Abstract
The incidence of woody breast (WB) is increasing on a global scale representing a significant welfare problem and economic burden to the poultry industry and for which there is no effective treatment due to its unknown etiology. In this study, using diffuse reflectance spectroscopy (DRS) coupled with iSTAT portable clinical analyzer, we provide evidence that the circulatory- and breast muscle-oxygen homeostasis is dysregulated [low oxygen and hemoglobin (HB) levels] in chickens with WB myopathy compared to healthy counterparts. Molecular analysis showed that blood HB subunit Mu (HBM), Zeta (HBZ), and hephaestin (HEPH) expression were significantly down regulated; however, the expression of the subunit rho of HB beta (HBBR) was upregulated in chicken with WB compared to healthy counterparts. The breast muscle HBBR, HBE, HBZ, and hypoxia-inducible factor prolyl hydroxylase 2 (PHD2) mRNA abundances were significantly down regulated in WB-affected compared to normal birds. The expression of HIF-1α at mRNA and protein levels was significantly induced in breasts of WB-affected compared to unaffected birds confirming a local hypoxic status. The phosphorylated levels of the upstream mediators AKT at Ser473 site, mTOR at Ser2481 site, and PI3K P85 at Tyr458 site, as well as their mRNA levels were significantly increased in breasts of WB-affected birds. In attempt to identify a nutritional strategy to reduce WB incidence, male broiler chicks (Cobb 500, n = 576) were randomly distributed into 48 floor pens and subjected to six treatments (12 birds/pen; 8 pens/treatment): a nutrient adequate control group (PC), the PC supplemented with 0.3% myo-inositol (PC + MI), a negative control (NC) deficient in available P and Ca by 0.15 and 0.16%, respectively, the NC fed with quantum blue (QB) at 500 (NC + 500 FTU), 1,000 (NC + 1,000 FTU), or 2,000 FTU/kg of feed (NC + 2,000 FTU). Although QB-enriched diets did not affect growth performances (FCR and FE), it did reduce the severity of WB by 5% compared to the PC diet. This effect is mediated by reversing the expression profile of oxygen homeostasis-related genes; i.e., significant down regulation of HBBR and upregulation of HBM, HBZ, and HEPH in blood, as well as a significant upregulation of HBA1, HBBR, HBE, HBZ, and PHD2 in breast muscle compared to the positive control.
Highlights
MATERIALS AND METHODSPoultry production supports the livelihoods and food security of billions of people worldwide
It is facing several challenges from a steep projected increase in global demand for high-quality animal proteins and the need to solve the problem associated with high incidence of metabolic disorders such as woody breast (WB) myopathy, which has garnered tremendous attention the last few years
We provide evidence that the circulatory and breast muscle oxygen homeostasis is dysregulated along with the activation of hypoxic signaling pathways in chickens with WB myopathy
Summary
Poultry production supports the livelihoods and food security of billions of people worldwide. Target analysis, bloods were collected in tubes containing TRIzol LS reagent according to manufacturer’s recommendations (Life Technologies Corporation, CA, United States). Total RNA was extracted from chicken blood and breast muscle samples by using TRIzol LS (for blood) and TRIzol (for muscle) reagent (Life Technologies Corporation, NY, United States). For the fluorescent western blot analysis, 100 mg breast muscle tissue was homogenized using an IKA (Germany) T10 ULTRATURRAX R homogenizer, fitted with a S10N-8G-ST probe, in 1 mL ice-cold radioimmunoprecipitaion assay (RIPA) buffer with Pierce phosphatase and protease inhibitors (Life Technology Corporation, NY, United States). The membrane was washed three times with TBST for 10 min incubated with 1/10,000 secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor 790, #ab186697) (Abcam, Cambridge, MA, United States) in 10 mL 5% goat serum in TBST at room temperature for 1 h. If ANOVA revealed significant effects, the means were compared by Tukey’s multiple range test using the Graph Pad Prism version 6.00 for Windows (Graph Pad Software, La Jolla, CA, United States), and differences were considered significant at P < 0.05
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