Influence of dietary vitamin E on phagocytic functions of macrophages in broilers.

Vitamin E (VE) is known for its antioxidant properties and has been shown to modulate immune system functions in various species. This study examined the influence of different levels of dietary VE (alpha-tocopherol acetate) on phagocytic functions of macrophages (abdominal exudate cells) in broiler chickens at 3, 5, and 7 wk. Birds were fed commercial diets containing 16 (control), 110, or 220 mg of VE/kg of feed. Macrophages were elicited into the abdominal cavity by injecting a 3% Sephadex solution prepared in PBS (G50-50, 1 mL/100 g of BW) 42 h prior to harvest. The percentage of phagocytically active macrophages and the number of SRBC phagocytosed per macrophage for unopsonized and antibody-opsonized SRBC were determined. These aspects of macrophage function were assessed based on 900 macrophages per sample. When unopsonized SRBC were used, dietary VE supplementation above control level did not affect phagocytic function of macrophages at wk 3, 5, or 7. With antibody-opsonized SRBC, the percentage of phagocytically active macrophages and the number of SRBC phagocytosed per macrophage were higher (P = 0.08 and P = 0.01, respectively) in 3-wk-old birds fed 110 and 220 mg of VE/kg of feed compared with age-matched controls. This enhancing effect of VE supplementation on macrophage function was not observed in 5- and 7-wk-old broilers. It appears from this study that supplemental VE enhances Fc-receptor-mediated macrophage phagocytic activity at early stages of broiler growth.

Pulmonary and hematological inflammatory responses to intravenous cellulose micro-particles in broilers.

When injected intravenously, cellulose micro-particles become lodged in pulmonary arterioles. The current study investigated the systemic and pulmonary inflammatory responses triggered by cellulose micro-particles at 3, 24, and 48 h postinjection in 6-wk-old broilers. Proportions and concentrations of circulating white blood cells were assessed in saline-injected (control group) and cellulose-injected (particle group) birds. Hematoxylin-eosin (HE)-stained cross-sections of the lungs were used to count the number of granuloma/lymphocyte aggregates, which is indicative of the severity of the inflammatory response to the trapped particles. The cellular components of the aggregates were identified by immunohistochemical staining of frozen cross sections of the lungs. Results showed that cellulose micro-particles trapped in the pulmonary vasculature initiated a dynamic, localized inflammatory response within the surrounding lung parenchyma. Monocytes and basophilic granulocytes closely surrounded the particles. CD4, CD8, TCR1, TCR2, and TCR3 subsets of T cells and B cells were present in the outer rim of the granuloma/lymphocyte aggregates. Circulating total white blood cell (WBC, leukocytes) concentrations were similar in both groups at all times postinjection, whereas at 48 h post-injection the percentages of eosinophils and basophils among circulating WBC were higher in the particle group than in the control group (P < or = 0.05). The circulating monocyte concentration also increased within 24 h postinjection (P < or = 0.05). These observations demonstrate that cellulose micro-particles trapped in the pulmonary vasculature initiated acute focal inflammatory responses in the lungs and that the proportions of WBCs in the blood are modulated within 48 h postinjection.

Effect of intravenous endotoxin on blood cell profiles of broilers housed in cages and floor litter environments.

Commercial broilers are constantly exposed to airborne microorganisms and endotoxin (lipopolysaccharide, LPS). It has been shown that microbial contamination of the air was higher in broiler houses using floor litter than in broiler houses using netting-type floors. The current study evaluated the effect of housing conditions on blood leukocyte profiles and tested the hypothesis that, when compared to broilers reared in clean stainless steel cages (Cage group), broilers raised on floor litter (Floor group) should experience a higher environmental challenge and have a desensitized immune system that may exhibit better tolerance/resistance to subsequent intravenous LPS challenge. Hematological parameters were evaluated prior to and following i.v. administration of 1 mg/kg BW Salmonella typhimurium LPS (dissolved at 1 mg/0.25 mL in PBS) or i.v. injection of 0.25 mL/kg BW PBS alone. The results showed that prior to LPS/PBS injection, broilers in the cage group had higher heterophil and monocyte concentrations, a higher B cell percentage within the lymphocyte population, and a higher heterophil to lymphocyte (H:L) ratio in the blood. The i.v. LPS injection resulted in 25% mortality in the cage group and 42% mortality in the floor group within 8 h post-injection. LPS reduced the concentrations of total white blood cells (WBC) and all differential WBC except eosinophils and increased thrombocyte concentrations within 1 h post-injection in both groups. All of these values returned to their respective pre-injection levels within 48 h post-injection in the surviving birds. The two groups exhibited similar overall hematological changes after LPS injection except that the cage group showed a higher H:L ratio at 8 h post-injection and a lower B-cell percentage within the lymphocyte population at 48 h post-injection when compared with the floor group. We concluded that the immune systems of broilers reared on floor litter were desensitized and exhibited less pronounced leukocyte responses to i.v. LPS when compared with those of broilers reared in clean stainless steel cages. However, such desensitization of the immune system did not help broilers survive subsequent i.v. LPS challenge.

Herpesvirus connection in the expression of autoimmune vitiligo in Smyth line chickens.

The Smyth line (SL) chicken is an animal model for human vitiligo, a common acquired depigmentary disorder affecting about 1-2% of people worldwide. The vitiligo-like depigmentation in SL chickens typically develops when the birds are between 6 and 14 weeks of age and may affect 70-95% of hatch mates. The development of SL vitiligo is considered to depend on two interacting components, namely an inherent melanocyte defect and an autoimmune reaction to melanocytes. Recently, a role for an environmental factor in the expression of vitiligo was suggested by the observation that only 10% of SL chicks imported from the University of Massachusetts (UM) and reared in isolation at biosecurity level 2 (BSL 2) at the University of Arkansas (UA) exhibited vitiligo. Following further assessment of environmental differences between UA and UM SL chickens, three environmental factors that may have influenced the expression of SL vitiligo were identified. Included were housing condition, status of Mycoplasma synoviae infection, and turkey herpesvirus (HVT) vaccination status. Studies were subsequently conducted at UA and UM to assess the role of these environmental factors in the expression of SL vitiligo. M. synoviae infection was not found necessary for vitiligo expression in SL chickens. However, HVT emerged as a strong candidate for an important environmental factor in SL vitiligo. The connection between HVT and SL vitiligo was confirmed for both BSL 2 and conventional housing. Therefore, the observations reported here suggest a strong causative link between HVT infection and SL vitiligo.

CD4, CD8 and TCR defined T-cell subsets in thymus and spleen of 2- and 7-week old commercial broiler chickens.

To better understand immune development and function in meat-type chickens (broilers), the proportions of T-cells expressing CD4, CD8, and T-cell receptors (TCR) in the thymus and spleen were determined by three-color fluorescence and flow cytometry in 2- and 7-week old broilers raised in commercial growing conditions. Broiler thymocytes consisted of single-(CD4+CD8- and CD4-CD8+) and double-(CD4+CD8+) positive subpopulations. Within these CD4+ and/or CD8+ thymocyte populations, all types of TCR (y delta, V beta 1 alpha beta, and V beta 2 alpha beta) could be identified. In the thymus, percentages of CD4+CD8- cells increased, CD4-CD8+ cells remained unchanged, and CD4+CD8+ cells decreased between 2 and 7 weeks of age. In the spleen, in addition to single-positive lymphocytes, double-positive populations were identified, expressing either y delta or alpha beta TCR. The percentage of CD4+CD8- splenocytes decreased, and the percentages of both CD4-CD8+ and CD4+CD8+ splenocytes increased between 2 and 7 weeks of age. Age-associated shifts in TCR usage (the proportion of cells expressing a certain type of TCR) were observed in the single-positive, but not in the double-positive, T-cell populations of both thymus and spleen. This multiparameter cell population analysis in broilers demonstrates thymic and splenic T-cell subsets similar to those previously described in layers. Differences in the proportions among T-cell subsets between 2- and 7-week old broilers likely reflect a more competent immune system in the older birds.

Effects of dietary vitamin E on the immune system in broilers: altered proportions of CD4 T cells in the thymus and spleen.

To gain insight into the immunomodulatory effects of vitamin E (VE), immune cell population analyses were conducted using thymus and spleen from male broilers fed diets with various levels of VE supplementation (0, 17, 46, and 87 mg dl-alpha-tocopherol acetate/kg of feed). At 2 and 7 wk of age, the percentages of B cells, macrophages, and T cell subsets, delineated by the expression of CD4, CD8, and T cell receptor (TCR) isotype, in thymus and spleen were determined by flow cytometry. The percentages of thymic and splenic B cells and macrophages from 2- and 7-wk-old chickens, as well as the percentage of thymic T cells in 2-wk-old chickens, were unaffected by VE treatment. However, 7-wk-old broilers maintained on 87 mg VE/kg feed had a higher percentage of CD4+CD8- thymocytes, a higher CD4+CD8- to CD4-CD8+ thymocyte ratio, and a lower percentage of CD4+CD8+ thymocytes than chickens receiving no dietary VE supplementation. The VE-induced increase in the percentage of CD4+CD8- thymocytes was due to an increase in the TCR2+CD4+CD8- thymocyte subset, whereas the decrease in the percentage of CD4+CD8+ thymocytes involved all TCR defined T cell subsets. In the spleen, the percentage of CD4+CD8- T cells was lower in 2-wk-old chickens and higher in 7-wk-old chickens maintained on 87 mg/kg feed than in chickens receiving no dietary VE supplementation. The decrease in CD4+CD8- splenocytes at 2 wk of age was due to a decline in the percentage of TCR2+CD4+CD8- splenocytes, whereas the increase in CD4+CD8- splenocytes in 7-wk-old chicks was due to an increase in the percentages of all TCR defined CD4+CD8- T cell subsets. These data support an immunomodulatory effect of VE on CD4+CD8- T cells.

Effect of dietary dl-alpha-tocopherol on tissue alpha- and gamma-tocopherol and pulmonary hypertension syndrome (ascites) in broilers.

The objectives of this experiment were to determine the effects of high dietary levels of vitamin E on growth performance and pulmonary hypertension syndrome (PHS) mortality. Male broiler chicks (Cobb 500) were randomly assigned to one of four dietary treatments consisting of standard starter and grower diets supplemented with 0, 17, 46, and 87 mg dl-alpha-tocopherol acetate/kg. To encourage the development of PHS, air temperature in the house was 32 and 28 C for Weeks 1 and 2, dropped to 18 C during Week 3, and kept between 10 and 15 C during Weeks 4 through 7. Also, chicks were placed in floor pens on litter used for five previous flocks and ventilation reduced to increase dust and ammonia in the house. Ammonia levels increased from an initial 18 to 36 ppm on Day 42 with the increase in ammonia corresponding to an obvious increase in dust in the air. Lung and liver tissue obtained at 2, 5, and 7 wk of age were analyzed for tissue alpha- and gamma-tocopherol by liquid chromatography. Dietary vitamin E had no effect on body weight, feed intake, or feed efficiency. Cumulative PHS mortality through 7 wk of age was 21% and was also unaffected by dietary treatment. Liver and lung alpha-tocopherol concentrations exhibited a dose-response increase to dietary tocopherol and there was a high correlation between lung and liver tissue alpha-tocopherol (r = 0.72, P < 0.05). Whereas gamma-tocopherol concentrations in lung and liver were unaffected by dietary treatment, liver and lung exhibited age-dependent increases in both alpha- and gamma-tocopherol. Despite dose-dependent increases in tissue alpha-tocopherol, supplementation of diets with up to 87 mg dl-alpha-tocopherol acetate had no effect on growth performance or PHS mortality in broilers under the conditions used in this study.