| Excessive fat accretion is a crucial problem in broiler production. The molecular mechnism of Abdominal fat(AbF) deposition remains unclear. By transcriptome and miRNA sequencing of the F2 generation of Beijing-you and cobb chicken resource population of the Chinese Academy of Agricultural Sciences(CAAS), we identified critical miRNA and genes related AbF deposition. BNP was identified as a candidate gene associated with AbF depostion by integrated analysis of diffrentially expressed genes(DEGs) and our GWAS data that we got before this study. Further study was carried out for BNP on its effect on proliferation, differentiation and lipolysis of adipocyte, also we investigated the regulation of BNP on chicken fat deposition at cellullar level.Trial 1. Screening of the Genes and microRNAs Related to Chicken Abdominal Fat Deposition and Intergrated Analysis of miRNA/mRNA differential expressionThe Chinese Academy of Agricultural Science(CAAS) chicken F2 resource population was used. The population was derived from a crosses between Beijing-You(BJY) chickens and Cobb broilers(CB, Cobb-Vantress, Inc.). Abdominal fat weight(AbFW) was measured and abdominal fat percentage(AbFP), relative to eviscerated weight, was calculated after dissection. Based on the data of 183 females, cohorts with extreme phenotypes of abdominal fat percentage and abdominal fat weight, 2 phenotypic groups, each of 6 birds with high or low Ab F(HAbF, LAbF), were used to constructe HAb F and LAbF libraries and investigated the expression profiles of mRNA and mi RNA by RNA-seq and mi RNA-seq. Our results showd that 1) Compared to the LAbF library, 32 up-regulated and 30 down-regulated miRNA were identified. 2) 189 up-regulated and 114 down-regulated annotated genes were identified. 3) Integrated analysis of differentially expressed miRNAs(DEMs) and differentially expressed genes(DEGs) showed that 110 intersection genes which were both differentially expressed between the two groups and were targets of differentially expressed miRNAs were identified. 4) 18 intersection genes involved in lipid metabolism might be important candidate genes related to Ab F deposition. Among these genes, ACSL1、SCD、PECR and FADS2 might have critical roles in AbF deposition in chicken. 5) Corresponding mi RNAs for 18 intersection genes might be critical miRNAs related to AbF deposition, including gga-mi R-19b-3p, miR-19a-3p, miR-17-5p, miR-301b-3p and miR-215-5p et al. 6) BNP mRNA expression in HAbF was significant higher than that of LAbF, and changes of BNP were positively correlated with AbFW(R2= 0.79,P=0.03), where these results indicated that BNP might be an important candidate gene related with AbF deposition in chicken.Trial 2:The Effect of BNP on Lipid Metabolism in ChickenBNP knockdown and BNP hormone were carried out in preadipocytes of 2~5 week female Beijing-You chickens, we observed their influence on preadipocytes proliferation and differentiation using MTT method and Oil Red O staining. 1) Compared with control group, BNP knockdown resulted in decreasing cell proliferation and differentiation(P < 0.05)and in decreasing glycerin concentration in medium at 48 h and 96h(P< 0.01). 2) Treatments of cells with 10-6, 10-7 and 10-8 mol/L BNP hormone led to enhanced cell proliferation at 24 h and 48 h of prolification. BNP did not have a dose-dependent effect cell proliferation. BNP treatment resulted in increased cell differentiation and fat droplets deposition(P< 0.05) and enhanced glycerinconcentration(P< 0.01)and NPR1 transcription(P< 0.05) at 96 h of differentiation. In total, our results indicated that BNP could promote cell proliferation, differentiation and lipolysis by inducing the expression of NPR1. |
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