Iron plays a key role in oxygen transport, hematopoiesis, and hypoxia adaptation. This study aimed to explore the dynamic response mechanism of the iron regulatory network and key genes in Duroc piglets. Eighteen weaned piglets were randomly divided into three dietary intervention groups: low iron (0 mg/kg), conventional (100 mg/kg), and high iron (200 mg/kg). Transcriptomics technology was used to screen key liver iron regulatory genes under the influence of different dietary iron concentrations, and the expression of related genes was verified using primary pig liver cells. Fasting serum iron metabolism parameters were detected and iron content in organs was quantified. The results show, enrichment analysis highlighted immune–metabolic signaling, including NF-κB, PI3K-Akt, and TGF-β, and a total of 14 candidate genes (such as FGF21, SAA2/3, FNDC1, ETNPPL, TFR1) were identified. The study observed that these genes showed obvious dosage differentiation and nonlinear patterns. However, findings reflect mRNA-level changes and GO/KEGG over-representation, protein-level validation is planned in follow-up studies. Through the integration of in vitro and in vivo data, this study discovered new liver genes that may be related to pig iron homeostasis function, providing a theoretical basis for analyzing the regulatory mechanism of piglet iron response.