Project description:Chickens divergently selected for either high abdominal fat content (fat genotype) or low abdominal fat content (lean genotype) at SRA-INRA, France were used to profile pituitary gland gene expression during juvenile development (1 to 7 weeks of age) and to identify differentially expressed genes associated with genotype and age. The fat line (FL) and lean line (LL) chickens are different in various phenotypic and metabolic measurements, including abdominal fatness, plasma glycemia, and T3. The FL and LL chickens represent unique models for characterizing biomedical and agricultural traits. The Del-Mar 14K Chicken Integrated Systems microarrays were used for a transcriptional scan in the pituitary gland during juvenile development using a reference design for the hybridization of total RNA from the pituitary gland. A reference RNA pool was made from an equal amount of high-quality total RNA extracted from all of the samples. Each of the microarrays was co-hybridized with Cy3-labeled cDNA targets from one of the pituitary samples and Cy5-labled cDNA targets from the reference pituitary RNA pool. Log2-transformed fluorescence intensities were analyzed with a mixed model. Two-way ANOVA (SAS) of log2 ratios was used to detect significant (p<0.05) differences by line, age, and the line-by-age interaction. There were 1150 significantly different genes between the two lines, and 339 of these genes exhibited greater than 0.68-fold differences in their log2 ratios (highest group mean at least 160% of the lowest group mean). One thousand four hundred twenty nine genes were significantly different by age and of these, 583 exhibited fold changes greater than 0.68 in the log2 ratio. There were 145 genes that significantly differed in their line-by-age interaction, and 62 of these exhibited greater than 0.68-fold differences. There were 386 genes with significant differences by line or for line-by-age interaction with at least a 0.68-fold change in log2 ratio and n ≥ 2 for each experimental group.

Project description:Chickens divergently selected for either high abdominal fat content (fat genotype) or low abdominal fat content (lean genotype) at SRA-INRA, France were used to profile pituitary gland gene expression during juvenile development (1 to 7 weeks of age) and to identify differentially expressed genes associated with genotype and age. The fat line (FL) and lean line (LL) chickens are different in various phenotypic and metabolic measurements, including abdominal fatness, plasma glycemia, and T3. The FL and LL chickens represent unique models for characterizing biomedical and agricultural traits. The Del-Mar 14K Chicken Integrated Systems microarrays were used for a transcriptional scan in the pituitary gland during juvenile development using a reference design for the hybridization of total RNA from the pituitary gland. A reference RNA pool was made from an equal amount of high-quality total RNA extracted from all of the samples. Each of the microarrays was co-hybridized with Cy3-labeled cDNA targets from one of the pituitary samples and Cy5-labled cDNA targets from the reference pituitary RNA pool. Log2-transformed fluorescence intensities were analyzed with a mixed model. Two-way ANOVA (SAS) of log2 ratios was used to detect significant (p<0.05) differences by line, age, and the line-by-age interaction. There were 1150 significantly different genes between the two lines, and 339 of these genes exhibited greater than 0.68-fold differences in their log2 ratios (highest group mean at least 160% of the lowest group mean). One thousand four hundred twenty nine genes were significantly different by age and of these, 583 exhibited fold changes greater than 0.68 in the log2 ratio. There were 145 genes that significantly differed in their line-by-age interaction, and 62 of these exhibited greater than 0.68-fold differences. There were 386 genes with significant differences by line or for line-by-age interaction with at least a 0.68-fold change in log2 ratio and n ≥ 2 for each experimental group. Four biological replicates were analyzed, each consisting of a complete set of individual samples from pituitary glands of Fat and Lean line chickens at week 1, week 3, week 5, or week 7 of age. Amplified antisense RNA from each sample was analyzed using Del-Mar 14 K array in a reference design. Each experimental sample (Fat and Lean Weeks 1, 3, 5, and 7) was labeled with Cy3 and hybridized with an aliquot of the Cy5 reference pool, which was generated from all 32 RNA samples.

Project description:BACKGROUND: This descriptive study of the abdominal fat transcriptome takes advantage of two experimental lines of meat-type chickens (Gallus domesticus), which were selected over seven generations for a large difference in abdominal (visceral) fatness. At the age of selection (9 wk), the fat line (FL) and lean line (LL) chickens exhibit a 2.5-fold difference in abdominal fat weight, while their feed intake and body weight are similar. These unique avian models were originally created to unravel genetic and endocrine regulation of adiposity and lipogenesis in meat-type chickens. The Del-Mar 14K Chicken Integrated Systems microarray was used for a time-course analysis of gene expression in abdominal fat of FL and LL chickens during juvenile development (1-11 weeks of age). RESULTS: Microarray analysis of abdominal fat in FL and LL chickens revealed 131 differentially expressed (DE) genes (FDR?0.05) as the main effect of genotype, 254 DE genes as an interaction of age and genotype and 3,195 DE genes (FDR?0.01) as the main effect of age. The most notable discoveries in the abdominal fat transcriptome were higher expression of many genes involved in blood coagulation in the LL and up-regulation of numerous adipogenic and lipogenic genes in FL chickens. Many of these DE genes belong to pathways controlling the synthesis, metabolism and transport of lipids or endocrine signaling pathways activated by adipokines, retinoid and thyroid hormones. CONCLUSIONS: The present study provides a dynamic view of differential gene transcription in abdominal fat of chickens genetically selected for fatness (FL) or leanness (LL). Remarkably, the LL chickens over-express a large number of hemostatic genes that could be involved in proteolytic processing of adipokines and endocrine factors, which contribute to their higher lipolysis and export of stored lipids. Some of these changes are already present at 1 week of age before the divergence in fatness. In contrast, the FL chickens have enhanced expression of numerous lipogenic genes mainly after onset of divergence, presumably directed by multiple transcription factors. This transcriptional analysis shows that abdominal fat of the chicken serves a dual function as both an endocrine organ and an active metabolic tissue, which could play a more significant role in lipogenesis than previously thought.

Project description:Genetic selection for enhanced growth rate in meat-type chickens (Gallus domesticus) is usually accompanied by excessive adiposity, which has negative impacts on both feed efficiency and carcass quality. Enhanced visceral fatness and several unique features of avian metabolism (i.e., fasting hyperglycemia and insulin insensitivity) mimic overt symptoms of obesity and related metabolic disorders in humans. Elucidation of the genetic and endocrine factors that contribute to excessive visceral fatness in chickens could also advance our understanding of human metabolic diseases. Here, RNA sequencing was used to examine differential gene expression in abdominal fat of genetically fat and lean chickens, which exhibit a 2.8-fold divergence in visceral fatness at 7 wk. Ingenuity Pathway Analysis revealed that many of 1687 differentially expressed genes are associated with hemostasis, endocrine function and metabolic syndrome in mammals. Among the highest expressed genes in abdominal fat, across both genotypes, were 25 differentially expressed genes associated with de novo synthesis and metabolism of lipids. Over-expression of numerous adipogenic and lipogenic genes in the FL chickens suggests that in situ lipogenesis in chickens could make a more substantial contribution to expansion of visceral fat mass than previously recognized. Distinguishing features of the abdominal fat transcriptome in lean chickens were high abundance of multiple hemostatic and vasoactive factors, transporters, and ectopic expression of several hormones/receptors, which could control local vasomotor tone and proteolytic processing of adipokines, hemostatic factors and novel endocrine factors. Over-expression of several thrombogenic genes in abdominal fat of lean chickens is quite opposite to the pro-thrombotic state found in obese humans. Clearly, divergent genetic selection for an extreme (2.5-2.8-fold) difference in visceral fatness provokes a number of novel regulatory responses that govern growth and metabolism of visceral fat in this unique avian model of juvenile-onset obesity and glucose-insulin imbalance.

Project description:BackgroundThough intensive genetic selection has led to extraordinary advances in growth rate and feed efficiency in production of meat-type chickens, endocrine processes controlling these traits are still poorly understood. The anterior pituitary gland is a central component of the neuroendocrine system and plays a key role in regulating important physiological processes that directly impact broiler production efficiency, though how differences in pituitary gland function contribute to various growth and body composition phenotypes is not fully understood.ResultsGlobal anterior pituitary gene expression was evaluated on post-hatch weeks 1, 3, 5, and 7 in male broiler chickens selected for high (HG) or low (LG) growth. Differentially expressed genes (DEGs) were analyzed with gene ontology categorization, self-organizing maps, gene interaction network determination, and upstream regulator identification to uncover novel pituitary genes and pathways contributing to differences in growth and body composition. A total of 263 genes were differentially expressed between HG and LG anterior pituitary glands (P ≤ 0.05 for genetic line-by-age interaction or main effect of line; ≥1.6-fold difference between lines), including genes encoding four anterior pituitary hormones. Genes involved in signal transduction, transcriptional regulation, and vesicle-mediated transport were differentially expressed and are predicted to influence expression and secretion of pituitary hormones. DEGs involved in immune regulation provide evidence that inflammation and response to cellular stressors may compromise pituitary function in LG birds, affecting their ability to adequately produce pituitary hormones. Many DEGs were also predicted to function in processes that regulate organ morphology and angiogenesis, suggesting pituitary gland structure differs between the divergently selected lines.ConclusionsThe large number of DEGs within the anterior pituitary gland of birds selected for high or low body weight highlights the importance of this gland in regulating economically important traits such as growth and body composition in broiler chickens. Intracellular signaling, transcriptional regulation, and membrane trafficking are important cellular processes contributing to proper hormone production and secretion. The data also suggest that pituitary function is intimately tied to structure, and organization of the gland could influence hypothalamic and systemic metabolic inputs and delivery of hormones regulating growth and metabolism into peripheral circulation.

Project description:Several processes governing growth and metabolism are regulated by the neuroendocrine system, particularly the trophic hormones produced in the anterior pituitary gland. However, many of the differences that exist in pituitary gene expression and functional gene networks associated with divergent growth characteristics are still largely unknown. Using the Del-Mar 14K Chicken Integrated Systems microarray, this experiment evaluated global gene expression in the anterior pituitary gland during the first 7 weeks of post-hatch growth in two lines of chickens genetically selected for high growth (HG) or low growth (LG) at SRA-INRA, France. The HG and LG chickens differ in body weight, abdominal fat content, and plasma hormone and metabolite levels, making them a valuable model for both biomedical and agricultural traits. Amplified total RNA extracted from individual anterior pituitary glands collected from 4 birds from each line at each age (n=4) was produced. Cy-3 labeled cDNA from these samples was hybridized with an aliquot of Cy-5 labeled cDNA made from a pool of total RNA from all 32 samples. Data were analyzed as log2-(Cy3intensity (norm)/Cy5intensity (raw)) by two-way ANOVA using a linear mixed model procedure (PROC MIXED; SAS institute Inc., Cary, NC), and a total of 2,310 spots were significantly different (P<0.05) by line (1,100), age (1,058), or the line-by-age interaction (353). Of these, we identified 291 spots representing 263 genes as differentially expressed between HG and LG birds, where there was a >160% difference between the highest expression in one line and the lowest expression in the other and a significant line-by-age (110) or line (181) effect. Four anterior pituitary hormones were contained in this set of differentially expressed genes: thyroid-stimulating hormone ?-subunit, luteinizing hormone ?-subunit, and follicle-stimulating hormone ?-subunit mRNA levels were higher in the HG line; and growth hormone mRNA was elevated in the LG line. Pathway analysis revealed differences in expression between HG and LG lines associated with molecular transport, endocrine system development and function, organ and tissue morphology, and cellular compromise. Potential alteration in pituitary function between the two lines was highlighted in a network containing genes associated with hormone expression and secretion, as well as those potentially involved in pituitary gland morphology and vascularization.

Project description:Decades of intensive genetic selection in the domestic chicken (Gallus gallus domesticus) have enabled the remarkable rapid growth of today's broiler (meat-type) chickens. However, this enhanced growth rate was accompanied by several unfavorable traits (i.e., increased visceral fatness, leg weakness, and disorders of metabolism and reproduction). The present descriptive analysis of the abdominal fat transcriptome aimed to identify functional genes and biological pathways that likely contribute to an extreme difference in visceral fatness of divergently selected broiler chickens.We used the Del-Mar 14 K Chicken Integrated Systems microarray to take time-course snapshots of global gene transcription in abdominal fat of juvenile [1-11 weeks of age (wk)] chickens divergently selected on bodyweight at two ages (8 and 36 wk). Further, a RNA sequencing analysis was completed on the same abdominal fat samples taken from high-growth (HG) and low-growth (LG) cockerels at 7 wk, the age with the greatest divergence in body weight (3.2-fold) and visceral fatness (19.6-fold).Time-course microarray analysis revealed 312 differentially expressed genes (FDR ? 0.05) as the main effect of genotype (HG versus LG), 718 genes in the interaction of age and genotype, and 2918 genes as the main effect of age. The RNA sequencing analysis identified 2410 differentially expressed genes in abdominal fat of HG versus LG chickens at 7 wk. The HG chickens are fatter and over-express numerous genes that support higher rates of visceral adipogenesis and lipogenesis. In abdominal fat of LG chickens, we found higher expression of many genes involved in hemostasis, energy catabolism and endocrine signaling, which likely contribute to their leaner phenotype and slower growth. Many transcription factors and their direct target genes identified in HG and LG chickens could be involved in their divergence in adiposity and growth rate.The present analyses of the visceral fat transcriptome in chickens divergently selected for a large difference in growth rate and abdominal fatness clearly demonstrate that abdominal fat is a very dynamic metabolic and endocrine organ in the chicken. The HG chickens overexpress many transcription factors and their direct target genes, which should enhance in situ lipogenesis and ultimately adiposity. Our observation of enhanced expression of hemostasis and endocrine-signaling genes in diminished abdominal fat of LG cockerels provides insight into genetic mechanisms involved in divergence of abdominal fatness and somatic growth in avian and perhaps mammalian species, including humans.

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Transcriptional profiling of hypothalamus during development of adiposity in genetically selected fat and lean chickens.

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Transcriptional profiling of hypothalamus during development of adiposity in genetically selected fat and lean chickens.

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