Faculty of Medicine, Dentistry and Health Sciences Department of Physiology

Introduction

Muscle wasting and weakness are common in many disease states and conditions including ageing, cancer cachexia, denervation, disuse, unloading/microgravity, and the muscular dystrophies. Although the maintenance of muscle mass is generally regarded as a simple balance between protein synthesis and protein degradation, these mechanisms are not strictly independent but in fact are coordinated by a number of different and sometimes complementary signalling pathways. Our laboratory is interested in examining these different molecular pathways and their role in the etiology of muscle wasting disorders.

Research from this laboratory has led to the discovery that…

• Sarcopenia can be ameliorated in rats by treatment with b₂-adrenoceptor agonists
• Treatment with the cytokine Interleukin-15 or the growth factor insulin-like growth factor-I, improves the function of muscles from dystrophic mice
• Inhibition of the calcineurin signalling cascade impairs muscle structure and function in young dystrophic mice
• b₂-adrenoceptors are involved in the repair process of muscles after injury and adrenoceptor stimulation via an agonist can speed up recovery from injury

In addition, work from this laboratory has resulted in a fully optimised electroporation protocol for studying gene expression in skeletal muscle

2009 B&CM Laboratory Research Profiles

Basic and Clinical Myology Laboratory (2009) - [Back row, from left] Mr. Tim Naim (research assistant), Mr. Nic Tirtaatmadja (PhD student), Dr. Chris van der Poel (Research Officer), Dr. René Koopman (Research Fellow), Mr. Tim Sayer (Honours student), Mr. Ben Gleeson (research assistant); [Middle row, from left] Ms. Jennifer Trieu (research assistant), Ms Annabel Chee (Honours student), Prof. Gordon Lynch (laboratory head), Ms. Fiona Colarossi (laboratory manager), Mr. Marc Nicolas (PhD student), Dr. Kate Murphy (Research Fellow); [Front row, from left] Dr. Jarrod Church (Senior Research Officer), Mr. Daniel Ham (PhD student), Dr. Brendan Adams (Research Officer), Mr. Stefan Gehrig (PhD student), Dr. Bertrand Leger (Honorary Research Fellow). Absent - Ms. Leah Harcourt (PhD student), Miss. Sarah Turpin (PhD student), .

Latest News

• December 2008
• Congratulations to Dr. Robert Mayne for successful completion of his Doctor of Clinical Dentistry.
• Dr. Jonathan Schertzer was awarded the Chancellor’s Prize (Medicine) from the University – based on the quality of the research published from his Ph.D. thesis.
• Dr. Jonathan Schertzer was awarded the Dean’s Award for Excellence in a Ph.D. thesis.
• Dr. Jonathan Schertzer was awarded the 2008 Postdoctoral Publication Award from the Australian Physiological Society.
• Dr. James Ryall was awarded the 2008 A.K. McIntyre Medal from the Australian Physiological Society for outstanding contributions to Australian physiology by a young investigator over their predoctoral and early postdoctoral years.
• Stefan Gehrig was awarded the best Student Speaker Award at the Australian Physiological Society annual meeting in Melbourne, December 2008
• Stefan Gehrig was awarded a National Heart Foundation Postgraduate Scholarship
• Dr. René Koopman was successful in being awarded an Ajinomoto Amino Acid Research Program (3ARP), Ajinomoto, Japan. Project title: Citrulline as a pharmaco-nutrient to attenuate muscle wasting.
• Dr. Jarrod Church and Prof. Gordon Lynch have been awarded a Rebecca L. Cooper Medical Research Foundation award. Project title: Investigating the potential for erectile dysfunction drugs to enhance the repair of injured skeletal muscles in the elderly.
• Dr. Kate Murphy was awarded an Australian Association of Gerontology Inc. R.M. Gibson Scientific Research Fund award. Project title: Smad7 over-expression: a novel therapy for attenuating ageing-related muscle weakness.
• Stefan Gehrig awarded second prize in the Under the Coverslip ~ Imaging and Microscopy Competition for "Where's Wally"
• Congratulations to Dr. Brendan Adams on successful completion of his PhD.
• November 2008
• Dr. René Koopman was awarded the MSD Prijs voor Sportgeneeskunde, for best scientific thesis in the field of Sports Medicine in the Netherlands, Noordwijkerhout, The Netherlands, 2008
• Dr. René Koopman was awarded the NUTRIM thesis award, for best scientific thesis 2006-2007, Nutrition and Toxicology Research Institute Maastricht, Maastricht, The Netherlands, 2008
• October 2008
• HUGE congratulations to all 4 honours students for an excellent honours year that was reflected by 4 H1's. Congratulations to both Radhika and Sarah for getting into Medicine at Deakin, and Jo for accepting an APA scholarship to begin her PhD investigating multiple sclerosis. All the best girls.
• September 2008
• Congratulations to Dr. Jarrod Church who has been awarded an Early Career Research Grant from the University of Melbourne
• August 2008
• We are extremely proud to report that Dr. Jonathan Schertzer has been awarded the Chancellor’s Prize for Excellence in the Ph.D. Thesis for the Faculties of Medicine, Dentistry and Health Sciences and Veterinary Sciences. Jon’s thesis, "Non-viral gene transfer of growth factors in skeletal muscle: implications for injury, regeneration and disease", has generated many exceptional, high-quality international publications.  The Chancellor's Prize for Excellence in the PhD Thesis is awarded on an annual basis and consists of a medal for desk display. Jon’s success follows on from that of Dr. James Ryall who was awarded the 2007 Chancellor’s Prize for Excellence in the Ph.D. Thesis. Congratulations Jon!

2009 Student Research Projects

Ms. Leah Harcourt (PhD project) - Investigating novel agents for ameliorating the pathophysiology of muscular dystrophy.

Miss. Sarah Turpin (PhD project) - Lipid-induced apoptosis in skeletal muscle.

Mr. Stefan Gehrig (PhD project) - The role of IGF binding proteins in IGF-I mediated signalling in the mdx dystrophic mouse.

Mr. Nicolin Tirtaatmadja (PhD project) -Biomolecule Delivery Systems for Tissue Engineering and Skeletal Muscle Regeneration.

Mr. Daniel Ham (PhD project) - Skeletal muscle modifications with exercise and ergogenic supplements.

Mr. Marc Nicolas (PhD project) - Role of beta-adrenergic signalling in skeletal muscle regeneration.

Mr. Tim Sayer (Honours project) - Effect of Hsp70 overexpression on EC coupling of dystrophic mice.

Ms Annabel Chee (Honours project) - Effect of myostatin inhibition on attenuating cancer cachexia.

A single muscle fibre expressing green fluorescent protein. Nuclei have been labelled with ethidium bromide and appear yellow.

Collaborators

• Dr. Paul Gregorevic (Baker IDI Heart and Diabetes Institute). Gene therapy and skeletal muscle.
• Dr. Craig Harrison (Prince Henry’s Institute of Medical Research, Victoria, Australia). TGF-beta signalling in muscular dystrophy.
• Dr. Aaron Russell (School of Exercise and Nutrition Sciences, Deakin University, Victoria, Australia). Role of Notch signalling in muscle health and disease.
• Dr. Stéphane Walrand & Prof. Yves Boirie (Human Nutrition, Unit, INRA Clermont-Ferrand, France). Citrulline as a pharmaco-nutrient to attenuate muscle wasting.
• Prof. Luc Cynober (Nutrition Laboratory, Pharmacy Faculty, University of Paris). Citrulline as a pharmaco-nutrient to attenuate muscle wasting.
• Prof. João Bizario (Associação de Amigos dos Portadores de Distrofia Muscular, São Paulo, Brazil). Characterisation of the Golden Retriver model of muscular dystrophy.
• Prof. Miranda Grounds (University of Western Australia). Functional regeneration in skeletal muscles during ageing and in muscular dystrophy.
• Prof. Mark Hargreaves (Department of Physiology, The University of Melbourne). Functional and metabolic characteristics of artificial skeletal muscles.
• Dr. Glenn McConnell (Department of Physiology, The University of Melbourne). nNOS signalling in skeletal muscle regeneration.
• Assoc. Prof. Edna Hardeman & Dr. Anthony Kee (Department of Anatomy, School of Medical Sciences, University of New South Wales, Sydney). Role of cytoskeletal proteins in skeletal muscle contraction.
• Prof. David A. Williams (Department of Physiology, The University of Melbourne). In situ and in vitro skeletal muscle calcium imaging.
• Prof. Bruce E. Kemp, Dr. Belinda Michell & Dr. Greg Steinberg (St. Vincent’s Hospital Research Institute, The University of Melbourne). AMP-kinase during intense muscle activity; calcineurin activity in skeletal muscle; aging and skeletal muscle metabolism.
• Prof. Wayne A. Morrison (Bernard O’Brien Institute for Microsurgery, The University of Melbourne). Improving skeletal muscle repair after ischaemia-reperfusion injury.
• Prof. George E. Muscat (Institute for Molecular Bioscience, The University of Queensland). Nuclear hormone receptors and skeletal muscle.
• Prof. Eric Olson & Assoc. Prof. Rhonda Bassel-Duby (University of Texas-Southwestern, U.S.A.). Role of the calcineurin signal transduction pathway in muscle regeneration and muscular dystrophy.
• Prof. Martin N. Sillence (Queensland University of Technology, Qld.). b₂-adrenoceptors, b₂-agonists and skeletal muscle function.
• Prof. Geoff W. Stevens & Dr. Andrea O'Connor (Department of Chemical & Biomolecular Engineering, The University of Melbourne). Interactions between the interfaces of solid-liquid and liquid-liquid systems for the dispersion of novel growth factors.
• Assoc. Prof. Matthew J. Watt (Department of Physiology, Monash University). Molecular and cell biology of lipid metabolism.
• Prof. Michael G. Woods (School of Dentistry, The University of Melbourne). Functional properties of human facial muscles and their relation to clinical dentistry.
• Prof. Jeffrey Zajac & Dr. Helen Maclean (Department of Medicine Austin and Repatriation Hospital). Androgens and skeletal muscle function.
• Assoc. Prof. Luc Gosselin (Exercise and Nutrition Sciences, University at Buffalo, The State University of New York). Ageing, inflammation and muscle repair.
• Prof. Mark Febbraio (Baker IDI Heart and Diabetes Institute). Role of heat shock proteins in muscle damage and repair in muscular dystrophy.

Current Funding Sources

NHMRC

G.S. Lynch, P. Gregorevic

The University of Melbourne

C. van der Poel

The University of Melbourne

K.T. Murphy

2007

ARC

G.S. Lynch, D.A. Williams

NHMRC

G.S. Lynch, W.A. Morrison, G.W. Stevens

The University of Melbourne

J.G. Ryall

2006

MDA (USA)

G.S. Lynch

Pfizer Inc. (USA)

G.S. Lynch

ARC

G.S. Lynch

Recent B&CM Laboratory Publications

2009

Porrello ER, Bell JR, Schertzer JD, Curl CL, McMullen JR, Mellor KM, Ritchie RH, Lynch GS, Harrap SB, Thomas WG, Delbridge LM. (2009). HERITABLE PATHOLOGIC CARDIAC HYPERTROPHY IN ADULTHOOD IS PRECEDED BY NEONATAL CARDIAC GROWTH RESTRICTION. Am J Physiol Regul Integr Comp Physiol. (in press)

2008

Lynch GS. (2008). Update on emerging drugs for sarcopenia - age-related muscle wasting. Expert Opin Emerg Drugs.13(4): 655-73.

Vlahovich N, Kee AJ, Van der Poel C, Kettle E, Hernandez-Deviez D, Lucas C, Lynch GS, Parton RG, Gunning PW, Hardeman EC. (2008). Cytoskeletal Tropomyosin Tm5NM1 Is Required for Normal Excitation-contraction Coupling in Skeletal Muscle. Mol Biol Cell. (in press).

van der Poel C, Edwards JN, Macdonald WA, Stephenson DG. (2008). Effect of temperature-induced reactive oxygen species production on excitation-contraction coupling in mammalian skeletal muscle. Clin Exp Pharmacol Physiol. 35(12):1482-7.

MacLean, H.E., Chiu, M.W.S., Notini, A.J., Axell, A-M, Davey, R.A., McManus, J.F., Ma, C., Plant, D.R., Lynch, G.S. & Zajac, J.D. (2008). Impaired skeletal muscle mass and function in male but not female androgen receptor knockout mice. FASEB J. 22: 2676-2689

Dzamko N, Schertzer JD, Ryall JG, Steel R, Macaulay SL, Wee S, Chen ZP, Michell BJ, Oakhill JS, Watt MJ, Jørgensen SB, Lynch GS, Kemp BE, Steinberg GR. (2008). AMPK-independent pathways regulate skeletal muscle fatty acid oxidation. J Physiol. 586(Pt 23): 5819-31.

Pearen, M.A., Myers, S., Ryall, J.G., Lynch, G.S. & Muscat, G.E.O. (2008). The orphan nuclear receptor, NOR-1, regulates gene expression that controls oxidative metabolism in skeletal muscle. Endocrinology. 149: 2853-2865

Ryall JG, Lynch GS. (2008). The potential and the pitfalls of beta-adrenoceptor agonists for the management of skeletal muscle wasting. Pharmacol Ther. 120(3) :219-32.

Schertzer JD. & Lynch GS. (2008). Plasmid-based gene transfer in mouse skeletal muscle by electroporation. Methods Mol Biol. 433: 115-25.

Gehrig SM, Ryall JG, Schertzer JD, Lynch GS. (2008). IGF-I analogue protects muscles of dystrophic mdx mice from contraction-mediated damage. Exp. Physiol. (in press).

Lynch GS, Faulkner JA, Brooks SV. (2008). Force deficits and breakage rates after single lengthening contractions of single fast fibers from unconditioned and conditioned muscles of young and old rats. Am. J. Physiol. Cell Physiol. 295: C249-C256.

Grounds MD, Radley HG, Lynch GS, Nagaraju K, De Luca A. (2008). Towards developing standard operating procedures for pre-clinical testing in the mdx mouse model of Duchenne muscular dystrophy. Neurobiol Dis. 31: 1-19.

Lynch GS, Schertzer JD, Ryall JG. (2008). Anabolic agents for improving muscle regeneration and function after injury. Clin. Exper. Pharmacol. Physiol. 35: 852-858.

Ryall JG, Schertzer JD, Murphy KT, Allen AM & Lynch GS (2008). Chronic b₂-adrenoceptor stimulation impairs cardiac relaxation via reduced SR Ca²⁺-ATPase activity. Am. J. Physiol. Heart Circ. Physiol.294: H2587-H2595.

Ryall JG, Schertzer JD, Alabakis TM, Gehrig SM, Plant DR & Lynch GS (2008). Intramuscular b₂-agonist administration enhances regeneration and functional repair after myotoxic injury in rat skeletal muscle. J. Appl. Physiol.105: 165-172.

**Ryall JG, Schertzer JD & Lynch GS (2008). Cellular and molecular mechanisms underlying age-related skeletal muscle wasting and weakness. Biogerontol.9: 213-228.

Stupka N, Schertzer JD, Bassel-Duby R, Olson EN & Lynch GS (2008). Stimulation of calcineurin-Aa activity attenuates muscle pathophysiology in mdx dystrophic mice. Am. J. Physiol. Reg. Int. Comp. Physiol. 294: R983-R992.

Schertzer JD*, van der Poel C*, Shavlakadze T, Grounds MD & Lynch GS (2008). Muscle specific overexpression of IGF-I improves E-C coupling in skeletal muscle fibers from dystrophic mdx mice. Am. J. Physiol. Cell. Physiol. 294:C161-168. [*- these authors contributed equally to this manuscript]

**Lynch GS & Ryall JG (2008). Role of b-adrenoceptor signaling in skeletal muscle: Implications for muscle wasting and disease. Physiol. Rev. 88:729-767.

Book Chapters

Schertzer JD, & Lynch GS (2008). Methods for optimising non-viral ectopic gene expression in normal and regenerating muscle. In (ed) Le Doux JM Gene Therapy Protocols Humana Press, 115-125.

2007

Beitzel F, Sillence MN & Lynch GS (2007). b-adrenoceptor signaling in regenerating skeletal muscle after b-agonist administraion. Am. J. Physiol. Endocrinol. Metab. 293: E932-E940.

Schertzer JD*, Gehrig SM*, Ryall JG & Lynch GS (2007). Modulation of IGF-I and IGFBP interactions enhances skeletal muscle regeneration and ameliorates the dystrophic pathology in mdx mice. Am. J. Pathol. 171: 1180-1188. [*- these authors contributed equally to this manuscript]

Ryall JG, Schertzer JD & Lynch GS (2007). Attenuation of age-related muscle wasting and weakness in rats after formoterol treatment: therapeutic implications for sarcopenia. J. Gerontol. Series A. 62: 813-823.

Stupka N, Schertzer JD, Bassel-Duby R, Olson EN & Lynch GS (2007). Calcineurin-Aa activation enhances the structure and function of regenerating muscles after myotoxic injury. Am. J. Physiol. Reg. Int. Comp. Physiol. 293: R686-R694.

Lynch GS, Schertzer JD & Ryall JG (2007). Therapeutic approaches for muscle wasting disorders. Pharmacol. Ther. 113: 461-487.

**Harcourt LJ, Schertzer JD, Ryall JG & Lynch GS (2007). Low dose formoterol administration improves muscle function in dystrophic mdx mice without increasing fatigue. Neuromusc. Disord. 17: 47-55.

Experimental Techniques

Muscle functional testing

Intact skeletal muscles – We examine the contractile properties of different muscles of rats or mice using in situ (with an intact nerve and blood supply) or in vitro methods. The muscles most commonly assessed are the extensor digitorum longus (EDL) representative of a fast-twitch muscle and the soleus muscle which is representative of a slow-twitch muscle. We typically assess the tibialis anterior muscle using an in situ approach and study the contractile properties of isolated diaphragm muscle strips in vitro.

Single muscle fibres (cells) – Segments of single muscle cells (i.e. muscle fibres) can be carefully dissected from intact muscles (of rats and mice) or from open- or needle-biopsies from animals or humans. We attach these isolated muscle fibre segments to sensitive force transducers and servomotors to assess the contractile properties of skeletal muscle at the cellular level.

Cell lines - (C2C12 & L6) and primary myoblasts

Immunohistochemistry - intracellular localization of proteins

Immunocytochemistry - intracellular localization of proteins

Myoblasts differentiating into myotubes in culture. To form new muscle, myoblasts (muscle precursor cells) proliferate and then fuse together to form multinucleated muscle fibres. We can examine the ability of various therapeutic compounds to enhance either myoblast proliferation or myoblast fusion.

Current Animal Models

The laboratory utilises a number of animal models to study muscular dystrophy and ageing, including;

• mdx - the mdx mouse arose from a spontaneous mutation in the gene encoding the dystrophin gene (located on the X chromosome) in the C57Bl10 mouse. The mdx mouse is the most commonly studied model of Duchene muscular dystrophy (DMD).
• Adrb null - we have recently acquired mice that are gentically null for the β₁-adrenoceptor (Adrb1-/-), the β₂-adrenoceptor (Adrb2-/-), or both (Adrb1/2-/-)
• dko - the utrophin null dystrophic mdx mouse (mdx/utrn^-/-) known as the double knock out (dko) mouse. dko mice lack both dystrophin and utrophin (structural/signalling proteins linked to the muscle membrane) which causes muscular dystrophy similar to humans suffering DMD.
• F344 - the Fischer 344 (F344) rat is traditionally used in studies investigating the effect of ageing on skeletal muscle. The F344 rat is generally considered ‘old’ after the age of 24 months, and shows severe age related muscle wasting and weakness (termed ‘sarcopenia’) after the age of 28 months.

Alumni

Postdoctoral Researchers:

• Dr. Paul Gregorevic (Pfizer Australia Senior Research Fellow, Laboratory for Muscle Research and Therapeutics Development, The Baker Heart Research Institute, Melbourne, VIC, Australia)
• Dr. David R. Plant (Senior Regulatory Affairs Associate, CSL Limited, Melbourne, VIC, Australia)
• Dr. Jonathan D. Schertzer (NSERC Research Fellow, The Hospital for Sick Children, Ontario, Canada)
• Dr. James G. Ryall (CJ Martin Fellow, The Laboratory of Muscle Stem Cells and Gene Regulation, NIH, MD, USA)

Ph.D. students:

• Dr. Noel D. Duncan (Executive Director, Health1st Pty. Ltd. T/A Fitness2live, London, UK)
• Dr. Siun P. O'Sullivan (Lecturer, Biomedical Sciences, Victoria University of Technology, Melbourne, VIC, Australia)
• Dr. Paul Gregorevic (Pfizer Australia Senior Research Fellow, Laboratory for Muscle Research and Therapeutics Development, The Baker Heart Research Institute, Melbourne, VIC, Australia)
• Dr. David R. Plant (Senior Regulatory Affairs Associate, CSL Limited, Melbourne, VIC, Australia)
• Dr. James G. Ryall (CJ Martin Fellow, The Laboratory of Muscle Stem Cells and Gene Regulation, NIH, MD, USA)
• Dr. Nicole Stupka (Peter Doherty Fellow, Deakin University, Melbourne, VIC, Australia)
• Dr. Jonathan D. Schertzer (Postdoctoral Researcher, The Hospital for Sick Children, Ontario, Canada)
• Dr. Felice Beitzel (training for the Winter Olympics in Bend, Oregon)

Visitors/Sabbaticals

November-April 2000: Prof. Bartley R. Frueh, Kellogg Eye Center, Department of Ophthalmology, The University of Michigan.

November-April 2006: A/Prof. Luc Gosselin, Department of Exercise and Nutrition Sciences, University at Buffalo The State University of New York.

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