Metformin improves insulin resistance, liver healthy and abnormal hepatic glucolipid metabolism via IR/PI3K/AKT pathway in Ctenopharyngodon idella fed a high-carbohydrate diet.

The effects and underlying mechanisms of metformin which can improve glucose homeostasis of fish have rarely been explored. This experiment aimed to explore the influence of metformin on growth performance, body composition, liver health, hepatic glucolipid metabolic capacity and IR/PI3K/AKT pathway in grass carp (Ctenopharyngodon idella) fed high-carbohydrate diets. A normal diet (Control) and high carbohydrate diets with metformin supplementation (0.00 %, 0.20 %, 0.40 %, 0.60 % and 0.80 %) were configured. Six groups of healthy fish were fed with the experimental diet for eight weeks. The results showed that the growth performance of grass carp was impaired in high carbohydrate diet. Impairment of IR/PI3K/AKT signalling pathway reduced insulin sensitivity, while hepatic oxidative stress damage and decreased immunity affected liver metabolic function. The glycolysis and lipolysis decrease while the gluconeogenesis and fat synthesis increase, which triggers hyperglycaemia and lipid deposition in the body. Metformin supplementation restored the growth performance of grass carp. Metformin improved IR/PI3K/AKT pathway signalling and alleviated insulin resistance, while liver antioxidant capacity and immunity were enhanced resulting in the restoration of liver health. The elevation of glycolysis and lipolysis maintains glycaemic homeostasis and reduces lipid deposition, respectively. These results suggest that metformin supplementation restores liver health and activates the IR/PI3K/AKT signalling pathway, ameliorating insulin resistance and glucose-lipid metabolism disorders caused by a high-carbohydrate diet. As judged by HOMA-IR, the optimum supplementation level of metformin in grass carp (C. idella) fed a high-carbohydrate diet is 0.67 %.

Comprehensive analysis of butyric acid impact on immunology, histopathology, gene expression, and metabolomic responses in pacific shrimp experiencing cold stress.

In this study, our objective was to investigate the impact of dietary butyric acid (BA) on the homeostasis mechanism of Pacific shrimp under cold stress. Specifically, we analyzed its effects on immunity, antioxidant capacity, gene expression, and metabolomics response. To carry out this research, Litopenaeus vannamei were fed a diet supplemented with BA for 8 weeks. Following this feeding period, a total of 180 shrimp, with an average weight of 12.76 ± 0.38 g, were exposed to cold conditions, with the temperature decreasing from 28 °C to 14 °C within an hour. The results of our study revealed survival rates ranging from 90 % to 100 %. Shrimp that were fed a diet containing 1.5 % BA exhibited a significant increase in acid phosphatase (ACP) and alkaline phosphatase (AKP) activity. Conversely, the control groups showed an increase in aspartate aminotransferase (AST) and alanine transaminase (ALT) activity. Shrimp that consumed diets containing 1.5 % BA displayed the lowest malondialdehyde (MDA) levels with the highest superoxide dismutase (SOD) content. The shrimp fed the BA diet exhibited tightly organized hepatic tubules with a star-shaped lumen filled with numerous B and R cells. Furthermore, shrimp fed the BA diet demonstrated a significant increase in caspase 3 (CASP) expression. There were no significant variations in the expression levels of prophenoloxidase (ProPO), manganese superoxide dismutase (MnSOD), and glutathione S-transferase (GST) The metabolites of Dl-carnitine, acetyl-l-carnitine, propionylcarnitine, hexanoylcarnitine, palmitoylcarnitine, decanoylcarnitine, and Dl-carnitine exhibited significantly increased expression in shrimp that were fed BA, suggesting their role in the lipolysis process. Based on the findings, adding 2 % BA to the diet of Pacific shrimp helps reduce inflammation and oxidative stress when they are under cold stress.

Effects of hydroxyproline supplementation in low fish meal diets on collagen synthesis, myofiber development and muscular texture of juvenile Pacific white shrimp (Litopenaeus vannamei).

This experiment aimed to evaluate the impact of dietary hydroxyproline (Hyp) supplementation on the muscle quality of juvenile Pacific white shrimp (Litopenaeus vannamei) fed a low fishmeal diet. Six formulated diets included one high fishmeal (HF; 25% fishmeal content) and five low fishmeal diets (10% fishmeal content) with 0%, 0.2%, 0.4%, 0.6% and 0.8% Hyp (LF0, LF2, LF4, LF6 and LF8, respectively). Each diet was assigned to four replicates, and 40 shrimp (0.32 ± 0.00 g) per replicate were fed four times a day for 8 weeks. Dietary Hyp supplementation had little effects on growth performance, but increased the contents of Hyp, prolyl 4-hydroxylases (P4Hs), and collagen. The meat yield, springiness, hardness, chewiness, and cohesiveness of muscle were the highest in the LF4 group among the low fishmeal groups (P < 0.05). Cooking loss and freezing loss of muscle were the lowest in the LF4 group (P < 0.05). Dietary supplementation with 0.4% Hyp increased the myofiber density and decreased the myofiber diameter of muscle (P < 0.05). Supplementation of Hyp in the diet up-regulated the mRNA expression of smyhc5, smyhc15, col1a1, col1a2, igf-1f, tgf-ß and tor and down-regulated the mRNA expression of smyhc 1, smyhc 2, smyhc 6a (P < 0.05). Supplementation of Hyp in the diet up-regulated the protein expression of P-4E-BP1, P-AKT, AKT and P-AKT/AKT (P < 0.05). These results suggested that the addition of 0.4% Hyp to low fishmeal diets improved the muscle quality of L. vannamei.

Single-Cell Transcriptomic Analysis Revealed the Cell Population Changes and Cell-Cell Communication in the Liver of a Carnivorous Fish in Response to High-Carbohydrate Diet.

BACKGROUND: Carnivorous fish have a low carbohydrate utilization ability, and the physiologic and molecular basis of glucose intolerance has not been fully illustrated. OBJECTIVES: This study aimed to use largemouth bass as a model to investigate the possible mechanism of glucose intolerance in carnivorous fish with the help of single-nuclei RNA sequencing (snRNA-seq). METHODS: Two diets were formulated, a low-carbohydrate (LC) diet and a high-carbohydrate (HC) diet. The feeding trial lasted for 6 wk, and then, growth performance, biochemical parameters, liver histology, and snRNA-seq were performed. RESULTS: Growth performance of fish was not affected by the HC diet, while liver glucolipid metabolism disorder and liver injury were observed. A total of 13,247 and 12,848 cells from the liver derived from 2 groups were isolated and sequenced, and 7 major liver cell types were annotated by the marker genes. Hepatocytes and cholangiocytes were lower and hepatic stellate cells (HSCs) and immune cells were higher in the HC group than those in the LC group. Reclustering analysis identified 7 subtypes of hepatocytes and immune cells, respectively. The HSCs showed more cell communication with other cell types, and periportal hepatocytes showed more cell communication with other hepatocyte subtypes. Cell-cell communication mainly focused on cell junction-related signaling pathways. Uncovered by the pseudotime analysis, midzonal hepatocytes were differentiated into 2 major branches-biliary epithelial hepatocytes and hepatobiliary hybrid progenitor. Cell junction and liver fibrosis-related genes were highly expressed in the HC group. HC diet induced the activation of HSCs and, therefore, led to the liver fibrosis of largemouth bass. CONCLUSIONS: HC diet induces liver glucolipid metabolism disorder and liver injury of largemouth bass. The increase and activation of HSCs might be the main reason for the liver injury. In adaption to HC diet, midzonal hepatocytes differentiates into 2 major branches-biliary epithelial hepatocytes and hepatobiliary hybrid progenitors.

Protective effects of butyric acid during heat stress on the survival, immune response, histopathology, and gene expression in the hepatopancreas of juvenile pacific shrimp (L. Vannamei).

This study looked at the effects of adding butyric acid (BA) to the diets of juvenile Pacific shrimp and how it affected their response to survival, immunity, histopathological, and gene expression profiles under heat stress. The shrimp were divided into groups: a control group with no BA supplementation and groups with BA inclusion levels of 0.5 %, 1 %, 1.5 %, 2 %, and 2.5 %. Following the 8-week feeding trial period, the shrimp endured a heat stress test lasting 1 h at a temperature of 38 °C. The results showed that the control group had a lower survival rate than those given BA. Interestingly, no mortality was observed in the group receiving 1.5 % BA supplementation. Heat stress had a negative impact on the activities of alkaline phosphatase (AKP) and acid phosphatase (ACP) in the control group. Still, these activities were increased in shrimp fed the BA diet. Similar variations were observed in AST and ALT fluctuations among the different groups. The levels of triglycerides (TG) and cholesterol (CHO) increased with high temperatures but were reduced in shrimp-supplemented BA. The activity of an antioxidant enzyme superoxide dismutase (SOD) increased with higher BA levels (P < 0.05). Moreover, the groups supplemented with 1.5 % BA exhibited a significant reduction in malondialdehyde (MDA) content (P < 0.05), suggesting the potential antioxidant properties of BA. The histology of the shrimp's hepatopancreas showed improvements in the groups given BA. Conversely, the BA significantly down-regulated the HSPs and up-regulated MnSOD transcript level in response to heat stress. The measured parameters determine the essential dietary requirement of BA for shrimp. Based on the results, the optimal level of BA for survival, antioxidant function, and immunity for shrimp under heat stress is 1.5 %.

The Supplementation of Berberine in High-Carbohydrate Diets Improves Glucose Metabolism of Tilapia (Oreochromis niloticus) via Transcriptome, Bile Acid Synthesis Gene Expression and Intestinal Flora.

Berberine is an alkaloid used to treat diabetes. This experiment aimed to investigate the effects of berberine supplementation in high-carbohydrate diets on the growth performance, glucose metabolism, bile acid synthesis, liver transcriptome, and intestinal flora of Nile tilapia. The six dietary groups were the C group with 29% carbohydrate, the H group with 44% carbohydrate, and the HB1-HB4 groups supplemented with 25, 50, 75, and 100 mg/kg of berberine in group H. The results of the 8-week trial showed that compared to group C, the abundance of Bacteroidetes was increased in group HB2 (p < 0.05). The cholesterol-7α-hydroxylase (CYP7A1) and sterol-27-hydroxylase (CYP27A1) activities were decreased and the expression of FXR was increased in group HB4 (p < 0.05). The pyruvate carboxylase (PC) and phosphoenolpyruvate carboxykinase (PEPCK) activities was decreased in group HB4 (p < 0.05). The liver transcriptome suggests that berberine affects carbohydrate metabolic pathways and primary bile acid synthesis pathways. In summary, berberine affects the glucose metabolism in tilapia by altering the intestinal flora structure, enriching differentially expressed genes (DEGs) in the bile acid pathway to stimulate bile acid production so that it promotes glycolysis and inhibits gluconeogenesis. Therefore, 100 mg/kg of berberine supplementation in high-carbohydrate diets is beneficial to tilapia.

Effects of Dietary Aflatoxin B1 on Hybrid Grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂) Growth, Intestinal Health, and Muscle Quality.

This study investigated the effects of varying doses of dietary aflatoxin B1 (AFB1) on the growth, intestinal health, and muscle quality of hybrid grouper. Four diets with varying AFB1 concentrations (0, 30, 445, and 2,230 µg kg-1) were used. Elevating AFB1 concentrations led to a decline in growth indexes, specifically the weight gain rate and the specific growth rate, although the survival rate remained unchanged. Morphological indicators showed a dose-dependent decline with AFB1 exposure. Intestinal MDA content and hindgut reactive oxygen species (ROS) levels increased, while antioxidant indexes and digestive enzymes decreased with higher AFB1 levels. AFB1 negatively influenced hindgut tight junction protein and antioxidant-related gene expression while promoting inflammation-related gene expression. The presence of AFB1 in the experiment led to a decrease in beneficial intestinal bacteria, such as Prevotella, and an increase in harmful intestinal bacteria, such as Prevotellaceae_NK3B31_group. Muscle lipid and unsaturated fatty acid content significantly decreased, while muscle protein and liver AFB1 content increased dramatically with higher AFB1 concentrations. AFB1 caused myofibrillar cleavage and myofilament damage, leading to increased spaces between muscle fibers. In conclusion, diets with AFB1 levels exceeding 30 µg kg-1 inhibited hybrid grouper growth, while levels surpassing 445 µg kg-1 resulted in hindgut ROS accumulation, inflammation, elevated intestinal permeability, reduced digestive enzyme activity, and compromised muscle quality.

Effects of α-Lipoic Acid Supplementation on Growth Performance, Liver Histology, Antioxidant and Related Genes Expression of Hybrid Grouper (Epinephelus fuscoguttatus ♀ × E. lanceolatus ♂).

This study aimed to assess the impact of α-lipoic acid on the growth performance, antioxidant capacity and immunity in hybrid groupers (♀ Epinephelus fuscoguttatus × â™‚ E. lanceolatus) fed with a high-lipid diet. Groupers (8.97 ± 0.01 g) were fed six different diets, with α-lipoic acid content in diets being 0, 400, 800, 1200, 1600, and 2000 mg/kg, named S1, S2, S3, S4, S5, and S6, respectively. The results show that the addition of 2000 mg/kg α-lipoic acid in the diet inhibited the growth, weight gain rate (WGR), and specific growth rate (SGR), which were significantly lower than other groups. In serum, catalase (CAT) and superoxide dismutase (SOD) were significantly higher in the S5 group than in the S1 group. In the liver, CAT, SOD and total antioxidative capacity (T-AOC) levels were significantly increased in α-lipoic acid supplemented groups. α-lipoic acid significantly upregulated liver antioxidant genes sod and cat, anti-inflammatory factor interleukin 10 (il10) and transforming growth factor ß (tgfß) mRNA levels. Conclusion: the addition of 2000 mg/kg of α-lipoic acid inhibits the growth of hybrid groupers. In addition, 400-800 mg/kg α-lipoic acid contents improve the antioxidant capacity of groupers and have a protective effect against high-lipid-diet-induced liver oxidative damage.

Effects of replacing fishmeal with different proportions of mixed protein source in the diet of largemouth bass (Micropterus salmoides).

Fishmeal is an important protein source for largemouth bass (Micropterus salmoides). However, the production of fishmeal is decreasing each year and the price of fishmeal is rising. Therefore, it is necessary to find new high-quality and suitable protein sources. This study used a mixed animal protein source (chicken meal:blood meal:shrimp meal:brewer's yeast = 50:12.5:25:12.5) to replace fishmeal. Using a 48 % fishmeal group as the control, five diets with different fishmeal levels (FM48, FM44, FM40, FM36, FM32) were established to determine the effects on largemouth bass growth performance, liver health and intestinal health. There were no significant differences in the percentage weight gain, specific growth rate, feed conversion rate, and condition factor of largemouth bass, but the hepatosomatic and viscerosomatic indexes were significantly decreased when the dietary fishmeal level was reduced to 40 %. The content of taurine, glycine, and histidine was significantly reduced in the muscle of largemouth bass fed the FM32 diet compared with those fed the FM48 and FM44 diets. Mixed protein feed reduced the total bile acid content and increased the low-density lipoprotein cholesterol content in the plasma of largemouth bass. The replacement of fishmeal with the mixed protein source inhibited the expression of tnf-α and caspase 3 and enhanced the expression of apoa1 in the liver, as well as enhancing the protein expression of FXR and SREBP and inhibiting the protein expression of P-PPARA in the liver. The intestinal pparα expression was suppressed when dietary fishmeal was replaced. When dietary fishmeal decreased, the mucosal folds height and muscle layer thickness also decreased. In conclusion, partial replacement of fishmeal with the mixed protein source did not affect the growth performance, while lipid metabolism and intestinal health were negatively affected when dietary fishmeal levels were below 36 %.