Angus primary muscle cells grow faster than primary skeletal muscle cells derived from Hereford and WagyuX cattle.

<p>The number of cells (cell index) was measured in culture using xCELLigence technology during two defined phases of growth. A. Proliferation Phase I (5–20 hours in culture) and B. Proliferation Phase II (20–72 hours in culture). (NOTE: Standard error bars in A and B are displayed as either positive or negative SE values only so the means are clearly visible and there is no overlap in error bars).</p

Expression of muscle regulatory factors in un-differentiated <i>bovine</i> primary skeletal muscle cells isolated from Angus, Hereford and WagyuX cattle.

<p>Expression of <i>PAX7</i> (A), <i>MYF5</i> (B), <i>MYOD</i> (C) and <i>MYOG</i> (D) are expressed as mean normalised gene expression (<b>MNE</b>) using <i>bovine HPRT</i> (hypoxanthineguanine phosphoribosyltransferase) as the housekeeping gene. Error bars represent SEM.</p

Liveweights (kg) and carcass weights (kg) of 6 months old Angus (n = 4), Hereford (n = 5) and Wagyu X Angus (n = 4) Angus cattle sampled for <i>bovine</i> primary skeletal muscle cells¹.

<p>¹Data was kindly obtained Casino abattoir near Lismore, New South Wales. Australia.</p><p>Liveweights (kg) and carcass weights (kg) of 6 months old Angus (n = 4), Hereford (n = 5) and Wagyu X Angus (n = 4) Angus cattle sampled for <i>bovine</i> primary skeletal muscle cells<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124468#t002fn001" target="_blank">¹</a>.</p

Determination of myogenic cell population.

<p>A. Expression of the myogenic marker desmin and nuclei (DAPI) in undifferentiated primary skeletal muscle cells from WagyuX, Angus and Hereford cattle. B. Mean ± SEM proportion of the total number of cells positive for desmin (myogenic cells). A good proportion of cells were positive for desmin for all breeds. There was no difference in proportion of desmin positive cells between WagyuX (97.33 ± 2.0), Angus (98.12 ± 2.0) and Hereford (95.67 ± 4.3). Scale bar is equal to 60μm.</p

Proliferation rate of <i>bovine</i> primary skeletal muscle cells <i>in vitro</i> positively correlates with carcase weight.

<p>Rate of proliferation of <i>bovine</i> primary skeletal muscle cells <i>in vitro</i> versus carcase weight (6-months old) for A. Phase I (5–20 hours in culture)(R² = 0.34) and B. Phase II (20–72 hours in culture)(R² = 0.23)(B). P < 0.05 for linear regressions in A and B.</p

Primer sequences used for qPCR analysis of myogenic gene expression in <i>bovine</i> primary muscle cells.

<p>^<b>1</b> Hypoxanthineguanine phosphoribosyltransferase.</p><p>Primer sequences used for qPCR analysis of myogenic gene expression in <i>bovine</i> primary muscle cells.</p

Proliferation rate of <i>bovine</i> primary skeletal muscle cells <i>in vitro</i> is positively correlated with liveweight of cattle at slaughter.

<p>A. Rate of proliferation of <i>bovine</i> primary skeletal muscle cells <i>in vitro</i> versus liveweight of cattle at slaughter (6-months old) for Phase I (5–20 hours in culture)(R² = 0.43) and B. Phase II (20–72 hours in culture)(R² = 0.30). P < 0.05 for linear regressions in A and B.</p

Impacts of the Callipyge Mutation on Ovine Plasma Metabolites and Muscle Fibre Type

<div><p>The ovine Callipyge mutation causes postnatal muscle hypertrophy localized to the pelvic limbs and torso, as well as body leanness. The mechanism underpinning enhanced muscle mass is unclear, as is the systemic impact of the mutation. Using muscle fibre typing immunohistochemistry, we confirmed muscle specific effects and demonstrated that affected muscles had greater prevalence and hypertrophy of type 2X fast twitch glycolytic fibres and decreased representation of types 1, 2C, 2A and/or 2AX fibres. To investigate potential systemic effects of the mutation, proton NMR spectra of plasma taken from lambs at 8 and 12 weeks of age were measured. Multivariate statistical analysis of plasma metabolite profiles demonstrated effects of development and genotype but not gender. Plasma from Callipyge lambs at 12 weeks of age, but not 8 weeks, was characterized by a metabolic profile consistent with contributions from the affected hypertrophic fast twitch glycolytic muscle fibres. Microarray analysis of the perirenal adipose tissue depot did not reveal a transcriptional effect of the mutation in this tissue. We conclude that there is an indirect systemic effect of the Callipyge mutation in skeletal muscle in the form of changes of blood metabolites, which may contribute to secondary phenotypes such as body leanness.</p></div

Metabolic pathways involved in postnatal lamb development.

<p>Metabolites indentified in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099726#pone-0099726-t002" target="_blank">Table 2</a> are summarized according to their occurrence in metabolic pathways as annotated by KEGG metabolic pathways. ↓, metabolites decreasing with age (blue); ↑, metabolites increasing with age (red). TMAO, trimethylamine N-oxide; HCHO, formaldehyde; CH₄, methane. The reactions contained in the boxed section are likely to be derived from rumen microbial metabolism, except for reactions denoted by **, which occur in mammalian tissues <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099726#pone.0099726-Asatoor1" target="_blank">[63]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099726#pone.0099726-Yeung1" target="_blank">[84]</a>, and reactions denoted by *, which can occur both in mammalian tissues or the gut microbiome <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099726#pone.0099726-Zhang1" target="_blank">[85]</a>. This representation of the data recognises that plasma metabolites report the combined metabolic activities of all major tissues as well as the rumen microbiome.</p

Metabolic pathway analysis.

<p>The metabolic pathways are represented as circles according to their scores from enrichment (vertical axis) and topology analyses (pathway impact, horizontal axis) using MetaboAnalyst 2.0 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099726#pone.0099726-Xia1" target="_blank">[56]</a>. Darker circle colors indicate more significant changes of metabolites in the corresponding pathway. The size of the circle corresponds to the pathway impact score and is correlated with the centrality of the involved metabolites. The metabolic pathways involved in age differences are shown in panels A and B, and the pathways perturbed due to the Callipyge mutation are summarized in panels C and D. Panels A and C were generated using the <i>Homo sapiens</i> library, while the <i>Bos taurus</i> library was selected for production of panels B and D, respectively. Pathways were annotated by numbering when the <i>P</i> values calculated from the enrichment analysis were <0.05. The annotated pathways include: 1, Aminoacyl-tRNA biosynthesis; 2, Arginine and proline metabolism; 3, Biotin metabolism; 4, Cyanoamino acid metabolism; 5, D-Glutamine and D-glutamate metabolism; 6, Galactose metabolism; 7, Glycine, serine and threonine metabolism; 8, Lysine degradation; 9, Methane metabolism; 10, Nitrogen metabolism; 11, Synthesis and degradation of ketone bodies. The color of each metabolic pathway is related to the <i>P</i> value obtained from enrichment analysis and its size represents the fold enrichment score <i>i.e.</i> −ln(<i>P</i>).</p