Metabolomic and transcriptomic profiling reveals the effect of dietary protein and lipid levels on growth performance in loach (Paramisgurnus dabryanus).

The subject of this study was to explore the optimum requirements of loach (Paramisgurnus dabryanus) regarding dietary proteins and lipids and discuss the underlying mechanism. We designed nine diets to determine the effects of different levels of dietary crude protein (CP: 30%, 35%, and 40%) and ether extract (EE: 6%, 10%, and 14%) on the growth performance and metabolism of P. dabryanus. In total, 2160 healthy P. dabryanus (5.19 ± 0.01 g) were divided into nine groups with four replications at 60 fish per barrel stocking density. The trial lasted for eight weeks. Serum and liver samples were gathered for metabolomic and transcriptomic analyses. The results showed that the specific growth rate of P. dabryanus in the CP40EE10 group was the fastest and notably higher than that in other groups (P< 0.05). Analysis of the metabolome results found that the mTOR signaling pathway, glycerophospholipid metabolism, D-arginine and D-ornithine metabolism were significantly enriched pathways in the CP40EE10 group compared with the other groups (P< 0.05). Moreover, the transcriptomic analysis of differentially expressed genes (DEGs) showed that the expression of ARG (arginase) involved in protein synthesis was significantly upregulated in the CP40EE10 group compared to the slowest growing group (P< 0.05). Additionally, the expression of SPLA2 (secretory phospholipase A2) involved in lipid metabolism and FBP (fructose-1,6-bisphosphatase) involved in glucose metabolism were all significantly downregulated in the CP30EE6 group compared with the CP40EE10 group (P< 0.05). Furthermore, the analysis of differentially expressed metabolites (DEMs) and DEGs co-enriched in the KEGG pathway revealed that the significantly enriched pathways were arginine and proline metabolism, glycerophospholipid metabolism, and glycolysis/gluconeogenesis in CP40EE10 compared with other groups (P< 0.05). We conclude that including 40% CP and 10% EE in the P. dabryanus diet could result in a better growth rate. We hypothesized from metabolomic and transcriptomic analyses that the CP40EE10 diet might promote the growth of P. dabryanus by promoting protein synthesis, lipid metabolism, and energy production.

Effect of acute exposure to nitrite on physiological parameters, oxidative stress, and apoptosis in Takifugu rubripes.

In the present study, we evaluated the effects of nitrite exposure on hematological parameters, oxidative stress, and apoptosis in juvenile Takifugu rubripes. The fish were exposed to nitrite (0, 0.5, 1, 3, and 6 mM) for up to 96 h. In the high nitrite concentration groups (i.e., 3 and 6 mM), the concentrations of methemoglobin (MetHb), cortisol, glucose, heat shock protein (Hsp)-70, Hsp-90, and potassium (K+) were significantly elevated. Whereas, the concentrations of hemoglobin (Hb), triglyceride (TG), total cholesterol (TC), and sodium (Na+) and chloride (Cl-) ions were significantly decreased. Compared with those of the control groups, the concentrations of the antioxidant enzymes, namely, superoxide dismutase (SOD), catalase (CAT), glutathione (GSH), and glutathione peroxidase (GPx), in the gills were considerably elevated at 12 and 24 h after exposure to nitrite (1, 3, and 6 mM), but reduced at 48 and 96 h. The increase in the antioxidant enzymes may contribute to the elimination of reactive oxygen species (ROS) induced by nitrite during early nitrite exposure, when the antioxidant system is not sufficiently effective to eliminate or neutralize excessive ROS. In addition, we found that nitrite exposure could alter the expression patterns of some key apoptosis-related genes (Caspase-3, Caspase-8, Caspase-9, p53, Bax, and Bcl-2). This indicated that the caspase-dependent apoptotic pathway and p53-Bax-Bcl-2 pathway might be involved in apoptosis induced by nitrite exposure. Furthermore, our study provides insights into how acute nitrite exposure affects the physiological responses and potential molecular mechanism of apoptosis in marine fish. The results can help elucidate the mechanisms involved in nitrite-induced aquatic toxicology in marine fish.

Impact of nitrite exposure on plasma biochemical parameters and immune-related responses in Takifugu rubripes.

Nitrite is a major environmental pollutant in aquatic environments that negatively affects aquatic species. In this study, we investigated the impact of nitrite exposure on plasma biochemical parameters and immune responses in Takifugu rubripes. Fish were exposed to various concentrations of nitrite (0, 0.5, 1, 3, and 6 mM) for 96 h. After 0, 12, 24, 48, and 96 h of exposure, fish blood samples were collected to assay the levels of total protein (TP), albumin (Alb), glutamic-oxaloacetic transaminase (GOT), glutamic-pyruvic transaminase (ALT), complement C3 (C3), complement C4 (C4), immunoglobulin (IgM), and lysozyme activity (LZM). The gills were sampled to analyze the mRNA levels of heat shock protein 70 (hsp70), heat shock protein 90 (hsp90), tumor necrosis factor α (tnf-α), B-cell activating factor (baff), interleukin-6 (il-6), and interleukin-12 (il-12). Levels of GOT, ALT, C3, and C4 were significantly enhanced in the high nitrite concentration group (3 and 6 mM), whereas those of TP, Alb, LZM, and IgM decreased significantly with the same treatments. Nitrite significantly upregulated hsp70, hsp90, tnf-α, il-6, il-12, and baff mRNA levels after 96 h of exposure. These results indicated that nitrite exposure altered the blood physiological status and immune system response, resulting in dysfunction and immunotoxicity in T. rubripes. Furthermore, our results reveal the possible mechanism of aquatic-nitrite-induced toxicity in fish.

Exposure to nitrite alters thyroid hormone levels and morphology in Takifugu rubripes.

Nitrite (NO2-) can act as a toxic nitrogenous compound with the potential to disrupt endocrine systems in fish. The aim of the present study was to investigate the effects of nitrite on the thyroid endocrine system of Takifugu rubripes. Fish were exposed to 0, 0.5, 1, 3, and 6 mM nitrite concentrations. Blood was collected to assay the concentrations of thyroid-stimulating hormone (TSH), thyroxine (T4), triiodothyronine (T3), free thyroxine (FT4), free triiodothyronine (FT3), and 3,3,5'-triiodothyronine (rT3), as well as the activity of iodothyronine deiodinases (Dio1, Dio2, and Dio3,) after 0, 12, 24, 48, and 96 h of exposure to nitrite. The first branchial arch to the third branchial arch of T. rubripes were sampled and fixed, and thyroid morphology was observed. The results showed that exposure to nitrite significantly increased the concentrations of TSH, T3, FT3, and reduced the concentrations of T4, FT4, and rT3. The activity of Dio1 and Dio2 increased significantly, whereas Dio3 activity decreased significantly. Additionally, thyroid follicles degenerated and became blurred and most colloid material disappeared 96 h after exposure to high nitrite concentrations. Based on these results, high nitrite concentration exposure can disturb thyroid hormone homeostasis, alter thyroid follicle morphology, and result in thyroid endocrine toxicity.

Effects of nitrite exposure on haematological parameters, oxidative stress and apoptosis in juvenile turbot (Scophthalmus maximus).

Nitrite (NO2(-)) is commonly present as contaminant in aquatic environment and toxic to aquatic organisms. In the present study, we investigated the effects of nitrite exposure on haematological parameters, oxidative stress and apoptosis in juvenile turbot (Scophthalmus maximus). Fish were exposed to various concentrations of nitrite (0, 0.02, 0.08, 0.4 and 0.8mM) for 96 h. Fish blood and gills were collected to assay haematological parameters, oxidative stress and expression of genes after 0, 24, 48 and 96 h of exposure. In blood, the data showed that the levels of methemoglobin (MetHb), triglyceride (TG), potassium (K(+)), cortisol, heat shock protein 70 (HSP70) and glucose significantly increased in treatments with higher concentrations of nitrite (0.4 and/or 0.8mM) after 48 and 96 h, while the levels of haemoglobin (Hb) and sodium (Na(+)) significantly decreased in these treatments. In gills, nitrite (0.4 and/or 0.8mM) apparently reduced the levels of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and glutathione (GSH), increased the formation of malondialdehyde (MDA), up-regulated the mRNA levels of c-jun amino-terminal kinase (JUK1), p53, caspase-3, caspase-7 and caspase-9 after 48 and 96 h of exposure. The results suggested caspase-dependent and JUK signaling pathways played important roles in nitrite-induced apoptosis in fish. Further, this study provides new insights into how nitrite affects the physiological responses and apoptosis in a marine fish.