Peptide biomarkers of myocardial ischemia
Supervisors:  Dr Stuart Cordwell (Lab Head), Associate Professor Brett Hambly and Dr Melanie White
Contact Details: Room 704/705 Biochemistry Building G08; email: s.cordwell@mmb.usyd.edu.au; phone: 9351 6050
Project Description:  Cardiovascular disease (CVD) results in approximately 7 million deaths per annum world-wide and is the most significant cause of death in Australians. Many of these result from sequelae following myocardial ischemia / reperfusion (I/R) injury. Reduction or cessation of blood flow (ischemia) generally results from the formation of atherosclerotic lesions in the coronary arteries. Reintroduction of blood-flow (reperfusion) by thrombolysis or primary percutaneous coronary artery intervention remains the best strategy for resolving ischemia and preventing cell death and permanent cardiac dysfunction (infarction). Morbidity and mortality from acute myocardial infarction (AMI) remain significant. The endogenous or ‘native’ peptidome is the full complement of natural, low molecular mass (<10kDa) peptides, as well as those created by the proteolysis of larger proteins, contained within a cell, tissue or body fluid. Pathology is often underpinned by protein damage, particularly following protease activation. The peptidome is therefore a rich source of putative disease biomarkers. This project will utilize chromatography coupled with mass spectrometry to identify peptide biomarkers of I/R injury in animal models and thus act as a model for developing new and more effective strategy for the rapid diagnosis of myocardial ischemia.

The role of calcium overload in myocardial I/R injury
Supervisors:  Dr Stuart Cordwell (Lab Head) and Associate Professor B. Hambly
Contact Details: Room 704/705 Biochemistry Building G08; email: s.cordwell@mmb.usyd.edu.au; phone: 9351 6050
Project Description: At the cardiomyocyte level, I/R injury is characterized by Ca2+ overload and the generation of ROS. Both are highly reactive molecules and are able to interact with almost any biological substrate. Lipids and proteins are possible targets, with potentially wide-reaching implications – lipid damage will affect the integrity and permeability of cell membranes, while protein damage may change the functionality of regulatory enzymes or contractile mediators. We have previously investigated the role of ROS on protein post-translational modification in I/R injury. In this project, proteomics will be used to investigate the effects of Ca2+ overload on proteins within myocardial tissue during I/R. The project will involve animal studies (Langendorf perfusion surgery), as well as biochemical assays including two-dimensional electrophoresis and mass spectrometry. These studies will identify proteins damaged during I/R and potentially lead to clinical therapies.

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VASCULAR IMMUNOLOGY UNIT

General Context
Fatal malaria is one of the most destructive and potentially correctable disease burdens in the world, affecting mainly children. A major complication is cerebral malaria (CM). Current therapies are limited by our lack of knowledge of the pathophysiological events. Our studies aim at increasing the understanding of pathogenetic mechanisms of CM, and may therefore lead to new approaches to the prevention or prompt treatment of potentially fatal malaria.

General Background
We recently demonstrated that microparticles (MP), which we found to be present in greatly increased concentrations in the peripheral blood of Malawian children with CM (JAMA 2004), are crucial elements of pathogenesis in experimental murine CM (Am J Pathol 2005). Based on these findings, we now aim, in vitro and in vivo, to demonstrate that by modulating the process of vesiculation (i.e. by reducing the production or release of MPs, or by blocking their toxic effects) we can reduce the pathological processes characteristic of CM. The ultimate aim is to reach a novel intervention for preventing the progression of severe malaria towards a fatal outcome and for hastening recovery from severe malarial disease.

In our current projects, we therefore intend to unravel the mechanisms of MP production and action, to delineate pharmacological ways to interfere with these mechanisms, and to define the pathophysiological consequences of excessive MP production.

Assessing microparticles as effectors in the microvascular lesion of cerebral malaria.
Supervisors:  Professor Georges Grau (Pathology), Dr Valéry Combes (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: ggrau@med.usyd.edu.au; Phone: 9036 3260
Project Description:  Co-cultures of human brain microvascular endothelial cells with parasites and blood cells, in which purified microparticles will be added, to assess the functional consequences. Analysis of immunological and physiological parameters, such as monolayer permeability and trans-endothelial electrical resistance.

Investigating mechanisms of microparticle production by microvascular endothelial cells and defining clinically useful inhibitors.
Supervisors:  Professor Georges Grau (Pathology) éry Combes (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: ggrau@med.usyd.edu.au; Phone: 9036 3260
Project Description:  Co-cultures of human brain microvascular endothelial cells with parasites and blood cells, in which various inhibitors of intracellular pathways will be added, to assess their ability to reduce the number of MP released upon stimulation by parasites, or to alter their phenotype. Apart from classical inhibitors, we already have new candidate molecules with obvious effects on MP. Analysis of immunological and haemostatic parameters of the MP produced.

Defining the implication of hypoxia in the pathology of cerebral malaria
Supervisors:  Professor Georges Grau (Pathology) and Professor Nick Hunt (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: ggrau@med.usyd.edu.au; Phone: 9036 3260
Project Description: Using our well characterised model of cerebral malaria (Trends Immunol 2003), several immunological and pathological pathways have been unravelled but numerous questions remain unsolved. In particular, the extent to which the obstruction of brain microvessels by parasites and host cells lead to hypoxia in the surrounding parenchyma. Drs. V. Combes and S. Parekh in our lab have shown in preliminary experiments that tissue hallmarks of hypoxia can be detected in mice that are genetically susceptible to CM, but not in strains known to be resistant. Injection of probes in mice and follow up of tissue changes, notably in the brain, by immunohistochemistry. Comparison, using quantitative image analysis, of mice infected with the CM-inducing parasite, PbA or with the non-encephalitogenic parasite, K173. Also, comparison of brains from mouse strains exhibiting various genetic susceptibility towards CM.

Determining the interactions of MP with their target cells in the host
Supervisors:  Dr. Valéry Combes (Pathology) and Professor Georges Grau (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: vcombes@med.usyd.edu.au; Phone: 9036 3260
Project Description:  Microparticles (MP) are known to not only bind to cells but also interact with them and are responsible for various downstream effects such as activation (as evidenced by up-regulation of surface molecules, or triggering of signalling pathways), induction of apoptosis, increase binding to other cells, release of soluble molecules.  We will address the following questions:

• What is the outcome of a pathogen-induced MP once it reaches its target cell and is there a difference according to the vesiculation agonist?
• What is the consequence of the interaction between MP and target cells at the level of the latter?

We have preliminary data, generated by confocal microscopy and flow cytometry, which show that platelet-derived MP are able to bind to endothelial cells and activate them.
For this project we will use two cell types (endothelial cells and monocytes) and study the interactions of their derived-MP with either endothelial cells or monocytes.
The techniques will include cell culture, flow cytometry, fluorescence micoscopy and electron microscopy.  The ImageStream® will also be used to visualise the interactions MP-target cell.  The ImageStream® is a benchtop imaging flow cytometer designed for quantitative image-based analyses.  It allows the collection of large numbers of digital images and combines cell information provided by fluorescence microscopy with the statistical significance of large sample size common to flow cytometry.

Deciphering the Involvement of TCTP Proteins in Malarial Pathogenesis
Supervisors:  Dr. Ronan Jambou (Pathology) and Professor Georges Grau (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: rjambou@med.usyd.edu.au; Phone: 9036 3260
Project Description:  During cerebral malaria, the alteration of endothelium in cerebral vessels is a major element of pathogenesis.  Several mechanisms have been described, leading to apoptosis of endothelial cells and/or to the production of pro-inflammatory microparticles.  In culture, these alterations can be induced by P. falciparum-parasited red blood cells, but the proteins and the mechanisms involved in this process are not identified.  Two approaches are proposed within this framework:

1. Study of the role of histamine and TCTP proteins in endothelial alterations.
   The role of parasitic proteins and of mediators of allergy in the formation of the lesions (especially in the opening of the tight junctions) will be studied in cultures of the human microvascular endothelial cell line, HBEC-5i, using electrophysiology, microscopy and molecular approaches.
2. Study of the production of microparticles by cells, upon stimulation by TCTP proteins.
   The in vitro production of microparticles by endothelial cells, monocytes and platelets will be analysed by flow cytometry after stimulation of the cells by recombinant parasitic proteins.  In vivo studies also will be undertaken in mice infected by P. berghei ANKA associated or not with the injection of recombinant proteins.

Defining intracellular signalling pathways of microvascular endothelium stimulation by parasite and host cells
Supervisors:  Dr. Angeles Sanchez-Perez (Pathology) and Professor Georges Grau (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: angeles@med.usyd.edu.au; Phone: 9036 3260
Project Description:  Adherence of Plasmodium falciparum-infected erythrocytes (iRBC) to brain microvessels is believed to be a critical step in the development of cerebral malaria (CM), an illness that kills around 2 million people, mostly children, every year.  The malarial parasite modfies the infected erythrocyte by transporting proteins into its membrane.  PfEMP1, a parasite protein expressed on the erythrocyte surface, binds to receptors in endothelial cells and promotes sequestration of the iRBCs.
Co-cultures of human brain microvascular endothelial cells with parasites and blood cells will be used to mimic the conditions found in CM and to investigate the intracellular pathways triggered by the parasite in the endothelial cells.

Dissecting the molecular mechanisms of platelet-endothelial interactions
Supervisors:  Dr. Manuela Florido and Professor Georges Grau (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25); email: ggrau@med.usyd.edu.au; Phone: 9036 3260
Project Description:  In cerebral malaria PBRCs and platelets accumulate and induce deleterious functional alterations in the endothelium of brain microvessels.  S-1-P is a sphingolipid that is released in high amounts by activated platelets and is involved in the regulation of the endothelial barrier function.  The role of this molecule in the induction of microvessel lesions in the context of cerebral malair will be studied using an in vitro platelet-endothelial cell co-culture system already established in this lab.  We will analyze the effect of the modulation of the S-1-P signalling pathway by selective chemical agonists/inhibitors or by RNA interference in the co-cultured cells.

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MACROMOLECULAR STRUCTURE UNIT

Immunological Mechanisms in Atherosclerosis
Supervisors:  Associate Professor Brett Hambly and Associate Professor Bob Bao
Contact Details: Room 557 Blackburn Building D06; email: bretth@pathology.usyd.edu.au or bobbao@pathology.usyd.edu.au; phone: 9351 3059 or arrange a meeting through the Department Executive Assistant Ms Lorraine Rhind (9351 2414)
Project Description: The immune response is important in modulating the inflammation that occurs in atherosclerosis. We have developed a mouse model of atherosclerosis to examine the process of neointimal proliferation (thickening) of the walls of arteries that have been mechanical damaged. This model allows us to probe immune mechanisms using genetically altered mice. These studies involve the use of microsurgical techniques, immunohistochemistry and mRNA quantitation using real time PCR.

Protein defects causing Familial Hypertrophic Cardiomyopathy
Supervisors:  Associate Professor Brett Hambly
Contact Details: Room 557 Blackburn Building D06; email: bretth@pathology.usyd.edu.au; phone: 9351 3059 or arrange a meeting through the Department Executive Assistant Ms Lorraine Rhind (9351 2414)
Project Description:  The commonest form of inherited heart disease is familial hypertrophic cardiomyopathy (FHC). We know that about 30% of all families with this disease have a mutation in a gene expressing the heart protein myosin binding protein-C (MyBP-C). This protein is an important protein in the heart that is responsible for adjusting the contraction of the heart in response to stress (adrenergic stimulation). MyBP-C does this by modifying the interaction between the key contractile proteins actin and myosin within the cardiomyocytes. Mutations that occur in FHC in MyBP-C interfere with this modulatory function, but the structural basis for these abnormalities is very poorly understood. These studies involve the use of DNA coding for MyBP-C to express parts of the MyBP-C protein, to allow us to study the protein, mainly using spectroscopic techniques, especially fluorescence spectroscopy.

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MOLECULAR IMMUNOPATHOLOGY UNIT

Understanding a key new enzyme in the kynurenine pathway
Supervisors:  Dr Helen Ball (Pathology), Professor Nick Hunt (Pathology)
Contact Details: Level 1, Medical Foundation Building helenb@med.usyd.edu.au; phone: 9036 3238
Project Description:  Indoleamine 2,3-dioxygenase (IDO) catalyses the first step in the kynurenine pathway of tryptophan metabolism. This pathway is involved in numerous physiological and pathophysiological processes including immunomodulation and central nervous system disorders. We recently discovered a second enzyme (IDO2) that performs the same reaction as IDO (now IDO1) but which is found in different anatomical locations. This project will further investigate the expression pattern of IDO2 by looking at subcellular localisation and expression during development and disease states. The biochemical properties of IDO2 will be examined by determining whether IDO2 enzymatic activity in intact cells is inhibited by nitric oxide, an important mechanism for regulation of IDO1 activity. This project will possibly also include investigating phenotypical characteristics of an IDO2 gene knockout mouse e.g. susceptibility to infectious disease (malaria and meningitis). Laboratory techniques involved in this project include immunohistochemistry, immunofluorescence, quantitative RT-PCR, cell culture, transfection, biochemical assays, mouse models of disease.

What causes death and disability in bacterial meningitis?
Supervisors:  Professor Nick Hunt and Dr Helen Ball (Pathology)
Contact Details: Level 1, Medical Foundation Building (K25); email: nhunt@pathology.usyd.edu.au; helenb@med.usyd.edu.au 
Project Description:   Bacterial Meningitis kills over 150,000 people every year and many more suffer neurological sequelae following infection.  We have exetensive experience in uncovering the immunopathological mechanisms in cerebral malaria. We have recently established a mouse model of bacterial meningitis and our preliminary evidence suggests that the cytokine interferon gamma (IFNg) plays an important role in determining whether mice with bacterial meningitis succumb to the disease. IFNg is a key regulator of indoleamine 2,3-dioxygenase-1 (IDO1) expression and thus activity of the kynurenine pathway. Activation of this pathway is observed in a number of central nervous system disorders including cerebral malaria.
This project will further investigate the role of IFNg in bacterial meningitis. Gene expression during bacterial meningitis in IFNg gene knockout and wildtype mice will be compared using gene arrays to identify pathways regulated by IFNg in bacterial meningitis. IDO1 expression is upregulated in bacterial meningitis in wildtype mice and IDO1 expression will also be investigated in the IFNg gene knockout mice. The levels of kynurenine pathway metabolites during bacterial meningitis will be measured in both mouse strains. The neurological sequelae in IFNg gene knockout mice that recover from bacterial meningitis will be measured in behavioural studies that compare these mice with both uninfected and antiobiotic-treated mice. The progression and outcome of bacterial meningitis in IDO1 gene knockout mice will also be investigated. Laboratory techniques involved in the project include mouse models of disease, mouse behavioural studies, quantitative RT-PCR and gene arrays, histopathological analysis, biochemical assays.

VIRAL IMMUNOPATHOLOGY UNIT

Background
Flaviviruses are single-stranded, RNA viruses transmitted by mosquitoes and ticks. This group of viruses is a major cause of illness and death throughout the world. Well known diseases include Yellow fever, dengue, Japanese encephalitis and Murray Valley encephalitis.  Another flavivirus, West Nile virus (WNV), which causes encephalitis in humans and animals has recently spread for the first time throughout the continental USA, Mexico and Canada since a single epidemic outbreak in New York in August 1999.  This disease causes 10-15% death in clinical cases of encephalitis and survivors may end up with permanent nervous system damage.
Importantly, this disease is likely to be due to the over vigorous activity of the immune response.  Indeed, infection of cells with WNV, causes a massive increase in the cell surface molecules that help to induce a successful immune response.  These molecules include the class I major histocompatibility complex (MHC-I) and MHC-II molecules, intracellular adhesion molecule-1 (ICAM-1) (CD54), VCAM-1 (CD106) and E-selectin (CD62E), as well as the co-stimulatory molecule, B-7 (CD80), on Langerhans cells.  These molecules are the major targets for cytotoxic T cell recognition.  Thus, not surprisingly, these changes are associated with an increase in killing of infected cells by antiviral T cells during the immune response.

Flavivirus-mediated increases in the immune response may constitute a novel virus survival strategy.  However, it may also be the cause of the excessive immune system-mediated damage (immunopathology) in disease caused by these viruses.  The mechanisms involved in these dramatic interactions are unknown and investigation of these issues is the main thrust of this laboratory.
We use a laboratory adapted strain of WNV, that has been used in Australia for about 40 years.  Our laboratory has used this virus for 20 years.  All students are trained in the use of sterile and safe techniques in virus handling.  The following are brief outlines of a few possible projects in this laboratory for Honours (or PhD) in 2008.  Should you wish to discuss these or others please contact Prof Nicholas King (nickk@pathology.usyd.edu.au) to make a time to talk about them in more detail.

Changes in gene expression in neurons and microglia in WNV encephalitis.
Supervisors:  Professor Nicholas King and Professor Iain Campbell (Molecular and Microbial Sciences)
Contact Details: Room 260, Level 2 Blackburn Building nickk@pathology.usyd.edu.au(best); phone: 9351 4553
Project Description:  We have shown that in WNV encephalitis, resting microglia (the macrophages of the brain) differentiate into a highly activated phenotype.  They also increase MHC and adhesion molecule expression and proliferate and migrate to surround infected neurones.  In addition, significant numbers of blood-derived inflammatory macrophages migrate to the brain.  We think this may be the way in which WNV is cleared from neurones, since in interferon-gamma gene knockout mice, which exhibit greater activation of microglia, animals survive 3-fold better.  However, our work indicates that these macrophages may also contribute to the immunopathology occurring in WNV encephalitis.  The aim of this project is to examine the expression of various genes in neurones and microglia during infection, to determine the progress of events by looking at changes in various cell populations and determine how WNV is cleared from the brain.

Infiltrating leukocytes in WNV encephalitis - their role in mortality.
Supervisors:  Professor Nicholas King and Professor Iain Campbell (Molecular and Microbial Sciences)
Contact Details: Room 260, Level 2 Blackburn Building nickk@pathology.usyd.edu.au(best); phone: 9351 4553
Project Description:  Part of the response to WNV infection of neurones is an infiltration of leukocytes into the brain on day 5 after infection.  In interferon-gamma gene knockout mice, this infiltration is much reduced.  This suggests that the increased survival in these mice may be because a particular subset of leukocytes does not migrate to the brain.  This subset may also be involved in attracting inflammatory macrophages (see above).  The aim of this project is to examine these subsets in the brain in both the wild type and gene knockout mice and look at their cytokine and chemokine expression.  This will tell us why an increased leukocyte infiltration is associated with increased mortality in the wild type mouse.

The role of mucosal dendritic cells in controlling virus infection in epithelium.
Supervisors:  Professor Nicholas King
Contact Details: Room 260, Level 2 Blackburn Building nickk@pathology.usyd.edu.au(best); phone: 9351 4553
Project Description:  In a sexually transmitted disease (STD) model of virus infection in the mouse we find that infection by this route results in excellent immunity and very low mortality, compared to other routes on infection.  Why does this happen?  We are therefore examining what factors are associated with innate resistance to infection and what conditions best facilitate the generation of specific and effective anti-viral responses.  This is important for the development of vaccines against STD's.
Using this model we are analysing the steps involved in generating long-lasting antiviral immune memory responses.  For example, have found that the dendritic cells (DC) in the vaginal tissue accumulate where the virus infection is greatest in the epithelium and these DC disperse once the infection is eradicated.  However, DC gather more quickly upon a second challenge with the same virus.  This suggests the presence of factors that allow them to accumulate in this accelerated manner.  We are measuring when and how these cells migrate from the vaginal epithelium to the local draining lymph nodes after infection to enable the antiviral immune response to be initiated and how different this is on the second challenge with the same virus.  We are also looking at the migration of naive and immune lymphocytes along these pathways.
From these results we may be able to better understand how these immune response are initiated and understand better the differences in antiviral immune responses after primary and secondary infection.  This will also allow us and determine when, where and how STD vaccines should be administered to obtain the most effective immunity.

Maintenance of epithelial barriers in cell defence against virus infection
Supervisors:  Professor Nicholas King and Dr Michelle Madigan (Save Sight Institute)
Contact Details: Room 260, Level 2 Blackburn Building nickk@pathology.usyd.edu.au(best); phone: 9351 4553
Project Description: In the eye, a major cellular seal against the 'outside world' is the retinal pigment epithelium, which protects the cells inside the eye from infectious organisms, such as viruses, and even immune cells that may invade it.  Infection of these cells with WNV in culture results in changes in the cells that may prevent the transmission of virus into the eye itself.  We are investigating the factors involved in controlling these changes.  They have an important bearing on the way a crucial organ like the eye avoids being destroyed by virus infection or the resulting over-vigorous immune responses.

The role of Langerhans and dendritic cells in initiating WNV immune responses
Supervisors:  Professor Nicholas King
Contact Details: Room 260, Level 2 Blackburn Building nickk@pathology.usyd.edu.au(best); phone: 9351 4553
Project Description: We are also looking at the induction of the immune response via the skin where WNV gains its first access in vivo.  We have already shown that Langerhans cells in the epidermis migrate to the local draining lymph nodes faster in response to WNV infection than in response to infection by other skin-acquired viruses.  However, in addition, dendritic cells also migrate from the dermis to the lymph nodes and also seem to take part in the accelerated initiation of immune response to viruses.  These cells originally migrate from the bone marrow as monocytes and change into dendritic cells once they get to the lymph node draining the infected site. They also migrate to the infected skin site and change into dendritic cells there.  It is unknown how they change into dendritic cells and what stimuli are associated with this change. The aim of this project is therefore to examine the changes that occur in association with migration of dendritic cells to the local draining lymph node and the infected site at various times after infection.

The role of microparticles in the immune response to WNV
Supervisors:  Professor Nicholas King and Professor Georges Grau (Pathology, Medical Foundation Building)
Contact Details: Room 260, Level 2 Blackburn Building nickk@pathology.usyd.edu.au(best); phone: 9351 4553
Project Description: Recent research shows that endothelial and other cells can produce small membrane vesicles or microparticles (MP) in response to inflammatory conditions.  MP may worsen disease, but they may also be instrumental in enabling the efficient initiation and/or modulation of the immune response to infectious agents, for example, by interacting with dendritic cells.  This project will investigate the production of MP by WNV-infected cells in vitro and in vivo.  It will investigate when such particles are produced, what effect they have on other uninfected cells and if they can enhance the initiation of immune response in infected animals.

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NEUROPATHOLOGY UNIT - DEMENTIA, NEURODEGENERATION & AGING

The substrate of memory impairment in Frontotemporal dementia (FTD)
Supervisors:  Professor Jillian Kril
Contact Details: Level 4 Blackburn Building (D06); email: jilliank@med.usyd.edu.au;
Project Description: FTD is a form of dementia which is as common as Alzheimer’s disease (AD) in the <65y age group and accounts for 12-20% of all dementia. It is characterised by disturbances in social and interpersonal conduct and language abnormalities. One feature used to distinguish FTD from other forms of dementia is the relative preservation of memory function early in FTD. Nevertheless a proportion of FTD patients do present with amnesia and these are commonly mis-diagnosed as AD.
Pathologically FTD shows marked degeneration of the cerebral cortex and hippocampus. Neuropathology in the hippocampus correlates well with memory deficits in AD, however the same does not hold true for patients with FTD. This project will investigate the pathological substrate of memory impairment in FTD.
Human postmortem tissue from a prospective brain donor program is available for study. They type, severity and distribution of pathology will be examined in cortical brain regions known to have a role in memory function and the findings from patients with and without amnesia compared.

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NEUROPATHOLOGY UNIT - THE STACEY MOTOR NEURON DISEASE LABORATORY

Looking for the cause of motor neuron disease
Supervisors:  Associate Professor Roger Pamphlett
Contact Details: Room 502A, Level 5 Blackburn Building (D06); email: rogerp@pathology.usyd.edu.au; phone: 9351 3381
Project Description: Motor neuron disease causes a progressive loss of motor neurons in adults, and leads to death in about 3 years. The cause is unknown, but we think it is due to a gene defect that is found only in cells of the central nervous system. We are therefore looking (in brain tissue from people who have died of motor neuron disease) for novel genetic mechanisms that could underlie this disease. If you are interested in neurology and genetics, and expect to get a WAM of over 80, we would be very interested in discussing our projects with you further.

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IRON METABOLISM AND CHELATION PROGRAM

Development of New Iron Chelators as Novel Drugs Against Cancer
Supervisors:  Professor Des R. Richardson and Dr. David Lovejoy
Contact Details: Room 555B Blackburn Building D06; email: d.richardson@med.usyd.edu.au; Phone: 9036-6548
Project Description: This project will use a combination of techniques that are implemented in chemistry, biochemistry, physiology, molecular biology and pharmacology to design and assess the activity of novel drugs for the treatment of cancer. Your project will be multi-faceted and will involve growing human tumour cells in tissue culture and assessing the effects of chelators on gene expression. This will be done using Western analysis, RT-PCR and microarray analysis. The lab has considerable experience in these cutting-edge techniques and you will be taught the intricacies of their use. Feel free to contact Prof. Des Richardson or Dr. David Lovejoy (dlovejoy@med.usyd.edu.au) to have a chat about whether the project matches your interests.

Transport of Nitric Oxide in Cells and its Interaction with Iron Containing Proteins in Tumour Cells
Supervisors:  Professor Des R. Richardson
Contact Details: Room 555B Blackburn Building D06; email: d.richardson@med.usyd.edu.au; Phone: 9036-6548
Project Description:  This project will use a combination of techniques that are used in biochemistry, physiology, cell biology, molecular biology and pharmacology. Your project will be multi-faceted and will involve growing human tumour cells in tissue culture and assessing the effects of nitric oxide on the iron transport. This will be done using a wide variety of techniques including Western analysis, RT-PCR and microarray analysis. The lab has considerable experience in these cutting-edge techniques and you will be taught the intricacies of their use. Feel free to contact Prof. Des Richardson to have a chat about whether the project matches your interests.

The Function of the Malignant Melanoma Tumour Antigen, Melanotransferrin (p97), in Tumourigenesis
Supervisors:  Professor Des R. Richardson
Contact Details: Room 555B Blackburn Building D06; email: d.richardson@med.usyd.edu.au; Phone: 9036-6548
Project Description: This project will use a combination of techniques that are used in biochemistry, physiology and molecular biology. Your project will be multi-faceted and will involve growing human tumour cells in tissue culture and the use of a number of knockout and transgenic mouse models developed in our laboratory. A wide variety of techniques will be used including Western analysis, RT-PCR and microarray analysis. The lab has considerable experience in these cutting-edge techniques and you will be taught the intricacies of their use. Feel free to contact Prof. Des Richardson or Mr. Yohan Rahmanto (yohans@med.usyd.edu.au) to have a chat about whether the project matches your interests.

The Role of Iron in the Pathogenesis of the Crippling Neurodegenerative Disease, Friedreich’s Ataxia
Supervisors:  Professor Des R. Richardson
Contact Details: Room 555B Blackburn Building D06; email: d.richardson@med.usyd.edu.au; Phone: 9036-6548
Project Description: This project will use a combination of techniques that are used in biochemistry, physiology, pharmacology and molecular biology. Your project will be multi-faceted and will involve growing cells in tissue culture and the use of a unique knockout mouse model. A wide variety of techniques will be used including Western analysis, RT-PCR and microarray analysis. The lab has considerable experience in these cutting-edge techniques and you will be taught the intricacies of their use. Feel free to contact Prof. Des Richardson or Miss Megan Whitnall (meganw@med.usyd.edu.au) to have a chat about whether the project matches your interests.

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CENTRE FOR VASCULAR RESEARCH

Role of indoleamine 2,3-dioxygenase in control of blood vessel tone in inflammation
Supervisors:  Professors Roland Stocker and Nick Hunt, and Dr Yutang Wang (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25), Email: rstocker@med.usyd.edu.au, Phone: 9036 3207
Project Description: Using a mouse model we are studying the role of the tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO) in various aspects of the pathogenesis and related physiological changes of cerebral malaria infection. We were the first to show that endothelial cells located in small blood vessels of the brain and other tissues express IDO during malaria infection, in a process strictly dependent on interferon gamma. We recently (unpublished) observed that IDO contributes to the control of vascular tone (blood pressure) in malaria-infected mice.
In this joint project between the two laboratories, the blood vessel relaxing properties of IDO-derived products will be studied, with the aim to establish the active component derived from tryptophan and the mode of action of this compound. We also will examine the role of IDO in the regulation of vascular tone in other mouse models of inflammation, including atherosclerosis. The techniques used will include measurement of blood pressure in and isolation of blood vessels from mice, and in vitro vascular function studies using physiological myobath systems.

The role of heme oxygenase as an antioxidant defense
Supervisors:  Professors Roland Stocker (Pathology) and Ian Dawes (UNSW), and Dr Emma Collinson (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25), Email: rstocker@med.usyd.edu.au, Phone: 9036 3207
Project Description: Heme oxygenase catalyzes the oxidative degradation of heme and plays a key role in iron homeostasis by facilitating the ‘return’ of heme-derived iron to the bone marrow where it can be used for hematopoiesis. A large body of recent literature suggests that one of the isozymes of heme oxygenase (heme oxygenase-1) has a number of protective activities in diseases associated with increased oxidative stress, such as cardiovascular, neurodegenerative and inflammatory conditions. The mechanisms underlying these protective effects are largely unknown, although antioxidant protection has been put forward as one likely possibility.
Recently, a functionally active yeast heme oxygenase has been identified. The role of HMX1 in controlling intracellular heme levels has been elucidated. However, little is known about whether it offers protection against oxidative stress and whether its expression is redox regulated. This project will use standard yeast molecular techniques and biochemistry to address these questions.

Control of growth of vascular cells by heme oxygenase-1
Supervisors:  Dr Konstanze Beck and Professor Roland Stocker (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25), Email: kbeck@med.usyd.edu.au, Phone: 9036 3212
Project Description: Atherosclerosis is the major single cause of cardiovascular disease (CVD) that itself remains the single major cause of death in Western countries including Australia. While lipid-lowering drugs (e.g., statins) have been extremely successful in lowering CVD, there nevertheless is an urgent need for the development of novel drugs that protect against atherosclerotic vascular disease by means other than lipid lowering. We recently identified heme oxygenase 1 (HO 1) as a target of the anti-atherosclerotic action of an old, now uncommonly used, drug (probucol) and are developing novel probucol analogs that share the beneficial but not undesirable side effects of probucol. Regulation of HO 1 by probucol and the novel drugs we are developing provides several protective effects in vivo, including the promotion of re-endothelialization and the inhibition of excessive proliferation of vascular smooth muscle cells. In vitro studies confirm that the novel drugs induce HO 1 mRNA, protein and activity in vascular smooth muscle cells, and this is directly responsible for the inhibition of excessive growth of these cells. However, at present, we know much less about how the novel drugs exert this striking cell-specific effect, i.e., they promote (rather than inhibit) the growth of endothelial cells. As hydrogen peroxide (H2O2) is the only agent known to have such cell type-specific effect, this project will investigate the effect of the novel drugs on enzymes involved in H2O2 synthesis and metabolism. This will be examined in both endothelial and vascular smooth muscle cells. The techniques involved include cell culture, molecular techniques (e.g., to increase and decrease HO 1 expression) and biochemical methods (to relate differences in HO 1 activity to differences in peroxidase activity).

The role of heme oxygenase-1 in cerebral malaria infection
Supervisors:  Professors Roland Stocker (Pathology) and Nick Hunt (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25), Email: rstocker@med.usyd.edu.au, Phone: 9036 3207
Project Description: High activity of heme oxygenase 1 (HO 1) in host brain and in tissue macrophages has recently been reported to protect against experimental cerebral malaria, and this protective effect could be recapitulated by administration of carbon monoxide (CO), a metabolic product of heme oxygenase activity. It is thought that CO binds to hemoglobin released from erythrocytes undergoing rupture during the blood stage of the parasite life cycle, and by doing so prevents hemoglobin from releasing the toxic heme that otherwise damages endothelial cells and contributes to the adherence to, and hence accumulation of, CD8+ T cells in cerebral blood vessels. The latter event has been intimately linked to the pathogenesis of cerebral malaria. High endogenous activity of heme oxygenase, or induction of HO-1 by pharmaceutical agents, is thought to protect against cerebral malaria because it results in enhanced generation of CO.
As heme plays a central role in the proposed new function of HO-1, this project will examine (i) the potential role of the heme-binding protein hemopexin in cerebral malaria, (ii) where precisely HO-1 is induced during infection, and (iii) whether metabolic products in addition to CO may impact on disease outcome. For this, the mouse model of cerebral malaria established in the laboratory of Professor Hunt will be used in conjunction with techniques available in the laboratory of Professor Stocker. The outcome of malarial infection will be assessed by survival, neurological and biochemical parameters.

Imaging redox regulation of cellular signalling systems
Supervisors:  Dr Savine Wimmer-Kleikamp and Professor Roland Stocker (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25), Email: swimmer@med.usyd.edu.au, Phone: 9036 3212
Project Description: If you chose this project you will become familiar with a number of cutting edge microscopy and imaging technologies, in combination with biochemical and molecular biology methods. Receptor tyrosine kinases (RTKs) regulate key cellular processes like cell migration, -differentiation and -proliferation. Abnormal signalling of many family members has been linked to diseases, such as cancer and vascular disease. The aim of this project is to identify oxidants and enzymes that affect the redox state and redox processes following receptor tyrosine kinase activation on endothelial cells. We will approach this question from both a cellular biology and biochemical perspective and test the functional relevance of our findings in experimental animal models. This multifaceted project will provide a better understanding of these complex processs, which may allow the development of novel therapies to target abnormal redox-modulated pathways in endothelial disease.

Role of cytochrome b5 in the reductive activation of indoleamine 2,3-dioxygenase
Supervisors:  Dr Gus Maghzal and Professor Roland Stocker (Pathology)
Contact Details: Level 2, Medical Foundation Building (K25), Email: rstocker@med.usyd.edu.au, Phone: 9036 3207
Project Description: Human indoleamine 2,3-dioxygenase (IDO) is an intracellular heme enzyme that catalyzes the oxidative metabolism of L-Tryptophan (L-Trp) along the kynurenine (kyn) pathway.  The induction of IDO and the formation of kynurenine pathway metabolites have been implicated in processes such as immune regulation, neuropathology, microbial and tumor defense and more recently by our group in the regulation of vascular tone.
IDO requires to be activated via reduction of its ferric heme.  For the last 30 years, the dogma has been that IDO requires and consumes superoxide anion radical (O2-.) to metabolise L-Tryp to Kyn. We observed that O2-. can also activate recombinant human IDO. However, the extent of this activation is modest, and small changes in the cellular concentration of O2-. barely affect IDO activity, suggesting that O2-. play s only a minor role. Instead, we have recently obtained evidence for a role of microsomal cytochrome b5 and NAPDH cytochrome P450 reductase in the activation of cellular IDO. In this project, we will investigate the mechanisms of cytochrome b5-induced activation of IDO by examining whether a physical interaction is required. We will also investigated if the mitochondrial form of cytochrome b5 is also involved in the activation of IDO.

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REDOX BIOLOGY LABORATORY

Post translational changes to key cardiac proteins in the hearts of diabetic rats after experimental heart attack
Supervisors:  Dr Paul Witting (Pathology) and Dr Aisling McMahon (Biogerontology Group, ANZAC Research Institute)
Contact Details: Room 509 Blackburn Building, Email: pwitting@med.usyd.edu.au
Project Description:
The major causes of morbidity and mortality in diabetes are the cardiovascular complications of the disease.  Of these, the development of diabetic cardiomyopathy is of particular concern, as it increases the risk and severity of acute myocardial infarction (heart attack) compared with the general population.  The proposed project will provide information on the role for oxidative stress and inflammation in promoting the severity of myocardial infarct in an animal model of diabetic cardiomyopathy.  We plan to use an established rat model of diabetes and monitor post translational changes to key cardiac proteins through a proteomic approach combining 2D gel and mass spectrometry peptide mass mapping.  An understanding of the role for oxidative stress and inflammation following acute myocardial infarct in diabetics may allow the development of new therapeutic approaches targeted at slowing down the progression of cardiomyopathy and thereby potentially improving heart function and quality of life in diabetes sufferers.  This would be viewed as a major advance.

AFFILIATES

MOLECULAR ONCOLOGY LABORATORY, ONCOLOGY RESEARCH UNIT, THE CHILDREN'S HOSPITAL AT WESTMEAD

In vitro models for the role of D52 protein in tumourigenesis
Supervisors:  Associate Professor Jennifer Byrne
Contact Details: Oncology Research Unit, The Children's Hospital at Westmead, Email: JennifeB@chw.edu.au
Project Description:
Our group studies the functions of a family of proteins which we have identified in human cancer cells, called D52-like proteins.  We and other investigators have shown that the D52 gene and/or protein is upregulated in most breast, prostate and ovarian cancers, and that this is frequently associated with gene amplification.  Overexpression of D52 may contribute to the early stages of tumour initiation or progression through increasing cell proliferation.  However, it is likely that D52 overexpression also contributes to cancers forming or progressing in other ways.  For example, we have recently found that expressing D52-like proteins in cancer cells gives rise to multinucleated cells.
To examine the phenotypic consequences of expressing D52 and the related protein D53 in cell lines, we have constructed mouse 3T3 fibroblast cell lines expressing either human protein, and have sujected these cell lines to a variety of phenotypic analyses.  These have indicated only D52 expression increasesd proliferation in both culture dishes and soft agar, but that expression of D52 and D53 similarly alters cell morphology.  These results represent the first confirmation at the cellular level that D52-like proteins have both redundant and isoform-restricted functions at the molecular level.  The current project involves the continued analysis of these cell lines, and others already derived from the MDA-MB-231 breast cancer cel line, to determine additional phenotypes associated with D52 and/or D53 expression.  We will be particularly focussed upon testing hypotheses which have recently emerged from assessing the clinical significance of D52 expression in human breast cancer.  These analyses represent the first to compare the functions of individual D52-like genes in a single system, and we predict that they will indentify functional differences between D52-like proteins which will be relevant to their roles in cancer cells.
Techniques:  Cell culture, fixing and immunofluorescently staining cultured cells, RNA and/or protein extraction from cultured cells, RT-PCR and/or Western blot analyses, with scope to include antibody preparation and testing.

CANCER BIOLOGY GROUP, DIVISION OF MEDICINE (ENDOCRINOLOGY) AND SYDNEY CANCER CENTRE

Arachidonic acid pathway and prostate cancer
Supervisors:  Dr Qihan Dong
Contact Details: Room 371 and 394, Blackburn Building D06, Email: qhd@med.usyd.edu.au
Project Description:
Prostate cancer is the second most commonly-diagnosed malignancy after skin cancer and second only to lung cancer in males in terms of cancer-related death. The strategies we have taken are to (1) identify the altered signalling pathways in human prostate cancer tissue; (2) verify the consequence of the alteration in signalling on cell growth; (3) determine the cellular mechanism leading to the change in cell growth; (4) evaluate the therapeutic potential of targeting the pathway and (5) conduct clinical trials.  We have established that an aberrant activation of the arachidonic acid (AA) pathway occurs in human prostate cancer tissue.  This project aims to target the AA in a collective manner as a therapeutic strategy for the treatment of advanced prostate cancer.  Under the guidance of postdoctoral or senior PhD student, Honours student will use the techniques of molecular biology (qRT-PCR and Western), biochemistry (AA release) and cell biology (cell culture, immunostaining and reporter assay) to understand the role of key players of the AA pathway in the growth of advanced prostate cancer.  This project is unique in the following aspects; (i) it tests a number of chemicals never before tested in prostate cancer cells, (ii) it explores relationship between AA and other key pathways in the same experimental paradigm and (iii) in-vitro effect will be validated in animal models of prostate cancer.

HEPATITIS C VIRUS PATHOGENESIS GROUP, STORR LIVER UNIT, WESTMEAD MILLENNIUM INSTITUTE

Hepatitis C Virus and Steatosis
Supervisors: Dr Mark Douglas
Contact Details: Storr Liver Unit, Westmead Millennium Institute. Email: mark.douglas@usyd.edu.au
Project Description:
There is increasing evidence that cellular lipid pathways are fundamentally involved in the replication cycle of hepatitis C virus (HCV). Around 50% of patients infected with HCV develop steatosis (fatty liver), and this may accelerate fibrosis and cirrhosis. Steatosis is more common in patients infected with HCV genotype 3, suggesting a direct, genotype-specific viral effect. HCV core protein causes accumulation of lipid droplets within hepatic cells, and we have observed redistribution of lipid droplets in cells expressing core protein. The importance of this process for lipid metabolism in hepatocytes is not clear, but in adipocytes redistribution of lipid droplets is critical for regulation of lipid flux. This project would look at the effects of HCV proteins from different HCV genotypes on lipid metabolism in hepatic cells, as a model for steatosis. Various systems would be used to express HCV proteins in hepatic cells, including recombinant baculovirus and unique intergenotype chimeras, based on the new infectious HCV cell culture model. A better understanding of HCV and steatosis may lead to improved treatments for HCV-induced steatosis and for HCV infection itself.

How does Hepatitis C virus Cause Insulin Resistance?
Supervisors:  Dr Mark Douglas
Contact Details: Storr Liver Unit, Westmead Millennium Institute. Email: mark.douglas@usyd.edu.au
Project Description:
It was recently discovered that patients infected with HCV have an increased rate of insulin resistance, making HCV the first non-lifestyle cause of diabetes mellitus. Insulin resistance is more common in patients infected with HCV genotypes 1 or 2, and predicts faster disease progression and non-response to anti-viral therapy. The exact mechanism is not known, but it is likely that HCV proteins interact with molecules in the insulin signalling pathway. In our project we will use various novel models to study the effects of HCV proteins on hepatic cells in vitro, including a recently developed infectious cell culture model of HCV. We will use inter-genotype chimeras to express proteins from different HCV genotypes in HCV-infected cells and study their effects on insulin signalling. We will also study the effects of HCV proteins on immortalised human hepatocytes, a more physiologically relevant cell line, using a recombinant baculovirus system. We expect that the results of this study will suggest new treatment strategies for people infected with HCV.

HEPATOCELLULAR CANCER GROUP, STORR LIVER UNIT, WESTMEAD MILLENNIUM INSTITUTE

Understanding the role of adiponectin in liver disease and cancer
Supervisors: Dr Lionel Hebbard
Contact Details: Storr Liver Unit, Westmead Millennium Institute. Email: lionel_hebbard@mail.wmi.usyd.edu.au
Project Description:
Adiponectin is a growth factor produced by adipocytes. Mouse models have shown adiponectin to have protective functions in reducing the risk for obesity-related metabolic problems, including insulin resistance and Type 2 diabetes, hypertension, coronary heart disease and stroke. Additionally, recent clinical studies from the Storr Liver Unit have shown a link between lowered serum levels of adiponectin and liver disease progression. AdipoR1, AdipoR2 and T-cadherin (CDH13) are the three receptors that have been identified to bind adiponectin. We hypothesize that adiponectin and binding to its receptors are important mediators in liver disease and ultimately liver cancer formation. Therefore, our studies concern elucidating which receptor mediates the action of adiponectin in liver disease. We will do this utilizing knock-out mice for each of the adiponectin receptors and a well-published rapid model of liver injury, regeneration and fibrogenesis. The successful applicant will receive extensive training and support from a senior scientist and two research assistants, who collectively have extensive experience in mouse models, histology, biochemical techniques and isolating primary cells. The Westmead Millennium Institute is part of the University of Sydney Western Clinical School and offers an extremely collaborative research environment. The completion of this project will lead to a better understanding of the pathogenesis of liver injury and fibrosis, and ultimately better and more targeted therapies for the treatment of liver disease.

VASCULAR BIOLOGY RESEARCH CENTRE, WESTMEAD HOSPITAL

Redefining the role of monocytes in atherosclerosis
Supervisors:  Dr Heather Medbury / Dr Sarah Tarran
Contact Details: Vascular Biology Research Centre, Westmead Hospital, Westmead, Email: hmedbury@med.usyd.edu.au
Project Description:
Atherosclerosis continues to be a major cause of morbidity and mortality in Australia.  With current treatments offering only a temporary solution - what is urgently needed is a new class of drugs that address the key cause of clinical events; plaque rupture.  With ruptures more likely to occur in unstable atherosclerotic plaques (ie ones that have a relatively thing fibrous cap and large fatty core), the therapeutic goal is thus to convert unstable plaques to stable plaques.
We propose that the stability of the atherosclerotic plaque hinges on the differentiation of the monocytes. While monocytes have traditionally been known to play a detrimental role in cardiovascular disease (and indeed there is a wealth of literature in this area), we have recently shown (in clinical samples) that monocytes contribute to the formation of the atherosclerotic cap by transforming into a smooth muscle-like cell called a fibrocytes.
Aim:  To further investigate the role of the monocyte in plaque stability we have developed a unique mouse model, the Macgreen /ApoE-/- mouse, an atherosclerotic mouse whose monocyte and monocyte-derived cells fluoresce green.  Our aim to track monocyte contribution to formation of the fibrous cap, determining when and where it transforms, under what conditions and to what degree it contributes to the cap versus other cell types.  Students will be given their own distinct angle of this project to investigate.
Significance: Understanding monocyte transformation will help us to identify targets for therapeutic manipulation that could stabilize the disease.  This could result in a reduction in clinical events such as heart attack and stroke, thus decreasing the morbidity and mortality associated with the disease.
Techniques used:  The main techniques that will be used include immunohistochemistry, immunofluorescence and confocal microscopy.

PHARMACOGENOMICS GROUP, MEDICAL FOUNDATION BUILDING

Regulation of human renal organic anion transporter genes by bradykinin
Supervisors:               Prof Michael Murray and Dr Fanfan Zhou (Pharmacogenomics, Pharmacy)
Contact details:        Level 1, Medical Foundation Building (K25), Email: michaelm@pharm.usyd.edu.au or fzhou@pharm.usyd.edu.au, Phone: 9351 2326 (Michael Murray) or 9036 3015 (Fanfan Zhou)
Project description:
Human organic anion transporters (hOATs) are a family of membrane-bound proteins that control the flux of drugs and toxins through the kidney. Factors that regulate their expression in cells may influence the pathogenesis of renal disease by altering toxin elimination. Bradykinin (BK) is an important member of the kinin group of proteins that regulates many renal genes including transporters, but the details are unclear at this stage.  In this project the regulatory effects of BK on the expression and function of individual hOAT genes will be investigated for the first time in human kidney cell lines. The major techniques involved in this study will include cell culture, RNA analysis by real-time PCR, immunoblotting and transporter functional assays. Training in appropriate methods will be provided by Dr Zhou and postdoctoral scientists within the group. The findings could aid our understanding of the pathogenesis of kidney diseases in humans and provide a basis for novel treatments.

Antitumour effects of w-3 polyunsaturated fatty acids in breast cancer cells
Supervisors:               Prof Michael Murray and Dr Sarah Cui (Pharmacogenomics, Pharmacy)
Contact details:        Level 1, Medical Foundation Building (K25), Email: michaelm@pharm.usyd.edu.au or pcui@pharm.usyd.edu.au, Phone: 9351 2326 (Michael Murray) or 9036 3245 (Sarah Cui)
Project description:
From epidemiological studies the dietary intake of w-3 polyunsaturated fatty acids (w-3 PUFA) has a range of health benefits, including a decrease in the spread of breast and other cancers. In contrast, w-6 PUFA, which are prevalent in western diets, accelerate tumour growth. Recent findings from our laboratory and others suggest that PUFA regulate the tumour cell cycle by altering the expression of important cyclin genes. This project will study the mechanism by which cyclins are modulated in breast cancer cells by different types of PUFA found in diets. Major techniques will include cell culture, RNA analysis by real-time PCR, immunoblotting and cell cycle kinetics. Training in appropriate methods will be provided by Dr Cui and other senior scientists within the group. The findings will increase our understanding of the health benefits produced by w-3 PUFA and could identify new targets for anticancer therapies.

MOLECULAR GENETICS LABORATORY & SUPAMAC, CENTRAL CLINICAL SCHOOL

Detecting Splice-Affecting DNA Variants in Familial Hypertrophic Cardiomyopathy
Supervisors:  Dr Bing Yu (Molecular Genetics) and Dr Julia Morahan (Pathology)
Contact Details: Ground floor, Medical Foundation Building K25, Email: bingy@med.usyd.edu.au, Phone: 9515 5016
Project Description:
    Familial hypertrophic cardiomyopathy (FHC) is a common, inherited heart disease that is caused by more than 250 mutations in genes encoding sarcomere proteins. Most of the mutations previously detected have focused on the coding region and have assessed the impact of these mutations based on protein changes. This project, on the other hand, will explore the splice-affecting nature of FHC mutations, irrespective of coding or non-coding changes, using in silico tools and cell culture. DNA variants, including rare DNA changes under the umbrella of “polymorphisms”, will be screened. Selected variants with plausible splice-affecting characteristics will be confirmed using a splicing functional assay in cell culture. This project is multi-faceted and will involve the collection and in silico screening of FHC mutations as “dry” laboratory work. It will also have traditional “wet” experimental components such as minigene construction, growing cells, transfection, RT-PCR and transcript characterisation. The student will have opportunities to be involved in one or more of these activities and will gain experience in many cutting-edge techniques in the Molecular Genetics laboratory.

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SIDS & SLEEP APNEA LABORATORY

Effects of nicotine exposure on mRNA expression of nicotinic receptors in the brainstem.
Supervisors: Dr. Rita Machaalani & A/Prof. Karen Waters
Contact Details:  Room 206, Blackburn Building (D06), USYD, Phone: 9351 3851; Email: ritam@med.usyd.edu.au
Project Description:
The major focus of the laboratory is on the neuropathology of the Sudden infant death syndrome (SIDS). SIDS is the leading cause of death among infants (1-12 months of age) in developed countries. SIDS victims die suddenly during a sleep period, and the cause is still unknown, although a strong hypothesis is that it is a brainstem disorder of the cardiorespiratory system. SIDS is also associated with risk factors, the two most common being the prone sleep position and exposure to cigarette smoke. This project focuses on identifying changes in nicotinic receptor mRNA levels in a piglet model of nicotine exposure (model designed to mimic passive smoke exposure). The project will involve non-radioactive in-situ hybridisation staining for the expression of nicotinic receptors alpha 7 and beta 2 in nicotine exposed vs non-exposed piglet brainstem. Staining will then be quantified using image analysis systems at the Electron Microscope Unit.

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