Hormone Signaling, Gene Expression, and Mitochondrial Hormone Receptor Expression in Avian Muscle (Cells)

Mitochondria are vital to the proper growth and function of muscle cells since they’re responsible for the majority of ATP production used for cellular energy. Previous studies have investigated how differences in mitochondrial function affects feed efficiency (FE) in broilers phenotyped for High and Low FE. Low FE broilers have been shown to have increased levels of reactive oxygen species (ROS), thus contributing to higher levels of oxidative stress and damage seen in these birds. Global gene and protein expression studies conducted on breast muscle of the High FE and Low FE phenotypes have suggested that differences in mitochondrial function and hormone signaling play a role in feed efficiency. In mammalian muscle cells, hormones such as the neuropeptide orexin are known to affect mitochondrial function. Therefore, the focus in this study was to determine whether hormones can affect mitochondrial dynamics in avian muscle cells, compare the expression of genes involved in muscle growth and insulin signaling in the High FE and Low FE phenotypes, and determine whether hormone receptors are present in the mitochondria of avian muscle cells. The actions of hormones and their receptors play an important role in the regulation of growth and metabolism. Investigation of orexin expression in avian muscle cells revealed that the hormone and its receptor are expressed in muscle. Orexin was also shown to be secreted by muscle cells and caused differential expression of a number of mitochondrial-related genes. Based on predictions generated by the results obtained from global expression studies, qRT-PCR analysis revealed several differentially expressed genes between the High and Low FE phenotype that are associated with muscle growth/development and the insulin signaling pathway. Lastly, due to the lack of scientific literature concerning the expression of hormone receptors in the mitochondria of avian muscle cells, studies were conducted that do indicate the presence of receptors in muscle mitochondria

Orexin System and Avian Muscle Mitochondria

In mammals, orexin A and B (also known as hypocretin 1 and 2) are two orexigenic peptides produced primarily by the lateral hypothalamus that signal through two G-protein-coupled receptors, orexin receptors 1/2, and have been implicated in the regulation of several physiological processes. However, the physiological roles of orexin are not well defined in avian (non-mammalian vertebrate) species. Recently, we made a breakthrough by identifying that orexin and its related receptors 1/2 (ORXR1/2) are expressed in avian muscle tissue and cell line, and appears to be a secretory protein. Functional in vitro studies showed that orexin A and B differentially regulated expression of the orexin system, suggesting that orexins might have autocrine, paracrine, and/or endocrine roles. Administration of recombinant orexin modulated mitochondrial biogenesis, dynamics, function, and bioenergetics. In this chapter, we include a brief overview of the (patho) physiological role of orexin, comparative findings between mammalian and avian orexin, and in-depth analysis of orexin’s action on avian muscle mitochondria

Progesterone signalling in broiler skeletal muscle is associated with divergent feed efficiency

Background: We contrast the pectoralis muscle transcriptomes of broilers selected from within a single genetic line expressing divergent feed efficiency (FE) in an effort to improve our understanding of the mechanistic basis of FE. Results: Application of a virtual muscle model to gene expression data pointed to a coordinated reduction in slow twitch muscle isoforms of the contractile apparatus (MYH15, TPM3, MYOZ2, TNNI1, MYL2, MYOM3, CSRP3, TNNT2), consistent with diminishment in associated slow machinery (myoglobin and phospholamban) in the high FE animals. These data are in line with the repeated transition from red slow to white fast muscle fibres observed in agricultural species selected on mass and FE. Surprisingly, we found that the expression of 699 genes encoding the broiler mitoproteome is modestly–but significantly–biased towards the high FE group, suggesting a slightly elevated mitochondrial content. This is contrary to expectation based on the slow muscle isoform data and theoretical physiological capacity arguments. Reassuringly, the extreme 40 most DE genes can successfully cluster the 12 individuals into the appropriate FE treatment group. Functional groups contained in this DE gene list include metabolic proteins (including opposing patterns of CA3 and CA4), mitochondrial proteins (CKMT1A), oxidative status (SEPP1, HIG2A) and cholesterol homeostasis (APOA1, INSIG1). We applied a differential network method (Regulatory Impact Factors) whose aim is to use patterns of differential co-expression to detect regulatory molecules transcriptionally rewired between the groups. This analysis clearly points to alterations in progesterone signalling (via the receptor PGR) as the major driver. We show the progesterone receptor localises to the mitochondria in a quail muscle cell line. Conclusions: Progesterone is sometimes used in the cattle industry in exogenous hormone mixes that lead to a ~20% increase in FE. Because the progesterone receptor can localise to avian mitochondria, our data continue to point to muscle mitochondrial metabolism as an important component of the phenotypic expression of variation in broiler FE

Gene Expression Essential for Myostatin Signaling and Skeletal Muscle Development Is Associated With Divergent Feed Efficiency in Pedigree Male Broilers

Background: Feed efficiency (FE, gain to feed) is an important genetic trait as 70% of the cost of raising animals is due to feed costs. The objective of this study was to determine mRNA expression of genes involved in muscle development and hypertrophy, and the insulin receptor-signaling pathway in breast muscle associated with the phenotypic expression of FE.Methods: Breast muscle samples were obtained from Pedigree Male (PedM) broilers (8 to 10 week old) that had been individually phenotyped for FE between 6 and 7 week of age. The high FE group gained more weight but consumed the same amount of feed compared to the low FE group. Total RNA was extracted from breast muscle (n = 6 per group) and mRNA expression of target genes was determined by real-time quantitative PCR.Results: Targeted gene expression analysis in breast muscle of the high FE phenotype revealed that muscle development may be fostered in the high FE PedM phenotype by down-regulation several components of the myostatin signaling pathway genes combined with upregulation of genes that enhance muscle formation and growth. There was also evidence of genetic architecture that would foster muscle protein synthesis in the high FE phenotype. A clear indication of differences in insulin signaling between high and low FE phenotypes was not apparent in this study.Conclusion: These findings indicate that a gene expression architecture is present in breast muscle of PedM broilers exhibiting high FE that would support enhanced muscle development-differentiation as well as protein synthesis compared to PedM broilers exhibiting low FE

Enrichment of Autophagy and Proteosome Pathways in Breast Muscle of Feed Efficient Pedigree Male Broilers

Background: Feed efficiency (FE) is an important genetic trait in poultry and livestock. Autophagy (self-eating) and proteosomes are cellular processes that remove damaged cell components (e.g., proteins, organelles). As evidence of extensive protein oxidation was observed in Pedigree Male (PedM) broilers exhibiting a low FE (LFE) phenotype compared to a high FE (HFE) phenotype, the main goal of this study was to assess gene and protein expression of the autophagy and proteosome pathways in breast muscle obtained in PedM broilers exhibiting HFE and LFE phenotypes.Methods: Feed efficiency was calculated as weight gain divided by feed intake gain in individual PedM broilers that were measured between 6 and 7 weeks of age. Targeted gene expression was conducted on breast muscle using quantitative real-time polymerase chain reaction (qPCR) to determine mRNA expression of genes associated with the autophagy pathway; AMP-activated protein kinase alpha 1 (AMPKα1), mammalian target of rapamycin (mTOR), Beclin 1, and autophagy genes (Atg) 3, Atg7, and Atg16L1. Binomial distribution analysis was conducted on transcriptomic and data obtained by RNAseq and shotgun proteomics, respectively on the same set of tissues for genes associated with autophagy, vacuole formation, and proteosome expression.Results: Greater efficiency was attained in the HFE PedM broilers by greater weight gain on the same amount of feed consumed resulting in FEs of 0.65 ± 0.01 and 0.46 ± 0.01 in the HFE and LFE phenotypes, respectively. Targeted mRNA expression analysis revealed significant (P < 0.05) elevations in AMPKa1, mTOR, Atg16L1, and Atg7 and a marginal (P = 0.07) elevation in Beclin1. Binomial distribution analysis transcriptomic and proteomic data revealed significant skews favoring autophagy-, vacuole-, and proteosome-related genes in the HFE phenotype. These results indicate that the autophagy and proteosome expression is enhanced in the HFE compared to the LFE pedigree male broiler phenotype suggesting that protein and organelle quality control may be enhanced in high feed efficiency

Regulacija sile hidrauličke preše pomoću programa LabVIEW i uređaja NI-myRIO

U ovom završnom radu načinjen je eksperiment regulacije sile hidrauličke preše pomoću programa LabVIEW i uređaja NI myRIO. LabVIEW je grafički programski jezik namijenjen lakšem sakupljanju podataka iz ljudske okoline sa raznih instrumenata. Najčešće se koristi za prikupljanje, analizu i obradu podataka te kontrolu instrumenata i opreme. Programi rađeni pomoću LabVIEW alata nazivaju se Virtualni Instrumenti, skraćeno VI (engl. Virtual Instruments), jer njihov izgled i rad imitira stvarne instrumente. Svaki VI se sastoji od prednjeg panela i blok dijagrama. Prednji panel (engl. Front panel) namijenjen je za izradu grafičkog sučelja koji korisnik vidi kada je sustav pokrenut, a blok dijagram (engl. Block diagram) sadrži programske elemente preko kojih se odvija obrada podataka. S elektro-hidrauličkim servo sustavom povezan je uređaj NI myRIO na kojem se izvršava upravljački program za regulaciju sile hidrauličke preše primjenom servo ventila. NI myRIO je laboratorijski uređaj koji omogućuje izvršavanje algoritma u stvarnom realnom vremenu. Za vizualizaciju i nadzor rada elektro-hidrauličkog servo sustava koristi se prijenosno računalo na kojem se nalazi grafičko sučelje

Orexin regulates mitochondrial dynamics in avian muscle

The growing obesity epidemic has sparked numerous studies on the identification of molecular signatures that regulate energy homeostasis using different experimental animal models. Orexin, which acts via two G-protein coupled receptors, orexin receptor 1 and 2, has been originally identified as feeding-related hypothalamic neuropeptide that regulate energy balance in mammals. Recently, using chicken, non-mammalian species that are characteristically hyperglycemic and prone to obesity, we made a breakthrough by identifying the orexin system in avian muscle and unraveling its effect on mitochondrial dynamics and function. Therefore, understanding orexin signaling and function may help to identify novel therapeutic opportunities for treating metabolic disorders related to mitochondrial dysfunction

Tissue distribution, gender- and genotype-dependent expression of autophagy-related genes in avian species.

As a result of the genetic selection of broiler (meat-type breeders) chickens for enhanced growth rate and lower feed conversion ratio, it has become necessary to restrict feed intake. When broilers are fed ad libitum, they would become obese and suffer from several health-related problems. A vital adaptation to starvation is autophagy, a self-eating mechanism for recycling cellular constituents. The autophagy pathway has witnessed dramatic growth in the last few years and extensively studied in yeast and mammals however, there is a paucity of information in avian (non-mammalian) species. Here we characterized several genes involved in autophagosome initiation and elongation in Red Jungle fowl (Gallus gallus) and Japanese quail (coturnix coturnix Japonica). Both complexes are ubiquitously expressed in chicken and quail tissues (liver, leg and breast muscle, brain, gizzard, intestine, heart, lung, kidney, adipose tissue, ovary and testis). Alignment analysis showed high similarity (50.7 to 91.5%) between chicken autophagy-related genes and their mammalian orthologs. Phylogenetic analysis demonstrated that the evolutionary relationship between autophagy genes is consistent with the consensus view of vertebrate evolution. Interestingly, the expression of autophagy-related genes is tissue- and gender-dependent. Furthermore, using two experimental male quail lines divergently selected over 40 generations for low (resistant, R) or high (sensitive, S) stress response, we found that the expression of most studied genes are higher in R compared to S line. Together our results indicate that the autophagy pathway is a key molecular signature exhibited gender specific differences and likely plays an important role in response to stress in avian species