Detection of chromosomal regions showing differential gene expression in human skeletal muscle and in alveolar rhabdomyosarcoma

Background  

Rhabdomyosarcoma is a relatively common tumour of the soft tissue, probably due to regulatory disruption of growth and differentiation of skeletal muscle stem cells. Identification of genes differentially expressed in normal skeletal muscle and in rhabdomyosarcoma may help in understanding mechanisms of tumour development, in discovering diagnostic and prognostic markers and in identifying novel targets for drug therapy.     

Identification of mycorrhiza-regulated genes with arbuscule development-related expression profile

Suppressive subtractive hybridisation was applied to the analysis of late stage arbuscular mycorrhizal development in pea. 96 cDNA clones were amplified and 81, which carried fragments more than 200 nt in size, were sequence analysed. Among 67 unique fragments, 10 showed no homology and 10 were similar to sequences with unknown function. RNA accumulation of the corresponding 67 genes was analysed by hybridisation of macro-arrays. The cDNAs used as probes were derived from roots of wild type and late mutant pea genotypes, inoculated or not with the AM fungus Glomus mosseae. After calibration, a more than 2.5-fold mycorrhiza-induced RNA accumulation was detected in two independent experiments in the wild type for 25 genes, 22 of which seemed to be induced specifically during late stage AM development. Differential expression for 7 genes was confirmed by RT-PCR using RNA from mycorrhiza and from controls of a different pea cultivar. In order to confirm arbuscule-related expression, the Medicago truncatula EST data base was screened for homologous sequences with putative mycorrhiza-related expression and among a number of sequences with significant similarities, a family of trypsin inhibitor genes could be identified. Mycorrhiza-induced RNA accumulation was verified for five members by real-time PCR and arbuscule-related activation of the promoter could be shown in transgenic roots for one of the genes, MtTi1.     

Investigating the genetic regulation of the expression of 63 lipid metabolism genes in the pig skeletal muscle

A comprehensive and systematic view of the genetic regulation of lipid metabolism genes is still lacking in pigs. Herewith, we have investigated the genetic regulation of 63 porcine genes with crucial roles in the uptake, transport, synthesis and catabolism of lipids. With this aim, we have performed an expression QTL (eQTL) scan in 104 pigs with available genotypes for the Illumina Porcine SNP60 chip and microarray measurements of gene expression in the gluteus medius muscle. Analysis of the data with gemma software revealed 13 cis‐ and 18 trans‐eQTL modulating the expression of 19 loci. Genes regulated by eQTL participated in a wide array of lipid metabolism pathways such as the β‐oxidation of fatty acids, lipid biosynthesis and lipolysis, fatty acid activation and desaturation, lipoprotein uptake, apolipoprotein assembly and cholesterol trafficking. These data provide a first picture of the genetic regulation of loci involved in porcine lipid metabolism.     

Dietary l-arginine supplementation differentially regulates expression of lipid-metabolic genes in porcine adipose tissue and skeletal muscle

Obesity is a major health crisis worldwide and new treatments are needed to fight this epidemic. Using the swine model, we recently reported that dietary l-arginine (Arg) supplementation promotes muscle gain and reduces body-fat accretion. The present study tested the hypothesis that Arg regulates expression of key genes involved in lipid metabolism in skeletal muscle and white adipose tissue. Sixteen 110-day-old barrows were fed for 60 days a corn- and soybean-meal-based diet supplemented with 1.0% Arg or 2.05% l-alanine (isonitrogenous control). Blood samples, longissimus dorsi muscle and overlying subcutaneous adipose tissue were obtained from 170-day-old pigs for biochemical studies. Serum concentrations of leptin, alanine and glutamine were lower, but those for Arg and proline were higher in Arg-supplemented pigs than in control pigs. The percentage of oleic acid was higher but that of stearic acid and linoleic acid was lower in muscle of Arg-supplemented pigs, compared with control pigs. Dietary Arg supplementation increased mRNA levels for fatty acid synthase in muscle, while decreasing those for lipoprotein lipase, glucose transporter-4, and acetyl-coenzyme A carboxylase-α in adipose tissue. Additionally, mRNA levels for hormone sensitive lipase were higher in adipose tissue of Arg-supplemented pigs compared with control pigs. These results indicate that Arg differentially regulates expression of fat-metabolic genes in skeletal muscle and white adipose tissue, therefore favoring lipogenesis in muscle but lipolysis in adipose tissue. Our novel findings provide a biochemical basis for explaining the beneficial effect of Arg in improving the metabolic profile in mammals (including obese humans).     

Comparison of false discovery rate methods in identifying genes with differential expression

Current high-throughput techniques such as microarray in genomics or mass spectrometry in proteomics usually generate thousands of hypotheses to be tested simultaneously. The usual purpose of these techniques is to identify a subset of interesting cases that deserve further investigation. As a consequence, the control of false positives among the tests called "significant" becomes a critical issue for researchers. Over the past few years, several false discovery rate (FDR)-controlling methods have been proposed; each method favors certain scenarios and is introduced with the purpose of improving the control of FDR at the targeted level. In this paper, we compare the performance of the five FDR-controlling methods proposed by Benjamini et al., the qvalue method proposed by Storey, and the traditional Bonferroni method. The purpose is to investigate the "observed" sensitivity of each method on typical microarray experiments in which the majority (or all) of the truth is unknown. Based on two well-studied microarray datasets, it is found that in terms of the "apparent" test power, the ranking of the FDR methods is given as Step-down相似文献   

Exploration of genes showing intramuscular fat deposition-associated expression changes in musculus longissimus muscle

Marbling, as defined by the amount of intramuscular fat, is an economically important trait in beef cattle. Intramuscular fat deposition is postulated to arise mainly from a series of adipogenic events in intramuscular adipocyte-lineage cells and in the physiological or anatomical milieux surrounding them. This study was designed to investigate gene-expression patterns associated with fat deposition in musculus longissimus muscle, including adipocyte-lineage cells and part of the milieux. Differential-display PCR (ddPCR) was used to examine expression differences between low-marbled and high-marbled steer groups at 8, 10, 12 and 14 months of age, encompassing the time that marbling starts to appear. Seventy-four of 2114 total bands on ddPCR gel-bands were significant (P < 0.05) for the group effect, the interaction effect between group and age, or both the group and the interaction effects. Sequence analysis of 72 of these bands revealed 77 genes, including 35 annotated genes and 42 novel sequences. Among the 35 annotated genes, 6 (BTG2, PDHB, SORBS1, TRDN, TTN and MGP) have been related to changes in intramuscular fat deposition, possibly by exerting effects on adipocyte-lineage cells or on the milieux surrounding them.     

Bacteria associated with skeletal tissue growth anomalies in the coral Platygyra carnosus

Scleractinian corals with growth anomalies, often referred to as 'tumors', have been reported globally. A recent survey of Hong Kong waters showed that > 60% of Platygyra carnosus colonies developed tumors. Here we report for the first time, the bacterial community associated with tumors in P. carnosus over different seasons and locations in Hoi Ha Wan Marine Park and Port Shelter. Culture-based methods for strain isolation and molecular techniques of 16S rRNA analysis for strain identification were used, as well as the culture-independent technique terminal-restriction fragment length polymorphism. We tested the hypothesis that the community composition would be considerably different between healthy and tumor corals and aimed to investigate whether potential differences because of tumors would override the seasonal and spatial influences. Our analysis detected only minor differences between the communities associated with the healthy and tumor corals, indicating that tumors are not associated with major changes in the bacterial community structure. In contrast, community structure was strongly influenced by the location and season, with greater Alphaproteobacteria diversity in the winter than in the summer. This study demonstrated that the coral-associated bacterial community composition was more related to environmental variables (i.e. season and location) than to disease (i.e. tumor).     

Identification of genes showing altered expression in preimplantation and early postimplantation parthenogenetic embryos

Uniparental embryos have been instrumental in studying imprinting because contributions from the parental genomes can be determined unambiguously. In this study, we set out to identify imprinted genes showing differential expression between parthenogenetic and fertilized embryos during preimplantation and early postimplantation stages of development. We identified three genes-apolipoprotein E, pyruvate kinase-3, and protein phosphatase 1 gamma-that represent excellent candidates for imprinted genes, based on the results of the differential screen, their function in differentiation and the cell cycle, and their location within imprinted chromosomal regions. In addition, two novel genes expressed in trophoblast were identified, 1661 and RA81. These genes, together with four known imprinted genes, H19, Igf2r, Igf2, and Snrpn, showed evidence of expression from both parental alleles in early stage embryos, indicating a role for postfertilization processes in regulating imprinted gene function. © 1995 Wiley-Liss, Inc.     

Atypical expression of circadian clock genes in denervated mouse skeletal muscle

Muscle force production and power output in active males, regardless of the site of measurement (hand, leg, or back), are higher in the evening than in the morning. This diurnal variation is attributed to motivational, peripheral and central factors, and higher core and, possibly, muscle temperatures in the evening. This study investigated whether increasing morning rectal temperatures to evening resting values, by active or passive warm-ups, leads to muscle force production and power output becoming equal to evening values in motivated subjects. Ten healthy active males (mean ± SD: age, 21.2 ± 1.9 yrs; body mass, 75.4 ± 8 kg; height, 1.76 ± .06 m) completed the study, which was approved by the University Ethics Committee. The subjects were familiarized with the techniques and protocol and then completed four sessions (separated by at least 48 h): control morning (07:30 h) and evening (17:30 h) sessions (with an active 5-min warm-up) and then two further sessions at 07:30 h but proceeded by an extended active or passive warm-up to raise rectal temperature to evening values. These last two sessions were counterbalanced in order of administration. During each trial, three measures of handgrip strength, isokinetic leg strength measurements (of knee flexion and extension at 1.05 and 4.19 rad.s^?1 through a 90° range of motion), and four measures of maximal voluntary contraction (MVC) on an isometric ergometer (utilizing the twitch-interpolation technique) were performed. Rectal and intra-aural temperatures, ratings of perceived exertion (RPE) and thermal comfort (TC) were measured. Measurements were made after the subjects had reclined for 30 min and after the warm-ups and prior to the measurement of handgrip and isokinetic and isometric ergometry. Muscle temperature was taken after the warm-up and immediately before the isokinetic and MVC measurements. Warm-ups were either active (cycle ergometer at 150 W) or passive (resting in a room at 35°C, relative humidity 45%). Data were analyzed using analysis of variance models with repeated measures. Rectal and intra-aural temperatures were higher at rest in the evening (.56°C and .74°C; p < .05) than in the morning, but there were no differences after the active or passive warm-ups, the subjects' ratings of thermal comfort reflecting this. Muscle temperatures also displayed significant diurnal variation, with higher values in the evening (～.31°C; p < .05). Grip strength, isokinetic knee flexion for peak torque and peak power at 1.05 rad.s^?1, and knee extension for peak torque at 4.19 rad.s^?1 all showed higher values in the evening. All other measures of strength or power showed a trend to be higher in the evening ( .10 > p > .05). There was no significant effect of active or passive warm-ups on any strength or power variable, and subjects reported maximal values for effort for each strength measure. In summary, effects of time of day were seen in some measures of muscle performance but, in this population of motivated subjects, there was no evidence that increasing morning rectal temperature to evening values by active or passive warm-up increased muscle strength to evening values. (Author correspondence: B.J.Edwards@ljmu.ac.uk)     

Differential expression of lipid metabolism-related genes and myosin heavy chain isoform genes in pig muscle tissue leading to different meat quality

The aim of this study was to investigate the variations in meat quality, lipid metabolism-related genes, myosin heavy chain (MyHC) isoform genes and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) gene mRNA expressions in longissimus dorsi muscle (LM) of two different pig breeds. Six Rongchang and six Landrace barrows were slaughtered at 161 days of age. Subsequently, meat quality traits and gene expression levels in LM were observed. Results showed that Rongchang pigs not only exhibited greater pH, CIE a*_{24 h} and intramuscular fat content but also exhibited lower body weight, carcass weight, dressing percentage, LM area and CIE b*_{24 h} compared with Landrace pigs (P<0.05). Meanwhile, the mRNA expression levels of the lipogenesis (peroxisome proliferator-activated receptor gamma, acetyl-CoA carboxylase and fatty acid synthase) and fatty acid uptake (lipoprotein lipase)-related genes were greater in the Rongchang (P<0.05), whereas the lipolysis (adipose triglyceride lipase and hormone sensitive lipase) and fatty acid oxidation (carnitine palmitoyltransferase-1B)-related genes were better expressed in the Landrace. Moreover, compared with the Landrace, the mRNA expression levels of MyHCI, MyHCIIa and MyHCIIx were greater, whereas the mRNA expression levels of MyHCIIb were lower in the Rongchang pigs (P<0.05). In addition, the mRNA expression levels of PGC-1α were greater in Rongchang pigs than in the Landrace (P<0.05), which can partly explain the differences in MyHC isoform gene expressions between Rongchang and Landrace pigs. Although the small number of samples does not allow to obtain a definitive conclusion, we can suggest that Rongchang pigs possess better meat quality, and the underlying molecular mechanisms responsible for the better meat quality in fatty pigs may be partly due to the higher mRNA expression levels of lipogenesis and fatty acid uptake-related genes, as well as the oxidative and intermediate muscle fibers, and due to the lower mRNA expression levels of lipolysis and fatty acid oxidation-related genes, as well as the glycolytic muscle fibers.     

Identification of sarcolemma-associated antigens with differential distributions on fast and slow skeletal muscle fibers

We have identified three sarcolemma-associated antigens, including two antigens that are differentially distributed on skeletal muscle fibers of the fast, fast/slow, and slow types. Monoclonal antibodies were prepared using partially purified membranes of adult chicken skeletal muscles as immunogens and were used to characterize three antigens associated with the sarcolemma of muscle fibers. Immunofluorescence staining of cryosections of adult and embryonic chicken muscles showed that two of the three antigens differed in expression by fibers depending on developmental age and whether the fibers were of the fast, fast/slow, or slow type. Fiber type was assigned by determining the content of fast and slow myosin heavy chain. MSA-55 was expressed equally by fibers of all types. In contrast, MSA-slow and MSA-140 differed in their expression by muscle fibers depending on fiber type. MSA-slow was detected exclusively at the periphery of fast/slow and slow fibers, but was not detected on fast fibers. MSA-140 was detected on all fibers but fast/slow and slow fibers stained more intensely suggesting that these fiber types contain more MSA-140 than fast fibers. These sarcolemma-associated antigens were developmentally regulated in ovo and in vitro. MSA-55 and MSA-140 were detected on all primary muscle fibers by day 8 in ovo of embryonic development, whereas MSA-slow was first detected on muscle fibers just before hatching. Those antigens expressed by fast fibers (MSA-55 and MSA-140) were expressed only after myoblasts differentiated into myotubes, but were not expressed by fibroblasts in cell culture. Each antigen was also detected in one or more nonskeletal muscle cell types: MSA-55 and MSA-slow in cardiac myocytes and smooth muscle of gizzard (but not vascular structures) and MSA-140 in cardiac myocytes and smooth muscle of vascular structures. MSA-55 was identified as an Mr 55,000, nonglycosylated, detergent-soluble protein, and MSA-140 was an Mr 140,000, cell surface protein. The Mr of MSA-slow could not be determined by immunoblotting or immunoprecipitation techniques. These findings indicate that muscle fibers of different physiological function differ in the components associated with the sarcolemma. While the function of these sarcolemma-associated antigens is unknown, their regulated appearance during development in ovo and as myoblasts differentiate in culture suggests that they may be important in the formation, maturation, and function of fast, fast/slow, and slow muscle fibers.