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. 2024 Dec 9;15(12):1585.
doi: 10.3390/genes15121585.

Immune-Related Gene Expression Responses to In Ovo Stimulation and LPS Challenge in Two Distinct Chicken Genotypes

Affiliations

Immune-Related Gene Expression Responses to In Ovo Stimulation and LPS Challenge in Two Distinct Chicken Genotypes

Anna Slawinska et al. Genes (Basel). .

Abstract

Background: In ovo stimulation introduces bioactive compounds, such as prebiotics, probiotics, or synbiotics into incubating eggs to enhance gut health and immune system development in chickens. This study aimed to determine the genetic and environmental effects modulating responses to in ovo stimulation in commercial broilers and Green-legged Partridge-like (GP) native chickens.

Methods: Eggs were stimulated on day 12 of incubation with prebiotics (GOS-galactooligosaccharides), probiotics (Lactococcus lactis subsp. cremoris), or synbiotics (GOS + L. lactis), with controls being mock-injected. Hatched chicks were reared in group pens and challenged with lipopolysaccharide (LPS) on day 42 post-hatching. Cecal tonsils (CT) and spleens were harvested 2 h post-challenge. RT-qPCR was used to analyze the relative gene expression of cytokine genes: IL-1β, IL-2, IL-4, IL-6, IL-10, IL-12p40, and IL-17.

Results: The results show that genotype influenced the expression of all immune-related genes, with broiler chickens exhibiting stronger innate inflammatory responses than native chickens. LPS induced both mucosal (CT) and systemic (spleen) immune responses in broilers but only systemic (spleen) responses in native chickens.

Conclusions: In ovo stimulation had less of an impact on cytokine gene expression than LPS challenge. Broilers expressed higher inflammatory immune responses than GP native chickens.

Keywords: broilers; cytokine; gene expression; mucosal immune responses; native chickens; systemic.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Gene expression responses to in ovo stimulation and LPS challenge determined in the CT and spleen of broiler chickens. In ovo stimulation was carried out on day 12 of egg incubation using three bioactive compounds: prebiotic (GOS—galactooligosaccharides), probiotic (L. lactis) and synbiotic (GOS + L. lactis). Controls were mock-injected. Hatched chicks were reared in the group pens. On day 42 post-hatching, immune challenge was applied (intraperitoneal injection of LPS—lipopolysaccharides). Controls were mock-injected. Samples of CT were harvested 2 h post-challenge for relative gene expression analysis of cytokine genes. Relative gene expression was performed using RT-qPCR and SYBR green chemistry. Cytokine genes IL-1β, IL-2, IL-4, IL-6, IL-10, IL-12p40, and IL-17 were the target genes. ACTB and UB were used as reference genes. Calculations were based on the ΔΔCt method. Down-regulated data were transformed using the formula 2/-FC. Significance levels: p < 0.05 (*) and p < 0.001 (***). Graph prepared with GraphPad Prism 7 (GraphPad, La Jolla, CA, USA).
Figure 2
Figure 2
Gene expression responses to in ovo stimulation and LPS challenge determined in the CT and spleen of native chickens. In ovo stimulation was carried out on day 12 of egg incubation using three bioactive compounds: prebiotic (GOS—galactooligosaccharides), probiotic (L. lactis) and synbiotic (GOS + L. lactis). Controls were mock-injected. Hatched chicks were reared in group pens. On day 42 post-hatching, immune challenge was applied (intraperitoneal injection of LPS—lipopolysaccharides). Controls were mock-injected. Samples of CT were harvested 2 h post-challenge for the relative gene expression analysis of cytokine genes. Relative gene expression performed using RT-qPCR and SYBR green chemistry. Cytokine genes IL-1β, IL-2, IL-4, IL-6, IL-10, IL-12p40, and IL-17 were target genes. ACTB and UB were used as reference genes. Calculatio were based on the ΔΔCt method. Down-regulated data were transformed using the formula 2/-FC. Significance levels: p < 0.05 (*), p < 0.01 (**) and p < 0.001 (***). Graph prepared with GraphPad Prism 7 (GraphPad, La Jolla, CA, USA).
Figure 3
Figure 3
Experimental outline. The two trials were conducted on broiler and native chickens. The eggs were set in an incubator and on day 12 of incubation the bioactive compounds were injected into the air cell. C—physiological saline (control), PRE—galactooligosaccarides (3.5 mg/egg), PRO—L. lactis subsp. cremoris (105 CFU/egg), SYN (combination of PRE and PRO). On day 42 of rearing, the chickens were immunized with lipopolysaccharide (LPS) or mock-immunized with physiological saline. Two hours post-immunization, all chickens were slaughtered and two tissues (CT and spleen) were harvested for immune-related gene expression analysis.

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References

    1. Siwek M., SlawinsNSka A., Stadnicka K., Bogucka J., Dunislawska A., Bednarczyk M. Prebiotics and synbiotics—In ovo delivery for improved lifespan condition in chicken. BMC Vet. Res. 2018;14:402. doi: 10.1186/s12917-018-1738-z. - DOI - PMC - PubMed
    1. Olah I., Nagy N., Vervelde L. Avian Immunology. Academic Press; Cambridge, MA, USA: 2014. Structure of the Avian Lymphoid System; pp. 11–44.
    1. Smith A.L., Powers C., Beal R.K. Avian Immunology. Academic Press; Cambridge, MA, USA: 2014. The Avian Enteric Immune System in Health and Disease; pp. 227–250.
    1. Koenen M.E., Boonstra-Blom A.G., Jeurissen S.H.M. Immunological differences between layer- and broiler-type chickens. Vet. Immunol. Immunopathol. 2002;89:47–56. doi: 10.1016/S0165-2427(02)00169-1. - DOI - PubMed
    1. Simon K., Arts J.A.J., de Vries Reilingh G., Kemp B., Lammers A. Effects of early life dextran sulfate sodium administration on pathology and immune response in broilers and layers. Poult. Sci. 2016;95:1529–1542. doi: 10.3382/ps/pew074. - DOI - PubMed

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