Redox status regulates eggshell color by modulating protoporphyrin IX biosynthesis via the SIRT1/PGC-1α/ALAS1 axis in brown-shelled hens.

BACKGROUND: This study investigated the molecular mechanisms by which redox status regulates protoporphyrin IX (PpIX) biosynthesis and eggshell coloration in brown-shelled laying hens. This study consisted of two experiments involving 48 and 32 healthy 60-week-old Hy-Line Brown hens, respectively. The hens exhibited either dark (L* = 51.99 ± 2.08) or light (L* = 64.12 ± 3.02) brown eggshell colors. In Exp. 1, light brown-shelled hens were fed a basal diet (Lb group), while dark brown-shelled hens received either a basal diet (Db group) or a basal diet with 10 mg/kg ammonium metavanadate (Dbv group) for 20 d. In Exp. 2, light brown-shelled hens received either a basal diet (Lbc group) or a basal diet supplemented with 200 mg/kg resveratrol (Lbr group) for 12 weeks. RESULTS: Compared to the Db group, eggshell L* values increased, and PpIX concentrations in both eggshell and uterus decreased in Dbv and Lb groups. These groups also showed oxidative stress, as indicated by reduced hepatic T-SOD and CAT activities. Uterine redox status changes were further confirmed by increased T-AOC level (Dbv) and reduced CAT gene expression (Lb). These redox disturbances led to reduced expression of ND4 and COX1 mtDNA, decreased ATP production and CS activity, along with upregulation of IR, PI3K, HK, and PK gene expression, reflecting altered mitochondrial energy metabolism. Notably, the SIRT1/PGC-1α signaling cascade and its downstream target ALAS1 were significantly downregulated at both mRNA and protein levels in Dbv and Lb groups. Compared to the Lbc group, the Lbr group exhibited higher antioxidant capacity by increasing hepatic CAT activity and uterine T-SOD and GSH-Px activities, and reducing MDA levels. Moreover, the Lbr group restored mitochondrial function and PpIX biosynthesis by upregulating ND4 and COX1 mtDNA, CS and SDHA gene expression, and SIRT1/PGC-1α/ALAS1 signaling, while downregulating LDH activity and the expression of IR and PI3K, thereby alleviating eggshell color fading. CONCLUSION: Oxidative stress induces eggshell depigmentation by impairing mitochondrial function and downregulating the SIRT1/PGC-1α/ALAS1 pathway, leading to reduced PpIX biosynthesis. Specifically, vanadium-induced or endogenous oxidative stress disrupts mitochondrial energy metabolism and suppresses key components of this pathway, while resveratrol alleviates oxidative damage and restores mitochondrial function and ALAS1-driven PpIX synthesis through reactivation of the SIRT1/PGC-1α axis.