The role of insulin and IGF2 signalling on metabolic pathways in prostate cancer progression

Prostate cancer (CaP) is the most commonly diagnosed cancer in males in Australia, North America, and Europe. If found early and locally confined, CaP can be treated with radical prostatectomy or radiation therapy; however, 25-40% patients will relapse and go on to advanced disease. The most common therapy in these cases is androgen deprivation therapy (ADT), which suppresses androgen production from the testis. Lack of the testicular androgen supply causes cells of the prostate to undergo apoptosis. However, in some cases the regression initially seen with ADT eventually gives way to a growth of a population of cancerous cells that no longer require testicular androgens. This phenotype is essentially fatal and is termed castrate resistant prostate cancer (CRPC). In addition to eventual regression, there are many undesirable side effects which accompany ADT, including development of a metabolic syndrome, which is defined by the U.S. National Library of Medicine as “a combination of medical disorders that increase the risk of developing cardiovascular disease and diabetes.” This project will focus on the effect of ADT induced hyperinsulinemia, as mimicked by treating androgen receptor positive CaP cells with insulin in a serum (hormone) deprived environment. While this side effect is not widely explored, in this thesis it is demonstrated for the first time that insulin upregulates pathways important to CaP progression. Our group has previously shown that during CaP progression, the enzymes necessary for de novo steroidogenesis are upregulated in the LNCaP xenograft model, total steroid levels are increased in tumours compared to pre castrate levels, and de novo steroidogenesis from radio-labelled acetate has been demonstrated. Because of the CaP dependence on AR for survival, we and other groups believe that CaP cells carry out de novo steroidogenesis to survive in androgen deprived conditions. Because (a) men on ADT often develop metabolic syndrome, and (b) men with lifestyle-induced obesity and hyperinsulinemia have worse prognosis and faster disease progression, and because (c) insulin causes steroidogenesis in other cell lines, the hypothesis that insulin may contribute to CaP progression through upregulation of steroidogenesis was explored. Insulin upregulates steroidogenesis enzymes at the mRNA level in three AR positive cell lines, as well as upregulating these enzymes at the protein level in two cell lines. It has also been demonstrated that insulin increases mitochondrial (functional) levels of steroid acute regulatory protein (StAR). Furthermore, insulin causes increased levels of total steroids in and induction of de novo steroid synthesis by insulin has been demonstrated at levels induced sufficient to activate AR. The effect of insulin analogs on CaP steroidogenesis in LNCaP and VCaP cells has also been investigated because epidemiological studies suggest that some of the analogs developed may have more cancer stimulatory effects than normal insulin. In this project, despite the signalling differences between glargine, X10, and insulin, these analogs did not appear to induce steroidogenesis any more potently that normal insulin. The effect of insulin of MCF7breast cancer cells was also investigated with results suggesting that breast cancer cells may be capable of de novo steroidogenesis, and that increase in estradiol production may be exacerbated by insulin. Insulin has also been long known to stimulate lipogenesis in the liver and adipocytes, and has been demonstrated to increase lipogenesis in breast cancer cells; therefore, investigation of the effect of insulin on lipogenesis, which is a hallmark of aggressive cancers, was investigated. In CaP progression sterol regulatory element binding protein (SREBP) is dysregulated and upregulates fatty acid synthase (FASN), acetyl CoA-carboxylase, and other lipogenesis genes. SREBP is important for steroidogenesis and in this project has been shown to be upregulated by insulin in CaP cells. Fatty acid synthesis provides building blocks of membrane growth, provides substrates for acid oxidation, the main energy source for CaP cells, provides building blocks for anti-apoptotic and proinflammatory molecules, and provides molecules that stimulate steroidogenesis. In this project it has been shown that insulin upregulates FASN and ACC, which synthesize fatty acids, as well as upregulating hormone sensitive lipase (HSL), diazepam-binding inhibitor (DBI), and long-chain acyl-CoA synthetase 3 (ACSL3), which contribute to lipid activation of steroidogenesis. Insulin also upregulates total lipid levels and de novo lipogenesis, which can be suppressed by inhibition of the insulin receptor (INSR). The fatty acids synthesized after insulin treatment are those that have been associated with CaP; furthermore, microarray data suggests insulin may upregulate fatty acid biosynthesis, metabolism and arachidonic acid metabolism pathways, which have been implicated in CaP growth and survival. Pharmacological agents used to treat patients with hyperinsulinemia/ hyperlipidemia have gained much interest in regards to CaP risk and treatment; however, the scientific rationale behind these clinical applications has not been examined. This thesis explores whether the use of metformin or simvastatin would decrease either lipogenesis or steroidogenesis or both in CaP cells. Simvastatin is a 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) inhibitor, which blocks synthesis of cholesterol, the building block of steroids/ androgens. It has also been postulated to down regulate SREBP in other metabolic disorders. It has been shown in this thesis, in LNCaP cells, that simvastatin inhibited and decreased insulin induced steroidogenesis and lipogenesis, respectively, but increased these pathways in the absence of insulin. Conversely, metformin, which activates AMP-activated protein kinase (AMPK) to shut down lipogenesis, cholesterol synthesis, and protein synthesis, highly suppresses both steroidogenesis and lipogenesis in the presence and absence of insulin. Lastly, because it has been demonstrated to increase steroidogenesis in other cell lines, and because the elucidation of any factors affecting steroidogenesis is important to understanding CaP, the effect of IGF2 on steroidogenesis in CaP cells was investigated. In patient samples, as men progress to CRPC, IGF2 mRNA and the protein levels of the receptors it may signal through are upregulated. It has also been demonstrated that IGF2 upregulates steroidogenic enzymes at both the mRNA and protein levels in LNCaP cells, increases intracellular and secreted steroid/androgen levels in LNCaPs to levels sufficient to stimulate the AR, and upregulated de novo steroidogenesis in LNCaPs and VCaPs. As well, inhibition of INSR and insulin-like growth factor 1 receptor (IGF1R), which IGF2 signals through, suggests that induction of steroidogenesis may be occurring predominantly through IGF1R. In summary, this project has illuminated for the first time that insulin is likely to play a large role in cancer progression, through upregulation of the steroidogenesis and lipogenesis pathways at the mRNA and protein levels, and production levels, and demonstrates a novel role for IGF-II in CaP progression through stimulation of steroidogenesis. It has also been demonstrated that metformin and simvastatin drugs may be useful in suppressing the insulin induction of these pathways. This project affirms the pathways by which ADT- induced metabolic syndrome may exacerbate CaP progression and strongly suggests that the monitoring and modulation of the metabolic state of CaP patients could have a strong impact on their therapeutic outcomes.

Mapping Services to Support a Patient's Journey through Evidence-Based Care Pathways After a Cardiac Event

Background: There are inequalities in geographical access and delivery of health care services in Australia, particularly for cardiovascular disease (CVD), Australia's major cause of death. Analyses and models that can inform and positively influence strategies to augment services and preventative measures are needed. The Cardiac-ARIA project is using geographical spatial technology (GIS) to develop a national index for each of Australia's 13,000 population centres. The index will describe the spatial distribution of CVD health care services available to support populations at risk, in a timely manner, after a major cardiac event. Methods: In the initial phase of the project, an expert panel of cardiologists and an emergency physician have identified key elements of national and international guidelines for management of acute coronary syndromes, cardiac arrest, life-threatening arrhythmias and acute heart failure, from the time of onset (potentially dial 000) to return from the hospital to the community (cardiac rehabilitation). Results: A systematic search has been undertaken to identify the geographical location of, and type of, cardiac services currently available. This has enabled derivation of a master dataset of necessary services, e.g. telephone networks, ambulance, RFDS, helicopter retrieval services, road networks, hospitals, general practitioners, medical community centres, pathology services, CCUs, catheterisation laboratories, cardio-thoracic surgery units and cardiac rehabilitation services. Conclusion: This unique and innovative project has the potential to deliver a powerful tool to both highlight and combat the burden of disease of CVD in urban and regional Australia.

Book Review: "Mark Considine, Jenny Lewis and Damon Alexander Networks, Innovation and Public Policy : Politicians, Bureaucrats and the Pathways to Change inside Government Houndsmills, UK: Palgrave Macmillan, 2009"

Having wrung the most from workforce and workplace productivity initiaitves, innovation has come to the fore as a key goal and directive for public sector organisations to become more efficient. This clarion call for innovation can be heard all around the world, with public services everywhere taking up the message to develop better, smarter, novel, more innovative processes, programs and policies. In the current push for innovation, networks are considered to be a superior vehicle through which collective knowledge can be shared and leveraged; replacing or at least supplementing the role function previously provided by inventive leaders...

Pathways for developing a 'smart' workforce : does the achievement of formal workplace qualifications encourage participation in higher education?

The purpose of this paper is to investigate a public workforce education initiative in the context of State and agency policies designed to enhance employee capabilities to adapt to a volatile and changing environment. In particular, we are concerned with public employees’ experience of a higher educational pathway that resulted in their obtaining a Diploma level qualification. In addition to understanding the employees’ experience of this pathway we were interested in whether the experience contributed to their openness to the prospect of university level education. We conducted telephone interviews with a sample of participants from the program. Employees reported very positive experience of the program; in particular employees reported enhanced efficacy beliefs, a strong sense of achievement, and a feeling of recognition. This experience is explained by four main factors; (1) a program design that was well aligned with the employees learning needs, (2) strong support by organisational staff who delivered and assessed participants on capability criteria, (3) strong management support for employees’ participation, (4) an academic ceremony that provided participants with public recognition of their achievement by valued others. Participants’ motivation to participate was primarily intrinsic rather than extrinsic. Participants in the study reported that their experience in this educational pathway gave them the confidence to consider the possibility of university level education. The paper also discusses the practitioner-academic collaboration that led to the development of this paper.

Joined-up practice : five areas of exemplary practice for social workers and educators to re-engage homeless youth

Young people seen as ‘at risk’ are a substantial focus across a wide range of policy and practice fields in national and international contexts. This article addresses two of those fields, youth homelessness and youth failing to obtain a basic education that will give them access to employment and full community participation as active citizens. By comparing solutions to the problems of youth homelessness and youth educationally at risk, the article distils key meta-characteristics useful for both social workers and educators in mutually supporting some of the most at risk young people in our communities today. This is what the authors term ‘a joined-up practice’.

Expression of PTRF in PC3 cells modulates cholesterol dynamics and the actin cytoskeleton impacting secretion pathways

Expression of caveolin-1 is up-regulated in prostate cancer metastasis and is associated with aggressive recurrence of the disease. Intriguingly, caveolin-1 is also secreted from prostate cancer cell lines and has been identified in secreted prostasomes. Caveolin-1 is the major structural component of the plasma membrane invaginations called caveolae. Co-expression of the coat protein Polymerase I and transcript release factor (PTRF) is required for caveolae formation. We recently found that expression of caveolin-1 in the aggressive prostate cancer cell line PC-3 is not accompanied by PTRF, leading to noncaveolar caveolin-1 lipid rafts. Moreover, ectopic expression of PTRF in PC-3 cells sequesters caveolin-1 into caveolae. Here we quantitatively analyzed the effect of PTRF expression on the PC-3 proteome using stable isotope labeling by amino acids in culture and subcellular proteomics. We show that PTRF reduced the secretion of a subset of proteins including secreted proteases, cytokines, and growth regulatory proteins, partly via a reduction in prostasome secretion. To determine the cellular mechanism accounting for the observed reduction in secreted proteins we analyzed total membrane and the detergent-resistant membrane fractions. Our data show that PTRF expression selectively impaired the recruitment of actin cytoskeletal proteins to the detergent-resistant membrane, which correlated with altered cholesterol distribution in PC-3 cells expressing PTRF. Consistent with this, modulating cellular cholesterol altered the actin cytoskeleton and protein secretion in PC-3 cells. Intriguingly, several proteins that function in ER to Golgi trafficking were reduced by PTRF expression. Taken together, these results suggest that the noncaveolar caveolin-1 found in prostate cancer cells generates a lipid raft microenvironment that accentuates secretion pathways, possibly at the step of ER sorting/exit. Importantly, these effects could be modulated by PTRF expression.

The effect of silicate ions on proliferation, osteogenic differentiation and cell signalling pathways (WNT and SHH) of bone marrow stromal cells

Silicon (Si) is a trace element, which plays an important role in human bone growth. Si has been incorporated into biomaterials for bone regeneration in order to improve their osteogenic potential, both in vitro and in vivo. Little is known, however, as to how Si ions elicit their biological response on bone-forming cells. The aim of this study was to investigate the effect of Si ions on the proliferation, differentiation, bone-related gene expression and cell signalling pathways of bone marrow stromal cells (BMSCs) by comparing the BMSC responses to different concentrations of NaCl and Na2SiO3, while taking into account and excluding the effect of Na ions. Our study showed that Si ions at a concentration of 0.625 mM significantly enhanced the proliferation, mineralization nodule formation, bone-related gene expression (OCN, OPN and ALP) and bone matrix proteins (ALP and OPN) of BMSCs. Furthermore, Si ions at 0.625 mM could counteract the effect of the WNT inhibitor (W.I.) cardamonin on the osteogenic genes expression, (OPN, OCN and ALP), WNT and SHH signalling pathway-related genes in BMSCs. These results suggest that Si ions by themselves play an important role in regulating the proliferation and osteogenic differentiation of BMSCs, with the involvement of WNT and SHH signalling pathways. Our study provides evidence to explain possible molecular mechanisms whereby Si ions released from Si-containing biomaterials can acquire enhanced bioactivity at desired concentration.

S-allylmercaptocysteine reduces carbon tetrachloride-induced hepatic oxidative stress and necroinflammation via nuclear factor kappa B-dependent pathways in mice.

Purpose To study the protective effects and underlying molecular mechanisms of SAMC on carbon tetrachloride (CCl4)-induced acute hepatotoxicity in the mouse model. Methods Mice were intraperitoneally injected with CCl4 (50 μl/kg; single dose) to induce acute hepatotoxicity with or without a 2-h pre-treatment of SAMC intraperitoneal injection (200 mg/kg; single dose). After 8 h, the blood serum and liver samples of mice were collected and subjected to measurements of histological and molecular parameters of hepatotoxicity. Results SAMC reduced CCl4-triggered cellular necrosis and inflammation in the liver under histological analysis. Since co-treatment of SAMC and CCl4 enhanced the expressions of antioxidant enzymes, reduced the nitric oxide (NO)-dependent oxidative stress, and inhibited lipid peroxidation induced by CCl4. SAMC played an essential antioxidative role during CCl4-induced hepatotoxicity. Administration of SAMC also ameliorated hepatic inflammation induced by CCl4 via inhibiting the activity of NF-κB subunits p50 and p65, thus reducing the expressions of pro-inflammatory cytokines, mediators, and chemokines, as well as promoting pro-regenerative factors at both transcriptional and translational levels. Conclusions Our results indicate that SAMC mitigates cellular damage, oxidative stress, and inflammation in CCl4-induced acute hepatotoxicity mouse model through regulation of NF-κB. Garlic or garlic derivatives may therefore be a potential food supplement in the prevention of liver damage.

Stromal upregulation of lateral epithelial adhesions : gene expression analysis of signalling pathways in prostate epithelium

BACKGROUND: Stromal signalling increases the lateral cell adhesions of prostate epithelial cells grown in 3D culture. The aim of this study was to use microarray analysis to identify significant epithelial signalling pathways and genes in this process. METHODS: Microarray analysis was used to identify genes that were differentially expressed when epithelial cells were grown in 3D Matrigel culture with stromal co-culture compared to without stroma. Two culture models were employed: primary epithelial cells (ten samples) and an epithelial cell line (three experiments). A separate microarray analysis was performed on each model system and then compared to identify tissue-relevant genes in a cell line model. RESULTS: TGF beta signalling was significantly ranked for both model systems and in both models the TGF beta signalling gene SOX4 was significantly down regulated. Analysis of all differentially expressed genes to identify genes that were common to both models found several morphology related gene clusters; actin binding (DIAPH2, FHOD3, ABLIM1, TMOD4, MYH10), GTPase activator activity (BCR, MYH10), cytoskeleton (MAP2, MYH10, TMOD4, FHOD3), protein binding (ITGA6, CD44), proteinaceous extracellular matrix (NID2, CILP2), ion channel/ ion transporter activity (CACNA1C, CACNB2, KCNH2, SLC8A1, SLC39A9) and genes associated with developmental pathways (POFUT1, FZD2, HOXA5, IRX2, FGF11, SOX4, SMARCC1). CONCLUSIONS: In 3D prostate cultures, stromal cells increase lateral epithelial cell adhesions. We show that this morphological effect is associated with gene expression changes to TGF beta signalling, cytoskeleton and anion activity.

Health service pathways for patients with chronic leg ulcers : identifying effective pathways for facilitation of evidence based wound care

Background: Chronic leg ulcers cause long term ill-health for older adults and the condition places a significant burden on health service resources. Although evidence on effective management of the condition is available, a significant evidence-practice gap is known to exist, with many suggested reasons e.g. multiple care providers, costs of care and treatments. This study aimed to identify effective health service pathways of care which facilitated evidence-based management of chronic leg ulcers. Methods: A sample of 70 patients presenting with a lower limb leg or foot ulcer at specialist wound clinics in Queensland, Australia were recruited for an observational study and survey. Retrospective data were collected on demographics, health, medical history, treatments, costs and health service pathways in the previous 12 months. Prospective data were collected on health service pathways, pain, functional ability, quality of life, treatments, wound healing and recurrence outcomes for 24 weeks from admission. Results: Retrospective data indicated that evidence based guidelines were poorly implemented prior to admission to the study, e.g. only 31% of participants with a lower limb ulcer had an ABPI or duplex assessment in the previous 12 months. On average, participants accessed care 2–3 times/week for 17 weeks from multiple health service providers in the twelve months before admission to the study clinics. Following admission to specialist wound clinics, participants accessed care on average once per week for 12 weeks from a smaller range of providers. The median ulcer duration on admission to the study was 22 weeks (range 2–728 weeks). Following admission to wound clinics, implementation of key indicators of evidence based care increased (p<0.001) and Kaplan-Meier survival analysis found the median time to healing was 12 weeks (95% CI 9.3–14.7). Implementation of evidence based care was significantly related to improved healing outcomes (p<0.001). Conclusions: This study highlights the complexities involved in accessing expertise and evidence based wound care for adults with chronic leg or foot ulcers. Results demonstrate that access to wound management expertise can promote streamlined health services and evidence based wound care, leading to efficient use of health resources and improved health.

Social adjustment and symptomatology in two types of homeless adolescents : runaways and throwaways

Previous research on homeless adolescents has largely ignored the distinction between those who have left home on their volition (runaways) and those who have been forced to leave (throwaways). Fifty-two homeless adolescents in Brisbane, Australia, were assessed to compare male and female runaways and throwaways for social adjustment and symptomatology. Differences for social adjustment (antisocial tendencies and aggression) and symptomatology (social isolation and depression) were predicted. Results indicated that male runaways were significantly more hostile than male throwaways (p < 0,001), and significantly more socially isolated than female runaways (p < 0,025). Female throwaways, however, were significantly more hostile than male throwaways (p < 0,025) and female runaways (p < 0,025). Yet homeless males overall had a significantly stronger urge to act out hostility than homeless females (p < 0,025). In addition, female throwaways were significantly more antisocial than male throwaways (p < 0,001). There were no significant differences for depression. A theory of inner social control (Hirschi, 1969), postulating absence of bonding in earlier socialization, was supported.

Empower me : social innovation design for homeless families : collective design creativity

Design Creativity has largely been explored as an individual expression of design cognition rather than as the collective manifestation of interaction in context. Recent approaches to design with an emphasis on co-design suggest that the problem-solution space co-evolves through social interaction. Socially Responsive Design for Social Innovation constitutes the most recent and perhaps the most promising domain of application for design thinking practices that emphasize collaborative innovation. In this paper, we describe the ideation of a service design solution for homeless families (Em.power.me), developed through consultation with a range of stakeholders over a three month period. This service design innovation aimed to visualise how such a service would operate and identify the potential benefits for all stakeholders. We focus here on the phases leading to the ideation of the service design.