| Six Genes (GHR1、GHR2、TRαA、TRα、TRβ、LeptinA) and their expression after salinity regulation were studied in seabass. Hematological physiological parameters are detected after alinity regulation.1. Gene clone of growth and metabolization-related geneGHRs, as the center of GH/IGF axis, play key roles in individual endocrine. cDNA sequence of seabass (Lateolabrax maculatus) two GHRs were cloned by rapid amplification of cDNA cends (RACE). GHR1 and GHR2 were 2436 bp and 2940 bp in length, which encoded 637 and 582 amino acidrespectively. GHR1 and GHR2 contained different putative signal peptide, extracellular domain, transmembrane domain and intracellular domain. Thyroid hormones (THs), mediated through thyroid hormones receptors (TRs), plays key roles in growth, development and metabolism. TRaA, TRαB and TRβ were cloned in seabass. The full-length TRaA cDNA was 1851 bp, which encoded a predicted 416 amino acid protein. The full-length TRaB cDNA was 2370 bp, which encoded a predicted 409 amino acid protein. The full-length TRβ cDNA was 3343 bp, which encoded a predicted 395 amino acid protein. Leptin plays key roles in food intake, body weight regulation, reproduction, immunity and energy metabolism in mammals. LepA cDNA was 643bp in length, which contained a CDs of 161 amino acid residules, containing a signal peptide, and four a-helices. Deduced amino sequence showed low identity to Leps from other fish. However, Three-dimensional structure modeling of Leps showed high conservation.2.Tissue expression of growth and metabolization-related gene and expression after alinity regulationExpression level of GHR1 were higher than GHR2’s in brain, kindey and gill, while it is contrary in muscle, pituitary, liver, caecum, thymus and heart. These results suggested GHR1 of seabass was somatolactin receptor probably. Compared with TRuB and TRβ,the level of TRαA mRNA was low. relatively higher in brain, liver and heart. The level of TRab mRNA was relatively higher in pltuitary, followed by brain, kidney and heart. The level of TRβ mRNA was relatively higher in brain, followed by pituitary, kidney, stomach, caecus. liver and gill. TRaB and TRβ might be both have function on the regulation of TSH activity by TH feedback in seabass. Expression of LepA was relatively high in liver, a little less level was observed in brain.Salinity regulation experiment contained three group:seawater group, brackish water group and fresh water group. Hepatic GHRs, IGF-1, hypophyseal GH,TRs and PRLR in kidney and gill were determined after Id,2d,4d,6d,8d by qPCR. GHR1 mRNA was Constant, while the levels of GHR2, GH and IGF-1 decreased at 1d. Compared with seawater group,there were higher expression of GHRs, IGF-1 and lower expression of GH in brackish water group and fresh water group with following salinity regulation. GH/IGF axis participates in osmoregulation, probably in the way of improving the levels of GHRs, furthermore activated the expression of IGF-1. Three TRs subtypes expressed differently after salinity regulation in kidney and gill. TRαB may be the major TR isoform in osmoregulatory in seabass. Compared with seawater group, LepA mRNA decreased significantly in brackish water group and fresh water group at 2d, which reduced to 33.38 and 26.86 percent of seawater group respectively. Significant decrease of LepA mRNA expression was induced in all groups after 4d.These results suggested seabass LepA may have function in salinity regulation. Compared with seawater group and brackish water group, there were higher PRLR mRNA level in kidney and gill of fresh water group with following salinity regulation. PRLR play key roles in salinity regulation.3.Effect of salinity regulation on hematological physiological parameters of seabass and molecular mechanismThe study also examined the effects of salinity regulation on blood biochemical parameters of seabass. Compared with seawater group and brackish water group, there were higher level of red blood cell and hemoglobin in fresh water group with following salinity regulation. The study demonstrated that lower salinity stress improve metabolism and the need of oxygen, leading to higher level of red blood cell and hemoglobin in order to meet energy needs in seabass.Compared with brackish water group, there were higher level of glucose in seawater group and fresh water group. Content of urea of fresh water group and brackish water group rised in the later stage of salinity regulation. Nutrients in the blood raised in order to meet improved metabolism, leading to higher level of metabolic wastes.Bilirubin levels changed imperceptibly; nevertheless, alanine aminotransferase and aspartate aminotransferase levels increased before decreasing in fresh water group, indicating that salinity regulation harm liver imperceptibly, amino acids metabolism was improved with salinity regulation to meet needs of energy. Changed expression of genes maybe contact with hematological physiology through metabolism regulation in salinity regulation. |
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