Supplementation of a Protected Complex of Biofactors and Antioxidants on the Gene Expression and Kinomics in Broiler Chickens Undergoing Early Life Stress

Early life stress may have a negative impact on the performance of broiler flocks. It has been observed that a severe cold stress (12°C below the normal temperature) throughout the life cycle of broilers negatively impacts production parameters (Su et al., 2020) and immune system development (Zhao et al., 2013). The use of vitamins and other molecules with antioxidant properties have been investigated in poultry undergoing different challenge models (Ghazi Harsini et al., 2012; Hu et al., 2020). An imbalance between the pro-oxidant and antioxidant systems of the body is likely to drive damage to cellular components (Estévez, 2015), including immune cells, that may impair immune responses. The dietary supplementation of antioxidants and vitamins may improve growth performance and has the function of protecting the cells from reactive oxygen species (ROS; Ghazi Harsini et al., 2012) that are naturally generated due to the immune response or exacerbated in stress situations. Additionally, the supplementation of antioxidant vitamins and biofactors may induce immune-metabolic alterations in different tissues that could help the birds to cope with early life stressors. We hypothesised that the supplementation of different antioxidant molecules through feed would alleviate the negative effects of early life stress in broiler chickens by modulating the immune and antioxidant systems. Therefore, we evaluated the dietary supplementation of a protected complex of biofactors and antioxidants P(BF+AOx), which is a protected source of vitamins and fermentation extract, on the expression of immune-related genes, and immunometabolic phenotypic changes in the intestine and liver of broilers submitted to early life stressors simulated by the combination of a double Infectious Bronchitis (IB) vaccination followed by a temperature stress in their early life.

The experiment consisted of two treatments: feed supplemented with or without P(BF+AOx) (Jefo Nutrition Inc., Canada) from 1 to 14 d of age, distributed to a total of 720 one-day old male Ross 308 chickens placed in pens of 30 birds (12 replicates per treatment). When the period of supplementation ended (d 14), all the birds were fed the control diet (without supplementation) until d 35. The P(BF+AOx) is a complex of vitamins and fermentation extract microencapsulated in a matrix of triglycerides from hydrogenated vegetable oil (Jefo Nutrition Inc., Canada). Birds were double vaccinated against IB (MILDVAC-Ma5^TM) by spray at the hatchery (d 0) and submitted, on d 3, to an acute cold stress with a temperature drop from 32°C to 20-23°C during 48 h. Feed intake (FI), body weight gain (BWG), and feed conversion ratio (FCR) were calculated weekly. On d 7 and 15, one bird/pen was euthanised, and samples of jejunum, ileum, and liver were collected for expression of immune-related genes and kinome array analysis (Kogut and Arsenault, 2015). All the data were analyzed by ANOVA using the software SAS (SAS 9.4; P < 0.05).

The dietary supplementation of P(BF+AOx) improved the growth performance of the birds on d 21, 28, and 35 (Bortoluzzi et al., 2020). Additionally, on day 7, it was observed that the dietary supplementation upregulated the expression of IL-6 (P = 0.03) in the liver. On d 15, the dietary supplementation upregulated the expression of IL-6 in the ileum (P = 0.04) and IL-10 in the liver (P = 0.001) and tended to upregulate IL-6 in the liver (P = 0.08). The results of the kinome peptide array performed on jejunum and liver tissues showed that the treatment had significant effect on oxidative stress resistance on d 15. Catalase was activated via decreased phosphorylation, and the phosphorylation of immunoregulatory or proinflammatory proteins was decreased. We also observed that the supplementation promoted a decrease in the activation of proinflammatory proteins in the FOXO (Forkhead box protein O) pathway, including the transcriptional regulator nuclear factor-kB (NF- kB), and increased phosphorylation of IL-6R and TGF-beta receptor on their active sites. These results, along with increased IL-10 expression in the liver, may be an indication of reduced inflammation or antiinflammatory responses. In conclusion, the supplementation of P(BF+AOx) improved the growth performance of broiler chickens undergoing early life stress. Further analyses presented herein demonstrated that this novel feed additive was beneficial in modulating the immune and antioxidant defense systems of the birds. Overall, the kinome data functionally agreed with the gene expression and antioxidant results and indicate a general anti-inflammatory and antioxidant response in birds supplemented with P(BF+AOx).