annual report - O'Brien Institute

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expression is increased following trauma, and in which cells it is expressed. We found that, as in animals, LIF is expressed within injured human nerves by a variety of cells, including Schwann cells and inflammatory cells, and that chronic neuromas continue to contain high levels of LIF for several years. These data suggest further examination is warranted to determine whether continued expression of LIF in unhealed injured nerves promotes or exacerbates the development of neuromas, or is simply a response to continued ‘trauma’. LIF and Schwann cell survival Schwann cells are cells which line the nerve fibres and play a major role in promoting nerve survival and regeneration after injury. They are the major source of LIF, and GDNF, at the injury site. Our own research has shown that Schwann cells require LIF for survival in culture, and that when we blocked LIF function within the entubulation repair site the survival of Schwann cells was markedly reduced. This suggests that one of the actions of LIF following nerve trauma is to prevent injury-induced cell death. Role of NOS 2 in nerve regeneration The nitric oxide producing protein NOS 2 is thought to play a role in blood vessel formation. It is also thought to be regulated in some tissues by LIF expression. We used the NOS 2 inhibitor AET administered for one month following nerve repair to determine whether NO-derived from NOS 2 played a role in nerve regeneration in our model, and whether the addition of LIF at the time of repair could overcome the effects of NOS 2 inhibition. When NOS 2 production is blocked with AET, in addition to repairing the nerve stumps, re-vascularisation and nerve regeneration are delayed within the silicone tube for up to 4 weeks. Addition of LIF to the tube at the time of repair failed to overcome the effects of the AET. This series of experiments suggest that nitric oxide is important to angiogenesis and subsequent axon growth and may be required for the action of LIF. The treatment of pain resulting from a neuroma Neuromas are bulbous swellings which occur at the cut end of nerves, and these can become extremely painful, resulting in neuropathic pain and sometimes loss of work and, in extreme cases, suicidal tendencies. We are investigating the characteristics of the neuron population and local resident cells which contribute to the neuroma. Once a nerve is injured or cut and not repaired, a neuroma bulb forms at the cut end and persists for the life of the nerve. We have now shown that many uninjured nerve fibres grow into this end bulb neuroma, in addition to the injured nerves. We have commenced a very detailed study of motor nerves cells before injury, and after neuroma formation and will compare the effects of this procedure on the contralateral uninjured nerve cells. Alternative nerve repair methods In some circumstances it is not possible to repair an injured nerve branch by using the parent nerve and so an unrelated healthy nerve is used instead. Two methods of joining the injured nerve to the healthy nerve are possible. In the first, the healthy nerve is partially cut through and the ends of the injured nerve fibres are sutured to the cut ends of the healthy donor nerve fibres (end-end repair). In the other method, the injured nerve is sutured to the side of the uninjured nerve (end-side repair). We have shown that both these techniques result in a significant return of function to the denervated muscles. However, the end to end repair results in decreased function of the donor “muscles” whereas these muscles are spared by using end to side repair. Using a nerve tracer method, we determined that recovery of the denervated muscles is mediated by regeneration from nerve cells in the donor nerve and not from of the original nerve cells. When the distal stump of a severed nerve branch is rejoined to the parent nerve at a site distant to the point of severence, the denervated muscle regains some of its original function. We have shown that a number of nerve fibres grow down the parent nerve in newly formed tracks rather than joining the existing tracks of nerves. These nerve tracks originate at the level of the branch nerve injury and most likely include some of the original cut branch nerves fibres. Helen M Schutt Vascular Research Laboratory Senior Scientists: Geraldine Mitchell, Peter Vadiveloo. Clinical Research Fellow: Tony Penington. Members of the Laboratory: Angela Arvanitis, Mirna Boujaude, Rob Donato, Tanya Harkom, Peter Meagher, Wayne Morrison, Rosalind Romeo, Arthur Smardencas, and Debra Zafiropoulos. ...and Major Collaborators: Brain Cooke d , Alastair Stewart a aDepartment of Pharmacology, University of Melbourne dDepartment of Microbiology, Monash University, Clayton, Vic. Blood vessel biology Successful outcomes for surgery depend upon a good blood supply to nourish repaired and transplanted tissue. In this laboratory we are trying to get a better idea of how blood vessels work, and how to manipulate blood vessels. The knowledge generated from this work will not only impact in areas of surgery but will also be relevant to diseases such as cancer, arthritis and atherosclerosis since changes in blood vessel growth and shape are important features of these diseases. Angiogenesis The process of new blood vessel formation is called angiogenesis. Until recently there has not been a clinically relevant experimental model. The scientists and surgeons at BOBIM have now developed such a model in rats and mice. This is an important step since we can now use the Director’s Report Bernard O’Brien Institute of Microsurgery Director Professor Wayne Morrison, MD, BS, FRACS Reconstruction following injury, tumour resection, burns or congenital deformity involves the transfer of tissues from one part of the body to another. Virtually every body part or tissue is capable of being transferred, e.g. skin, muscle, bone and joints, nerves, fat, or composite structures such as toes, scalp, etc. At the Bernard O’Brien Institute research was originally directed towards the role of microsurgery in this process of tissue transfer. Many of these techniques are now in common use throughout the world and for this reason our research is moving into the new frontier of tissue engineering. This is a natural progression in the sophistication of reconstructive techniques and of microsurgery where only miniscule amounts of the patient’s tissues are required to manufacture the derived product. Microsurgery plays an integral role in the process, firstly by supplying the core vascular pedicle on which the tissue is grown and then, if need be, in transferring the product to the desired site for reconstruction. Tissue engineering demands collaborations with experts in several fields, especially cell culture including stem cells, matrix biology, vascular biology and bioengineering. We are very fortunate to have in Melbourne many experts in these fields who are generously contributing their ideas. Tissue engineering offers the potential to repair defects without the need to sacrifice other areas of the body. It is an alternative to free tissue transfer, artificial prostheses and organ and tissue transplants. MICROSURGERY FOUNDATION The Institute also has a major interest in nerve repair following injury and in parallel with the pain management service under the direction of Dr Andrew Muir is interested in nerve pain, especially neuroma formation. Research includes the development of neuroma models and studies mapping the sources of nerve fibres which migrate into developing neuromas. Studies have shown in human neuromas that the growth factor (leukaemia inhibitory factor) is abundantly produced. The role of this growth factor in nerve repair and in pathological processes is being investigated. The Schwann cell which forms a sheath around nerve fibres is a major producer of this growth factor and other factors involved in the maintenance of nerve function and in repair processes following injury. Much of our research is directed towards understanding the behaviour of this cell. Injury to tissues following interference with blood supply (ischaemia) has particular relevance to our clinical work which involves reattachment of amputated parts and the transferring of tissues from one part of the body to another. Investigations understanding the nature of this injury and methods to minimise its effect have been a long time interest in our laboratory. Inflammation is a fundamental body response to injury and is integral to the repair process. Macrophage function is one of our special interests and research continues into the molecular mechanisms which activate macrophages. Scarring following injury is a major source of morbidity and compromises the results of operations, particularly in organs which require movement for function, such as the hand. We are investigating agents which prevent collagen formation and are hopeful that these will have a role in reducing the morbidity associated with injury and surgery. 26 15
Many other research topics are being investigated and they are detailed under the section headed “Scientific Research”. It has been a pleasure to be involved in the many projects with cross-links with more and more colleagues and other research institutes both in Australia and overseas. These collaborators have been very generous of their time and facilities and their institutes are acknowledged elsewhere in this report. Fellows Research Fellows from overseas and Australia, many doing postgraduate degrees, contribute to the research program and participate in clinical work at St. Vincent’s Hospital. Each undertakes a fellowship of one year or more and gains a fellowship certificate in hand and microsurgery. During the period of this report the following held appointments at the Bernard O’Brien Institute of Microsurgery: Oliver Cassell, Plastic Surgeon, United Kingdom - Tissue engineering models and matrix. Rob Donato, Plastic Surgeon, Australia - Extracorporeal circulation. Slobodan Djurickovic, Orthopedic Surgeon, Canada. Sunao Furuta, Plastic Surgeon, Japan - In vivo model of angiogenesis and role of nitric oxide. Hans Mark, Plastic Surgeon, Sweden - Model of bone non-union. Peter Meagher, Plastic Surgeon, Ireland - Tissue engineering - muscle generation in stem cells. David McCombe, Plastic Surgeon, Australia - Tendon healing. Kanit Sananpanich, Plastic Surgeon, Thailand - End-to-side nerve repair. These fellows are the lifeblood of the Institute. Not only do they learn microsurgery and partake in research programs, but they contribute enormously from an intellectual, cultural and social viewpoint. Since the Institute began more than 200 fellows from all corners of the globe have trained in research and in clinical microsurgery at St. Vincent’s Hospital. Many have returned to their home country to establish microsurgery centres and become leaders in their field. A highlight of the year was the appointment of Allan MacLeod, Head of Plastic Surgery at St. Vincent’s Hospital and a Director of the Microsurgery Foundation, as Associate Professor, acknowledging his major contributions in clinical and research plastic and microsurgery. This is a great honour and we congratulate Allan on his achievement. Alumni A large alumni group now exists as a consequence of the large international fellowship that has developed through training at the Institute. Many fellows have become lifelong friends with their Australian colleagues and also with those who trained with them from other parts of the world. It is very gratifying, when attending international meetings to meet up again with so many colleagues and recount tales, tall or true, of their experience at St. Vincent’s and the Institute and on a personal note, how enjoyable it is to be made so welcome on visiting these fellows in their own countries. The hospitality has been extraordinary and I consider it a great privilege to have been able to share in their development and interests. This year I was particularly privileged to meet many of our alumni at the Asian Pacific Federation of Societies for Surgery of the Hand meeting in Madras and at the South African Society for Surgery of the Hand. Our friend and colleague Julian Pribaz, Professor of Plastic Surgery at Brigham and Women’s Hospital, Harvard, Boston, was appointed the B.K. Rank Travelling Professor for the Annual Scientific Meeting of the Royal Australasian College of Surgeons in Melbourne in May. Julian trained in plastic surgery at St. Vincent’s and was a member of our plastic surgery team before moving to Boston. We were inspired by his clinical and research presentations. Acknowledgements The Board of Directors of the Microsurgery Foundation under the Chairmanship of Mr Ronald Walker have been again instrumental in financing our research efforts. They give generously of their time and expertise and it is impossible to under estimate their contributions. Alan Skurrie, past Chairman of Microsurgery Foundation and long term member of the Board retired this year. It is with great regret that we say goodbye to Alan who over many years has been a tower of strength, especially through the lean years when with perseverance, wisdom and charm he enabled us to gain many major grants which The Jack Brockhoff Nerve and Muscle Laboratory The Jack Brockhoff Laboratory is dedicated to researching the mechanisms by which injured nerves and muscles regenerate and to identifying chemical factors and various administration protocols which enhance the repair process. Senior Scientists: Aurora Messina, Bruce Dowsing Clinical Research Fellow: Tim Bennett. Members of Laboratory: Richard Brouwer, Carl Byers, Rob Donato, Ruitong Fan, Sachiko Maeda, Marian Todaro, Rosalind Romeo, Kanit Sananpanich, Tim Shakespeare, and Wayne Morrison. …and Major Collaborators: Lawrie Austin 1 , Alan Hayes 2 , Nic Nicola 3 , Mary Galea 4 , Dr. Paul Patterson 5 , and Catherine Sangster 4 . 1Melbourne Neuromuscular Research Centre, St. Vincent’s Hospital, Melbourne. 2Department of Biological Sciences, Victoria University of Technology, Footscray, Vic. 3Walter and Eliza Hall Institute for Medical Research, Melbourne. 4School of Physiotherapy, University of Melbourne. 5California Institute of Technology, USA. Nerve repair and regeneration Microsurgeons are often faced with the task of repairing injured nerves. Unless successfully repaired, nerve injuries lead to dysfunction of their target organs, resulting in permanent disability such as loss of muscle use (motor nerves), or loss of sensation to a limb (sensory nerves) or pain. On average nerve injuries occur in young adults and hence, failure to recover has a tremendous economic impact. Modern microsurgical techniques enable accurate repair of most nerves but the functional outcome is often poor, due to death of the nerve cells (neurons), inadequate regeneration of axons (nerve fibers), and/or deterioration and scarring of the target prior to reinnervation. Neurons growing axons and target organs depend on continuing input from each other and supporting cells for growth, maintenance and survival, without these they atrophy and/or die. These actions are mediated by chemical factors (proteins) known as growth factors. The recent discovery and the availability of various growth factors, such as leukemia inhibitory factor (LIF), have provided a means of treating regenerating nerves and their targets to improve recovery, with very encouraging results! MICROSURGERY FOUNDATION Entubulation repair of transected nerves is the standard model, used in our laboratory, for studying peripheral nerve regeneration and recovery. It involves removing a small length of nerve, thereby leaving a gap between the proximal and distal ends of the nerve. This gap is bridged by plugging these stumps into either end of a hollow silicone tube which spans the gap, thus forming a sealed chamber. Within this chamber, proteins (survival factors, etc.) and cells that collect in response to nerve injury can be studied, test substances can be administered and their effects on the nerve repair process determined. Hand and limb injuries often involve trauma to their supplying nerves and despite current surgical techniques the degree of function recovered is generally unsatisfactory. Our research aims to understand the mechanisms of regeneration of nerve and muscle following injury and to evaluate agents which may enhance the repair process. Improving Nerve Repair with neurotrophic factors Following nerve injury cells near to the trauma site produce various proteins, called growth factors, that encourage healing and nerve regeneration. The availability of some of these growth factors, such as leukemia inhibitory factor (LIF) and glial cell derived neurotrophic factor (GDNF), has enabled us to develop methods to administer these factors to the site of injury and to evaluate their efficacy as nerve regeneration promoters. We found that the repair of cut or crushed rat nerves in conjunction with either LIF or GDNF results in significantly enhanced nerve regeneration, the muscles they re-connect with do not waste as much, and eventually their function is significantly improved. However, addition of these factors together does not further enhance the recovery obtained with either factor alone. Before these promising factors can be used in the clinical setting, a relatively simple and cheap method of administering them at the time of surgery must be found. In collaboration with AMRAD Pty Ltd we are about to test a method to administer LIF to the site of nerve repair, and if successful we believe we will be ready to commence clinical trials. Expression of LIF following human nerve injury To date the expression profile of LIF within nerves has been confined to animal models such as rats and mice. We therefore determined whether LIF is expressed in normal human nerves and whether its 16 25
Page 1 and 2: For the betterment of humanity MICR
Page 3 and 4: Restoring people’s confidence, ho
Page 5 and 6: 2000 Microsurgery Annual Report Cli
Page 7 and 8: CASE 2 John McKinnon Hope Fatherhoo
Page 9 and 10: CASE 3 Martin Turner Identity Commu
Page 11 and 12: Collaborators Bernard O’Brien Ins
Page 13 and 14: Patrons Microsurgery Foundation Pat
Page 15 and 16: Chairman’s Report Microsurgery Fo
Page 17: Overview Bernard O’Brien Institut
Page 21 and 22: Barbara Walker Centre for Pain Mana
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annual report - O'Brien Institute

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