Wen-Jing Pan

Identification of Differentially Expressed Micrornas Associate with Glucose Metabolism in Different Organs of Blunt Snout Bream (Megalobrama amblycephala)

Blunt snout bream (Megalobrama amblycephala) is a widely favored herbivorous fish species and is a frequentlyused fish model for studying the metabolism physiology. This study aimed to provide a comprehensive illustration of the mechanisms of a high-starch diet (HSD) induced lipid metabolic disorder by identifying microRNAs (miRNAs) controlled pathways in glucose and lipid metabolism in fish using high-throughput sequencing technologies. Small RNA libraries derived from intestines, livers, and brains of HSD and normal-starch diet (NSD) treated M. amblycephala were sequenced and 79, 124 and 77 differentially expressed miRNAs (DEMs) in intestines, livers, and brains of HSD treated fish were identified, respectively. Bioinformatics analyses showed that these DEMs targeted hundreds of predicted genes were enriched into metabolic pathways and biosynthetic processes, including peroxisome proliferator-activated receptor (PPAR), glycolysis/gluconeogenesis, and insulin signaling pathway. These analyses confirmed that miRNAs play crucial roles in glucose and lipid metabolism related to high wheat starch treatment. These results provide information on further investigation of a DEM-related mechanism dysregulated by a high carbohydrate diet.

Dietary arginine affects the insulin signaling pathway, glucose metabolism and lipogenesis in juvenile blunt snout bream Megalobrama amblycephala

This study evaluated the mechanisms governing insulin resistance, glucose metabolism and lipogenesis in juvenile fish fed with graded levels of dietary arginine. The results showed that, compared with the control group (0.87%), 2.31% dietary arginine level resulted in the upregulation of the relative gene expression of IRS-1, PI3K and Akt in the insulin signaling pathway, while 2.70% dietary arginine level led to inhibition of these genes. 1.62% dietary arginine level upregulated glycolysis by increasing GK mRNA level; 2.70% dietary arginine level upregulated gluconeogenesis and resulted in high plasma glucose content by increasing PEPCK and G6P mRNA level. Furthermore, 2.70% dietary arginine level significantly lowered GLUT2 and increased PK mRNA levels. 1.62% dietary arginine level significantly upregulated ACC, FAS and G6PDH mRNA levels in the fat synthesis pathway and resulted in high plasma TG content. These results indicate that 1.62% dietary arginine level improves glycolysis and fatty acid synthesis in juvenile blunt snout bream. However, 2.70% dietary arginine level results in high plasma glucose, which could lead to negative feedback of insulin resistance, including inhibition of IRS-1 mRNA levels and activation of gluconeogenesis-related gene expression. This mechanism seems to be different from mammals at the molecular level.

Effect of nitrite exposure on the antioxidant enzymes and glutathione system in the liver of bighead carp, Aristichthys nobilis.

&NA; Nitrite (NO2−) can cause oxidative stress in aquatic animal when it accumulates in the organism, resulting in different toxic effects on fish. In the present study, we investigated the effects of nitrite exposure on the antioxidant enzymes and glutathione system in the liver of Bighead carp (Aristichthys nobilis). Fish [Initial average weight: (180.05 ± 0.092) g] were exposed to 48.634 mg/L nitrite for 96 h, and a subsequent 96 h for the recovery test. Fish livers were collected to assay antioxidant enzymes activity, hepatic structure and expression of genes after 0 h, 6 h, 12 h, 24 h, 48 h, 72 h, 96 h of exposure and 12 h, 24 h, 48 h, 72 h, 96 h of recovery. The results showed that the activity of glutathione peroxidase (GSH‐Px), glutathione S‐transferase (GST), and glutathione reductase (GR) increased significantly in the early stages of nitrite exposure. The study also showed that nitrite significantly up‐regulated the mRNA levels of glutathione peroxidase (GSH‐Px), glutathione S‐transferase (GST), and glutathione reductase (GR) after 6, 48, and 72 h of exposure respectively. Nitrite also increased the formation of malondialdehyde (MDA), oxidized glutathione (GSSG), and the activity of catalase (CAT). Nitrite was observed to reduce the activity of superoxide dismutase (SOD) and the level of glutathione (GSH). In the recovery test, GSH and the GSSG recovered but did not return to pre‐stress levels. The results suggested that the glutathione system played important roles in nitrite‐induced oxidative stress in fish. The bighead carp responds to oxidative stress by enhancing the activity of GSH‐Px, GST, GR and up‐regulating the expression level of GSH‐Px, GST, GR, a whilst simultaneously maintaining the dynamic balance of GSH/GSSG. CAT was also indispensable. They could reduce the degree of lipid peroxidation, and ultimately protect the body from oxidative damage. Graphical abstract Figure. No caption available. HighlightsNitrite exposure induced oxidative stress and tissue damaged on the liver of Aristichthys nobilis.Upregulation of GSH‐Px, GST, GR mRNA expressions and enzyme activities maintained the dynamic balance of GSH/GSSG system.The GSH/GSSG system fighted against oxidative injury caused by nitrite stress.

Comparative transcriptome analysis reveals the gene expression profiling in bighead carp (Aristichthys nobilis) in response to acute nitrite toxicity

Objective: Nitrite exposure induces growth inhibition, metabolic disturbance, oxidative stress, organic damage, and infection‐mediated mortality of aquatic organism. This study aimed to investigate the mechanism in responses to acute nitrite toxicity in bighead carp (Aristichthys nobilis, A. nobilis) by RNA‐seq analysis. Methods: Bighead carps were exposed to water with high nitrite content (48.63mg/L) for 72h, and fish livers and gills were separated for RNA‐seq analysis. De novo assembly was performed, and differentially expressed genes (DEGs) between control and nitrite‐exposed fishes were identified. Furthermore, enrichment analysis was performed for DEGs to annotate the molecular functions. Results: A total of 406,135 transcripts and 352,730 unigenes were tagged after de novo assembly. Accordingly, 4108 and 928 DEGs were respectively identified in gill and liver in responses to nitrite exposure. Most of these DEGs were up‐regulated DEGs. Enrichment analysis showed these DEGs were mainly associated with immune responses and nitrogen metabolism. Conclusions: We suggested that the nitrite toxicity‐induced DEGs were probably related to dysregulation of nitrogen metabolism and immune responses in A. nobilis, particularly in gill. HIGHLIGHTSNitrite exposure caused DEGs in bighead carp gill and liver.DEGs in gill and liver responded to nitrite stress were associated with immune response.Nitrite exposure‐induced pathomechanism associated with immune responses and infections.Gills of bighead carp are more susceptible to nitrite stress than liver.

Bioinformatic prediction and analysis of glucolipid metabolic regulation by miR-34a in Megalobrama amblycephala

The objective of this study was to analyze the target genes and regulatory function of miR-34a in Megalobrama amblycephala using second-generation high-throughput sequencing and bioinformatic tools. Functional enrichment analysis was performed by gene ontology. MiR-34a and target gene expression levels were measured in M. amblycephala fed normal and high-carbohydrate diets. The results revealed that miR-34a was highly conserved in several species, and miR-34a of M. amblycephala has a close evolutionary relationship to that of zebrafish and common carp. miRanda, TargetScan, RNAhybrid predicted 5,185, 6,282 and 2,168 target genes, respectively, and 645 target genes were in common. According to annotation information, the target genes were enriched in phosphate metabolism, glycerophospholipid metabolism, Golgi vesicle transport, cell division, and other biological processes (P < 0.05). Pathway enrichment analysis revealed that these target genes were mainly enriched in alpha-linolenic acid and linoleic acid metabolism, ether lipid metabolism, VEGF signaling pathway, Fc epsilon RI signaling pathway, GnRH signaling pathway, and MAPK signaling pathway (P < 0.05). The regulatory role of miR-34a was more significant in the liver than in the brain of M. amblycephala. MiR-34a regulates glucose lipid homeostasis induced by high glucose diets by upregulating hepatic PI3K/Akt, FOXO, and TOR signaling pathways.

Regulation mechanism of oxidative stress induced by high glucose through PI3K/Akt/Nrf2 pathway in juvenile blunt snout bream (Megalobrama amblycephala)

ABSTRACT This study was conducted to investigate the effects of oral administration of a high concentration of glucose on the respiratory burst, antioxidant status, and hepatic gene expression of heme oxygenase‐1 (ho1) and PI3K/Akt/Nrf2‐related signaling molecules in juvenile blunt snout bream (Megalobrama amblycephala). Blunt snout bream juveniles with an initial body weight of 19.94 ± 0.58 g were orally fed with a high concentration of glucose (3 g/kg body weight). The results indicated that plasma glucose exhibited a biphasic response. Acute and persistent hyperglycemia due to the oral glucose administration significantly reduced (P < 0.05) the white blood cell count, red blood cell count, and hemoglobin content and caused oxidative stress (significantly increased alanine aminotransferase, aspartate transaminase, alkaline phosphatase, and glucose levels) and early apoptosis of hepatocytes in the fish. Hepatic superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase activities increased rapidly (P < 0.05) as protection from oxidative stress and were downregulated (P < 0.05) because of persistent hyperglycemia. Blood respiratory burst was significantly reduced (P < 0.05) because of hyperglycemia and showed a trend that was opposite to that of plasma glucose. Slight upregulation of nrf2 mRNA and antioxidants acts as a compensative protection mechanism, and the downregulated PI3K/Akt pathway blocked this function of Nrf2. In conclusion, the PI3K/Akt pathway and Nrf2 mediated the antioxidative mechanism independently in the blunt snout bream juveniles subjected to the oral administration of a high glucose concentration. HighlightsOral glucose administration affected blood respiratory burst in M. amblycephala.Oral glucose administration induced blood glucose fluctuation and oxidative stress.Upregulation of Nrf2 against oxidative stress injury due to hyperglycemia.Inhibition of PI3K, Akt, and HO‐1 induced inflammation due to hyperglycemia.

Publisher Correction: Dietary arginine affects the insulin signaling pathway, glucose metabolism and lipogenesis in juvenile blunt snout bream Megalobrama amblycephala

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

In Vivo Analysis of miR-34a Regulated Glucose Metabolism Related Genes in Megalobrama amblycephala

The Megalobrama amblycephala (M. amblycephala) is one of the most important economic freshwater fish in China. The molecular mechanism under the glucose intolerance responses which affects the growth performance and feed utilization is still confused. miR-34a was reported as a key regulator in the glucose metabolism, but how did the miR-34a exert its function in the metabolism of glucose/insulin in M. amblycephala was still unclear. In this study, we intraperitoneally injected the miR-34a inhibitor (80 nmol/100 g body weight) into M. amblycephala (fed with high starch diet, 45% starch) for 12 h, and then analyzed the gene expression profiling in livers by RNA-seq. The results showed that miR-34a expression in M. amblycephala livers was inhibited by injection of miR-34a inhibitor, and a total of 2212 differentially expressed genes (DEGs) were dysregulated (including 1183 up- and 1029 downregulated DEGs). Function enrichment analysis of DEGs showed that most of them were enriched in the peroxisome proliferator-activated receptor (PPAR), insulin, AMP-activated protein kinase (AMPK) and janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathways, which were all associated with the glucose/lipid metabolic and biosynthetic processes. In addition, we examined and verified the differential expression levels of some genes involved in AMPK signaling pathway by qRT-PCR. These results demonstrated that the inhibition of miR-34a might regulate glucose metabolism in M. amblycephala through downstream target genes.

High glucose affected respiratory burst activity of peripheral leukocyte via G6PD and NOX inhibition in Megalobrama amblycephala

Abstract High glucose levels are known to impair growth and immune function in fish. Here we investigated the role of glucose‐6‐phosphate dehydrogenase (G6PD) and NADPH oxidase (NOX) in high glucose‐associated impairment of leukocyte respiratory burst activity in Megalobrama amblycephala. We cultured peripheral leukocytes isolated from M. amblycephala with media containing no glucose (non‐glucose group), 11.1 mmol/L d‐glucose (physiologic glucose group), 22.2 mmol/L d‐glucose (high‐glucose group), or 11.1 mmol/L d‐glucose + 100 &mgr;mol/L dehydroepiandrosterone (DHEA) (DHEA‐treated group). After 24 h, we assayed production of reactive oxygen species (ROS) as a measure of respiratory burst function as well as activity of G6PD and NOX. The high‐glucose group and DHEA‐treated group showed significantly reduced respiratory burst function, reduced production of ROS, and reduced G6PD and NOX activity at 24 h, compared to the non‐glucose and physiologic glucose groups (P < 0.05). The degree of impairment was similar between high‐glucose and DHEA‐treated groups (P > 0.05). These findings suggest that reduced NADPH availability likely underlies the suppression of respiratory burst function in M. amblycephala leukocytes exposed to high glucose levels. HighlightsHigh glucose and dehydroepiandrosterone (DHEA) reduce respiratory burst function in Megalobrama amblycephala leukocytes.High glucose and DHEA reduce activity of NOX and G6PD, decreasing NADPH availability.Reduced NADPH availability likely underlies impaired respiratory burst function.High glucose and DHEA induce MPO activity, reflecting leukocyte inflammation.