Hypoxia severely threatens the survival and growth of fish, thereby greatly restricting the development of aquaculture. Rainbow trout (Oncorhynchus mykiss) is highly sensitive to hypoxia, but the regulatory mechanism of rainbow trout spleen under different levels of hypoxia is not yet clear. In this study, we analyzed the effects of moderate hypoxia (DO: 4.5 ± 0.1 mg/L) and severe hypoxia (DO: 3.0 ± 0.1 mg/L) on the biochemical parameters of rainbow trout spleen at various time points (4, 8, 12, 24 h, 1 month, and re-oxygenation for 12 and 24 h). Additionally, we comparatively analyzed the transcriptomic changes in moderate and severe hypoxia at 12 h, as well as in normoxic controls. The activities of antioxidant enzymes (T-AOC, T-SOD, GOT, GPT, CAT) and immune-related enzymes (AKP, ACP) were significantly decreased under moderate and severe hypoxia, while the content of MDA was significantly increased. After reoxygenation, the activities of antioxidant enzymes gradually increased, and the content of MDA gradually decreased. One-month moderate hypoxia resulted in a significant decrease in the activities of GOT, GPT, and T-SOD, along with a significant increase in the activities of AKP, T-AOC, and the content of MDA. In contrast, 1-month severe hypoxia led to a significant reduction in the activities of AKP, CAT, GOT, and T-SOD, as well as a significant elevation in the activities of ACP, T-AOC, and the content of MDA. Transcriptomic analysis identified 5766 DEGs. At the molecular level, numerous hypoxia-related, immune-related, and metabolism-related genes (including hif-1α, acly, egln1/2/3, fn1, fn1a, cd22, ccl7, ccl36.1, cd209a, cd209d, ighv3-43, ighv4-39, igkv1-37, igkv4-1, ldha, plpp2, plpp3, aldoa, pk, g6pc, eno1a, acaca, pfkfb3, and pfkpa) showed significant changes in expression. These genes were primarily enriched in the B cell receptor signaling, Glycolysis/Gluconeogenesis, Notch signaling, and PI3K-Akt signaling pathway demonstrating their crucial roles in the rainbow trout’s response to hypoxic stress. The analysis of dynamic expression of hypoxia-associated genes revealed that hif-1α, acly, egln2, plpp3, fn1, and cd22 were upregulated under moderate hypoxia, whereas aldoa and ldha were downregulated, with their expression levels recovering during reoxygenation. Egln1, egln2, egln3, aldoa, plpp3, pk, g6pc, and eno1a were upregulated during severe hypoxia stress, and their expression returned to baseline levels following reoxygenation. These results improve our comprehension of the molecular processes involved in hypoxia stress in rainbow trout.

Integrated biochemical and transcriptomic profiling reveals the graded response mechanisms of rainbow trout (Oncorhynchus mykiss) spleen to moderate and severe hypoxia

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发表日期：

2026年3月28日

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Kangkang Liu,
Jinqiang Huang,
Yongjuan Li,
Lu Zhao,
Qi Wang,
Zhijia Guo,
Tongzhen Sun,
Manqiang Sun

摘要

Hypoxia severely threatens the survival and growth of fish, thereby greatly restricting the development of aquaculture. Rainbow trout (Oncorhynchus mykiss) is highly sensitive to hypoxia, but the regulatory mechanism of rainbow trout spleen under different levels of hypoxia is not yet clear. In this study, we analyzed the effects of moderate hypoxia (DO: 4.5 ± 0.1 mg/L) and severe hypoxia (DO: 3.0 ± 0.1 mg/L) on the biochemical parameters of rainbow trout spleen at various time points (4, 8, 12, 24 h, 1 month, and re-oxygenation for 12 and 24 h). Additionally, we comparatively analyzed the transcriptomic changes in moderate and severe hypoxia at 12 h, as well as in normoxic controls. The activities of antioxidant enzymes (T-AOC, T-SOD, GOT, GPT, CAT) and immune-related enzymes (AKP, ACP) were significantly decreased under moderate and severe hypoxia, while the content of MDA was significantly increased. After reoxygenation, the activities of antioxidant enzymes gradually increased, and the content of MDA gradually decreased. One-month moderate hypoxia resulted in a significant decrease in the activities of GOT, GPT, and T-SOD, along with a significant increase in the activities of AKP, T-AOC, and the content of MDA. In contrast, 1-month severe hypoxia led to a significant reduction in the activities of AKP, CAT, GOT, and T-SOD, as well as a significant elevation in the activities of ACP, T-AOC, and the content of MDA. Transcriptomic analysis identified 5766 DEGs. At the molecular level, numerous hypoxia-related, immune-related, and metabolism-related genes (including hif-1α, acly, egln1/2/3, fn1, fn1a, cd22, ccl7, ccl36.1, cd209a, cd209d, ighv3-43, ighv4-39, igkv1-37, igkv4-1, ldha, plpp2, plpp3, aldoa, pk, g6pc, eno1a, acaca, pfkfb3, and pfkpa) showed significant changes in expression. These genes were primarily enriched in the B cell receptor signaling, Glycolysis/Gluconeogenesis, Notch signaling, and PI3K-Akt signaling pathway demonstrating their crucial roles in the rainbow trout’s response to hypoxic stress. The analysis of dynamic expression of hypoxia-associated genes revealed that hif-1α, acly, egln2, plpp3, fn1, and cd22 were upregulated under moderate hypoxia, whereas aldoa and ldha were downregulated, with their expression levels recovering during reoxygenation. Egln1, egln2, egln3, aldoa, plpp3, pk, g6pc, and eno1a were upregulated during severe hypoxia stress, and their expression returned to baseline levels following reoxygenation. These results improve our comprehension of the molecular processes involved in hypoxia stress in rainbow trout.