This study assessed the effects of dietary honeycomb supplementation on laying performance, intestinal inflammation, ovarian function, and yolk precursor synthesis in laying hens. A total of 320 Dawu Golden Phoenix laying hens (288-d-old) were randomly assigned into four treatment groups with eight replicates of 10 hens each. Hens were provided diets containing 0, 0.5, 1.0, or 2.0 g/kg honeycomb for 30 d. Dietary honeycomb significantly increased average egg weight, average daily feed intake, and laying rate, while decreasing the feed conversion ratio (p < 0.05). It enhanced serum antioxidant capacity, as reflected by higher GSH, SOD, and CAT activity (p < 0.05). Meanwhile, honeycomb modulated the expression of Nrf2/Keap1 pathway-related genes, accompanied by increased expression of downstream antioxidant-related genes. Additionally, honeycomb downregulated intestinal inflammatory- and apoptosis-related factors (p < 0.05), which effectively alleviated intestinal inflammation responses. In the liver, honeycomb showed favorable histological changes, including fewer lipid droplets and less vacuolar degeneration. Furthermore, it decreased serum ALT, AST, and TG levels and promoted yolk precursor synthesis by upregulating genes associated with lipid transport and vitellogenesis (p < 0.05). In the ovary, honeycomb optimized ovarian morphology and follicle development; elevated serum FSH, E2, and MLT levels; and enhanced the expression of steroidogenesis-related genes (p < 0.05), thereby promoting steroid hormone synthesis and reducing follicular atresia. In conclusion, dietary honeycomb supplementation could maintain intestinal health, modulate hepatic and ovarian metabolism functions, promote yolk precursor synthesis, improve follicle development, and consequently, enhance the laying performance of hens.

Objective: Accumulating evidence demonstrates that melatonin is involved in modulating granulosa cell function and follicular development. lncRNAs (long non-coding RNAs) and circRNAs (circular RNAs) have been reported to participate in multiple biological processes. This study aimed to explore the candidate circRNAs and lncRNAs related to molecular mechanisms when exploring the role of melatonin in regulating ovarian function. Methods: Bovine ovary granulosa cells were collected 48 h after treatment with melatonin at 10⁻⁷ M. The lncRNA and circRNA profiles of bovine granulosa cells were further explored using high-throughput sequencing in the absence/presence of melatonin. The differentially expressed lncRNAs and circRNAs were analyzed through the annotation information of source transcripts for GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes). Results: We identified 99 differentially expressed lncRNAs and 28 differentially expressed circRNAs. Enrichment analysis of differentially expressed lncRNAs and circRNAs showed they were enriched in multiple pathways involved in development, apoptosis, and reproductive function, such as the mTOR (mammalian Target of Rapamycin) signaling pathway, FoxO (Forkhead box O) signaling pathway, MAPK (Mitogen-Activated Protein Kinase) signaling pathway, Hippo signaling pathway, TGF-beta (Transforming Growth Factor-β) signaling pathway, PI3K-Akt (Phosphatidylinositol 3-Kinase-Akt) signaling pathway, apoptosis, and Rap1 (Ras-related protein 1), most of which were mainly related to granulosa cell function and the crosstalk between granulosa cells and oocytes. The present analysis indicated the potential role of melatonin in granulosa cell function by regulating lncRNA and circRNA expression and, thus, mediating follicular development. An lncRNA/circRNA and miRNA regulatory network was also constructed to take their interactions into account. Conclusions: Our study offers details of lncRNA and circRNA expression in bovine granulosa cells and further provides insight into the potential role of melatonin in regulating reproduction by modulating lncRNA and circRNA expression.