of Avian Leukocytes In Vitro

Abstract Treatment of splenic leukocytes from Cornell K strain male chickens (homozygous at the B 15 locus of the major histocompatibility complex) with ovine corticotropin-releasing factor (oCRF), before their co-incubation with naive chicken adrenal cells, resulted in an increase in corticosterone production. Supernatants from the oCRF-treated splenic leukocytes caused a time-dependent increase in corticosterone production when incubated with chicken adrenal cells. Adding oCRF directly to chicken adrenal cells did not increase corticosterone production. Pretreatment of peripheral leukocytes with oCRF increased their activity in a concanavalin A mitogen assay. Thus, chicken leukocytes stimulated with corticotropin releasing factor appear to increase the production of an “adrenocorticotropin-like” substance (adrenocorticotropin-like because it increases corticosterone production by adrenal cells), and increased their cell-mediated immune activity.

Effects of corticosterone in vivo and in vitro on adrenocorticotropic hormone production by corticotropin releasing factor-stimulated leukocytes.

Abstract Corticotropin releasing factor (CRF) and corticosterone have been shown to affect immune cell function. Previously, we have shown that CRF stimulates immunoreactive adrenocorticotropic hormone (ACTH) production by leukocytes. In this study, splenic leukocytes from corticosterone-injected chickens failed to show a CRF-induced increase in ACTH production. In addition, corticosterone in vitro inhibited the production of leukocyte ACTH as well as the stimulatory effect of CRF on splenic leukocyte ACTH production. These findings show that, as with anterior pituitary ACTH production, CRF-stimulated leukocyte ACTH production is inhibited by glucocorticoids.

Dietary Cholesterol Metabolism in Japanese Quail Lines Selected for Plasma Cholesterol Levels

Abstract Dietary cholesterol metabolism was studied, using a single dose of emulsion, per os (test meal), in lines of Japanese quail that were divergently selected for high (HL) and low (LL) plasma total cholesterol. The meal contained [3H] cholesterol, [14C] β-sitosterol, unlabeled cholesterol, triolein, and bile salt. Recovery of the non-absorbable β-sitosterol in the excreta permitted determination of the percentage of cholesterol absorbed. The amounts of [3H] in the plasma, egg yolks, and the excreta neutral and acid sterols were determined. A line-x-time interaction for [3H] in plasma indicated that the level of plasma cholesterol derived from the test meal declined more rapidly in the LL than in the HL. The higher [3H] detected in the excreta acidic sterols of the LL 12 hr after the test meal indicated that bile acid excretion of cholesterol was greater in the LL than in the HL. There were no differences in cholesterol absorption between lines or sexes. Cumulative [3H] radioactivity in the eggs over 18 days following the test meal was higher in the HL yolks; however, this line effect was due to the grater number of eggs produced by the HL. Thus, one of the mechanisms by which the LL maintains low plasma cholesterol levels is by an enhanced excretion of bile acid compared with the HL. The data also suggest that the more severe atherogenic effect of dietary cholesterol observed in the HL could be, in part, due to the longer residence time of cholesterol in circulation.

Instrumentation for Measuring Continuous Activity, Feed and Water Consumption, and Growth Rate of Chickens

ABSTRACT STRAIN-GAGE, load beams and solid-state circuitry were designed and used to measure, accumulate, and record locomotor activity, feed and water consumption, and growth rate of undisturbed chickens from 1 day to 7 wk of age in six small isolation cabinets. Static loads such as weights of the feed, water, and floor were measured with cantilever beams—each instrumented with four active strain gages. Activity was measured with a circuit whose output was proportional to the frequency of floor vibration when the vibration amplitude was above a preset adjustable reference level. The load beams and circuitry were used successfully over a period of several weeks to measure the above variables. The resolutions of the beam and circuit configurations used were 1.6 g to 2.9 g for the floor beams—depending on the position of the load, 1.6 g for the feed beam and 0.8 g for the water beam. The activity circuit permitted the detection of motion from as few as 1 or 2 1-day old chicks at the highest sensitivity, and was suitable for measuring the activity of several chicks up to 7 wk of age at a medium sensitivity.

Epitopes for chicken monoclonal antibodies in spleens of selected Japanese quail lines.

A line of Japanese quail selected for high plasma cholesterol is highly susceptible to diet-induced atherosclerosis. Lymphocyte epitopes recognized by mouse anti-chicken monoclonal antibodies (c-mAb), TCR-1, TCR-2, TCR-3. CD-3, CD-4, CD-8, and BU-1a/b were reacted with spleens from quail selected for high (HL) and low (LL) plasma total cholesterol and their nonselected controls (CL). Cross reactivity to c-mAb and effect of line and gender were immunohistochemically evaluated. Chicken spleens were positive controls. Quail were immunologically stimulated with either sheep red blood cells (SRBC) or Brucella abortus 2 weeks before spleens were removed. Quail spleen epitopes of all lines recognized TCR-3 and CD-8 c-mAb, but no other c-mAb. Number of reacting cells and staining intensity to the TCR-3 c-mAb were greater in the HL than in the LL regardless of the stimulating Ag or dose used. For the CD-8 c-mAb, there were no differences among lines in birds receiving SRBC. In B. abortus-immunized birds, sex x line interactions indicated that males of the HL and CL had lower responses than females but LL males were not different than females. TCR-3 and CD8 c-mAb may be useful in studying immunological mechanisms for atherosclerosis in Japanese quail.