1- Department of Animal Science, Faculty of Animal Sciences and Fisheries, Sari Agricultural Sciences and Natural Resources University, Sari, Iran
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Extended Abstract
Background: Generally, respiratory infections, environmental stressors, chronic heat stress, intestinal parasitic infections, nutritional imbalances, liver diseases, and increased ovulation frequency and microbial invasion can contribute to chronic ovarian inflammation. The severity of ovarian inflammation tends to increase with age in laying hens. Chronic inflammation is defined as a prolonged and sustained host response to inflammatory stimuli. Microstructural changes in ovarian follicles, along with increased inflammation and oxidative stress, have been reported with advancing age in laying hens. Age-related inflammatory complications in broiler breeder hens may impair reproductive performance, egg quality, and metabolic status through imbalances between pro-inflammatory genes (such as IL-1β, IL-6, TNF-α, and COX-2) and anti-inflammatory genes (including IL-10 and TGF-β). In recent years, non-steroidal strategies, such as willow bark extract (containing salicin), propolis, and the selective COX-2 inhibitor celecoxib, have attracted attention for reducing inflammation and modulating the expression of inflammation-related genes. Salix extract and some of its constituents possess potent anti-inflammatory, anti-proliferative, and antimicrobial properties. Their adverse effects appear to be less pronounced than those associated with conventional nonsteroidal anti-inflammatory drugs (NSAIDs), including aspirin. Propolis is a resinous substance collected by worker honeybees from plant exudates and mixed with wax and other components. In general, propolis exerts anti-inflammatory effects by suppressing and regulating pro-inflammatory cytokines such as IL-1β and IL-6. On the other hand, celecoxib selectively inhibits cyclooxygenase-2 (COX-2), thereby reducing prostaglandin synthesis and modulating the expression pattern of inflammatory genes toward the attenuation of detrimental inflammatory responses. Nonsteroidal anti-inflammatory drugs are widely used, and the lower gastrointestinal toxicity of coxibs represents a potentially important advantage over non-selective NSAIDs. This study aimed to evaluate the effectiveness of these three strategies in promoting anti-inflammatory factors and controlling inflammation during the late laying period of broiler breeder hens.
Methods: The experiment was conducted on 80 hens and 10 roosters of the Ross strain (52 weeks old; average hen body weight: 3860 g) for 5 weeks, including 1 week of adaptation and 4 weeks of treatments. The study was carried out in the poultry breeding hall of Sari Agricultural Sciences and Natural Resources University using a completely randomized design with four treatments and four replicates. The treatments included 1) a control group receiving a basal diet, 2) a group receiving propolis alcoholic extract at 100 mg/kg metabolic body weight per hen per day, administered orally, 3) a group receiving celecoxib at 15 mg/kg metabolic body weight per hen per day, administered orally, and 4) a group receiving alcoholic willow bark extract at 150 mg/kg metabolic body weight per hen per day, administered orally. Powdered propolis extract and willow bark extract, together with celecoxib, were incorporated into the basal diet on a daily basis to minimize oxidation. Furthermore, the hens were maintained under a photoperiod of 14 h light and 10 h dark throughout the experimental period. On the final day, 2 mL of blood was collected from the brachial vein of one hen per replicate. The collected samples were immediately transferred to the Physiology Laboratory under cold-chain conditions and stored at −80°C until gene expression analysis. The evaluated genes included IGFBP2, IL-1, IL-6, IL-10, IGF-1, IGF-2, IGF-1R, and TNF-α. Relative gene expression was measured using Real-Time PCR with gene-specific primers. The threshold cycle (Ct) value was determined for each sample using a Real-Time PCR system. Amplification reactions were performed using specific primers and corresponding kits (Sinaclon and Yekta Tajhiz Co.) in a thermal cycler equipped with an optical detection system. The expression of the GAPDH gene was used as the reference (housekeeping) gene. The obtained data were entered into Microsoft Excel for the calculation of relative gene expression levels. Relative gene expression was initially analyzed using the 2^−ΔΔCt method. Statistical analyses were performed using the General Linear Model (GLM) procedure of SAS software (version 9.1). Mean comparisons were carried out using Duncan’s multiple range test at a 5% significance level.
Results: Willow bark extract significantly increased the relative expression of IL-6 and IL-10 (P < 0.05). Propolis extract significantly increased IGF-1 expression compared with the other treatments (P < 0.05). None of the treatments, including the extracts and celecoxib, significantly affected on the expression of IGF-1R, IGF-2, or IGFBP2 compared with the control group.
Conclusion: The findings of the present study indicate that willow bark extract may serve as a promising natural anti-inflammatory agent for alleviating age-related inflammatory responses in broiler breeder hens during the late laying period. However, further investigations involving different dosage levels, longer supplementation periods, and evaluations across various production stages are required to confirm its efficacy and establish optimal application protocols. Given the advanced age of the hens, the elevated baseline inflammatory status, and the presence of environmental stressors, the administered doses may have been insufficient to elicit significant changes in the expression of most insulin-like growth factor-related and pro-inflammatory genes. Therefore, higher supplementation levels or longer treatment durations may be necessary to achieve measurable effects on these molecular markers.
Background: Generally, respiratory infections, environmental stressors, chronic heat stress, intestinal parasitic infections, nutritional imbalances, liver diseases, and increased ovulation frequency and microbial invasion can contribute to chronic ovarian inflammation. The severity of ovarian inflammation tends to increase with age in laying hens. Chronic inflammation is defined as a prolonged and sustained host response to inflammatory stimuli. Microstructural changes in ovarian follicles, along with increased inflammation and oxidative stress, have been reported with advancing age in laying hens. Age-related inflammatory complications in broiler breeder hens may impair reproductive performance, egg quality, and metabolic status through imbalances between pro-inflammatory genes (such as IL-1β, IL-6, TNF-α, and COX-2) and anti-inflammatory genes (including IL-10 and TGF-β). In recent years, non-steroidal strategies, such as willow bark extract (containing salicin), propolis, and the selective COX-2 inhibitor celecoxib, have attracted attention for reducing inflammation and modulating the expression of inflammation-related genes. Salix extract and some of its constituents possess potent anti-inflammatory, anti-proliferative, and antimicrobial properties. Their adverse effects appear to be less pronounced than those associated with conventional nonsteroidal anti-inflammatory drugs (NSAIDs), including aspirin. Propolis is a resinous substance collected by worker honeybees from plant exudates and mixed with wax and other components. In general, propolis exerts anti-inflammatory effects by suppressing and regulating pro-inflammatory cytokines such as IL-1β and IL-6. On the other hand, celecoxib selectively inhibits cyclooxygenase-2 (COX-2), thereby reducing prostaglandin synthesis and modulating the expression pattern of inflammatory genes toward the attenuation of detrimental inflammatory responses. Nonsteroidal anti-inflammatory drugs are widely used, and the lower gastrointestinal toxicity of coxibs represents a potentially important advantage over non-selective NSAIDs. This study aimed to evaluate the effectiveness of these three strategies in promoting anti-inflammatory factors and controlling inflammation during the late laying period of broiler breeder hens.
Results: Willow bark extract significantly increased the relative expression of IL-6 and IL-10 (P < 0.05). Propolis extract significantly increased IGF-1 expression compared with the other treatments (P < 0.05). None of the treatments, including the extracts and celecoxib, significantly affected on the expression of IGF-1R, IGF-2, or IGFBP2 compared with the control group.
Conclusion: The findings of the present study indicate that willow bark extract may serve as a promising natural anti-inflammatory agent for alleviating age-related inflammatory responses in broiler breeder hens during the late laying period. However, further investigations involving different dosage levels, longer supplementation periods, and evaluations across various production stages are required to confirm its efficacy and establish optimal application protocols. Given the advanced age of the hens, the elevated baseline inflammatory status, and the presence of environmental stressors, the administered doses may have been insufficient to elicit significant changes in the expression of most insulin-like growth factor-related and pro-inflammatory genes. Therefore, higher supplementation levels or longer treatment durations may be necessary to achieve measurable effects on these molecular markers.
Keywords: Anti-inflammatory, Broiler breeder, Gene expression, Insulin-like growth factor, Pro-inflammatory
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