ABSTRACT
Oncomodulins (OCMs), also known as non-α-parvalbumins, are small molecules known for their high-affinity binding of Ca2+ ions. They play crucial roles as Ca2+ buffers and participate in signaling pathways within muscle and neuron cells. In chickens, 3 oncomodulin molecules have been identified at the protein level and are named chicken oncomodulin 1 (OCM1), -3 (OCM3), and alpha-parvalbumin (PVALB). OCM4 was newly assigned by genome annotation. A gene cluster containing OCM1, OCM3, and OCM4 is located in chromosome 14, while a single gene of PVALB is on chromosome 1. The Ca2+ signaling pathway may be a potential contributor to the onset of chicken breast myopathies. However, chicken OCMs have not been extensively studied in muscle tissues. In this study, the genetic specifications, tissue-specific and differential expression of OCM1, OCM3, OCM4, and PVALB in the context of chicken breast myopathies were investigated. OCM1 exhibited moderate expression in the liver, intestine, and kidney. OCM3 was highly expressed in thymus and breast muscle. A long noncoding RNA (lncRNA) transcribed from the antisense strand of the OCM3 gene was found to be expressed in liver, lung, heart, intestine, and kidney tissues. OCM4 was barely expressed in thymus, thigh-, and breast muscle. PVALB exhibited high expression across all tissues examined. Results of quantitative PCR (qPCR) indicated that the expression of OCM3 was significantly increased (4.4 ± 0.7 fold; P-value = 0.03) in woody breast (WB) muscle and even greater (8.5 ± 0.6 fold; P-value = 0.004) in WB/white striping (WS) muscles. The expression of PVALB showed no difference in WB muscle, but it was notably higher (4.6 ± 0.7 fold; P-value = 0.054) in WB/WS muscle, although statistical significance was not reached. These findings suggest that increased expression of OCM3 and PVALB may be linked to chicken breast myopathies with regard to disruption of Ca2+ buffering.
Subject(s)
Avian Proteins , Chickens , Muscular Diseases , Poultry Diseases , Animals , Chickens/genetics , Poultry Diseases/genetics , Poultry Diseases/metabolism , Muscular Diseases/veterinary , Muscular Diseases/genetics , Muscular Diseases/metabolism , Avian Proteins/genetics , Avian Proteins/metabolism , Pectoralis Muscles/metabolism , Gene Expression Profiling/veterinaryABSTRACT
White striping (WS) is an emerging myopathy that results in significant economic losses as high as $1 billion (combined with losses derived from other breast myopathies including woody breast and spaghetti meat) to the global poultry industry. White striping is detected as the occurrence of white lines on raw poultry meat. The exact etiologies for WS are still unclear. Proteomic analyses of co-expressed WS and woody breast phenotypes previously demonstrated dysfunctions in carbohydrate metabolism, protein synthesis, and calcium buffering capabilities in muscle cells. In this study, we conducted shotgun proteomics on chicken breast fillets exhibiting only WS that were collected at approximately 6 h postmortem. After determining WS severity, protein extractions were conducted from severe WS meat with no woody breast (WB) condition (n = 5) and normal non-affected (no WS) control meat (n = 5). Shotgun proteomics was conducted by Orbitrap Lumos, tandem mass tag (TMT) analysis. As results, 148 differentially abundant proteins (|fold change|>1.4; p-value < 0.05) were identified in the WS meats compared with controls. The significant canonical pathways included BAG2 signaling pathway, glycogen degradation II, isoleucine degradation I, aldosterone signaling in epithelial cells, and valine degradation I. The potential upstream regulators include LIPE, UCP1, ATP5IF1, and DMD. The results of this study provide additional insights into the cellular mechanisms on the WS myopathy and meat quality.
Subject(s)
Avian Proteins , Chickens , Meat , Muscular Diseases , Pectoralis Muscles , Poultry Diseases , Proteomics , Animals , Muscular Diseases/veterinary , Muscular Diseases/pathology , Muscular Diseases/metabolism , Poultry Diseases/metabolism , Meat/analysis , Pectoralis Muscles/metabolism , Avian Proteins/metabolism , Avian Proteins/genetics , Proteome , Muscle Proteins/metabolism , Muscle Proteins/geneticsABSTRACT
OBJECTIVE: The aim of this study was to investigate the roles of ribonucleotide reductase subunit M2 (RRM2; subunit of ribonucleotide reductase) in severe woody breast (WB) and normal breast muscles. ANIMALS: 40 8-week-old male Ross-708 broiler chickens. METHODS: Quantitative PCR was performed to determine gene expression, and commercial ELISA/assay kits were used to obtain several enzymatic activities. RESULTS: Results showed that RRM2 activity (P = .0002) and RRM2 (P = .05) and hydroxymethylbilane synthase expression (impaired oxygen transport and metabolism, P = .002) were reduced in WB, while caveolin-3 (defected membrane integrity, P = .09), endoglin (increased fibrosis, P = .06), and secreted protein acidic rich in cysteine (metabolic dysregulation, P = .09) expression tended to increase in WB. WB tended to have increased levels of homocysteine (P = .06), aspartate aminotransferase mitochondria (P = .02), pyruvate kinase (P = .04), DNA damage (P = .06), creatine kinase (P = .05), and triglyceride (P = .002) but decreased ATPase activity (P = .01), all indicating mitochondria dysfunction and tissue damage. CLINICAL RELEVANCE: In this study, differences in various enzyme activities and increased DNA damage suggest that RRM2-mediated mitochondrial abnormalities may play a role in WB myopathy.
Subject(s)
Chickens , Mitochondrial Diseases , Animals , Male , DNA Damage , Mitochondrial Diseases/veterinaryABSTRACT
The wooden breast (WB) condition notably alters moisture content and water holding capacity (WHC) in broiler breast fillets. The purpose of this study was to investigate water properties during refrigerated storage from 4 h to 168 h postmortem using time domain nuclear magnetic resonance (TD-NMR). Water properties measured included mobility (T), proportion (P), and abundance per 100 g of meat (A). Changes in meat quality indicators including compression force, color, pH, cumulative purge loss, and proximate composition were also measured. Compression force and energy of the WB fillets were higher than normal fillets (P < 0.05). Slopes of changes in lightness of the WB and normal fillets were different in skin and bone side (P < 0.05). The slope of the purge loss from the WB fillets was higher than the normal fillets (P < 0.05). Time domain nuclear magnetic resonance analysis showed 4 water populations in intact broiler fillets with transverse relaxation time (T2) constants at approximately 4 to 5 milliseconds (ms) (designated as 2b, corresponding to hydration water or bound water), 40 to 60 ms (designated as 21, corresponding to intra-myofibrillar water or immobilized water), 80 to 210 ms (designated as 22a, corresponding to extra-myofibrillar water or free water with lower mobility) and 210 to 500 ms (designated as 22b, corresponding to extra-myofibrillar water or free water with higher mobility) during early postmortem storage (between 4 h and 72 h postmortem) and only 3 populations (2b, 21, and 22a) after 72 h postmortem. There were interaction effects (P < 0.05) between storage time and WB condition for all water properties except T2b, A2b/100 g, and T22b. The linear change of T21, P21, A21/100 g, T22a, A22a/100 g, P22b, and A22b/100 g in stored WB samples were different from the normal fillets (P < 0.05). During storage, P21 and A21/100 g of the WB fillets exhibited faster linear increases than those of the normal fillets, whereas T21 and T22a of the normal fillets and A22a/100 g, P22b, and A22b/100 g of the WB fillets showed faster linear decreases (P < 0.05). Our data demonstrate that the WB condition affects changes in water properties in broiler fillets during postmortem refrigerated storage.
Subject(s)
Pectoralis Muscles , Water , Animals , Water/analysis , Pectoralis Muscles/chemistry , Chickens , Meat/analysis , PressureABSTRACT
The cellular events leading to the development of the woody breast myopathy in broiler breast muscle are unclear. Affected woody breast muscle exhibits muscle fiber degeneration/regeneration, connective tissue accumulation, and adverse morphological changes in mitochondria. Ribonucleotide reductase (RNR) is an enzyme for the synthesis of dNTP, which is important for mitochondria DNA content (mtDNA). RNR consists of two subunits: RRM1/RRM2. A decrease in RRM2 is associated with a decrease in mtDNA and mitochondria proteins, leading to impaired ATP production. The objective of this study was to investigate potential RNR differences between woody breast (WB) and normal (N) breast muscle by examining RRM2 expression and associated pathways. Gene expression and enzyme activities were examined by qPCR and commercial kits. Results showed that RRM2 expression reduced for WB (p = 0.01) and genes related to mitochondria, including ATP6 (p = 0.03), COX1 (p = 0.001), CYTB (p = 0.07), ND2 (p = 0.001) and ND4L (p = 0.03). Furthermore, NDUFB7 and COX 14, which are related to mitochondria and ATP synthesis, tended to be reduced in WB. Compared to N, GLUT1 reduced for WB (p = 0.05), which is responsible for glucose transport in cells. Consequently, PDK4 (p = 0.0001) and PPARG (p = 0.008) increased in WB, suggesting increased fatty acid oxidation. Citric synthase activity and the NAD/NADH ratio (p = 0.02) both reduced for WB, while WB increased CHRND expression (p = 0.001), which is a possible indicator of high reactive oxygen species levels. In conclusion, a reduction in RRM2 impaired mitochondria function, potentially ATP synthesis in WB, by increasing fibrosis and the down-regulation of several genes related to mitochondria function.
ABSTRACT
Poultry meat is becoming one of the most important animal protein sources for human beings in terms of health benefits, cost, and production efficiency. Effective genetic selection and nutritional programs have dramatically increased meat yield and broiler production efficiency. However, modern practices in broiler production result in unfavorable meat quality and body composition due to a diverse range of challenging conditions, including bacterial and parasitic infection, heat stress, and the consumption of mycotoxin and oxidized oils. Numerous studies have demonstrated that appropriate nutritional interventions have improved the meat quality and body composition of broiler chickens. Modulating nutritional composition [e.g., energy and crude protein (CP) levels] and amino acids (AA) levels has altered the meat quality and body composition of broiler chickens. The supplementation of bioactive compounds, such as vitamins, probiotics, prebiotics, exogenous enzymes, plant polyphenol compounds, and organic acids, has improved meat quality and changed the body composition of broiler chickens.
ABSTRACT
To identify possible mechanisms involved in the development and progression of nonalcoholic fatty liver disease (NAFLD), we conducted shotgun proteomics analysis on liver of obese Zucker rats fed either casein (CAS) or soy protein isolate (SPI) for 8 and 16 weeks. Rats (7 weeks old, n = 8-9/group) were randomly assigned to either a CAS-based or an SPI-based diet. Rats were killed after 8 or 16 weeks of feeding and livers were stored at -80°C. Ingenuity Pathway Analysis (IPA) software was used to facilitate interpretation of proteomics data. Predictions of activation or inhibition of molecules in the data were made based on activation z-score and P value of overlap (P < .05). Activation z-scores ≥2.0 indicate that a molecule is predicted to be activated, whereas activation z-scores of less than or equal to -2.0 indicate that a target molecule is predicted to be inhibited. Upstream regulator analysis with IPA revealed Neuregulin 1 (NRG1) to be the top activated protein in (z-score = 2.48, P < .05), and MKNK1 as the top inhibited protein (z-score = -2.83, P < .05) in SPI diet compared with CAS diet after both 8 and 16 weeks of SPI feeding. Regulator effects analysis also predicted that some proteins would be participating, directly or indirectly, in the inhibition of immune response functions (such as leukocyte migration) and lipid metabolism (such as synthesis of lipids) in SPI-fed rats relative to CAS-fed rats. Our results suggest that SPI diet modifies the expression of proteins that could be involved in the reduction of NAFLD.
Subject(s)
Non-alcoholic Fatty Liver Disease , Animals , Liver/metabolism , Non-alcoholic Fatty Liver Disease/metabolism , Obesity/metabolism , Proteomics , Rats , Rats, Zucker , Soybean ProteinsABSTRACT
Prohibitin 1 (PHB1) is an evolutionarily conserved and ubiquitously expressed protein that stabilizes mitochondrial chaperone. Our previous studies showed that liver-specific Phb1 deficiency induced liver injuries and aggravated lipopolysaccharide (LPS)-induced innate immune responses. In this study, we performed RNA-sequencing (RNA-seq) analysis with liver tissues to investigate global gene expression among liver-specific Phb1-/-, Phb1+/-, and WT mice, focusing on the differentially expressed (DE) genes between Phb1+/- and WT. When 78 DE genes were analyzed for biological functions, using ingenuity pathway analysis (IPA) tool, lipid metabolism-related genes, including insulin receptor (Insr), sterol regulatory element-binding transcription factor 1 (Srebf1), Srebf2, and SREBP cleavage-activating protein (Scap) appeared to be downregulated in liver-specific Phb1+/- compared with WT. Diseases and biofunctions analyses conducted by IPA verified that hepatic system diseases, including liver fibrosis, liver hyperplasia/hyperproliferation, and liver necrosis/cell death, which may be caused by hepatotoxicity, were highly associated with liver-specific Phb1 deficiency in mice. Interestingly, of liver disease-related 5 DE genes between Phb1+/- and WT, the mRNA expressions of forkhead box M1 (Foxm1) and TIMP inhibitor of metalloproteinase (Timp1) were matched with validation for RNA-seq in liver tissues and AML12 cells transfected with Phb1 siRNA. The results in this study provide additional insights into molecular mechanisms responsible for increasing susceptibility of liver injuries associated with hepatic Phb1.
ABSTRACT
Woody breast (WB) myopathy results in poor muscle quality. The increasing incidence of WB over the last several years indicates a need for improved prediction or early diagnosis. We hypothesized that the use of body fluids, including blood, may be more suitable than breast muscle tissue in developing a minimally invasive diagnostic tool for WB detection. To identify potential early-age-biomarkers that may represent the potential onset of WB, blood samples were collected from 100, 4 wks old commercial male broilers. At 8 wks of age, WB conditions were scored by manual palpation. A total of 32 blood plasma samples (eight for each group of WB and non-WB control birds at two time points, 4 wks and 8 wks) were subjected to shotgun proteomics and untargeted metabolomics to identify differentially abundant plasma proteins and metabolites in WB broilers compared to non-WB control (Con) broilers. From the proteomics assay, 25 and 16 plasma proteins were differentially abundant (p < 0.05) in the 4 and 8 wks old samples, respectively, in WB compared with Con broilers. Of those, FRA10A associated CGG repeat 1 (FRAG10AC1) showed >2-fold higher abundance in WB compared with controls. In the 8 wks old broilers, 4 and 12 plasma proteins displayed higher and lower abundances, respectively, in WB compared with controls. Myosin heavy chain 9 (MYH9) and lipopolysaccharide binding protein (LBP) showed more than 2-fold higher abundances in WB compared with controls, while transferrin (TF) and complement C1s (C1S) showed more than 2-fold lower abundances compared with controls. From the untargeted metabolomics assay, 33 and 19 plasma metabolites were differentially abundant in birds at 4 and 8 wks of age, respectively, in WB compared with controls. In 4 wks old broilers, plasma 3-hydroxybutyric acid (3-HB) and raffinose concentrations showed the highest and lowest fold changes, respectively, in WB compared with controls. The blood plasma 3-HB and raffinose concentrations were confirmed with targeted biochemical assays. Blood biomarkers, such as 3-HB and raffinose, may be suitable candidate targets in the prediction of WB onset at early ages.
ABSTRACT
A study was conducted to understand the differentially expressed genes in Pectoralis (P) major under woody breast (WB) myopathy condition in a high yielding broiler strain using RNA-sequencing at the growing (d 21) and finishing (d 42 and d 56) grow-out ages. Follow-up study was conducted to understand the in vivo triglyceride (TG) synthesis (d 49) occurring in adipogenic tissues using deuterium oxide (2H2O) as a metabolic tracer. Results indicated the top physiological systems affected in myopathy broiler were related to the musculo-skeletal system (d 21, 42, and 56) and cardiovascular system (d 42 and 56). Ubiquitin-specific proteases are expressed higher in myopathy broiler at d 21 (OTUD1) and d 42 (SACS) that potentially indicated higher degradation of muscle protein occurring at those ages. While genes related to transcription factors and muscle cell differentiation (ZNF234, BTG2) and muscle growth (IGF1) were upregulated with myopathy broiler suggesting concurrent muscle fiber regeneration. The downregulation of PYGB and MGAM genes related to carbohydrate transport and metabolism at d 42 potentially indicated nutrient-deficient state of myopathy affected fibers; whereas the nutrient-deficient physiological state of cells seemed to be counteracted by up-regulation of genes related to carbohydrate (ALDOB, GPD1L2) at d 56. There was a reduced (P < 0.05) in vivo TG synthesis in liver of the myopathy broiler (0.123 %/hr) compared to non-myopathy broiler (0.197 %/hr). The majority of TG synthesized in liver with myopathy broiler could conceivably be delivered to P. major (rather than to abdominal fat pad storage) to fulfil the increased energy need of muscle cells (via TG lipolysis and fatty acid [FA] oxidation). The increased utilization of FAs in the WB affected muscle could result in reduced secretion of FAs into blood circulation leading to sub-optimal availability of FAs for re-esterification for TG synthesis in liver. Results indicated that myopathy broiler at later age (d 56) of grow-out period were synchronously going through adaptive physiological processes of feedback responses to adverse cellular states.
Subject(s)
Muscular Diseases , Poultry Diseases , Animals , Chickens/genetics , Follow-Up Studies , Gene Expression , Kinetics , Muscular Diseases/veterinary , Pectoralis Muscles , TriglyceridesABSTRACT
Mitigation of stress is of great importance in poultry production, as chronic stress can affect the efficiency of production traits. Selective breeding with a focus on stress responses can be used to combat the effects of stress. To better understand the genetic mechanisms driving differences in stress responses of a selectively bred population of Japanese quail, we performed genomic resequencing on 24 birds from High Stress (HS) and Low Stress (LS) lines of Japanese quail using Illumina HiSeq 2 × 150 bp paired end read technology in order to analyze Single Nucleotide Polymorphisms (SNPs) within the genome of each line. SNPs are common mutations that can lead to genotypic and phenotypic variations in animals. Following alignment of the sequencing data to the quail genome, 6,364,907 SNPs were found across both lines of quail. 10,364 of these SNPs occurred in coding regions, from which 2886 unique, non-synonymous SNPs with a SNP% ≥ 0.90 and a read depth ≥ 10 were identified. Using Ingenuity Pathway Analysis, we identified genes affected by SNPs in pathways tied to immune responses, DNA repair, and neurological signaling. Our findings support the idea that the SNPs found within HS and LS lines of quail could direct the observed changes in phenotype.
Subject(s)
Coturnix/genetics , Polymorphism, Single Nucleotide/genetics , Stress, Physiological/genetics , Animals , DNA Repair/genetics , Genome/genetics , Genomics/methods , Genotype , PhenotypeABSTRACT
Obesity can lead to several health disorders including nonalcoholic fatty liver disease (NAFLD), the aggregation of lipids within hepatocytes, and consequent inflammation of the liver tissue. Previously, we reported that feeding obese Zucker rats with soy protein isolate (SPI) can reduce liver steatosis. To understand how SPI reduced liver steatosis, we conducted global gene expression analysis on liver samples obtained from these rats after short- (8 weeks) and long-term SPI feeding (16 weeks). We compared and contrasted these data using Ingenuity Pathway Analysis (IPA) software. This study focused mainly on target molecules that could be participating in inflammation processes and lipid metabolism that are well-known components of NAFLD. Inflammatory response was predicted to be inhibited in animals fed the SPI diet at both 8 and 16 weeks of experiment. This general prediction was based on negative activation z scores obtained through IPA (z score < -2.0, P < .00001) for eight aspects of immune function/inflammatory response. Lipid metabolism was predicted to be strongly enhanced in rats fed the SPI diet for 16 weeks than for 8 weeks. This prediction was based on positive activation z scores (z scores >2.0, P < .00001) of eight functions involved in lipid transport and metabolism. We observed that the longer the rats were fed the SPI diet, the more beneficial it resulted against NAFLD. Based on our findings, the predicted reductions in inflammatory mechanisms while enhancing lipid transport out of the liver could be the reasons behind the reduction of liver steatosis.
Subject(s)
Non-alcoholic Fatty Liver Disease , Soybean Proteins , Animals , Inflammation/genetics , Liver , Non-alcoholic Fatty Liver Disease/genetics , Obesity/genetics , Rats , Rats, ZuckerABSTRACT
Probiotics often play an important role in improving gut health in chickens through multiple mechanisms, including enhancement of tight junctions, nutrient acquisition, niche colonization, or coaggregation with enteric pathogens. The objective of this study was to characterize lactic acid bacteria (LAB) isolated from the gut of healthy broiler chickens for a number of phenotypes that might be indicative of good probiotic potentials. A total 40 bacterial isolates were isolated from 3-week-old chickens using Man, Rogosa and Sharpe (MRS) agar plates. The bacterial isolates were evaluated in vitro for motility, autoaggregation, pathogen inhibition, pH of overnight culture, growth on different agar plates, and their impact on gut integrity. Selected isolates were genotyped by sequencing the 16S-23S rRNA gene intergenic region. Based on the phenotype and genotype, we identified 20 potential probiotic (PP) isolates that belong to LAB. Multivariate analysis showed that PP isolates were positively correlated with parameters such as growth on MRS agar plate (pH 5.5), pathogen inhibition, and autoaggregation. However, growth on MacConkey agar plates, supernatant pH, motility, and transepithelial electrical resistance were negatively correlated with the PP isolates. Furthermore, in vivo study needs to be performed for evaluation of the utility of these probiotic candidates in poultry production.
Subject(s)
Chickens/microbiology , Gastrointestinal Microbiome , Lactobacillales/physiology , Probiotics , Animals , Caco-2 Cells , Electric Impedance , Humans , Hydrogen-Ion Concentration , Lactobacillales/classification , Lactobacillales/genetics , Lactobacillales/growth & development , Movement , Phenotype , RNA, Ribosomal, 16S/genetics , RNA, Ribosomal, 23S/geneticsABSTRACT
Limited studies reported mechanisms by which microRNAs (miRNA) are interlinked in the etiology of fructose-induced non-alcoholic fatty liver disease (NAFLD). Here, we aimed to investigate the significance of miRNAs in fructose-induced NAFLD pathogenesis through unbiased approaches. In experiment I, C57BL/6N mice were fed either water or 34% fructose for six weeks ad libitum. In experiment II, time course effects of fructose intervention were monitored using the same conditions; mice were killed at the baseline, fourth, and sixth weeks. Bioinformatic analyses for hepatic proteomics revealed that SREBP1 is the most significant upstream regulator influenced by fructose; miR-33-5p (miR-33) was identified as the key miRNA responsible for SREBP1 regulation upon fructose intake, which was validated by in vitro transfection assay. In experiment II, we confirmed that the longer mice consumed fructose, the more severe liver injury markers (e.g., serum AST) appeared. Moreover, hepatic Srebp1 mRNA expression was increased depending upon the duration of fructose consumption. Hepatic miR-33 was time-dependently decreased by fructose while serum miR-33 expression was increased; these observations indicated that miR-33 from the liver might be released upon cell damage. Finally we observed that fructose-induced ferroptosis might be a cause of liver toxicity, resulting from oxidative damage. Collectively, our findings suggest that fructose-induced oxidative damage induces ferroptosis, and miR-33 could be used as a serological biomarker of fructose-induced NAFLD.
Subject(s)
Fructose/adverse effects , Lipogenesis/physiology , Liver/metabolism , MicroRNAs/metabolism , Non-alcoholic Fatty Liver Disease/blood , Animals , Biomarkers/blood , Diet , Female , Fructose/administration & dosage , Gene Expression Regulation/drug effects , Lipogenesis/genetics , Male , Mice , Mice, Inbred C57BL , MicroRNAs/genetics , Sterol Regulatory Element Binding Protein 1/genetics , Sterol Regulatory Element Binding Protein 1/metabolismABSTRACT
In an effort to understand the apparent trade-off between the continual push for growth performance and the recent emergence of muscle pathologies, shotgun proteomics was conducted on breast muscle obtained at ~8 weeks from commercial broilers with wooden breast (WB) myopathy and compared with that in pedigree male (PedM) broilers exhibiting high feed efficiency (FE). Comparison of the two proteomic datasets was facilitated using the overlay function of Ingenuity Pathway Analysis (IPA) (Qiagen, CA, USA). We focused on upstream regulator analysis and disease-function analysis that provides predictions of activation or inhibition of molecules based on (a) expression of downstream target molecules, (b) the IPA scientific citation database. Angiopoeitin 2 (ANGPT2) exhibited the highest predicted activation Z-score of all molecules in the WB dataset, suggesting that the proteomic landscape of WB myopathy would promote vascularization. Overlaying the FE proteomics data on the WB ANGPT2 upstream regulator network presented no commonality of protein expression and no prediction of ANGPT2 activation. Peroxisome proliferator coactivator 1 alpha (PGC1α) was predicted to be inhibited, suggesting that mitochondrial biogenesis was suppressed in WB. PGC1α was predicted to be activated in high FE pedigree male broilers. Whereas RICTOR (rapamycin independent companion of mammalian target of rapamycin) was predicted to be inhibited in both WB and FE datasets, the predictions were based on different downstream molecules. Other transcription factors predicted to be activated in WB muscle included epidermal growth factor (EGFR), X box binding protein (XBP1), transforming growth factor beta 1 (TGFB1) and nuclear factor (erythroid-derived 2)-like 2 (NFE2L2). Inhibitions of aryl hydrocarbon receptor (AHR), AHR nuclear translocator (ARNT) and estrogen related receptor gamma (ESRRG) were also predicted in the WB muscle. These findings indicate that there are considerable differences in upstream regulators based on downstream protein expression observed in WB myopathy and in high FE PedM broilers that may provide additional insight into the etiology of WB myopathy.
ABSTRACT
To determine how soy protein isolate (SPI) ameliorated liver steatosis in male obese Zucker rats, we conducted global transcriptomic expression (RNAseq) analysis on liver samples of male rats fed either the SPI or a control casein (CAS)-based diet (n = 8 per group) for 16 weeks. Liver transcriptomics were analyzed using an Ilumina HiSeq system with 2 × 100 base pair paired-end reads method. Bioinformatics was conducted using Ingenuity Pathway Analysis (IPA) software (Qiagen, CA) with P < 0.05 and 1.3-fold differential expression cutoff values. Regression analysis between RNAseq data and targeted mRNA expression analysis of 12 top differentially expressed genes (from the IPA program) using quantitative PCR (qPCR) revealed a significant regression analysis (r 2 = 0.69, P = 0.0008). In addition, all qPCR values had qualitatively similar direction of up- or down-regulation compared to the RNAseq transcriptomic data. Diseases and function analyses that were based on differentially expressed target molecules in the dataset predicted that lipid metabolism would be enhanced whereas inflammation was predicted to be inhibited in SPI-fed compared to CAS-fed rats at 16 weeks. Combining upstream regulator and regulator effects functions in IPA facilitates the prediction of upstream regulators (e.g., transcription regulators) that could play important roles in attenuating or promoting liver steatosis due to SPI or CAS diets. Upstream regulators that were predicted to be activated (from expression of down-stream targets) linked to increased conversion of lipid and transport of lipid in SPI-fed rats included hepatocyte nuclear factor 4 alpha (HNF4A) and aryl hydrocarbon receptor (AHR). Upstream regulators that were predicted to be activated in CAS-fed rats linked to activation of phagocytosis and neutrophil chemotaxis included colony stimulating factor 2 and tumor necrosis factor. The results provide clear indication that long-term SPI-fed rats exhibited diminished inflammatory response and increased lipid transport in liver compared to CAS-fed rats that likely would contribute to reduced liver steatosis in this obese Zucker rat model.
ABSTRACT
Diets high in red meats, particularly meats cooked at high temperature, increase the risk of colon cancer due to a production of heterocyclic aromatic amines (HAAs). Of the identified HAAs, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is the most mass abundant colon carcinogen in charred meat or fish. Here, we comprehensively examined sex-dependent colon transcriptome signatures in response to PhIP treatment to identify biological discrepancies. Eight-week-old male and female C57BL/6N mice were intraperitoneally injected with PhIP (10 mg/kg of body weight) and colon tissues were harvested 24 h after PhIP injection, followed by colon transcriptomics analysis. A list of differentially expressed genes (DEGs) was utilized for computational bioinformatic analyses. Specifically, overrepresentation test using the Protein Analysis Through Evolutionary Relationships tool was carried out to annotate sex-dependent changes in transcriptome signatures after PhIP treatment. Additionally, the most significantly affected canonical pathways by PhIP treatment were predicted using the Ingenuity Pathway Analysis. As results, male and female mice presented different metabolic signatures in the colon transcriptome. In the male mice, oxidative phosphorylation in the mitochondrial respiratory chain was the pathway impacted the most; this might be due to a shortage of ATP for DNA repair. On the other hand, the female mice showed concurrent activation of lipolysis and adipogenesis. The present study provides the foundational information for future studies of PhIP effects on underlying sex-dependent mechanisms.
Subject(s)
Aminopyridines , Colon/metabolism , Colorectal Neoplasms/metabolism , Imidazoles , Sex Characteristics , Transcriptome , Animals , Colon/drug effects , Colorectal Neoplasms/chemically induced , Colorectal Neoplasms/genetics , Female , Male , Mice, Inbred C57BLABSTRACT
Woody breast (WB) myopathy is significantly impacting modern broilers and is imposing a huge economic burden on the poultry industry worldwide. Yet, its etiology is not fully defined. In a previous study, we have shown that hypoxia and the activation of its upstream mediators (AKT/PI3K/mTOR) played a key role in WB myopathy, and supplementation of quantum blue (QB) can help to reduce WB severity via modulation of hypoxia-related pathways. To gain further insights, we undertook here a metabolomics approach to identify key metabolite signatures and outline their most enriched biological functions. Ultra performance liquid chromatography coupled with high resolution mass spectrometry (UPLC-HRMS) identified a total of 108 known metabolites. Of these, mean intensity differences at P < 0.05 were found in 60 metabolites with 42 higher and 18 lower in WB-affected compared to unaffected muscles. Multivariate analysis and Partial Least Squares Discriminant analysis (PLS-DA) scores plot displayed different clusters when comparing metabolites profile from affected and unaffected tissues and from moderate (MOD) and severe (SEV) WB muscles indicating that unique metabolite profiles are present for the WB-affected and unaffected muscles. To gain biologically related molecule networks, a stringent pathway analyses was conducted using IPA knowledge-base. The top 10 canonical pathways generated, using a fold-change -1.5 and 1.5 cutoff, with the 50 differentially abundant-metabolites were purine nucleotide degradation and de novo biosynthesis, sirtuin signaling pathway, citrulline-nitric oxide cycle, salvage pathways of pyrimidine DNA, IL-1 signaling, iNOS, Angiogenesis, PI3K/AKT signaling, and oxidative phosphorylation. The top altered bio-functions in term of molecular and cellular functions in WB-affected tissues included cellular development, cellular growth and proliferation, cellular death and survival, small molecular biochemistry, inflammatory response, free radical scavenging, cell signaling and cell-to-cell interaction, cell cycles, and lipid, carbohydrate, amino acid, and nucleic acid metabolisms. The top disorder functions identified were organismal injury and abnormalities, cancer, skeletal and muscular disorders, connective tissue disorders, and inflammatory diseases. Breast tissues from birds fed with high dose (2,000 FTU) of QB phytase exhibited 22 metabolites with significantly different levels compared to the control group with a clear cluster using PLS-DA analysis. Of these 22 metabolites, 9 were differentially abundant between WB-affected and unaffected muscles. Taken together, this study determined many metabolic signatures and disordered pathways, which could be regarded as new routes for discovering potential mechanisms of WB myopathy.
ABSTRACT
Myopathies (Woody Breast (WB) and White Striping (WS)) of broiler chickens have been correlated with fast growth. Recent studies reported that localized hypoxia and metabolic impairment may involve in these myopathies of birds. In order to better understand the stress response mechanisms affecting myopathies of broilers, the aim of this study was to examine effects of WB and both WB/WS on stress hormone corticosterone (CORT) levels and expressional changes of stress response genes including glucocorticoid (GC) receptor (GR), 11ß-Hydroxysteroid dehydrogenase type 1 (11ß-HSD1), DNA methylation regulators (DNMTs), and arginine vasotocin receptor 1a and 1b (V1aR, V1bR). Results of radioimmunoassay showed that CORT levels of WB and WB/WS birds were significantly higher compared to Con (p < 0.05), however, the combination of WB/WS was not significantly higher than WB birds, implying that the effects of WB and WS on CORT are not synergistic. Hepatic GR expression of both WB and WB/WS birds were significantly higher compared to Con (p < 0.05). However, GR expression levels in breast muscle of both WB and WB/WS birds were decreased compared to Con (p < 0.05). Hepatic 11ß-HSD1 expression was increased only in WB/WS birds compared to Con birds with no significant difference between Con and WB birds. 11ß-HSD1 expression was decreased and increased in WB and WB/WS birds compared to Con, respectively, in breast muscle (p < 0.05). DNMT1 expression was significantly decreased in both muscle and liver of WB birds, and in muscle of WB/WS birds, but not in liver of WB/WS birds, indicating differential effects of WS on the epigenetical stress response of muscle and liver compared to WB. V1aR expression was significantly increased in muscle of WB birds, and in liver of WB/WS birds compared to Con birds (p < 0.05). V1bR was not changed in muscle and liver of WB birds compared to Con birds. Taken together, results suggest that GC-induced myopathies occur in fast-growing broiler chickens and circulating CORT level might be a significant biochemical marker of myopathies (WB and WS) of birds. In addition, chronic stress responses in breast muscle and tissue-specific epigenetic changes of stress response genes by DNMTs may play a critical role in the occurrence of myopathies.
Subject(s)
Chickens/physiology , Muscular Diseases/physiopathology , Muscular Diseases/veterinary , Stress, Physiological , 11-beta-Hydroxysteroid Dehydrogenase Type 1/genetics , 11-beta-Hydroxysteroid Dehydrogenase Type 1/metabolism , Animals , Body Weight , Chickens/blood , Chickens/genetics , Corticosterone/blood , DNA Methylation/genetics , Female , Gene Expression Regulation, Developmental , Liver/metabolism , Mammary Glands, Animal/metabolism , Muscles/metabolism , Muscular Diseases/blood , Muscular Diseases/genetics , Organ Specificity , Receptors, Glucocorticoid/metabolism , Receptors, Vasopressin/genetics , Receptors, Vasopressin/metabolismABSTRACT
Obesity predisposes people to a variety of chronic metabolic diseases. Identification of natural factors that prevent the development of obesity is likely to be the most successful means of ameliorating the current obesity epidemic. Patchouli alcohol is a sesquiterpene alcohol found in Pogostemon cablin and possesses health benefit activities. This study was designed to examine if patchouli alcohol affects adipogenesis, and investigates the underlying mechanisms whereby patchouli alcohol exerts antiobesity effect. 3T3-L1 adipocytes were differentiated with treatment of different concentrations of patchouli alcohol. An in vivo study was performed to test the effect of patchouli alcohol gavage on a high-fat diet (HFD)-induced obesity. Treatment of patchouli alcohol reduced lipid accumulation in 3T3-L1 adipocytes in a dose-dependent manner without toxicity. Regarding mechanism, treatment of patchouli alcohol reduced expression of peroxisome proliferator-activated receptor-gamma (PPARγ) and CCAAT-enhancer-binding protein-alpha (C/EBPα) and increased expression of total and active ß-catenin in 3T3-L1 adipocytes. Oral gavage of patchouli alcohol led to a significant reduction of body weight and fat accumulation in the mice fed with HFD. Transcriptome analysis indicates that smad7 is most highly activated gene in patchouli alcohol-treated 3T3-L1 cells. Patchouli alcohol possesses health benefit effect through inhibiting adipogenesis and fat tissue development.