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Delta-like non-canonical Notch ligand 1 (DLK1), which inhibits the differentiation of precursor adipocytes, is a recognized marker gene for precursor adipocytes. Lipids play a crucial role in energy storage and metabolism as a vital determinant of beef quality. In this study, we investigated the mechanism of the DLK1 gene in lipid metabolism by constructing adipose tissue-specific knockout mice. We examined some phenotypic traits, including body weight, liver coefficient, fat index, the content of triglyceride (TG) and cholesterol (CHOL) in abdominal white adipose tissue (WAT) and blood. Subsequently, the fatty acid content and genes related to lipid metabolism expression were detected in DLK1−/− and wild-type mice via GC-MS/MS analysis and quantitative real-time PCR (qRT-PCR), respectively. The results illustrated that DLK1−/− mice exhibited significant abdominal fat deposition compared to wild-type mice. HE staining and immunohistochemistry (IHC) results showed that the white adipocytes of DLK1−/− mice were larger, and the protein expression level of DLK1−/− was significantly lower. Regarding the blood biochemical parameters of female mice, DLK1−/− mice had a strikingly higher triglyceride content (p < 0.001). The fatty acid content in DLK1−/− mice was generally reduced. There was a significant reduction in the expression levels of the majority of genes that play a crucial role in lipid metabolism. This study reveals the molecular regulatory mechanism of fat metabolism in mice and provides a molecular basis and reference for the future application of the DLK1 gene in the breeding of beef cattle with an excellent meat quality traits. It also provides a molecular basis for unravelling the complex and subtle relationship between adipose tissue and health. [ABSTRACT FROM AUTHOR]

Meat quality has a close relationship with fat and connective tissue; therefore, screening and identifying functional genes related to lipid metabolism is essential for the production of high-grade beef. The transcriptomes of the Longissimus dorsi muscle in Wagyu and Chinese Red Steppe cattle, breeds with significant differences in meat quality and intramuscular fat deposition, were analyzed using RNA-seq to screen for candidate genes associated with beef quality traits. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that the 388 differentially expressed genes (DEGs) were involved in biological processes such as short-chain fatty acid metabolism, regulation of fatty acid transport and the peroxisome proliferator-activated receptor (PPAR) signaling pathway. In addition, crystallin alpha B (CRYAB), ankyrin repeat domain 2 (ANKRD2), aldehyde dehydrogenase 9 family member A1 (ALDH9A1) and enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase (EHHADH) were investigated for their effects on intracellular triglyceride and fatty acid content and their regulatory effects on genes in lipogenesis and fatty acid metabolism pathways. This study generated a dataset from transcriptome profiling of two cattle breeds, with differing capacities for fat-deposition in the muscle, and revealed molecular evidence that CRYAB, ANKRD2, ALDH9A1 and EHHADH are related to fat metabolism in bovine fetal fibroblasts (BFFs). The results provide potential functional genes for maker-assisted selection and molecular breeding to improve meat quality traits in beef cattle. [ABSTRACT FROM AUTHOR]

Carnitine palmitoyltransferase 1B (CPT1B) is a candidate gene that regulates livestock animal lipid metabolism and encodes the rate-limiting enzyme in fatty acid β-oxidation. To explore the effect of this gene on lipid metabolism in cattle, this study examined CPT1B gene polymorphism in Chinese Simmental cattle and the effect of CPT1B on lipid metabolism. The results showed that the triglyceride content increased significantly with increasing CPT1B gene expression in bovine fetal fibroblasts (BFFs) (p < 0.05), while CPT1B knockout led to decreased CPT1B expression and a downward trend in triglyceride levels. Correlation analysis showed a significant association between the g.119896238 G > C locus and Chinese Simmental cattle backfat thickness (p < 0.05). Backfat thickness was significantly greater in individuals with the GC genotype (0.93 ± 0.67 cm) than in those with the CC genotype (0.84 ± 0.60 cm). The g.119889302 T > C locus was significantly correlated with arachidonic acid content in Chinese Simmental cattle (p < 0.05). The arachidonic acid content in the longissimus muscle was significantly higher in CC genotype beef cattle (0.054 g/100 g) than in those with the other two genotypes (0.046 g/100 g, 0.049 g/100 g). These molecular markers can be effectively used for marker-assisted selection in cattle breeding. [ABSTRACT FROM AUTHOR]

Mitochondrial glycerol-3-phosphate acyltransferase (GPAM) catalyses the initial and rate-regulated first-stage pathway of glycerol lipid synthesis and helps to allocate acyl-CoA (acyl-coenzyme A) to triglyceride (TG) synthesis and away from degradation pathways in animal lipometabolism-related pathways. In this study, RNA interference (RNAi) and GPAM gene overexpression were used to examine the correlation between the expression of GPAM and adipogenesis in bovine mammary epithelial cells (bMECs). Additionally, three novel polymorphisms were identified within the bovine key functional domain of GPAM with Sanger sequencing. The relationship between variants of the GPAM gene and milk quality traits of Chinese Holstein cows was then analysed using statistical methods. The results showed that knockdown of the GPAM gene significantly reduced the synthesis of triglycerides in the bMECs (p < 0.05), whereas the overexpression of the GPAM gene significantly increased the synthesis of TG (p < 0.05). In Chinese Holstein dairy cattle, the polymorphic locus of the GPAM gene E20-3386G > A was significantly correlated with fat, protein and somatic cell count (p < 0.05); I18-652A > G was significantly correlated with fat, total fat content, protein, dry matter and somatic cell count (p < 0.05); and I18-726A > G was significantly correlated with protein, milk yield, dry matter and somatic cell count (p < 0.05). Specifically, individuals with the AA genotype of the I18-652A > G and E20-3386G > A polymorphic loci had a higher milk fat percentage (p < 0.05). In summary, GPAM plays a pivotal role in the intracellular regulation of triglyceride, and its mutations could work as a competent molecular marker for selective breeding in dairy cattle. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) can regulate a variety of biological functions such as fat and cholesterol synthesis, translocation, and utilization by regulating degradation or by inhibiting target mRNA translation. In this study, the target gene of miR-21-3p was analyzed through bioinformatics, which predicted Elovl5. Furthermore, miR-21-3p was verified for regulation of triglyceride expression. The results showed that luciferase activity was significantly lower in cells cotransfected with miR-21-3p and Elovl5 reporter vector, pmiR-RB-REPORT-Elovl5-WT, than in cells with miR-21-3p and reporter control vectors, pmiR-RB-REPORT-Elovl5-mut and pmiR-RB-REPORT-Elovl5-si, indicating that target sites exist in the 3′UTR of Elovl5. Further analysis using qPCR and Western blotting showed that the expression of miR-21-3p negatively correlated with the levels of Elovl5 mRNA and protein, suggesting that miR-21-3p might play an important role in regulating Elovl5 gene expression. Finally, transfection with bta-miR-21-3p mimics, bta-miR-21-3p inhibitor, or miRNA-ShNC in mammary epithelial cells suggested that bta-miR-21-3p promotes triglyceride production, which might be attributed to the expression of the target gene Elovl5. [ABSTRACT FROM AUTHOR]

Unlike specific expression in the skin of wild mice, the agouti signaling protein (ASIP) is expressed widely in the tissue of cattle, including adipose and muscle tissue. Hence, it has been suggested that ASIP plays a role in bovine fat metabolism. An inserted L1-BT element was recently identified upstream of the ASIP locus which led to an ectopic expression of ASIP mRNA in cattle. In this study, we detected the indel of the L1-BT element at g. -14643 nt and three SNPs in introns of the ASIP gene (g. -568 A > G, g. -554 A > T, and g. 4805A > T) in a Chinese Simmental steer population. The association analysis between variants of ASIP and economic traits showed that the homozygous genotype of L1-BT element insertion, AA genotype of g. -568 A > G, and AT genotype of g. 4805A > T were significantly correlated with carcass and fat-related traits, such as live weight and back fat thickness. Moreover, three haplotypes (H1: AT; H2: AA; H3: GT) were identified by linkage disequilibrium analysis and formed six combined genotypes. Results indicated that Chinese Simmental steers with an H1H2 combined genotype had a higher measured value of fat-deposition-related traits (p<0.05), including thickness of back fat and percentage of carcass fat coverage, but a lower content of linoleic acid and α -linolenic acid (p<0.05). Individuals of an H3H3 combination had a lower marbling score, perirenal fat weight, and carcass weight (p<0.05). This suggests that these three SNPs and two combined haplotypes might be molecular markers for beef cattle breeding selection. [ABSTRACT FROM AUTHOR]

Bama Xiang pig (BM) and Tibetan mini-pig (TM) are used as experimental animals in China; however, the dwarf molecular mechanisms of these Chinese local pig breeds are unknown. IGFBP-3 affects animal growth, carcass and meat quality. The aim of this study was to identify the polymorphisms in the promoter of theIGFBP-3and analyse their effect on theIGFBP-3mRNA expression level in liver and muscle tissues. High-density single-nucleotide polymorphisms (SNPs) (31) and InDels (5) were detected in the promoter of theIGFBP-3in the BM, TM and Junmu No. 1 White (JM, control) pig breeds from 114 individuals by re-sequencing. A perfect Linkage disequilibrium consisted of 13 SNPs was observed in the promoter region and 2 main haplotypes were identified, of which theh1genotype (GCA-ATGTACATAT) was more prevalent in JM breed than in TM or BM breeds (P < .0001),h2(ATGTGCACG--CGC) was the dominant haplotype in TM and BM breeds (P < .0001). Expression analysis showed that haplotype of the promoter region is highly associated with theIGFBP-3mRNA expression level in liver and muscle tissues of pigs. TheIGFBP-3mRNA expression level was determined higher in the liver and muscle tissues of pigs withh2genotype as compared to that in pigs withh1genotype (P < .05). The results suggest that the SNPs and haplotypes in the promoter of theIGFBP-3gene may serve as useful molecular markers for the body growth traits and the breeding in swine. [ABSTRACT FROM AUTHOR]

microRNAs are involved in various biological processes by regulating the degradation or repressing the translation of target genes. In this study, the target genes of miR-29b were analyzed and predicted by bioinformatics. And lipoprotein lipase ( LPL) and thymine DNA glycosylase ( TDG) were selected for further validation by dual luciferase reporter assay. In addition, we investigated the effects of miR-29b on triglyceride synthesis and mammary epithelial cell (MEC) apoptosis. The result showed that luciferase activity was significantly lower in cells that miR-29b cotransfected with LPL and TDG gene reporter vectors ( pmiR-RB-REPORT-LPL-WT, pmiR-RB-REPORT-TDG-WT) than in cells of miR-29b cotransfected with gene reporter vectors ( pmiR-RB-REPORT-LPL-mut and pmiR-RB-REPORT-LPL-si; pmiR-RB-REPORT-TDG-mut and pmiR-RB-REPORT-TDG-si) ( p < 0.05), indicating that target sites existed in 3′UTR of LPL and TDG. Furthermore, the expressions of miR-29b were negatively correlated with levels of mRNA and protein of LPL and TDG gene using quantitative real-time polymerase chain reaction and western blot analysis, suggesting that miR-29b might play an important role in regulation of LPL and TDG gene expression. Finally, miR-29b promoted triglyceride production and suppressed apoptosis in MECs, which might be attributed to the expressions of target genes LPL and TDG. [ABSTRACT FROM AUTHOR]

Insulin-like growth factor binding protein 6 (IGFBP6) is a key factor in regulating the effects of insulin-like growth factor 2 (IGF2) on the animal growth and development, but the mechanism is far from known. In this study, the 5'-terminal sequence of theIGFBP6gene (from −920 to −1 bp) which may regulate gene expression was sequenced in the Bama mini-pigs, the Tibetan mini-pigs, the Landrace pigs, the Large White pigs and the Northeast wild boars to screen for the single nucleotide polymorphisms (SNPs) and analyze the relations with the body size traits of swine through a chi-square test analysis. The genotype frequencies of the SNPs in the 5'-terminal sequence have shown that c.-726C > T, c.-722T > C, c.-535C > A, c.-488T > C, c.-403A > G, and c.-378T > C may be related to the dwarf traits of the Bama mini-pig and the Tibetan mini-pigs (P < .05). A haplotype analysis of the 5'-terminal sequence of theIGFBP6gene in the Landrace pigs, the Large White pigs and the Northeast wild boars found the two SNPs at −403 nt and −378 nt were in linkage and formed three kinds of haplotypes; AT was the dominant haplotype and the haplotype block was not formed in the Bama mini-pigs and Tibetan mini-pigs. Above all, the SNPs and haplotypes of the 5'-terminal sequence of theIGFBP6gene may be involved in the regulation of swine body size. [ABSTRACT FROM AUTHOR]

ABSTRACT Growth-hormone-releasing peptide 6 (GHRP-6) plays an important role in animal growth. However, there have been few studies focusing on the effect of GHRP-6 on animal growth through controlled release systems. We synthesized the poly(lactic-co-glycolic acid) (PLGA)-poly(ethylene glycol) (PEG)-PLGA copolymer to investigate its controlled released effect on GHRP-6 in vitro and to study the effect of a GHRP-6-copolymer hydrogel on the growth of rex rabbits. The copolymer was synthesized with ring-opening copolymerization and characterized by 1H-NMR. The interaction between GHRP-6 and the copolymer was characterized by Fourier transform infrared spectroscopy and X-ray diffraction. The body weight, serum level of insulin-like growth factor 1 (IGF-1), and hair coat quality were studied in rex rabbits. The results show that hydrogen bonds formed between the NH group in GHRP-6 and the CO group in the copolymer. The release mechanism of GHRP-6 was a combination of a diffusion-controlled mechanism and an erosion-controlled mechanism in the copolymer. The serum level of IGF-1, hair coat quality, and body weight were all significantly higher in the GHRP-6-copolymer hydrogel group than in the other groups. These results indicate that the copolymer effectively controlled the release of GHRP-6. In addition, the GHRP-6-copolymer hydrogel increased the synthesis of IGF-1 for a prolonged period and, thereby, increased the rex rabbits' growth and hair coat quality. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014, 131, 40185. [ABSTRACT FROM AUTHOR]