Global gene expression analysis of the turkey hen hypothalamo-pituitary-gonadal axis during the preovulatory hormonal surge.

The preovulatory hormonal surge (PS) consists of elevated circulating luteinizing hormone (LH) and progesterone levels and serves as the primary trigger for ovarian follicle ovulation. Increased LH and progesterone, produced by the pituitary and the granulosa layer of the largest ovarian follicle (F1), respectively, result from hypothalamic stimulation and steroid hormone feedback on the hypothalamo-pituitary-gonadal (HPG) axis. The hypothalamus, pituitary, F1 granulosa, and granulosa layer of the fifth largest follicle (F5) were isolated from converter turkey hens outside and during the PS and subjected to RNA sequencing (n = 6 per tissue). Differentially expressed genes were subjected to functional annotation using DAVID and IPA. A total of 12, 250, 1235, and 1938 DEGs were identified in the hypothalamus, pituitary, F1 granulosa, and F5 granulosa respectively (q<0.05, |fold change|>1.5, FPKM>1). Gene Ontology (GO) analysis revealed key roles for metabolic processes, steroid hormone feedback, and hypoxia induced gene expression changes. Upstream analysis identified a total of 4, 42, 126, and 393 potential regulators of downstream gene expression in the hypothalamus, pituitary, F1G, and F5G respectively, with a total of 63 potential regulators exhibiting differential expression between samples collected outside and during the PS (|z-score|>2). The results from this study serve to increase the current knowledge base surrounding the regulation of the PS in turkey hens. Through GO analysis, downstream processes and functions associated with the PS were linked to identified DEGs, and through upstream analysis, potential regulators of DEGs were identified for further analysis. Linking upstream regulators to the downstream PS and ovulation events could allow for genetic selection or manipulation of ovulation frequencies in turkey hens.

Epistemic sabotage: The production and disqualification of evidence in disability benefit claims.

Disability benefits function by demarcating categories of need (the administrative category of disability) and determine eligibility using assessments of functioning. In the UK, these assessments are the Work Capability Assessment and PIP assessment. Inherently technical and abstruse processes, these assessments have been opportune sites for welfare reform in recent years. Disability benefits have also been a central point of contention between disability studies and sociology. Sociology has traditionally favoured an 'incomes approach' and called for more adequate financial support from the state. Early figures in the disabled people's movement rejected this position, and aligned with an oppression paradigm, argued for a more radical economic and social inclusion. We contend that this divide, set out in the Fundamental Principles of Disability, remains relevant for researching welfare reform today. This article treats benefits assessments as epistemic practices-interactional processes wherein claimants, their personal health professionals and commercial assessment providers come together in the production of knowledge about disability. Data include 50 in-depth interviews with benefit claimants and a discourse analysis of official texts directed at claimants, personal health professionals and commercial assessment providers. We outline a phenomenon we term 'epistemic sabotage', whereby the knowledge claims of claimants and their health professionals are systemically disqualified.

The Effect of Commercial Genetic Selection on Somatotropic Gene Expression in Broilers: A Potential Role for Insulin-Like Growth Factor Binding Proteins in Regulating Broiler Growth and Body Composition.

The somatotropic axis influences growth and metabolism, and many of its effects are a result of insulin-like growth factor (IGF) signaling modulated by IGF-binding proteins (IGFBPs). Modern commercial meat-type (broiler) chickens exhibit rapid and efficient growth and muscle accretion resulting from decades of commercial genetic selection, and it is not known how alterations in the IGF system has contributed to these improvements. To determine the effect of commercial genetic selection on somatotropic axis activity, two experiments were conducted comparing legacy Athens Canadian Random Bred and modern Ross 308 male broiler lines, one between embryonic days 10 and 18 and the second between post-hatch days 10 and 40. Gene expression was evaluated in liver and breast muscle (pectoralis major) and circulating hormone concentrations were measured post-hatch. During embryogenesis, no differences in IGF expression were found that corresponded with difference in body weight between the lines beginning on embryonic day 14. While hepatic IGF expression and circulating IGF did not differ between the lines post-hatch, expression of both IGF1 and IGF2 mRNA was greater in breast muscle of modern broilers. Differential expression of select IGFBPs suggests their action is dependent on developmental stage and site of production. Hepatic IGFBP1 appears to promote embryonic growth but inhibit post-hatch growth at select ages. Results suggest that local IGFBP4 may prevent breast muscle growth during embryogenesis but promote it after hatch. Post-hatch, IGFBP2 produced in liver appears to inhibit body growth, but IGFBP2 produced locally in breast muscle facilitates development of this tissue. The opposite appears true for IGFBP3, which seems to promote overall body growth when produced in liver and restrict breast muscle growth when produced locally. Results presented here suggest that paracrine IGF signaling in breast muscle may contribute to overall growth and muscle accretion in chickens, and that this activity is regulated in developmentally distinct and tissue-specific contexts through combinatorial action of IGFBPs.

Remodeling of Hepatocyte Mitochondrial Metabolism and De Novo Lipogenesis During the Embryonic-to-Neonatal Transition in Chickens.

Embryonic-to-neonatal development in chicken is characterized by high rates of lipid oxidation in the late-term embryonic liver and high rates of de novo lipogenesis in the neonatal liver. This rapid remodeling of hepatic mitochondrial and cytoplasmic networks occurs without symptoms of hepatocellular stress. Our objective was to characterize the metabolic phenotype of the embryonic and neonatal liver and explore whether these metabolic signatures are preserved in primary cultured hepatocytes. Plasma and liver metabolites were profiled using mass spectrometry based metabolomics on embryonic day 18 (ed18) and neonatal day 3 (nd3). Hepatocytes from ed18 and nd3 were isolated and cultured, and treated with insulin, glucagon, growth hormone and corticosterone to define hormonal responsiveness and determine their impacts on mitochondrial metabolism and lipogenesis. Metabolic profiling illustrated the clear transition from the embryonic liver relying on lipid oxidation to the neonatal liver upregulating de novo lipogenesis. This metabolic phenotype was conserved in the isolated hepatocytes from the embryos and the neonates. Cultured hepatocytes from the neonatal liver also maintained a robust response to insulin and glucagon, as evidenced by their contradictory effects on lipid oxidation and lipogenesis. In summary, primary hepatocytes from the embryonic and neonatal chicken could be a valuable tool to investigate mechanisms regulating hepatic mitochondrial metabolism and de novo lipogenesis.

Patient partners' perspectives of meaningful engagement in synthesis reviews: A patient-oriented rapid review.

BACKGROUND: A growing literature describes promising practices for patient-oriented research (POR) generally; however, those for systematic reviews are largely derived through the lens of a researcher. This rapid review sought to understand meaningful engagement in synthesis reviews from the patient partner (PP) perspective. DESIGN: The review team comprised PPs, librarians, SCPOR staff and academic faculty. We searched OVID MEDLINE and EMBASE, ProQuest Nursing and Allied Health, and core POR websites. Documents describing PP reflections on their involvement in synthesis reviews were included. Screening and data extraction were conducted by two independent reviewers. Thematic analysis was employed to identify themes in the data regarding PP perceptions of engagement in synthesis reviews. RESULTS: The literature search yielded 1386 citations. Eight journal articles and one blog post were included. Seven studies focused on conducting systematic reviews on a particular health or patient-related topic to which PP involvement was an important part and two studies focused specifically on the experience of including PP in synthesis reviews. PPs engaged in the review process through a variety of mechanisms, levels and stages of the review process. Three major themes emerged from the data: (1) foster partnerships through team development, (2) provide opportunities for outcomes valued by PP and (3) strengthen the research endeavour. CONCLUSION: Fostering partnerships through team development is foundational for meaningful engagement in synthesis reviews. It requires sensitively balancing of various needs (eg overburdening with contributions). Meaningful involvement in reviews has both personal and research benefits. PATIENT INVOLVEMENT: Patient partners were equal collaborators in all aspects of the review.

Transcriptome Analysis During Follicle Development in Turkey Hens With Low and High Egg Production.

Low and high egg producing hens exhibit gene expression differences related to ovarian steroidogenesis. High egg producing hens display increased expression of genes involved in progesterone and estradiol production, in the granulosa layer of the largest follicle (F1G) and small white follicles (SWF), respectively, whereas low egg producing hens display increased expression of genes related to progesterone and androgen production in the granulosa (F5G) and theca interna layer (F5I) of the fifth largest follicle, respectively. Transcriptome analysis was performed on F1G, F5G, F5I, and SWF samples from low and high egg producing hens to identify novel regulators of ovarian steroidogenesis. In total, 12,221 differentially expressed genes (DEGs) were identified between low and high egg producing hens across the four cell types examined. Pathway analysis implied differential regulation of the hypothalamo-pituitary-thyroid (HPT) axis, particularly thyroid hormone transporters and thyroid hormone receptors, and of estradiol signaling in low and high egg producing hens. The HPT axis showed up-regulation in high egg producing hens in less mature follicles but up-regulation in low egg producing hens in more mature follicles. Estradiol signaling exclusively exhibited up-regulation in high egg producing hens. Treatment of SWF cells from low and high egg producing hens with thyroid hormone in vitro decreased estradiol production in cells from high egg producing hens to the levels seen in cells from low egg producing hens, whereas thyroid hormone treatment did not impact estradiol production in cells from low egg producing hens. Transcriptome analysis of the major cell types involved in steroidogenesis inferred the involvement of the HPT axis and estradiol signaling in the regulation of differential steroid hormone production seen among hens with different egg production levels.

Characterization of hypothalamo-pituitary-thyroid axis gene expression in the hypothalamus, pituitary gland, and ovarian follicles of turkey hens during the preovulatory surge and in hens with low and high egg production.

Dysregulation of the preovulatory surge (PS) leads to lowered egg production. The hypothalamo-pituitary-thyroid (HPT) axis has been shown to influence plasma progesterone levels and follicle ovulation. The presence of thyroid hormone receptors (THR) in the reproductive axis suggests possible effects of thyroid hormone. To further understand the potential role of thyroid hormone on the PS, HPT axis plasma hormone concentrations and gene expression were characterized surrounding the PS in average egg producing hens (AEPH), low egg producing hens (LEPH), and high egg producing hens (HEPH) (n = 3 hens/group). Data were analyzed using the mixed models procedure of SAS, with significance indicated at P < 0.05. Average egg producing hens and HEPH displayed lower levels of triiodothyronine (T3) and higher levels of thyroxine (T4) inside of the PS, whereas LEPH showed inverse T3 and T4 levels relative to the PS. Expression of mRNA for hypothalamic thyrotropin-releasing hormone (TRH), pituitary thyrotropin (TSHB), and the main thyroid hormone metabolism enzyme (DIO2) were downregulated during the PS in AEPH and HEPH. Low egg producing hens displayed higher expression of mRNA for hypothalamic TRH as well as pituitary TSHB and DIO2 compared with HEPH. Average egg producing hens expression of THR mRNAs was upregulated during the PS in the hypothalamus but downregulated in the pituitary. High egg producing hens showed decreased expression of THR mRNAs in both the hypothalamus and pituitary when compared with LEPH. In ovarian follicles, THR mRNAs were more prevalent in the thecal layer of the follicle wall compared with the granulosa layer, and expression tended to decrease with follicle maturity. Minimal differences in follicular THR expression were seen between LEPH and HEPH, indicating that THR expression is unlikely to be responsible for steroid hormone production differences occurring between LEPH and HEPH. Generally, downregulation of the HPT axis was seen during the PS in AEPH and HEPH, whereas upregulation of the HPT axis was seen in LEPH. Further studies will be required to clarify the role of the HPT axis in the regulation of ovulation and egg production rates in turkey hens.

Differences in invitro responses of the hypothalamo-pituitary-gonadal hormonal axis between low- and high-egg-producing turkey hens.

Low-egg-producing hens (LEPH) ovulate less frequently than high-egg-producing hens (HEPH) and exhibit differences in mRNA levels for components of the hypothalamo-pituitary-gonadal (HPG) axis, suggesting differential responsiveness to trophic stimulation. Ovulation frequency is governed by the production of the pituitary gonadotropins and feedback of the ovarian follicle steroid hormones, which are regulated by HPG axis stimulation and inhibition at the hypothalamic level. The pituitary and follicle cells from LEPH and HEPH were subjected to in vitro hormonal treatments to stimulate or inhibit the HPG axis, followed by expression analysis of mRNA levels for HPG axis genes and radioimmunoassays for steroid hormone production. Statistical analysis was performed using the mixed models procedure of SAS. The pituitary cells from HEPH showed upregulation of genes associated with ovulation stimulation, whereas cells from LEPH showed upregulation of genes associated with inhibition of ovulation. High-egg-producing hens' follicle cells displayed a higher sensitivity and responsiveness to gonadotropin treatment. Level of egg production impacted ovulation-related gene expression in the pituitary cells as well as steroid hormone production in the follicle cells, with HEPH displaying a greater positive response to stimulation. These findings indicate that differences in egg production among turkey hens likely involve differential responsiveness of the cells within the HPG axis.

Effects of heat stress on performance, blood chemistry, and hypothalamic and pituitary mRNA expression in broiler chickens.

This study was conducted to evaluate potential hormonal mechanisms associated with the stress response, thermoregulation, and metabolic changes of broiler chickens exposed to high environmental temperature. Nine hundred 1-day-old male broiler chicks (Ross 708) were placed in floor pens and raised to 24 d. At 24 d, chicks were randomly assigned to 1 of 2 treatments, heat stress (HS) or no HS, and allocated into battery cages in 8 batteries (10 birds per cage, 2 cages per battery). On day 31, blood was collected prior to HS and analyzed using an iSTAT analyzer. Half of the batteries were then moved into 2 rooms with an elevated ambient temperature (35°C) for 8 h. The remaining batteries stayed in the thermoneutral rooms with an ambient temperature of 22°C. Beginning at 5 h after the initiation of HS, blood was collected and analyzed using an iSTAT analyzer, birds were euthanized, and hypothalamus and pituitary samples were collected (16 birds per treatment), flash frozen, and stored at -80°C until RNA extraction. Reverse transcription-quantitative PCR was used to compare mRNA levels of key corticotropic and thyrotrophic genes in the hypothalamus and pituitary. Levels of mRNA for each target gene were normalized to PGK1 (pituitary) and GAPDH (hypothalamus) mRNA. Differences were determined using mixed model ANOVA. HS decreased (P < 0.05) feed intake, BW, bicarbonate, potassium, CO2, and triiodothyronine, while it increased mortality, glucose, pH, plasma thyroxine, and corticosterone. Expression of pituitary corticotropin-releasing hormone receptor 1 was downregulated (P < 0.001), while corticotropin-releasing hormone receptor 2 mRNA levels were higher (P = 0.001) in HS birds. HS increased expression of thyroid hormone receptor ß (P = 0.01) (2.8-fold) and thyroid stimulating hormone ß (P = 0.009) (1.4-fold). HS did not affect levels of mRNA of genes evaluated in the hypothalamus. Results showed that HS significantly affected both the thyrotropic and corticotropic axes. Understanding the role and regulation of these pathways during HS will allow researchers to better evaluate management strategies to combat HS.

Transcriptome analysis of the hypothalamus and pituitary of turkey hens with low and high egg production.

BACKGROUND: High egg producing hens (HEPH) show increased hypothalamic and pituitary gene expression related to hypothalamo-pituitary-gonadal (HPG) axis stimulation as well as increased in vitro responsiveness to gonadotropin releasing hormone (GnRH) stimulation in the pituitary when compared to low egg producing hens (LEPH). Transcriptome analysis was performed on hypothalamus and pituitary samples from LEPH and HEPH to identify novel regulators of HPG axis function. RESULTS: In the hypothalamus and pituitary, 4644 differentially expressed genes (DEGs) were identified between LEPH and HEPH, with 2021 genes up-regulated in LEPH and 2623 genes up-regulated in HEPH. In LEPH, up-regulated genes showed enrichment of the hypothalamo-pituitary-thyroid (HPT) axis. Beta-estradiol was identified as an upstream regulator regardless of tissue. When LEPH and HEPH samples were compared, beta-estradiol was activated in HEPH in 3 of the 4 comparisons, which correlated to the number of beta-estradiol target genes up-regulated in HEPH. In in vitro pituitary cell cultures from LEPH and HEPH, thyroid hormone pretreatment negatively impacted gonadotropin subunit mRNA levels in cells from both LEPH and HEPH, with the effect being more prominent in HEPH cells. Additionally, the effect of estradiol pretreatment on gonadotropin subunit mRNA levels in HEPH cells was negative, whereas estradiol pretreatment increased gonadotropin subunit mRNA levels in LEPH cells. CONCLUSIONS: Up-regulation of the HPT axis in LEPH and upstream beta-estradiol activation in HEPH may play a role in regulating HPG axis function, and ultimately ovulation rates. Thyroid hormone and estradiol pretreatment impacted gonadotropin mRNA levels following GnRH stimulation, with the inhibitory effects of thyroid hormone more detrimental in HEPH and estradiol stimulatory effects more prominent in LEPH. Responsiveness to thyroid hormone and estradiol may be due to desensitization to thyroid hormone and estradiol in LEPH and HEPH, respectively, due to up-regulation of the HPT axis in LEPH and of the HPG axis in HEPH. Further studies will be necessary to identify possible target gene desensitization mechanisms and elicit the regulatory role of the HPT axis and beta-estradiol on ovulation rates in turkey hens.

Characterization of the hypothalamo-pituitary-gonadal axis in low and high egg producing turkey hens.

Variation in egg production exists in commercial turkey hens, with low egg producing hens (LEPH) costing more per egg produced than high egg producing hens (HEPH). Egg production correlates with ovulation frequency, which is governed by the hypothalamic-pituitary-gonadal (HPG) axis. Ovulation is stimulated by a preovulatory surge (PS) of progesterone and luteinizing hormone, triggered by gonadotropin releasing hormone release and inhibited by gonadotropin inhibiting hormone. Differences between LEPH and HEPH were characterized by determining HPG axis plasma hormone profiles and mRNA levels for key genes, both outside and inside of the PS (n = 3 per group). Data were analyzed with a 2-way ANOVA using the mixed models procedure of SAS. In the HPG axis, plasma progesterone levels were not affected by egg production level but were elevated during the PS. In contrast, plasma estradiol levels were higher in HEPH than in LEPH but were not associated with the PS. LEPH exhibited decreased gene expression associated with ovulation stimulation and increased gene expression associated with ovulation inhibition in the hypothalamus and pituitary. In ovarian follicle cells, LEPH displayed decreased gene expression associated with progesterone, androgen, and estradiol production in the F1 follicle granulosa cells, F5 theca interna cells, and small white follicle cells, respectively. Different degrees of stimulation and inhibition within all tissues of the HPG axis were noted between LEPH and HEPH turkey hens, with HEPH showing higher expression of genes related to ovulation and steroidogenesis.

Transcriptome analyses of liver in newly-hatched chicks during the metabolic perturbation of fasting and re-feeding reveals THRSPA as the key lipogenic transcription factor.

BACKGROUND: The fasting-refeeding perturbation has been used extensively to reveal specific genes and metabolic pathways that control energy metabolism in the chicken. Most global transcriptional scans of the fasting-refeeding response in liver have focused on juvenile chickens that were 1, 2 or 4 weeks old. The present study was aimed at the immediate post-hatch period, in which newly-hatched chicks were subjected to fasting for 4, 24 or 48 h, then refed for 4, 24 or 48 h, and compared with a fully-fed control group at each age (D1-D4). RESULTS: Visual analysis of hepatic gene expression profiles using hierarchical and K-means clustering showed two distinct patterns, genes with higher expression during fasting and depressed expression upon refeeding and those with an opposing pattern of expression, which exhibit very low expression during fasting and more abundant expression with refeeding. Differentially-expressed genes (DEGs), identified from five prominent pair-wise contrasts of fed, fasted and refed conditions, were subjected to Ingenuity Pathway Analysis. This enabled mapping of analysis-ready (AR)-DEGs to canonical and metabolic pathways controlled by distinct gene interaction networks. The largest number of hepatic DEGs was identified by two contrasts: D2FED48h/D2FAST48h (968 genes) and D2FAST48h/D3REFED24h (1198 genes). The major genes acutely depressed by fasting and elevated upon refeeding included ANGTPL, ATPCL, DIO2, FASN, ME1, SCD, PPARG, SREBP2 and THRSPA-a primary lipogenic transcription factor. In contrast, major lipolytic genes were up-regulated by fasting or down-regulated after refeeding, including ALDOB, IL-15, LDHB, LPIN2, NFE2L2, NR3C1, NR0B1, PANK1, PPARA, SERTAD2 and UPP2. CONCLUSIONS: Transcriptional profiling of liver during fasting/re-feeding of newly-hatched chicks revealed several highly-expressed upstream regulators, which enable the metabolic switch from fasted (lipolytic/gluconeogenic) to fed or refed (lipogenic/thermogenic) states. This rapid homeorhetic shift of whole-body metabolism from a catabolic-fasting state to an anabolic-fed state appears precisely orchestrated by a small number of ligand-activated transcription factors that provide either a fasting-lipolytic state (PPARA, NR3C1, NFE2L2, SERTAD2, FOX01, NR0B1, RXR) or a fully-fed and refed lipogenic/thermogenic state (THRSPA, SREBF2, PPARG, PPARD, JUN, ATF3, CTNNB1). THRSPA has emerged as the key transcriptional regulator that drives lipogenesis and thermogenesis in hatchling chicks, as shown here in fed and re-fed states.

Living with multimorbidity? The lived experience of multiple chronic conditions in later life.

Multimorbidity is defined biomedically as the co-existence of two or more long-term conditions in an individual. Globally, the number of people living with multiple conditions is increasing, posing stark challenges both to the clinical management of patients and the organisation of health systems. Qualitative literature has begun to address how concurrency affects the self-management of chronic conditions, and the concept of illness prioritisation predominates. In this article, we adopt a phenomenological lens to show how older people with multiple conditions experience illness. This UK study was qualitative and longitudinal in design. Sampling was purposive and drew upon an existing cohort study. In total, 15 older people living with multiple conditions took part in 27 in-depth interviews. The practical stages of analysis were guided by Constructivist Grounded Theory. We argue that the concept of multimorbidity as biomedically imagined has limited relevance to lived experience, while concurrency may also be erroneous. In response, we outline a lived experience of multiple chronic conditions in later life, which highlights differences between clinical and lay assumptions and makes the latter visible.

Performance management: a qualitative study of relational boundaries in personal assistance.

Personal assistance (PA) is a model of support where disabled people take control of recruiting, training and managing the people that support them. Personal assistance differs from other forms of care, such as domiciliary or informal care, because the disabled person is in control of how, when and by whom they are supported. With the advent of personal health budgets, PA is no longer limited to social care but is also central to future NHS services and funding arrangements. The aims of this study were to gain a deeper understanding of PA relationships, and to explore how both parties manage interpersonal challenges. We report on data from 58 qualitative interviews with disabled employers and personal assistants. Applying concepts from Goffman's (1959) scheme of impression management, we present an analysis of the relational dynamics that occur when two people cooperate in shared endeavours. Goffman's concepts of team members and non-persons, in addition to the themes of regions and information control, aid a more fundamental understanding of the relational dynamics that occur between disabled employers and their PAs.

Hepatic Mitochondrial Oxidative Metabolism and Lipogenesis Synergistically Adapt to Mediate Healthy Embryonic-to-Neonatal Transition in Chicken.

During the normal embryonic-to-neonatal development, the chicken liver is subjected to intense lipid burden from high rates of yolk-lipid oxidation and also from the accumulation of the yolk-derived and newly synthesized lipids from carbohydrates. High rates of hepatic lipid oxidation and lipogenesis are also central features of non-alcoholic fatty liver disease (NAFLD) in both rodents and humans, but is associated with impaired insulin signaling, dysfunctional mitochondrial energetics and oxidative stress. However, these adverse effects are not apparent in the liver of embryonic and neonatal chicken, despite lipid burden. Utilizing comprehensive metabolic profiling, we identify that steady induction of hepatic mitochondrial tricarboxylic acid (TCA) cycle and lipogenesis are central features of embryonic-to-neonatal transition. More importantly, the induction of TCA cycle and lipogenesis occurred together with the downregulation of hepatic ß-oxidation and ketogenesis in the neonatal chicken. This synergistic remodeling of hepatic metabolic networks blunted inflammatory onset, prevented accumulation of lipotoxic intermediates (ceramides and diacylglycerols) and reduced reactive oxygen species production during embryonic-to-neonatal development. This dynamic remodeling of hepatic mitochondrial oxidative flux and lipogenesis aids in the healthy embryonic-to-neonatal transition in chicken. This natural physiological system could help identify mechanisms regulating mitochondrial function and lipogenesis, with potential implications towards treatment of NAFLD.

Dynamics of transcriptome changes during subcutaneous preadipocyte differentiation in ducks.

BACKGROUND: Pekin duck is an important animal model for its ability for fat synthesis and deposition. However, transcriptional dynamic regulation of adipose differentiation driven by complex signal cascades remains largely unexplored in this model. This study aimed to explore adipogenic transcriptional dynamics before (proliferation) and after (differentiation) initial preadipocyte differentiation in ducks. RESULTS: Exogenous oleic acid alone successfully induced duck subcutaneous preadipocyte differentiation. We explored 36 mRNA-seq libraries in order to study transcriptome dynamics during proliferation and differentiation processes at 6 time points. Using robust statistical analysis, we identified 845, 652, 359, 2401 and 1933 genes differentially expressed between -48 h and 0 h, 0 h and 12 h, 12 h and 24 h, 24 h and 48 h, 48 h and 72 h, respectively (FDR < 0.05, FC > 1.5). At the proliferation stage, proliferation related pathways and basic cellular and metabolic processes were inhibited, while regulatory factors that initiate differentiation enter the ready-to-activate state, which provides a precondition for initiating adipose differentiation. According to weighted gene co-expression network analysis, pathways positively related to adipogenic differentiation are significantly activated at the differentiation stage, while WNT, FOXO and other pathways that inhibit preadipocyte differentiation are negatively regulated. Moreover, we identified and classified more than 100 transcription factors that showed significant changes during differentiation, and found novel transcription factors that were not reported to be related to preadipoctye differentiation. Finally, we manually assembled a proposed regulation network model of subcutaneous preadipocyte differentiation base on the expression data, and suggested that E2F1 may serve as an important link between the processes of duck subcutaneous preadipocyte proliferation and differentiation. CONCLUSIONS: For the first time we comprehensively analyzed the transcriptome dynamics of duck subcutaneous preadipocyte proliferation and differentiation. The current study provides a solid basis for understanding the synthesis and deposition of subcutaneous fat in ducks. Furthermore, the information generated will allow future investigations of specific genes involved in particular stages of duck adipogenesis.

Characterization of gene expression in the hypothalamo-pituitary-gonadal axis during the preovulatory surge in the turkey hen.

A preovulatory surge (PS) of luteinizing hormone (LH) and progesterone triggers follicle ovulation, which is the first step of egg production and is orchestrated by the hypothalamo-pituitary-gonadal (HPG) axis. In the HPG axis, hypothalamic peptides, gonadotropin releasing hormone, and gonadotropin inhibitory hormone, control the production of follicle stimulating hormone and LH by the pituitary, which subsequently regulate ovarian production of estradiol and progesterone, respectively. The goal of this study was to characterize the HPG axis function of average egg producing hens by assessing plasma hormone profiles and hypothalamic, pituitary, and follicle gene expression outside and during the PS (n = 3 per group). Results were analyzed by a one-way ANOVA using the mixed models procedure of SAS. Plasma estradiol was not affected by the PS (P > 0.05), but plasma progesterone levels increased 8-fold during the PS when compared to basal progesterone levels (P < 0.05). HPG axis gene expression related to ovulation stimulation (e.g., GNRH, GNRHR, and LHB) was down-regulated during the PS; whereas gene expression related to follicle development (e.g., FSHB) was up-regulated during the PS. Additionally, in the hypothalamus and pituitary, estradiol receptor expression was up-regulated during the PS, whereas progesterone receptor expression was down-regulated during the PS. In the follicle cells, gene expression pertaining to progesterone (e.g., STAR), androgen (e.g., HSD17B1), and estradiol (e.g., CYP19A1) production was up-regulated during the PS. Prior to this study, the HPG axis had yet to be characterized during the PS in the turkey hen. This study showed that the PS significantly impacted gene expression in the hypothalamus, pituitary, and ovarian follicles. These results provide a foundation for further research into the regulation of ovulation and egg production in turkey hens.

Transcriptional profiling and pathway analysis reveal differences in pituitary gland function, morphology, and vascularization in chickens genetically selected for high or low body weight.

BACKGROUND: Though 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. RESULTS: Global 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. CONCLUSIONS: The 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.

Delayed Feeding Alters Transcriptional and Post-Transcriptional Regulation of Hepatic Metabolic Pathways in Peri-Hatch Broiler Chicks.

Hepatic fatty acid oxidation of yolk lipoproteins provides the main energy source for chick embryos. Post-hatching these yolk lipids are rapidly exhausted and metabolism switches to a carbohydrate-based energy source. We recently demonstrated that many microRNAs (miRNAs) are key regulators of hepatic metabolic pathways during this metabolic switching. MiRNAs are small non-coding RNAs that post-transcriptionally regulate gene expression in most eukaryotes. To further elucidate the roles of miRNAs in the metabolic switch, we used delayed feeding for 48 h to impede the hepatic metabolic switch. We found that hepatic expression of several miRNAs including miR-33, miR-20b, miR-34a, and miR-454 was affected by delaying feed consumption for 48 h. For example, we found that delayed feeding resulted in increased miR-20b expression and conversely reduced expression of its target FADS1, an enzyme involved in fatty acid synthesis. Interestingly, the expression of a previously identified miR-20b regulator FOXO3 was also higher in delayed fed chicks. FOXO3 also functions in protection of cells from oxidative stress. Delayed fed chicks also had much higher levels of plasma ketone bodies than their normal fed counterparts. This suggests that delayed fed chicks rely almost exclusively on lipid oxidation for energy production and are likely under higher oxidative stress. Thus, it is possible that FOXO3 may function to both limit lipogenesis as well as to help protect against oxidative stress in peri-hatch chicks until the initiation of feed consumption. This is further supported by evidence that the FOXO3-regulated histone deacetylase (HDAC2) was found to recognize the FASN (involved in fatty acid synthesis) chicken promoter in a yeast one-hybrid assay. Expression of FASN mRNA was lower in delayed fed chicks until feed consumption. The present study demonstrated that many transcriptional and post-transcriptional mechanisms, including miRNA, form a complex interconnected regulatory network that is involved in controlling lipid and glucose molecular pathways during the metabolic transition in peri-hatch chicks.

Differential expression of hepatic genes with embryonic exposure to an environmentally relevant PCB mixture in Japanese quail (Coturnix japonica).

The upper Hudson River was contaminated with polychlorinated biphenyls (PCB) Aroclor mixtures from the 1940s until the late 1970s. Several well-established biomarkers, such as induction of hepatic cytochrome P450 monooxygenases, were used to measure exposure to PCBs and similar contaminants in birds. In the present study, Japanese quail eggs were injected with a PCB mixture based upon a congener profile found in spotted sandpiper eggs at the upper Hudson River and subsequently, RNA was extracted from hatchling liver tissue for hybridization to a customized chicken cDNA microarray. Nominal concentrations of the mixture used for microarray hybridization were 0, 6, 12, or 49 µg/g egg. Hepatic gene expression profiles were analyzed using cluster and pathway analyses. Results showed potentially useful biomarkers of both exposure and effect attributed to PCB mixture. Biorag and Ingenuity Pathway Analysis® analyses revealed differentially expressed genes including those involved in glycolysis, xenobiotic metabolism, replication, protein degradation, and tumor regulation. These genes included cytochrome P450 1A5 (CYP1A5), cytochrome b5 (CYB5), NADH-cytochrome b5 reductase, glutathione S-transferase (GSTA), fructose bisphosphate aldolase (ALDOB), glycogen phosphorylase, carbonic anhydrase, and DNA topoisomerase II. CYP1A5, CYB5, GSTA, and ALDOB were chosen for quantitative real-time polymerase chain reaction confirmation, as these genes exhibited a clear dose response on the array. Data demonstrated that an initial transcriptional profile associated with an environmentally relevant PCB mixture in Japanese quail occurred.