Abstract: This experiment aimed to investigate the effects of high dietary glycinin (60 g/kg) on liver histopathology, oxidative stress, and transcriptome profiles in hybrid yellow catfish (Pelteobagrus fulvidraco ♀ × P. vachelli ♂), and the hepatoprotective effects of selenium (Se) supplementation (0.30 mg/kg). The hybrid yellow catfish (8.00±0.25) g were fed four experimental diets for8 weeks: basal diet (CK group), glycinin-supplemented diet (Gl group), Se-supplemented diet (Se group), and glycinin plus Se-supplemented diet (Gl-Se group). The results showed that compared to the CK and Se groups, the Gl group exhibited characteristic hepatic tissue damage, including reduced hepatocyte numbers, cytomembrane dissolution, nuclear offset, and severe vacuolization, accompanied by a significant increase in the hepatic histopathology score (HHS). In contrast, the Gl-Se group alleviated the aforementioned liver structural damage. Compared with the CK group, the Gl group significantly increased serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, as well as elevated hepatic total cholesterol (TC) content, whereas hepatic ALT activity and total protein (TP) content were significantly decreased, suggesting that glycinin might compromise normal liver function. However, these changes were significantly alleviated in the Gl-Se group. Furthermore, compared with the CK group, the Gl group significantly suppressed hepatic antioxidant-related activities, such as total antioxidant capacity (T-AOC), total superoxide dismutase (T-SOD), and glutathione peroxidase (GSH-Px). In contrast, the levels of hydrogen peroxide (H₂O₂), malondialdehyde (MDA), and nitric oxide (NO) were remarkably elevated. Notably, the Gl-Se group markedly mitigated the hepatic oxidative stress caused by the Gl group. Transcriptomic results revealed a significant enrichment of the differentially expressed genes in the KEGG pathways, such as PPAR, MAPK, ascorbate and aldarate metabolism, fatty acid biosynthesis, and steroid biosynthesis. RT-qPCR validation demonstrated that the Gl group considerably up-regulated the expression of cholesterol synthesis-related gene (hmgcrb) while down-regulating genes involved in cholesterol clearance (acat1, cyp7a1, cyp8b1, and fabp1) compared to the CK group. Concurrently, the expression levels of genes associated with the MAPK/ERK signaling pathway (flt3, rasgrp4, rras, erk2, and dusp16) were remarkably increased in the Gl group. Nevertheless, the Gl-Se group significantly attenuated the glycinin-induced alterations in the expression of hmgcrb, acat1, cyp8b1, and fabp1genes and mitigated the activation of the MAPK/ERK pathway. Correlation analysis revealed that HHS and the levels of TC, H₂O₂, NO, and MDA of the liver were significantly positively correlated with the expression levels of MAPK/ERK pathway-related genes, suggesting that Se may participate in modulation of the MAPK/ERK signaling pathway, thereby protecting liver tissue from the damage of glycinin-induced oxidative stress and high cholesterol. In conclusion, these results demonstrated that 0.30 mg/kg dietary Se could ameliorate the hepatic oxidative stress and cholesterol metabolic disorder induced by high-level glycinin in hybrid yellow catfish.