title: Post-mortem stability of RNA in skeletal muscle and adipose tissue and the tissue-specific expression of myostatin, perilipin and associated factors in the horse.
abstract: Obesity, a major concern for equine welfare, is highly prevalent in the leisure horse population. Skeletal-muscle and adipose tissues are important determinants of maintenance energy requirements. The myostatin and perilipin pathways play key roles in the regulation of muscle mass and lipolysis respectively and have both been associated with obesity predisposition in other mammalian species. High quality samples, suitable for molecular biology, are an essential prerequisite for detailed investigations of gene and protein expression. Hence, this study has evaluated a) the post-mortem stability of RNA extracted from skeletal-muscle and adipose-tissues collected under commercial conditions and b) the tissue-specific presence of myostatin, the moystatin receptor (activin receptor IIB, ActRIIB), follistatin and perilipin, genes and proteins across a range of equine tissues. Objectives were addressed using tissues from 7 Thoroughbred horses presented for slaughter at a commercial abattoir; a) samples were collected at 7 time-points from Masseter muscle and perirenal adipose from 5 minutes to 6 hours post-mortem. Extracted RN was appraised by Optical Density analysis and agarose-gel electrophoresis. b) Quantitative real time PCR and Western Blotting were used to evaluate gene and protein expression in anatomically-defined samples collected from 17 tissues (6 organs, 4 skeletal muscles and 7 discrete adipose depots). The results indicate that, under the present collection conditions, intact, good quality RNA could be extracted from skeletal-muscle for up to 2 hours post-mortem. However, RNA from adipose tissue may be more susceptible to degradation/contamination and samples should be collected no later than 30 minutes post-mortem. The data also show that myostatin and ActRIIB genes and proteins were almost exclusively expressed in skeletal muscle. The follistatin gene showed a more diverse gene expression profile, with expression evident in several organs, adipose tissue depots and skeletal muscles. Perilipin gene and protein were almost exclusively expressed by adipose tissue.
authors: University of Liverpool, Department of Obesity and Endocrinology, Faculty of Health and Life Sciences, Leahurst Campus, Neston, Wirral, United Kingdom.
authors: Philippa K
authors: PK
authors: Morrison
authors: University of Liverpool, Department of Obesity and Endocrinology, Faculty of Health and Life Sciences, Leahurst Campus, Neston, Wirral, United Kingdom.
authors: Chen
authors: C
authors: Bing
authors: Equine Studies Group, WALTHAM Centre for Pet Nutrition, Waltham-on-the-Wolds, Melton Mowbray, Leicestershire, United Kingdom.
authors: Patricia A
authors: PA
authors: Harris
authors: University of Liverpool, Department of Obesity and Endocrinology, Faculty of Health and Life Sciences, Leahurst Campus, Neston, Wirral, United Kingdom.
authors: Charlotte A
authors: CA
authors: Maltin
authors: University of Liverpool, Department of Obesity and Endocrinology, Faculty of Health and Life Sciences, Leahurst Campus, Neston, Wirral, United Kingdom.
authors: Dai
authors: D
authors: Grove-White
authors: University of Liverpool, Department of Obesity and Endocrinology, Faculty of Health and Life Sciences, Leahurst Campus, Neston, Wirral, United Kingdom.
authors: Caroline McG
authors: CM
authors: Argo
mesh_terms: D030301:Activin Receptors, Type II / Q000235:genetics / Q000378:metabolism; D000273:Adipose Tissue / Q000378:metabolism*; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D005260:Female; D038681:Follistatin / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D020043:Genes, Essential; D006736:Horses / Q000235:genetics*; D008297:Male; D018482:Muscle, Skeletal / Q000378:metabolism*; D055435:Myostatin / Q000235:genetics* / Q000378:metabolism; D009928:Organ Specificity / Q000235:genetics*; D000070778:Perilipin-1; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism; D011180:Postmortem Changes; D020871:RNA Stability; D060888:Real-Time Polymerase Chain Reaction; D012984:Software; D013053:Spectrophotometry
chemical_list: D002352:Carrier Proteins; D038681:Follistatin; D055435:Myostatin; D000070778:Perilipin-1; D010750:Phosphoproteins; D030301:Activin Receptors, Type II; C425584:activin receptor type II-B