Qing-Ling Zhu

Differential induction of enzymes and genes involved in lipid metabolism in liver and visceral adipose tissue of juvenile yellow catfish Pelteobagrus fulvidraco exposed to copper.

The present study was conducted to determine the mechanism of waterborne Cu exposure influencing lipid metabolism in liver and visceral adipose tissue (VAT) of juvenile yellow catfish Pelteobagrus fulvidraco. Yellow catfish were exposed to four waterborne copper (Cu) concentrations (2 (control), 24 (low), 71 (medium), 198 (high) μg Cu/l, respectively) for 6 weeks. Waterborne Cu exposure had a negative effect on growth and several condition indices (condition factor, viscerosomatic index, hepatosomatic index and visceral adipose index). In liver, lipid content, activities of lipogenic enzymes (6-phosphogluconate dehydrogenase (6PGD), glucose-6-phosphate dehydrogenase (G6PD), malic enzyme (ME), isocitrate dehydrogenase (ICDH), and fatty acid synthase (FAS)) as well as mRNA levels of 6PGD, G6PD, FAS and sterol-regulator element-binding protein-1 (SREBP-1) genes decreased with increasing Cu concentrations. However, activity and mRNA level of lipoprotein lipase (LPL) gene in liver increased. In VAT, G6PD, ME and LPL activities as well as the mRNA levels of FAS, LPL and PPARγ genes decreased in fish exposed to higher Cu concentrations. The differential Pearson correlations between transcription factors (SREBP-1 and peroxisome proliferators-activated receptor-γ (PPARγ)), and the activities and mRNA expression of lipogenic enzymes and their genes were observed between liver and VAT. Thus, our study indicated that reduced lipid contents in liver and VAT after Cu exposure were attributable to the reduced activities and mRNA expression of lipogenic enzymes and their genes in these tissues. Different response patterns of several tested enzymes and genes to waterborne Cu exposure indicated the tissue-specific regulatory effect of lipid metabolism following waterborne Cu exposure. To our knowledge, the present study provides, for the first time, evidence that waterborne chronic Cu exposure can disturb the normal processes of lipid metabolism at both the enzymatic and molecular levels, and in two tissues (the liver and adipose tissue), which serves to increase our understanding of the mechanisms underlying lipid metabolism during Cu exposure.

Characterization and tissue distribution of leptin, leptin receptor and leptin receptor overlapping transcript genes in yellow catfish Pelteobagrus fulvidraco.

In the present study, full-length cDNA sequences of leptin (LEP), leptin receptor (LEPR) and leptin receptor overlapping transcript (LEPROT) were cloned from yellow catfish Pelteobagrus fulvidraco, and their tissue distribution profiles were determined. The validated cDNA of yellow catfish leptin (ycLEP), leptin receptor (ycLEPR) and LEPROT were 1119, 4195 and 827bp in length, encoding the peptide of 172, 1086 and 130 amino acid residues, respectively. The phylogenetic analysis revealed that fish LEP, LEPR and LEPROT were separated from tetrapod, and also ycLEPS were separated from other fish species. The ycLEP mRNA expression levels were highest in liver, followed by ovary, mesenteric fat and spleen, and lowest in intestine, heart, muscle, pituitary and testis. The ycLEPR mRNA levels were highest in pituitary, intermediate in mesenteric fat, liver, ovary, muscle and spleen, and lowest in heart, intestine and testis. The ycLEPROT mRNA levels were highest in pituitary, followed by spleen, mesenteric fat, heart, ovary, liver, muscle, testis and intestine. Identification and tissue distribution of yellow catfish LEP, LEPR and LEPROT genes provided initial step towards understanding their biological roles in yellow catfish.

Molecular cloning and expression pattern of 11 genes involved in lipid metabolism in yellow catfish Pelteobagrus fulvidraco

11 genes involved in lipid metabolism were cloned from liver of yellow catfish Pelteobagrus fulvidraco, including CPT 1A, CPT 1B, PPARα, PPARγ, SREBP-1, G6PD, 6PGD, FAS, acetyl-CoA ACCa, ACCb, and LPL. Phylogenetic analysis further identified these genes, and confirmed the classification and evolutionary status of yellow catfish. mRNA of all eleven genes was present in liver, muscle, mesenteric adipose, ovary and heart, but at varying levels. The present study will facilitate further studies on the regulation of lipid metabolism at the molecular level for the fish species.

Molecular characterization, tissue distribution and kinetic analysis of carnitine palmitoyltransferase I in juvenile yellow catfish Pelteobagrus fulvidraco.

Up to date, only limited information is available on genetically and functionally different isoforms of CPT I enzyme in fish. In the study, molecular characterization and their tissue expression profile of three CPT Iα isoforms (CPT Iα1a, CPT Iα1b and CPT Iα2a) and a CPT Iβ isoform from yellow catfish Pelteobagrus fulvidraco is determined. The activities and kinetic features of CPT I from several tissues have also been analyzed. The four CPT I isoforms in yellow catfish present distinct differences in amino acid sequences and structure. They are widely expressed in liver, heart, white muscle, spleen, intestine and mesenteric adipose tissue of yellow catfish at the mRNA level, but with the varying levels. CPT I activity and kinetics show tissue-specific differences stemming from co-expression of different isoforms, indicating more complex pathways of lipid utilization in fish than in mammals, allowing for precise control of lipid oxidation in individual tissue.

Differential effect of waterborne cadmium exposure on lipid metabolism in liver and muscle of yellow catfish Pelteobagrus fulvidraco

The present study was conducted to investigate the effect of waterborne cadmium (Cd) exposure on lipid metabolism in liver and muscle of juvenile yellow catfish Pelteobagrus fulvidraco. Yellow catfish were exposed to 0 (control), 0.49 and 0.95 mg Cd/l, respectively, for 6 weeks, the lipid deposition, Cd accumulation, the activities and expression level of several enzymes as well as the mRNA expression of transcription factors involved in lipid metabolism in liver and muscle were determined. Waterborne Cd exposure reduced growth performance, but increased Cd accumulation in liver and muscle. In liver, lipid content, the activities and the mRNA expression of lipogenic enzymes (6-phosphogluconate dehydrogenase (6PGD), glucose-6-phosphate dehydrogenase (G6PD), fatty acid synthetase (FAS)) and lipoprotein lipase (LPL) activity increased with increasing waterborne Cd concentrations. However, the mRNA expressions of LPL and peroxisome proliferators-activated receptor (PPAR) α were down-regulated by Cd exposure. Carnitine palmitoyltransferase 1 (CPT1) activity as well as the mRNA expressions of CPT1 and PPARγ showed no significant differences among the treatments. In muscle, lipid contents showed no significant differences among the treatments. The mRNA expression of 6PGD, FAS, CPT1, LPL, PPARα and PPARγ were down-regulated by Cd exposure. Thus, our study indicated that Cd triggered hepatic lipid accumulation through the improvement of lipogenesis, and that lipid homeostasis in muscle was probably conducted by the down-regulation of both lipogenesis and lipolysis. Different variation patterns of lipid metabolism to waterborne Cd exposure indicated the tissue-specific regulatory effect of lipid metabolism under waterborne Cd exposure. To our knowledge, the present study provides, for the first time, evidence that waterborne chronic Cd exposure can disturb the normal processes of lipid metabolism at both the enzymatic and molecular levels, and in two tissues (the liver and muscle).

Regulation of insulin on lipid metabolism in freshly isolated hepatocytes from yellow catfish (Pelteobagrus fulvidraco)

Although the metabolic actions of insulin in fish have been investigated widely in the past years, the regulatory effect of insulin on lipid metabolism has received little attention, especially in primary hepatocytes of fish. In the present study, freshly hepatocytes were isolated from yellow catfish, cultured and subjected to different insulin levels (0, 10, 100 and 1000nM) for 0h, 24h and 48h. Triglyceride (TG) content, activity and expression of several key enzymes involved in lipid metabolism, as well as mRNA levels of key transcription factors related to lipid metabolism, were assessed at 0h, 24h and 48h, respectively. Insulin incubation tended to increase the activities and expression of several lipogenic enzymes (such as FAS, G6PD, 6PGD). However, reduced CPT I gene expression was observed in hepatocytes following incubation treatment. Insulin administration also tended to up-regulate SREBP-1 expression but down-regulate PPARα mRNA levels. Insulin incubation enhanced lipogenesis and reduced lipolysis of freshly isolated hepatocytes of yellow catfish, in coincidence with increased TG content. Pearson correlations between expression of SREBP-1 and PPARα, and expression and activity of several enzymes were also observed, especially at 48-h insulin incubation. To the best of our knowledge, this is the first to study the effects of insulin on lipogenesis and lipolysis at both transcriptional and enzymatic levels using primary hepatocytes culture model in fish, which will help to understand the regulation of lipid metabolism by insulin in vivo, and will give us new insight into the insulin role in nutrient metabolism in fish.

Hormone-sensitive lipase in yellow catfish Pelteobagrus fulvidraco: molecular characterization, mRNA tissue expression and transcriptional regulation by leptin in vivo and in vitro.

Hormone-sensitive lipase (hsl) plays a pivotal role in regulation of lipolysis in mammals, but information is very scarce about its gene structure and function in fish. In this study, two distinct hsl cDNAs, designated hsl1 and hsl2, were firstly isolated and characterized from yellow catfish Pelteobagrus fulvidraco. The validated cDNAs encoding for hsl1 and hsl2 were 2739 and 2629bp in length, encoding peptides of 679 and 813 amino acid residues, respectively, and shared 57.7% amino acid identity. The phylogenetic analysis revealed that hsl1 and hsl2 derived from paralogous genes that might have arisen during a teleost-specific genome duplication event. Both hsl mRNAs were expressed in a wide range of tissues, but the abundance of each hsl mRNA showed the tissue- and developmental stage-dependent expression patterns. Intraperitoneal injection in vivo and incubation in vitro of recombinant human leptin (rb-hLEP) stimulated the mRNA expression of hsl2, but not hsl1, in the liver and hepatocytes of P. fulvidraco, respectively, suggesting that two hsl isoforms might serve different roles in lipid metabolism. To our knowledge, for the first time, the present study provides evidence that two hsl mRNAs are differentially expressed with and among tissues during different developmental stages and also differentially regulated by leptin both in vivo and in vitro, which serves to increase our understanding on hsl physiological function in fish.

Peroxisome proliferator-activated receptor gamma (PPARγ) in yellow catfish Pelteobagrus fulvidraco: Molecular characterization, mRNA expression and transcriptional regulation by insulin in vivo and in vitro

Peroxisome proliferator-activated receptor gamma (PPARγ) is ligand-inducible transcription factor and has important roles in lipid metabolism, cell proliferation and inflammation. In the present study, yellow catfish Pelteobagrus fulvidraco PPARγ cDNA was isolated from liver by RT-PCR and RACE, and its molecular characterization and transcriptional regulation by insulin in vivo and in vitro were determined. The generation of PPARγ1 and PPARγ2 was due to alternative promoter of PPARγ gene. PPARγ1 and PPARγ2 mRNA covered 2426 bp and 2537 bp, respectively, with an open reading frame (ORF) of 1584 bp encoding 527 amino acid residues. Yellow catfish PPARγ gene was organized in a manner similar to that of their mammalian homologs, implying a modular organization of the proteins domains. A comparison between the yellow catfish PPARγ amino acid sequence and the correspondent sequences of several other species revealed the identity of 55-76.2%. Two PPARγ transcripts (PPARγ1 and PPARγ2) mRNAs were expressed in a wide range of tissues, but the abundance of each PPARγ mRNA showed the tissue- and developmental stage-dependent expression patterns. Intraperitoneal injection of insulin in vivo significantly stimulated the mRNA expression of total PPARγ and PPARγ1, but not PPARγ2 in the liver of yellow catfish. In contrast, incubation of hepatocytes with insulin in vitro increased the mRNA levels of PPARγ1, PPARγ2 and total PPARγ. To our knowledge, for the first time, the present study provides evidence that PPARγ1 and PPARγ2 are differentially expressed with and among tissues during different developmental stages and also regulated by insulin both in vivo and in vitro, which serves to increase our understanding on PPARγ physiological function in fish.

Different effect of dietborne and waterborne Zn exposure on lipid deposition and metabolism in juvenile yellow catfish Pelteobagrus fulvidraco

Juvenile yellow catfish Pelteobagrus fulvidraco were exposed to 0.04 or 0.35 mg l(-1) waterborne Zn, 27.25 or 213.84 mg kg(-1) dietary Zn, singly or in combination for 42 days. Growth and lipid metabolism in juvenile yellow catfish were investigated. Growth and survival were significantly inhibited by single waterborne Zn exposure but not by dietary Zn exposure. Dietary Zn addition reduced but waterborne Zn exposure increased hepatic lipid content. In contrast, muscle lipid content was reduced by waterborne Zn exposure but not by dietborne Zn exposure. The single exposure also affected several lipogenic enzymatic activities and expression of genes (in this article gene expression is taken synonymous to mRNA expression) related to lipogenesis and lipolysis. Pearson correlations among lipid content, enzymatic activities and mRNA expression levels were also observed, suggesting that changes at molecular and enzymatic levels may underlie the patterns of lipid metabolism and accordingly affect lipid deposition. For the first time, our study demonstrates the differential effect of different Zn exposure pathways on lipid metabolism at the molecular level in fish, indicating that the exposure route is critical to lipid deposition and metabolism.

Differential effects of the chronic and acute zinc exposure on carnitine composition, kinetics of carnitine palmitoyltransferases I (CPT I) and mRNA levels of CPT I isoforms in yellow catfish Pelteobagrus fulvidraco.

The present study is conducted to determine the effect of acute and chronic zinc (Zn) exposure on carnitine concentration, carnitine palmitoyltransferases I (CPT I) kinetics, and expression levels of CPT I isoforms in liver, muscle and heart of yellow catfish Pelteobagrus fulvidraco. To this end, yellow catfish are subjected to chronic waterborne Zn exposure (0.05 mg Zn L(-1), 0.35 mg Zn L(-1) and 0.86 mg Zn L(-1), respectively) for 8 weeks and acute Zn exposure (0.05 mg Zn L(-1) and 4.71 mg L(-1)Zn, respectively) for 96 h, respectively. Reduced Michaelis-Menten constants (Km) and maximal reaction rates (Vmax) values in liver and muscle are observed in fish exposed to chronic Zn concentration. In contrast, Vmax and Km values in heart increase with increasing Zn concentration. Chronic Zn exposure also significantly influences the contents of free carnitine (FC), total carnitine (TC) and acylcarnitine (AC) in liver and heart, but not in muscle. The acute Zn exposure significantly increases FC, AC, TC contents in liver and muscle, but reduces their contents in heart. The chronic and acute Zn exposure influences the mRNA levels of four CPT I isoforms (CPT Iα1b, CPT Iβ, CPT Iα2a and CPT Iα1a) in liver, muscle and heart. Furthermore, correlations are observed in the mRNA levels between CPT I isoforms and Km, and between isoforms expression and activity of CPT I. Thus, chronic and acute Zn exposure shows differential effects on carnitine content, CPT I kinetics and mRNA levels of four CPT I isoforms in yellow catfish, which provides new mechanism for Zn exposure on lipid metabolism and also novel insights into Zn toxicity in fish.