Haokun Liu

Different regulation of insulin on glucose and lipid metabolism in 2 strains of gibel carp

To test the hypothesis that response to insulin by regulating glucose and lipid metabolism in gibel carp A strain may be different from that in DT strain, bovine insulin was injected into both strains of gibel carp after previous fasting for 48h. The results showed that insulin induced hypoglycemia at 3h in 2 strains, and that this was coupled with increased expression of glucose transporters (GLUT2 in the liver and GLUT1, GLUT4 in the muscle) and glycolytic enzyme (HK2 in the muscle) in both strains. Insulin induced increased glycolysis (GK) and fatty acid oxidation (ACO3 in the liver and CPT1a, ACO3 in the muscle) in the DT strain. Conversely, very strong lipogenic capacity, as indicated by higher mRNA levels of transcription factor of fatty acid anabolism (SREBP1) and lipogenic enzymes (ACC, ACLY, and FAS) and decrease lipolytic capacity as indicated by lower mRNA levels of fatty acid oxidation enzymes in the liver (ACO3) and muscle (CPT1a and ACO3) detected in the A strain after insulin injection. Higher plasma insulin levels and decreased plasma free fatty acid levels were detected at 8h post insulin injection in A strain induced hypoglycemia. However, plasma glucose levels returned to baseline and no effect on fatty acid levels in the DT strain was observed in response to insulin treatment at the same point in time. These insulin-strain interactions demonstrated that insulin induced different changes in glucose and lipid metabolism in these 2 strains as expected.

Different physiological roles of insulin receptors in mediating nutrient metabolism in zebrafish

Insulin, the most potent anabolic hormone, is critical for somatic growth and metabolism in vertebrates. Type 2 diabetes, which is the primary cause of hyperglycemia, results from an inability of insulin to signal glycolysis and gluconeogenesis. Our previous study showed that double knockout of insulin receptor a ( insra) and b ( insrb) caused β-cell hyperplasia and lethality from 5 to 16 days postfertilization (dpf) (Yang BY, Zhai G, Gong YL, Su JZ, Han D, Yin Z, Xie SQ. Sci Bull (Beijing) 62: 486-492, 2017). In this study, we characterized the physiological roles of Insra and Insrb, in somatic growth and fueling metabolism, respectively. A high-carbohydrate diet was provided for insulin receptor knockout zebrafish from 60 to 120 dpf to investigate phenotype inducement and amplification. We observed hyperglycemia in both insra-/- fish and insrb-/- fish. Impaired growth hormone signaling, increased visceral adiposity, and fatty liver were detected in insrb-/- fish, which are phenotypes similar to the lipodystrophy observed in mammals. More importantly, significantly diminished protein levels of P-PPARα, P-STAT5, and IGF-1 were also observed in insrb-/- fish. In insra-/- fish, we observed increased protein content and decreased lipid content of the whole body. Taken together, although Insra and Insrb show overlapping roles in mediating glucose metabolism through the insulin-signaling pathway, Insrb is more prone to promoting lipid catabolism and protein synthesis through activation of the growth hormone-signaling pathway, whereas Insra primarily acts to promote lipid synthesis via glucose utilization.

Effect of dietary cottonseed meal on growth performance, physiological response, and gossypol accumulation in pre-adult grass carp, Ctenopharyngodon idellus

Cottonseed meal (CM) was used at up to 36.95% content in the diet (replacing 60% of dietary fish meal protein) without any negative eff ects on growth performance of pre-adult grass carp (initial body weight, 761 g) under outdoor conditions. A culture trial was conducted in net cages installed in a large concrete pond. Seven isonitrogenous and isoenergetic diets containing a gradient of CM concentrations (0, 12.2%, 24.4%, 36.6%, 48.8%, 54.8%, and 61.0%) as replacement for dietary fish meal protein (0, 20%, 40%, 60%, 80%, 90%, and 100%) were formulated. Dietary non-resistant starch (from maize) was inverse to dietary CM. Growth performance and feed utilization of fish fed the diets containing CM replacing 0–40% fishmeal protein were not aff ected after the 6-week feeding trial. Accumulation of hepatopancreatic total gossypol in the hepatopancreas was significantly correlated with free gossypol content in the diets (HTG=88.6+1.5×DFG, R2=0.89, P<0.05). Intestinal α-amylase and γ-glutamyl transpeptidase activities rose along with increasing dietary CM level. The structure of the mid-intestinal tissues and the ultrastructure of the enterocyte microvilli were normal when dietary CM was <36.6% (60% protein replacement). Increasing dietary CM content increased serum alanine aminotransferase levels but decreased serum alkaline phosphatase, cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, triglycerides, and albumin (P<0.05).

Different roles of insulin receptor a and b in maintaining blood glucose homeostasis in zebrafish

An inability of insulin to signal glycolysis and gluconeogenesis would largely result in type 2 diabetes. In this study, the physiological roles of zebrafish insulin receptor a and b in maintaining blood glucose homeostasis were characterized. We observed that, though blood glucose in insra-/- fish and insrb-/- fish were comparable with the control siblings at 0 h postprandium (hpp), the most evident hyperglycemia have been observed in insra-/- fish from 1 hpp to 3 hpp. A mild increase of blood glucose in insrb-/- fish has been seen only at 1.5 hpp. The down-regulated expressions of glycolytic enzymes were observed in insra-/- fish and insrb-/- fish liver and muscle, together with the significantly decreased activities or concentrations of glycolytic enzymes. These results suggest that both Insra and Insrb were critical in glycolysis. Intriguingly, the up-regulated expressions of gluconeogenic enzymes, pck1 and g6pca.1, along with the elevated enzyme activities, were observed in insra-/- fish liver at 1 hpp and 1.5 hpp. Compared with the control fish, the elevated plasma insulin and lowered phosphorylated AKT were observed in insra-/- fish and insrb-/- fish, suggesting that there is an insulin resistance in insra-/- fish and insrb-/- fish. The increased levels of both transcriptions of foxo1a and Foxo1a protein abundance in the insra-/- fish liver have been found. When insra-/- fish treated with the Foxo1 inhibitor, the postprandial blood glucose levels could be normalized, accompanied with the normalized expression levels and enzyme activities of both pck1 and g6pca.1. Therefore, Insra and Insrb demonstrate a similar role in promoting glycolysis, but Insra is involved in inhibiting gluconeogenesis via down-regulating the expression of foxo1a. Our results indicate that Insra and Insrb exhibit diversified functions in maintaining glucose homeostasis in zebrafish.

Effects of glucose administration on glucose and lipid metabolism in two strains of gibel carp (Carassius gibelio)

We compared the glucose clearance ability of gibel carp CAS III (A strain) with gibel carp Dongting (DT strain). A previous study suggested that these two strains responded to insulin differently. As insulin plays an important role in glucose utilization, we hypothesized that the ability to eliminate excess glucose after a glucose load would differ between A strain and DT strain. To test this hypothesis, fasted specimens of both strains of gibel carp were injected with glucose. As expected, glucose induced hyperglycemia in both A strain and DT strain. In both strains, mRNA levels of the glycolytic enzyme 6-phosphofructokinase (6PFK) increased in the white skeletal muscle 8 h post-injection, while expression levels of glucose-6-phosphatase (G6Pase), fructose 1,6-bisphosphatase (FBPase), and phosphoenolpyruvate carboxykinase (PEPCK) decreased in the liver 8 h post-injection. In the DT strain, both GLUT4 expression and muscular glycolytic processes increased, as reflected by elevated hexokinase 2 (HK2) and pyruvate kinase (PK) mRNA expression levels. The DT strain also returned to basal glycemia more quickly than the A strain (within 6 h versus more than 12 h). The glycogen concentration in the liver of the DT strain was higher than that of the A strain, indicating that the DT strain was better able to store glucose as glycogen than the A strain. Overall, the DT strain was better able to clear excess blood glucose after the glucose tolerance test than the A strain.

Effects of dietary yeast culture on growth performance, immune response and disease resistance of gibel carp (Carassius auratus gibelio CAS III)

Abstract A 50‐day feeding trial was carried out to evaluate the partial replacement of fishmeal by yeast culture (YC) on growth performance, immune response and resistance against Aeromonas hydrophila in gibel carp CAS III (Carassius auratus gibelio). Four isonitrogenous and isoenergetic practical diets including a basal diet (the control diet containing 10% fish meal, D0) and three yeast culture diets (substituting 20%, 40%, 60% of the fishmeal in the basal diet, D20, D40 and D60, respectively) were formulated. Each diet was randomly allocated to quadruplicate fish groups (average initial body weight: 28.70 ± 0.03 g) reared in a recirculating system. After the growth trial, bacterial challenge test was conducted. The results showed that no noteworthy variations in feed intake, growth performance and morphology indices were found among groups (P > 0.05). YC Supplemented diet exerted little significant influence on plasma parameters including triglyceride, glucose, creatinine, total protein and urea nitrogen compared with the control group (P > 0.05). No obvious variations were found in activities of plasma lysozyme, IgM, MPO and SOD before challenge test among dietary treatments (P > 0.05), whereas considerable higher value of the foresaid indicators was discovered in D40 after bacteria challenge (P < 0.05). Transcriptional levels of Toll like receptor 2 (TLR2), myeloid differentiation factor 88 (MyD88), Toll/IL‐1 receptor domain‐containing adaptor protein (TIRAP) and interleukin‐1&bgr; (IL‐1&bgr;) in spleen after challenge were significantly up‐regulated in D40 compared with D0 (P < 0.05). Cumulative survival rate in D40 and D60 were significantly higher than those in D0 and D20 (P < 0.05). Taken together, yeast culture could be a suitable fishmeal alternative in diets of gibel carp and dietary inclusion of 4 g YC per 100 g diet enhanced the immunity and disease resistance of gibel carp partly via TLR2 pathway. HighlightsDietary inclusion of 4% of yeast culture improved the immunity of gibel carp associated with TLR2 pathway.Dietary supplementation of 4% or 6% of yeast culture enhanced the survival of fish against Aeromonas hydrophila.Yeast culture could be a suitable fishmeal alternative in diets of gibel carp based on growth and immune response.

Replacement of fishmeal by spirulina Arthrospira platensis affects growth, immune related-gene expression in gibel carp (Carassius auratus gibelio var. CAS III), and its challenge against Aeromonas hydrophila infection

ABSTRACT The present study examined the effect of dietary spirulina, Arthrospira platensis on growth performance, blood physiological indices, immune‐related gene expressions and resistance of juvenile gibel carp against Aeromonas hydrophila infection. Four isonitrogenous (360gkg−1) and isolipidic (90gkg−1) diets were formulated with containing different levels of spirulina powder of 0g (SP0, the control diet), 3.38g (SP3.38), 6.76g (SP6.76) and 13.52g (SP13.52) per 100g diet to replace 0%, 25%, 50% and 100% of fishmeal protein, respectively. And each diet was randomly assigned to triplicate tanks (150‐L capacity per each) and each tank was stocked with 22 fish (15.37±0.06g). Fish were fed one of the tested diets up to satiation twice a day for 46 days. A challenge test was carried out after the feeding trial by injecting Aeromonas hydrophila intraperitoneally for 7 days. The results showed that fish growth, feeding rate in groups SP3.38 and SP6.76 were significantly higher than those of groups SP0 and SP13.52 (P<0.05). Feed efficiency and protein retention rate had no significant difference among all tested groups. Plasma superoxide dismutase and phagocyte activity of blood leukocytes significantly increased in the spirulina‐fed fish groups at 12‐h post the bacterial challenge (P<0.05). Both pre and post challenge test, plasma lysozyme activities in spirulina‐fed groups were significantly higher than that in the control group (P<0.05). Plasma malondialdehyde got the lowest value in the SP13.52 group before and after the challenge test. The transcriptional levels of TLR2 (Toll like receptor 2), myeloid differentiation factor 88 (MyD88), Toll/IL‐1 receptor domain‐containing adaptor protein (TIRAP), interleukin‐1&bgr; (IL‐1&bgr;) and tumor necrosis factor‐&agr;1 (TNF‐&agr;1) in spleen and kidney significantly increased post the bacterial challenge compared to the pre challenge. And the relative expressions of the immune‐related genes of spirulina‐fed fish groups were higher than those of the control group before and after the challenge test. The 7‐day cumulative survival rate after the bacterial challenge was highest in the SP3.38 group (P<0.05). The present results indicated that low dietary inclusion of spirulina significantly enhanced the immune response of gibel carp partly through TLR2 pathway and 3.38% of dietary spirulina was recommended for the juveniles based on the growth and immune response. HIGHLIGHTSDietary inclusion of 3.38% spirulina improved the immune response and disease resistance of gibel carp.The immune enhancing effect of spirulina in gibel carp was partly through TLR2 signaling pathway.Dietary fishmeal protein could be completely replaced by spirulina with no adverse effects on gibel carp.