Mechanism of liver oxidative damage induced by hemolysis in grass carp (Ctenopharyngodon idella)

A variety of bacterial or viral diseases usually cause massive hemolysis in fish, and release a large amount of cytotoxic hemoglobin into the tissues. However, the damage mechanism of hemolysis to fish tissues is still not well known. Therefore, to explore the damage mechanism of hemolysis in fish to the tissues, Ctenopharyngodon idella was used to systematically study the damage mechanism of hemolysis to the liver. Firstly, the hemoglobin was injected into the C. idella, hematoxylin eosin (H.E) staining assay results showed that injection of hemoglobin caused the dead cells to increase obviously in liver, and Prussian blue staining revealed that more iron deposited in liver. Quantitative real-time PCR (qRT-PCR) detection results showed that the injection of hemoglobin activated the expression of iron metabolism related genes. In order to further explore the damage of hemolysis to the liver, we injected phenylhydrazine into C. idella, H.E and Prussian blue staining results showed that the hemolysis caused the liver cell necrosis and iron deposits to increase, and the content of hemoglobin and iron in the liver all significantly increased with time. Furthermore, the increase of iron also activated the expression of iron metabolism related genes. Then, we detected the expression of inflammatory cytokines in liver after the injection of phenylhydrazine, and qRT-PCR results showed that high dose of hemoglobin activated the expression of various cytokines, such as pro-inflammatory cytokines TNF-α, IL-1β and IL-6, anti-inflammatory cytokines IL-10, and chemokines IL-4 and IL-8. In order to further explore the oxidation damage of hemoglobin in liver, we tested the content of malondialdehyde (MDA) and lipid peroxide (LPO) and β - galactose glucoside enzyme in liver, and the results showed that the hemolysis significantly increased liver oxidative damage effect, at the same time, the qRT-PCR and enzyme activity test results revealed that the oxidative damage of hemolysis induced the occurrence of liver cell apoptosis. Finally, we also examined the expression of antioxidant enzymes in liver, and the results showed that in vivo hemolysis significantly activated the antioxidant system in liver. In conclusion, the present study revealed that massive hemorrhage in fish released the high oxidative activity of hemoglobin which activated the occurrence of inflammation, oxidative damage and apoptosis in liver, and simultaneously up-regulated the expression of antioxidant system. The results of this study will enrich the blood immunology of fish, and also provide theoretical reference for the healthy culture of C. idella.