External Honours Projects

The Department of Physiology on occasion accepts into its Honours Course students who are performing their research in laboratories outside the Department. The project must be closely allied to physiology and an internal supervisor who is familiar with the area must be prepared to act as the associate supervisor. The student should normally have undertaken at least one 3rd year course in Physiology and must attend the teaching sessions for Honours students which occur weekly within the Department.

Supervisors and laboratories which have indicated their interest in taking such students are listed below. You should contact the supervisor of the project directly to express your interest.

ANZAC Research Institute - Dr Colin Dunstan

The ANZAC Research Institute is a new purpose built research facility in the grounds of Concord Hospital, Concord. The Bone Biology Laboratory is a well equipped research facility. The research group includes three scientists with research assistants and students and is conducting research into bone metastasis and osteoporosis,

Project 1.

Breast Cancer Metastasis
This projects investigates the factors which drive the propensity of breast cancer cells to establish secondary tumours in bone. The student will use fluorescently labelled cells to track the targeting of human breast cancer cells to bone in mice. The bone metabolism in the mice will be altered using pharmacologic agents to determine if this changes breast cancer cell trafficking. This project will involve small animal handling, cell culture, histology and molecular biology.

Project 2.

Glucocorticoid regulation of osteoblasts
Primary cultures of osteoblasts will be derived from wild type and transgenic mice that have osteoblast specific inhibition of glucocorticoid (cortisol) signalling that results in altered ability to form bone. This project will use detailed cell based gene expression studies to determine the pathways impacted by blocking of glucocorticoid signalling. Techniques developed will be cell culture, molecular biology, and small animal handling.

For more information contact:

Dr Colin Dunstan
Principal Research Fellow
ANZAC Research Institute
(in the grounds of Concord Hospital, Concord)
+61 2 9767 9163
cdunstan@med.usyd.edu.au

VASCULAR BIOLOGY RESEARCH CENTRE
DEPT SURGERY, WESTMEAD HOSPITAL - Dr Heather Medbury

We are at the cutting edge of vascular research, recently identifying the presence and role of progenitor cells in the development of atherosclerosis. Our success comes from the direct interaction between surgeons and scientists; enabling our research to span from a basic fundamental level to a clinical research angle. We have both honours and PhD projects available.


THE ROLE OF PROGENITOR CELLS IN DISEASE


Our aim is to understand the role of progenitor cells in wound healing so that we can alter its progression to give a better outcome in various disease states. We are focusing in particular on atherosclerosis (heart disease) and the wound healing response (intimal hyperplasia) generated upon its treatment. Our work has attracted interest from other researchers and as such we have established collaborations with several other groups.
 
Our current research projects (which will continue next year) include:
1.    The role of progenitor cells in atherosclerosis (vascular disease)
We have recently identified the presence and role of a progenitor cell in atherosclerosis. We are currently comparing samples from patients versus controls to determine what factors effect differentiation of progenitor cells in atherosclerosis development.

2.    The pluripotent nature of progenitor cells in wound healing
Wound healing involves a range of cells interacting to restore tissue integrity. We are determining what range of cells the progenitor can differentiate into in this process.

3.    The role of platelets in progenitor cell differentiation
Our preliminary work has shown that platelets can effect the differentiation of the progenitor cells into different cell types depending on how they are allowed to interact. We are examining what factors from the platelets effect this.


Contact:
Dr Heather Medbury
Phone: 9845 7677
Email: heather_medbury@wsahs.nsw.gov.au
www.wmi.usyd.edu.au/reseacentres/surgery.html

Brain and Mind Research Institute

Honors projects (9 months): Pathogenesis of Alzheimer’s disease

Available tools and techniques:

·     Transgenic mouse models, transgenic brain bank, tissue culture lines
·     Molecular biology (DNA, RNA work)
·     Cell biology, histology
·     Animal handling (stereotaxic injections, production of transgenic mice, micromanipulation)
·     Proteomics, transcriptomics
·     Biochemistry (to some extent)

Possible project outlines:

Histological and functional validation of candidate proteins identified in models of Alzheimer’s disease by Functional Genomics
Using proteomics, we have identified differential protein spots in a mouse model of Alzheimer’s disease (AD). The aim of the project is to use antibodies to determine by immunohistochemistry and Western blot analysis which of the candidates is differentially regulated in human AD and control brains. Where applicable, functional assays will be performed (for example phosphorylation assays for kinases or depolarisation assays for mitochondrial proteins. Part of these assays may be done in our tissue culture systems).

Transport of Aβ in the mouse brain
The amyloid cascade hypothesis claims that Aβ induces the tau pathology in AD and by that, in part, exerts its toxicity. When added extracellularly, Aβ may exert its toxic effects by pore formation, uptake and transport, via contribution of the glial compartment, and by damage of nerve terminals. We propose to stereotaxically inject fibrillar preparations of either fluorescently or 3H-labelled Aβ42 into the CA1 region of the hippocampus and to determine whether Aβ42 is transported in vivo along projections from the amygdala to the CA1 region.

Specificity of the Aβ42-mediated induction of tau tangles (NFTs) in vivo
Tau tangle (NFT) formation characterizes many neurodegenerative disorders besides AD. Whereas in some diseases, they are the only type of proteinaceous aggregate, in others they co-occur with fibrillar aggregates such as Aβ in AD, the scrapie form of the prion protein (PrPsc) in prion disorders, or the amyloid-Bri (A-Bri) peptide in British dementia. Thus, with respect to tau-related pathogenesis, this project addresses the question whether the observed increase in NFT numbers in P301L mice caused by stereotaxic injection of fibrillar preparations of Aβ42 could be induced by other fibrillar aggregates such as PrPsc, A-Bri, or the amylin which is aggregating in diabetes.

Role of oxidative stress in transgenic mice with an Alzheimer-like tau pathology
A proteomic and functional analysis of P301L tau transgenic mice (our major Alzheimer model) revealed a mitochondrial dysfunction. The aim of this project is to expose age-matched hemizygous and homozygous P301L mice to oxidative stress, determine functional impairment and correlate these with the tau pathology in distinct brain areas using Western blotting and immuno-histochemistry

Dissection of the functional domains of tau by a transgenic approach
The tau protein is organized into functional domains termed projection domain, proline-rich region, repeat region and carboxy-terminal tail, with the latter possibly inhibiting tau aggregation. Some of the interaction partners of these domains have been identified. As tau pathology represents a common end point of several diseases collectively termed tauopathies it will be important to understand the role of the complex interactions of tau in vivo. The project involves the design of novel transgenic animal models where individual interactions are disturbed. In addition, specific amino acid substitutes will be introduced assisting in the analysis.

Transgenic mouse model with a brain pathology in the skin
The brain is difficult to access for experimental manipulation. The mainly neuronal protein tau aggregates in human disorders. Tau is also expressed at lower levels in the skin, and many signalling pathways with tau as a target are shared between the brain and the skin. To circumvent the limitations of the brain’s inaccessibility, we generated transgenic mice which express mutant tau in the skin. This project involves a histopathological and Western blot analysis of this mouse strain, combined with depilation to determine the hair cycle-dependent pathology.

Suggested reading:
·Götz J, Chen F, van Dorpe J, Nitsch RM (2001) Formation of neurofibrillary tangles in P301L tau transgenic mice induced by Abeta42 fibrils, Science 293: 1491-1495 [plus perspectives]
·Ferrari A, Hoerndli F, Baechi T, Nitsch RM, Götz J (2003) β-Amyloid induces PHF-like tau filaments in tissue culture, J. Biol. Chem. 278, 40162-40168
·Götz J, Streffer JR, David D, Schild A, Hoerndli F, Pennanen L, Kurosinski P, Chen F (2004) Transgenic animal models of Alzheimer’s disease and related disorders: Histopathology, behavior and therapy, Mol. Psychiatr. 9: 664-683 [cover image]
·Pennanen L, Welzl H, D’Adamo P, Nitsch RM, Götz J (2004) Accelerated extinction of conditioned taste aversion in P301L tau transgenic mice, Neurobiol. Dis. 15: 500-509
·Chen F, Wollmer A, Hoerndli F, Münch G, Kuhla B, Rogaev EI, Tsolaki M, Papassotiropoulos A, Götz J (2004) Role for glyoxalase I in Alzheimer’s disease, Proc. Natl. Acad. Sci. USA 101: 7687-7692
·Hoerndli F, Toigo M, Schild A, Götz J, Day P (2004) Reference genes identified in SH-SY5Y cells using custom-made gene arrays with validation by quantitative PCR, Anal. Biochem. 335: 30-41
·David D, Hauptmann S, Scherping I, Schuessel K, Keil U, Dröse S, Brandt U, Müller WE, Eckert A, and Götz J (2005) Proteomic and functional analysis reveal a mitochondrial dysfunction in P301L tau transgenic mice, J. Biol. Chem. 280: 23802-23814

Current staff:

• Dr. Lars Ittner
• Dr. Della David
• Dr. NN
• Further staff will be recruited within the next months

Prof. Jürgen Götz
Brain and Mind Research Institute
University of Sydney
100, Mallett St
Camperdown NSW 2050
Tel. 93510799
jgoetz@med.usyd.edu.au

ONCOLOGY RESEARCH UNIT, THE CHILDREN?fS HOSPITAL AT WESTMEAD - Dr Geraldine O'Neill

FOCAL ADHESION BIOLOGY GROUP: RESEARCH PROGRAM
Cellular interaction with the external environment through receptor-mediated adhesion to the extra-cellular matrix regulates many cellular processes including response to death stimuli and cell movement. These processes are characteristically altered in cancer cells. The Focal Adhesion Biology group research program is divided into studies of two key interrelated mechanisms: (1) adhesion mediated survival signalling and; (2) adhesion regulation of cancer cell migration. We have student projects available in both areas.

(1) Adhesion mediated survival signalling: a mechanism for resistance to anti-cancer therapy?
Our research in this area arises from the idea that the very events that promote cancerous transformation also can advantage the cells to resist anti-cancer therapies. This aspect of cancer cell biology is a major confounder in the ability to efficiently treat cancer. We are particularly interested in how different adhesion structures function as organizing centres to alternatively regulate down-stream activation of survival signals. Specifically we are examining the role of adhesion-associated signalling in response to anti-breast cancer treatments. Techniques to be employed in this project include culture of breast cancer cell models, molecular biological and biochemical analyses.

(2) Focal adhesion regulation of cancer cell migration.
Two core components in migration are the actin cytoskeleton and integrin-based adhesion to the extra-cellular matrix (ECM). Importantly, these two components are characteristically altered in cancer and therefore may well provide cancer-specific targets for therapy. We have initiated a program investigating the cytoskeletal regulation of adhesion structure and the consequent effects on cell migration. We are exploring this mechanism by employing live cell imaging techniques to investigate cytoskeletal regulation of adhesion dynamics. Our overall goal is to investigate both normal and pathological adhesion signalling mechanisms that regulate the stimulation of cell movement.

Contact: Dr Geraldine O'Neill
Group Leader, Focal Adhesion Biology group
Oncology Research Unit
The Children's Hospital at Westmead
Phone: 98453116

Muscle Development Unit, The Children's Medical Research Institute, Westmead - Dr Edna Hardeman

Project 1. Understanding the genetic basis of a human brain disorder: The pathophysiology of the neurodevelopmental disease, Williams Syndrome
Background: Williams Beuren syndrome (WBS) is a neurodevelopmental disorder characterised by physical, cognitive and very distinctive behavioural abnormalities. This syndrome is caused by a 1.6 Mb hemizygous deletion on 7q11.23. The behavioural and cognitive abnormalities are known as the ‘Williams syndrome cognitive profile’ (WSCP) and include a hypersocial personality, enhanced verbal skills, a mild mental retardation and specific phobias. The uniformity of the WSCP indicates that one or more of the 22 genes deleted in WBS are responsible for the development of specific neurological pathways controlling aspects of normal behaviour and cognition. Recently, several small atypical deletions within the WBS deleted region have been described. The study of the cognitive and behavioural profiles of multiple patients with different deletions suggests that it is the deletion of two genes that gives rise to the WSCP, GTF2IRD1 and GTF2I.
Hypothesis: Gtf2i expression is involved in specific cognitive and behavioural processes.
Aims
1. To determine in which regions of the brain and in what cell type(s) the Gtf2i gene is expressed. The results from this aim will help us to better predict what neuronal processes might be disrupted in WBS that cause aberrant behaviour and cognition. Approach: We have made a Gtf2i knock-out/GFP knock-in mouse in which a green fluorescent protein (GFP) coding sequence has been inserted into the Gtf2i gene. The GFP protein will be expressed in cells in which this gene is active. Brains from developing and mature mice will be sectioned and examined using fluorescence microscopy. The type of neuron in which GFP appears will be confirmed by immunohistochemistry using antibodies to proteins specific to the cell type. Overall brain morphology will be analysed to detect morphological abnormalities resulting from the loss of this gene.
2. To determine whether the reduction in Gtf2i expression in WBS is responsible for any features of the WSCP. Approach: Specialised behavioural and cognitive tests will be performed on knock-out and normal mice to identify abnormalities in sociability, anxiety/fear responses and learning.
3. To identify the genes which are regulated by Gtf2i in the brain. Approach: Affymetrix array analyses will be performed using RNA isolated from regions of the brain in which Gtf2i is highly expressed. Up- and down-regulated transcripts will be confirmed by quantitative RT-PCR.

Project 2. A novel cytoskeleton in skeletal muscle that may have a role in diabetes
Background: Defects in the glucose transport pathway lead to obesity and type II diabetes. Glucose uptake into skeletal muscle is acutely stimulated by insulin and exercise and involves the movement of the glucose transporter, GLUT4, to the plasma membrane. Understanding how GLUT4 is stored in skeletal muscle and how it gets to the plasma membrane upon stimulation is important to understanding what goes wrong in obesity and type II diabetes. Some of the other components of glucose transport are syntaxin-4, a membrane protein involved in fusion of transport vesicles with the plasma membrane, and the actin cytoskeleton. Recent data indicates that syntaxin-4 binds to or interacts with an isoform of the actin-associated protein tropomyosin, Tm5NM1. Tropomyosin (Tms) is a family of filamentous proteins consisting of over forty isoforms that play an important role in defining the function of the actin cytoskeleton. The finding that syntaxin-4 interacts with this specific Tm suggests that the actin filament that Tm5NM1 associates with is the one involved in trafficking GLUT4 to the plasma membrane. Disruptions of this filament system could contribute to obesity and type II diabetes. We have transgenic and knockout mice that overexpress or lack expression of Tm5NM1 in skeletal muscle, respectively. This project will use these start-of-the-art experimental models to examine the role of Tm5NM1 in glucose transport.
Hypothesis: Disruption of the Tm5NM1-defined actin filaments in Tm5NM1 transgenic and knockout mice will alter the exercise- and insulin-induced stimulation of the GLUT4 pathway.
Aims
1. To examine the changes in expression of GLUT4 and other members of the glucose transport pathway in Tm5NM1 transgenic and knockout mice following exercise and insulin treatment. Approach: Tm5NM1 transgenic and knockout mice will be exercised on a treadmill or injected with insulin and various skeletal muscles will be taken for analysis of expression of GLUT4, syntaxin-4 and other components of the glucose transport pathway. RNA will be extracted from the muscle tissue and analysed by Northern blot or RT-PCR to quantify transcript abundance. The levels of the proteins encoded by these transcripts will also be analysed in extracts from muscle using Western blots.
2. To examine insulin-stimulated glucose uptake in vivo in Tm5NM1 mice. Approach: Mice will be injected with insulin and the changes in blood glucose will be measured at various times after the injection. Glucose will be measured using a standard glucometer.

Project 3. Molecular mechanisms of muscle fibre adaptation that influence muscle performance
Background: Skeletal muscle is composed of individual muscle cells or fibres that have varying contraction speeds and metabolic properties. These performance differences are underpinned by the expression of gene sets that are controlled in a highly ordered way. Positive or negative stimuli such as exercise, disuse, nerve damage, aging and muscle disease can change the types of fibres in muscles by influencing the expression patterns of these gene sets. Certain genes are expressed specifically in fast or slow twitch muscle fibres. By studying the regulation of one fibre type-specific gene Troponin I slow – we aim to understand the molecular basis of the process that determines fibre type and the mechanism that permits fibre conversion in adult life. We have determined the DNA sequence of a critical element that is needed for the expression of Troponin I slow in slow-twitch fibres. By genetic manipulation, we have created mouse models in which fibre type distortions occur via mechanisms that are, as yet, unclear. Now we want to isolate protei ns that can bind and regulate the critical DNA element.
Hypothesis: Different molecular mechanisms exist that determine muscle fibre type specific gene expression.
Aims
1. The protein complex will be isolated that can bind to the critical DNA element in the Troponin I slow gene that is responsible for directing the transcriptional activity of this gene specifically in slow-twitch fibres. Approach: Protein extracts from muscle cell lines will be mixed with a DNA probe containing the critical element in the Troponin I slow gene. The mixture will be electrophoresed under conditions to preserve the protein-DNA complex, the gel sections containing the complexes excised and the identity of the proteins interacting with the critical element determined using mass spectrometry.
2. The signal transduction pathways regulating fibre type change involve the activation of calcineurin and one of its target substrates, the nuclear factors of activated T cells (NFATs) and the transcriptional co-activator PGC1a. We will address whether these pathways are used in the muscles of our mouse models where fibre type distortion occurs. Approach: Changes in the mRNA and protein levels for signalling and transcription factors that are known to regulate muscle fibre type genes will be measured. RNA will be extracted from the muscle tissue of transgenic mice and analysed by real time (RT)-PCR to quantify transcript abundance. The levels of the proteins encoded by these transcripts will be analysed in extracts from muscle tissues using western blot anlaysis.

Contact:Dr Edna Hardeman
Muscle Development Unit, The Children's Medical Research Institute, Westmead
E-mail: ehardeman@cmri.usyd.edu.au
Phone: (02) 9687 2800

Discipline of Psychological Medicine at the Northern Clinical School

Children's Hospital at Westmead

Doctor Karen Waters MBBS, PhD, FRACP

The programme has three clinical honours student projects available in 2008.
They are all to do with Sleep apnoea in children.

The projects are:
1. Measuring respiratory control in children with cleft palate.
2. Does pre-operative CPAP affect surgical outcomes in children with OSA?
3. A randomised clinical trial of treatments for children with mild OSA.

All have ethics approval and have had the groundwork done so the student would start straight away with clinical data collection - thus allowing plenty of time for completion of the project in the time frame of the acadmic year.

Supervisor: A/Prof Janet KEAST, Pain Management Research Institute, RNS Hospital Ph: 99267194; jkeast@med.usyd.edu.au

Our research team explores the structure and function of the nervous system, particularly how the nervous system is affected by injury or inflammation and also how nerve circuits develop to make appropriate contacts in the pre- and neonatal periods. We are interested in basic neurobiology as well as in processes that relate to specific disease processes or injury states (e.g., interstitial cystitis, pain, spinal cord injury, peripheral nerve damage). Honours projects would be suited to enthusiastic students who have very good motor and analytical skills, excellent visual and observational abilities, and a broad interest in the nervous system. In these projects we use microscopy, imaging and microsurgical techniques, as well as knockout mice, in vitro pharmacological assays, behavioral testing, neuronal cultures and molecular biology.

Project titles:

1. Developing new methods for improving nerve growth and function after injury.

We are particularly interested in understanding the effects of injury on pelvic autonomic nerves (which are often damaged during surgical procedures such as hysterectomy and prostatectomy). We are trying to develop ways of promoting regenerative processes in these nerves by investigating the actions of neurotrophic factors, guidance factors and endogenous steroids. In this project you will learn how to culture neurons, investigate the molecular mechanisms of neuronal growth and, for students interested in in vivo function, study nerve regeneration in knockout mice.

2. How do natural steroids affect pain?

We are interested in the mechanisms by which estrogens affect pain signalling neurons (nociceptors) and related spinal circuits, and also how inflammation triggers chronic pelvic pain. We hope to develop new ways to prevent or reverse these chronic pain states by manipulating signaling pathways specific to these neurons. Depending on the interests of the student, this project may involve molecular studies on signaling pathways and receptor trafficking in cultured nociceptors, behavioural studies of spinal reflex pathways, or neuroanatomical studies on structure and activity of nociceptors and central reflex pathways.

Brain and Mind Research Institute: Alzheimer’s Disease Cell Biology Laboratory

Project title: Roles of oxidative-stress and energy deprivation in regulation of Alzheimer’s disease related proteins

Supervisor: Dr. Claire Goldsbury. cgoldsbury@usyd.edu.au tel 9351 0878

Project description: Oxidative stress and reduced glucose metabolism occur in the Alzheimer’s disease brain and are associated with neurodegeneration. Evidence of oxidative stress has been found to precede the major development of senile plaques (comprised of beta-amyloid peptide deposits) and neurofibrillary tangles (comprised of hyperphosphorylated tau protein). The aim of this project is to determine effects of oxidative stress and energy deprivation on the generation of beta-amyloid peptides and tau phosphorylation in neurons. A combination of techniques will be used including primary neuronal cell culture, cell viability assays, immunoprecipitation, Western blotting and immunofluorescence.