| In this paper, golden pompano(Trachinotus ovatus) hypoxia tolerance, liver and gills histology and physiology as well hypoxia stress-related gene LDH-A(lactate dehydrogenase A) & MMP9(matrix metalloprotein 9) expression changes under acute and chronic hypoxia was studied,survival rate and suffocation point of golden pompano was analyzed under hypoxia stress, contrasted hypoxia tolerance capability with other fish, optical and electron microscopy was used and combined with histological analysis method to study different forms of hypoxia(acute and chronic) effect on golden pompano liver and gill tissue damage, the physiological metabolism related enzyme activity also was measured to analysis oxidative stress status and energy metabolism, 3’race, 5’race, PCR and Real-time PCR was used, cloned golden pompano hypoxia-related genes LDHA & MMP9 and explored its expression under the different hypoxia stress, the main results are as follows: 1 Golden pompano suffocation point and survival rate under hypoxiaReference to the experimental data statistics,(50-60) g body weight of Golden pompano suffocation point are(0.91-0.99) mg. L-1, this data provides a reference standard for setting the hypoxia Stress dissolved oxygen level in the following acute hypoxia and chronic stress. The survival rates are zero in acute hypoxia but declining in chronic, indicating that long-term of low dissolved oxygen for fish is deadly hazards. 2 Effect of hypoxia stress on histology of golden pompano liver and gillsThe results show that, liver tissue gradually appears vacuoles, blood sinus expands, and the hepatic lobule structure blur during acute hypoxia. In acute 24 h hypoxia, liver tissue even appears cell fusion and local cell necrosis. In 14 d chronic hypoxia, golden pompano liver tissue appears local cell necrosis, vacuolation are inside the cell, cell structure is decentralized, intercellular connection of cell is divide, cell membrane dissolve, organelles are inconspicuous, nucleus decomposition, blood sinus expansion appear, and only distinguish rough endoplasmic reticulum.In comparison with normoxia group, gills secondary gill lamellae pavement cell uplift, mitochondria-rich cells(MRC) decrease firstly and recover during acute hypoxia. But after 14 d chronic hypoxia, surface of gill respiratory epithelial cells is covered with ditch and ridge, pavement cell detaches with second gill filament, which result in an increasing of respiratory surface area, MRC significantly increase, volume of mitochondria in MRC increase too, and long diameter of blood cells(BC) increase also. as for the number of BC, which significantly increase both in acute and chronic hypoxia. 3 Hypoxia stress on golden pompano liver and gill oxidative stress and energy metabolism in the liverHypoxia stress significantly affected the antioxidant enzyme activity of golden pompano liver tissue. MDA rise firstly and then recover in acute hypoxia, but increase significantly in 14 d of hypoxia. In acute hypoxia, CAT activity continuously rise, SOD and GSH recover after increase. But in chronic hypoxia stress, SOD, GSH activity and MDA contents significantly increase, CAT activity significantly decline.Golden pompano gill antioxidant enzyme activity also varies significantly during acute hypoxia stress. With prolonged hypoxia stress CAT decrease and then increase,which is significantly higher than other groups at 24 h. GSH content and SOD activity all rise first and then decrease, SOD decrease to inter of control group and highest group at 24 h, GHS decrease to the level of the control group at 24 h, MDA content increase in 3 h of hypoxia and then keep on the level with slight volatility. Chronic hypoxic stress golden pompano liver antioxidant enzyme activity more significantly fluctuation. Under chronic hypoxia, CAT, SOD activity was significantly higher than the control group, MDA, GSH content also increased significantly. 4 LDH-A and MMP9 of golden pompanoGolden pompano LDH-A gene full-length is 2331 bp, including 81 bp of 5`UTR, 1141 bp of 3`UTR, 999 bp coding region encoding 332 amino acids, protein molecular weight is 36.14 kDa, an isoelectric point of 6.95.Golden pompano MMP9 gene full length is 2827 bp, including 188 bp 5`UTR, 507 bp 3`UTR and 2132 bp coding region encoding 684 amino acids, protein molecular weight is 77.04 kDa, isoelectric point is 5.41. 5 LDH-A and MMP9 of golden pompano expression under hypoxic stressDuring Acute hypoxia stress, golden pompano liver and gill LDH-A expression levels continued increase over time and then recovery, LDH-A expression levels in liver and gill are 5.91 and 2.63-fold compare to control group at 6 h, but returned to the level of control group at 24 h. The liver and gill LDH-A also increased significantly after 14 d chronic hypoxia stress, the LDH-A of liver was 3.42 times compared with the control group. No matter in acute hypoxia stress or chronic hypoxia stress, the expression of gill LDH-A is always less than the liver, liver LDH-A expression levels of chronic hypoxia stress is located the maximum value between the control group and acute.MMP9 expression levels under different forms of hypoxia stress changed significantly. In acute hypoxia stress, golden pompano liver and gill organs MMP9 expression levels decline and then increase. at 24 h hypoxia, liver MMP9 levels rise less than gills, gill MMP expression level has been restored to normal level. The expression of liver and gill organs MMP9 with 14 d chronic hypoxia stress show different changes, liver expression significantly decreases but gill tissue expression significantly increases. |
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