Hospital to home outreach for malnourished elders (HHOME) : a feasibility pilot

Objectives: Malnutrition is common in older hospitalised patients, and barriers to adequate intake in hospital limit the effectiveness of hospital-based nutrition interventions. This pilot study was undertaken to determine whether nutrition-focussed care at discharge and in the early post-hospital period is feasible and acceptable to patients and carers, and improves nutritional status. Design: Prospective cohort study Setting: Internal medicine wards of a tertiary teaching hospital in Brisbane, Australia Participants: Patients aged 65 and older admitted for at least 3 days, identified as malnourished or at risk of malnutrition using Mini Nutritional Assessment (MNA). Interventions: An interdisciplinary discharge team (specialist discharge planning nurse and accredited practicing dietitian) provided nutrition-focussed education, advice, service coordination and follow-up (home visits and telephone) for 6 weeks following hospitalisation Measurements: Nutritional intake, weight, functional status and MNA were recorded 6 and 12 weeks after discharge. Service intensity and changes to care were noted, and hospital readmissions recorded. Service feedback from patients and carers was sought using a brief questionnaire. Results: 12 participants were enrolled during the 6 week pilot (mean age 82 years, 50% male). All received 1-2 home visits, and 3-8 telephone calls. Four participants had new community services arranged, 4 were commenced on oral nutritional supplements, and 7 were referred to community dietetics services for follow-up. Two participants had a decline in MNA score of more than 10% at 12 week follow-up, while the remainder improved by at least 10%. Individualised care including community service coordination was valued by participants. Conclusion: The proposed model of care for older adults was feasible, acceptable to patients and carers, and associated with improved nutritional status at 12 weeks for most participants. The pilot data will be useful for design of intervention trials.

IS service quality as a multi-dimensional formative constuct

This study is motivated by, and proceeds from, a central interest in the importance of evaluating IS service quality and adopts the IS ZOT SERVQUAL instrument (Kettinger & Lee, 2005) as its core theory base. This study conceptualises IS service quality as a multidimensional formative construct and seeks to answer the main research questions: “Is the IS service quality construct valid as a 1st-order formative, 2nd-order formative multidimensional construct?” Additionally, with the aim of validating the IS service quality construct within its nomological net, as in prior service marketing work, Satisfaction was hypothesised as its immediate consequence. With the goal of testing the above research question, IS service quality and Satisfaction were operationalised in a quantitative survey instrument. Partial least squares (PLS), employing 219 valid responses, largely evidenced the validity of IS service quality as a multidimensional formative construct. The nomological validity of the IS service quality construct was also evidenced by demonstrating that 55% of Satisfaction was explained by the multidimensional formative IS service quality construct.

Properties of the bovine viral diarrhoea virus replicase in extracts of infected MDBK cells

An assay for the bovine viral diarrhoea virus (BVDV) replicase was developed using extracts from BVDV-infected cells. The replicase activity was maximal approximately 8 h post-infection as measured by the generation of a genomic length radiolabelled RNA. Using a semi-denaturing gel system, three virus-specific in vitro radiolabelled nascent RNA species were identified. A fast-migrating RNA was demonstrated to be the double-stranded replicative form (RF). A second form was shown to be a partially single-stranded/partially doublestranded RNA, characteristic of the replicative intermediate (RI). A third form, which was often undetectable, migrated between the RF and RI and was probably genomic viral RNA. The optimal replicase activity was dependent on 5–10mM Mg2+ and although it was also active in 1–2mM Mn2+ it was inhibited at higher concentrations. The optimum KCl concentration for labelling of the RI and RF were different, suggestive of at least two distinct replicase activities. These results are supportive of a semi-conservative model of BVDV RNA replication.

Evaluation of the iPhone with an acrylic sleeve versus the Scoliometer for rib hump measurement in scoliosis

Background. Vertebral rotation found in structural scoliosis contributes to trunkal asymmetry which is commonly measured with a simple Scoliometer device on a patient's thorax in the forward flexed position. The new generation of mobile 'smartphones' have an integrated accelerometer, making accurate angle measurement possible, which provides a potentially useful clinical tool for assessing rib hump deformity. This study aimed to compare rib hump angle measurements performed using a Smartphone and traditional Scoliometer on a set of plaster torsos representing the range of torsional deformities seen in clinical practice. Methods. Nine observers measured the rib hump found on eight plaster torsos moulded from scoliosis patients with both a Scoliometer and an Apple iPhone on separate occasions. Each observer repeated the measurements at least a week after the original measurements, and were blinded to previous results. Intra-observer reliability and inter-observer reliability were analysed using the method of Bland and Altman and 95% confidence intervals were calculated. The Intra-Class Correlation Coefficients (ICC) were calculated for repeated measurements of each of the eight plaster torso moulds by the nine observers. Results. Mean absolute difference between pairs of iPhone/Scoliometer measurements was 2.1 degrees, with a small (1 degrees) bias toward higher rib hump angles with the iPhone. 95% confidence intervals for intra-observer variability were +/- 1.8 degrees (Scoliometer) and +/- 3.2 degrees (iPhone). 95% confidence intervals for inter-observer variability were +/- 4.9 degrees (iPhone) and +/- 3.8 degrees (Scoliometer). The measurement errors and confidence intervals found were similar to or better than the range of previously published thoracic rib hump measurement studies. Conclusions. The iPhone is a clinically equivalent rib hump measurement tool to the Scoliometer in spinal deformity patients. The novel use of plaster torsos as rib hump models avoids the variables of patient fatigue and discomfort, inconsistent positioning and deformity progression using human subjects in a single or multiple measurement sessions.

Hydrogen-bonding in cyclic imides and amide carboxylic acid derivatives from the facile reaction of cis-cyclohexane-1,2-carboxylic anhydride with o- and p-anisidine and m- and p-aminobenzoic acids

The structures of the open chain amide carboxylic acid rac-cis-[2-(2-methoxyphenyl)carbamoyl]cyclohexane-1-carboxylic acid, C15H19NO4, (I) and the cyclic imides rac-cis-2-(4-methoxyphenyl)-3a,4,5,6,7,7-hexahydroisoindole-1,3-dione,C15H17NO3, (II), chiral cis-2-(3-carboxyphenyl)-3a,4,5,6,7,7a-hexahydroisoindole-1,3-dione, C15H15NO4,(III) and rac-cis-2-(4-carboxyphenyl)- 3a,4,5,6,7,7a-hexahydroisoindole-1,3-dione monohydrate, C15H15NO4. H2O) (IV), are reported. In the amide acid (I), the phenylcarbamoyl group is essentially planar [maximum deviation from the least-squares plane = 0.060(1)Ang. for the amide O atom], the molecules form discrete centrosymmetric dimers through intermolecular cyclic carboxy-carboxy O-H...O hydrogen-bonding interactions [graph set notation R2/2(8)]. The cyclic imides (II)--(IV) are conformationally similar, with comparable phenyl ring rotations about the imide N-C(aromatic) bond [dihedral angles between the benzene and isoindole rings = 51.55(7)deg. in (II), 59.22(12)deg. in (III) and 51.99(14)deg. in (IV). Unlike (II) in which only weak intermolecular C-H...O(imide) hydrogen bonding is present, the crystal packing of imides (III) and (IV) shows strong intermolecular carboxylic acid O-H...O hydrogen-bonding associations. With (III), these involve imide O-atom acceptors, giving one-dimensional zigzag chains [graph set C(9)], while with the monohydrate (IV), the hydrogen bond involves the partially disordered water molecule which also bridges molecules through both imide and carboxyl O-atom acceptors in a cyclic R4/4(12) association, giving a two-dimensional sheet structure. The structures reported here expand the structural data base for compounds of this series formed from the facile reaction of cis-cyclohexane-1,2-dicarboxylic anhydride with substituted anilines, in which there is a much larger incidence of cyclic imides compared to amide carboxylic acids.

Malnutrition screening tools : comparison against two validated nutrition assessment methods in older medical inpatients

Objective Although several validated nutritional screening tools have been developed to “triage” inpatients for malnutrition diagnosis and intervention, there continues to be debate in the literature as to which tool/tools clinicians should use in practice. This study compared the accuracy of seven validated screening tools in older medical inpatients against two validated nutritional assessment methods. Methods This was a prospective cohort study of medical inpatients at least 65 y old. Malnutrition screening was conducted using seven tools recommended in evidence-based guidelines. Nutritional status was assessed by an accredited practicing dietitian using the Subjective Global Assessment (SGA) and the Mini-Nutritional Assessment (MNA). Energy intake was observed on a single day during first week of hospitalization. Results In this sample of 134 participants (80 ± 8 y old, 50% women), there was fair agreement between the SGA and MNA (κ = 0.53), with MNA identifying more “at-risk” patients and the SGA better identifying existing malnutrition. Most tools were accurate in identifying patients with malnutrition as determined by the SGA, in particular the Malnutrition Screening Tool and the Nutritional Risk Screening 2002. The MNA Short Form was most accurate at identifying nutritional risk according to the MNA. No tool accurately predicted patients with inadequate energy intake in the hospital. Conclusion Because all tools generally performed well, clinicians should consider choosing a screening tool that best aligns with their chosen nutritional assessment and is easiest to implement in practice. This study confirmed the importance of rescreening and monitoring food intake to allow the early identification and prevention of nutritional decline in patients with a poor intake during hospitalization.

Eruption of Soufrière Hills (1995–2009) from an offshore perspective : insights from repeated swath bathymetry surveys

This contribution provides an analysis of the 1995–2009 eruptive period of Soufrière Hills volcano (Montserrat) from a unique offshore perspective. The methodology is based on five repeated swath bathymetric surveys. The difference between the 2009 and 1999 bathymetry suggests that at least 395 Mm3 of material has entered the sea. This proximal deposit reaches 95 m thick and extends ∼7km from shore. However, the difference map does not include either the finer distal part of the submarine deposit or the submarine part of the delta close to the shoreline. We took both contributions into account by using additional information such as that from marine sediment cores. By March 2009, at least 65% of the material erupted throughout the eruption has been deposited into the sea. This work provides an excellent basis for assessing the future activity of the Soufrière Hills volcano (including potential collapse), and other volcanoes on small islands.

Submarine deposition of volcaniclastic material from the 1995-2005 eruptions of Soufriere Hills volcano, Montserrat

Soufrière Hills volcano, Montserrat, has been erupting since 1995. During the current eruption, a large part of the material produced by the volcano has been transported into the sea, modifying the morphology of the submarine flanks of the volcano. We present a unique set of swath bathymetric data collected offshore from Montserrat in 1999, 2002 and 2005. From 1999 to 2002, pyroclastic flows associated with numerous dome collapses entered the sea to produce 100 Mm3 deposit. From 2002 to 2005, the 290 Mm3 submarine deposit is mainly from the 12–13 July 2003 collapse. These data allow us to estimate that, by May 2005, at least 482 Mm3 of material had been deposited on the sea floor since 1995. We compare on-land characteristics and volumes of dome collapse events with the submarine deposits and propose a new analysis of their emplacement on the submarine flanks of the volcano. The deposition mechanism shows a slope dependence, with the maximum thickness of deposit before the break in the slope, probably because of the type of the dense granular flow involved. We conclude that from 1995 to 2005 more than 75% of the erupted volume entered the sea.

Sport Management from a Legal Perspective

The purpose of this paper is to investigate the essential elements of sport management in Australia in the 1990's. The essential purpose is to view these elements from a legal perspective. In the past 12 months there has been at least three conferences in the sports law area. The majority of this paper has been allocated to the area of legal liability, especially the legal relationships evolving between the player and his co-participant, the player and his club, the player and his coach, and the duties and liabilities of the coach and the club. The area of insurance will also be discussed as it is a vital element in protecting the players, coaches and clubs in the event of any litigation. A well publicised case was that of Rogers v Bugden where the plaintiff Steven Rogers, who was a first grade rugby league football player for Cronulla, suffered a broken jaw and sued his co-participant Mark Bugden and Bugden's employer Canterbury/Bankstown District Rugby League Football Club. It was held that there was a contract of employment and Canterbury/Bankstown was found to be vicariously liable and was ordered to pay Rogers the sum of $68,154.00. The legal actions in tort and negligence are increasing. Sports managers will need to investigate thoroughly the protection available for their clients.

Evaluating the success of large-scale, integrated information systems through the lens of IS-impact and IS-support

This study proceeds from a central interest in the importance of systematically evaluating operational large-scale integrated information systems (IS) in organisations. The study is conducted within the IS-Impact Research Track at Queensland University of Technology (QUT). The goal of the IS-Impact Track is, "to develop the most widely employed model for benchmarking information systems in organizations for the joint benefit of both research and practice" (Gable et al, 2009). The track espouses programmatic research having the principles of incrementalism, tenacity, holism and generalisability through replication and extension research strategies. Track efforts have yielded the bicameral IS-Impact measurement model; the ‘impact’ half includes Organisational-Impact and Individual-Impact dimensions; the ‘quality’ half includes System-Quality and Information-Quality dimensions. Akin to Gregor’s (2006) analytic theory, the ISImpact model is conceptualised as a formative, multidimensional index and is defined as "a measure at a point in time, of the stream of net benefits from the IS, to date and anticipated, as perceived by all key-user-groups" (Gable et al., 2008, p: 381). The study adopts the IS-Impact model (Gable, et al., 2008) as its core theory base. Prior work within the IS-Impact track has been consciously constrained to Financial IS for their homogeneity. This study adopts a context-extension strategy (Berthon et al., 2002) with the aim "to further validate and extend the IS-Impact measurement model in a new context - i.e. a different IS - Human Resources (HR)". The overarching research question is: "How can the impacts of large-scale integrated HR applications be effectively and efficiently benchmarked?" This managerial question (Cooper & Emory, 1995) decomposes into two more specific research questions – In the new HR context: (RQ1): "Is the IS-Impact model complete?" (RQ2): "Is the ISImpact model valid as a 1st-order formative, 2nd-order formative multidimensional construct?" The study adhered to the two-phase approach of Gable et al. (2008) to hypothesise and validate a measurement model. The initial ‘exploratory phase’ employed a zero base qualitative approach to re-instantiating the IS-Impact model in the HR context. The subsequent ‘confirmatory phase’ sought to validate the resultant hypothesised measurement model against newly gathered quantitative data. The unit of analysis for the study is the application, ‘ALESCO’, an integrated large-scale HR application implemented at Queensland University of Technology (QUT), a large Australian university (with approximately 40,000 students and 5000 staff). Target respondents of both study phases were ALESCO key-user-groups: strategic users, management users, operational users and technical users, who directly use ALESCO or its outputs. An open-ended, qualitative survey was employed in the exploratory phase, with the objective of exploring the completeness and applicability of the IS-Impact model’s dimensions and measures in the new context, and to conceptualise any resultant model changes to be operationalised in the confirmatory phase. Responses from 134 ALESCO users to the main survey question, "What do you consider have been the impacts of the ALESCO (HR) system in your division/department since its implementation?" were decomposed into 425 ‘impact citations.’ Citation mapping using a deductive (top-down) content analysis approach instantiated all dimensions and measures of the IS-Impact model, evidencing its content validity in the new context. Seeking to probe additional (perhaps negative) impacts; the survey included the additional open question "In your opinion, what can be done better to improve the ALESCO (HR) system?" Responses to this question decomposed into a further 107 citations which in the main did not map to IS-Impact, but rather coalesced around the concept of IS-Support. Deductively drawing from relevant literature, and working inductively from the unmapped citations, the new ‘IS-Support’ construct, including the four formative dimensions (i) training, (ii) documentation, (iii) assistance, and (iv) authorisation (each having reflective measures), was defined as: "a measure at a point in time, of the support, the [HR] information system key-user groups receive to increase their capabilities in utilising the system." Thus, a further goal of the study became validation of the IS-Support construct, suggesting the research question (RQ3): "Is IS-Support valid as a 1st-order reflective, 2nd-order formative multidimensional construct?" With the aim of validating IS-Impact within its nomological net (identification through structural relations), as in prior work, Satisfaction was hypothesised as its immediate consequence. The IS-Support construct having derived from a question intended to probe IS-Impacts, too was hypothesised as antecedent to Satisfaction, thereby suggesting the research question (RQ4): "What is the relative contribution of IS-Impact and IS-Support to Satisfaction?" With the goal of testing the above research questions, IS-Impact, IS-Support and Satisfaction were operationalised in a quantitative survey instrument. Partial least squares (PLS) structural equation modelling employing 221 valid responses largely evidenced the validity of the commencing IS-Impact model in the HR context. ISSupport too was validated as operationalised (including 11 reflective measures of its 4 formative dimensions). IS-Support alone explained 36% of Satisfaction; IS-Impact alone 70%; in combination both explaining 71% with virtually all influence of ISSupport subsumed by IS-Impact. Key study contributions to research include: (1) validation of IS-Impact in the HR context, (2) validation of a newly conceptualised IS-Support construct as important antecedent of Satisfaction, and (3) validation of the redundancy of IS-Support when gauging IS-Impact. The study also makes valuable contributions to practice, the research track and the sponsoring organisation.

DRD2 C957T and TaqIA genotyping reveals gender effects and unique low-risk and high-risk genotypes in alcohol dependence

AIMS: As recent conflicting reports describe a genetic association between both the C- and the T-alleles of the dopamine D2 receptor (DRD2) C957T polymorphism (rs6277) in alcohol-dependent subjects, our aim was to examine this polymorphism and TaqIA (rs1800497) in Australian alcohol-dependent subjects. METHODS: The C957T polymorphism was genotyped in 228 patients with alcohol dependence (72 females and 156 males) and 228 healthy controls. RESULTS: The C-allele and C/C genotype of C957T was associated with alcohol dependence, whereas the TaqIA polymorphism was not. When analysed separately for C957T, males showed an even stronger association with the C-allele and females showed no association. The C957T and TaqIA haplotyping revealed a strong association with alcohol dependence and a double-genotype analysis (combining C957T and TaqIA genotypes) revealed that the relative risk of different genotypes varied by up to 27-fold with the TT/A1A2 having an 8.5-fold lower risk of alcohol dependence than other genotypes. CONCLUSION: Decreased DRD2 binding associated with the C-allele of the DRD2 C957T polymorphism is likely to be important in the underlying pathophysiology of at least some forms of alcohol dependence, and this effect appears to be limited to males only.

How important is length? : mechanical testing and measurement of a cemented, polished, tapered femoral implant

Total hip arthroplasty (THA) has a proven clinical record for providing pain relief and return of function to patients with disabling arthritis. There are many successful options for femoral implant design and fixation. Cemented, polished, tapered femoral implants have been shown to have excellent results in national joint registries and long-term clinical series. These implants are usually 150mm long at their lateral aspect. Due to their length, these implants cannot always be offered to patients due to variations in femoral anatomy. Polished, tapered implants as short as 95mm exist, however their small proximal geometry (neck offset and body size) limit their use to smaller stature patients. There is a group of patients in which a shorter implant with a maintained proximal body size would be advantageous. There are also potential benefits to a shorter implant in standard patient populations such as reduced bone removal due to reduced reaming, favourable loading of the proximal femur, and the ability to revise into good proximal bone stock if required. These factors potentially make a shorter implant an option for all patient populations. The role of implant length in determining the stability of a cemented, polished, tapered femoral implant is not well defined by the literature. Before changes in implant design can be made, a better understanding of the role of each region in determining performance is required. The aim of the thesis was to describe how implant length affects the stability of a cemented, polished, tapered femoral implant. This has been determined through an extensive body of laboratory testing. The major findings are that for a given proximal body size, a reduction in implant length has no effect on the torsional stability of a polished, tapered design, while a small reduction in axial stability should be expected. These findings are important because the literature suggests that torsional stability is the major determinant of long-term clinical performance of a THA system. Furthermore, a polished, tapered design is known to be forgiving of cement-implant interface micromotion due to the favourable wear characteristics. Together these findings suggest that a shorter polished, tapered implant may be well tolerated. The effect of a change in implant length on the geometric characteristics of polished, tapered design were also determined and applied to the mechanical testing. Importantly, interface area does play a role in stability of the system; however it is the distribution of the interface and not the magnitude of the area that defines stability. Taper angle (at least in the range of angles seen in this work) was shown not to be a determinant of axial or torsional stability. A range of implants were tested, comparing variations in length, neck offset and indication (primary versus cement-in-cement revision). At their manufactured length, the 125mm implants were similar to their longer 150mm counterparts suggesting that they may be similarly well tolerated in the clinical environment. However, the slimmer cement-in-cement revision implant was shown to have a poorer mechanical performance, suggesting their use in higher demand patients may be hazardous. An implant length of 125mm has been shown to be quite stable and the results suggest that a further reduction to 100mm may be tolerated. However, further work is required. A shorter implant with maintained proximal body size would be useful for the group of patients who are unable to access the current standard length implants due to variations in femoral anatomy. Extending the findings further, the similar function with potential benefits of a shorter implant make their application to all patients appealing.

Near infrared spectroscopy for non-destructive evaluation of articular cartilage

There is a need for an accurate real-time quantitative system that would enhance decision-making in the treatment of osteoarthritis. To achieve this objective, significant research is required that will enable articular cartilage properties to be measured and categorized for health and functionality without the need for laboratory tests involving biopsies for pathological evaluation. Such a system would provide the capability of access to the internal condition of the cartilage matrix and thus extend the vision-based arthroscopy that is currently used beyond the subjective evaluation of surgeons. The system required must be able to non-destructively probe the entire thickness of the cartilage and its immediate subchondral bone layer. In this thesis, near infrared spectroscopy is investigated for the purpose mentioned above. The aim is to relate it to the structure and load bearing properties of the cartilage matrix to the near infrared absorption spectrum and establish functional relationships that will provide objective, quantitative and repeatable categorization of cartilage condition outside the area of visible degradation in a joint. Based on results from traditional mechanical testing, their innovative interpretation and relationship with spectroscopic data, new parameters were developed. These were then evaluated for their consistency in discriminating between healthy viable and degraded cartilage. The mechanical and physico-chemical properties were related to specific regions of the near infrared absorption spectrum that were identified as part of the research conducted for this thesis. The relationships between the tissue's near infrared spectral response and the new parameters were modeled using multivariate statistical techniques based on partial least squares regression (PLSR). With significantly high levels of statistical correlation, the modeled relationships were demonstrated to possess considerable potential in predicting the properties of unknown tissue samples in a quick and non-destructive manner. In order to adapt near infrared spectroscopy for clinical applications, a balance between probe diameter and the number of active transmit-receive optic fibres must be optimized. This was achieved in the course of this research, resulting in an optimal probe configuration that could be adapted for joint tissue evaluation. Furthermore, as a proof-of-concept, a protocol for obtaining the new parameters from the near infrared absorption spectra of cartilage was developed and implemented in a graphical user interface (GUI)-based software, and used to assess cartilage-on-bone samples in vitro. This conceptual implementation has been demonstrated, in part by the individual parametric relationship with the near infrared absorption spectrum, the capacity of the proposed system to facilitate real-time, non-destructive evaluation of cartilage matrix integrity. In summary, the potential of the optical near infrared spectroscopy for evaluating articular cartilage and bone laminate has been demonstrated in this thesis. The approach could have a spin-off for other soft tissues and organs of the body. It builds on the earlier work of the group at QUT, enhancing the near infrared component of the ongoing research on developing a tool for cartilage evaluation that goes beyond visual and subjective methods.

The iatrogenic reflex and belief that repression can coercively suppress asymmetric coercive violence : does very robust action by the government decrease functional support for insurgents?

Non-state insurgent actors are too weak to compel powerful adversaries to their will, so they use violence to coerce. A principal objective is to grow and sustain violent resistance to the point that it either militarily challenges the state, or more commonly, generates unacceptable political costs. To survive, insurgents must shift popular support away from the state and to grow they must secure it. State actor policies and actions perceived as illegitimate and oppressive by the insurgent constituency can generate these shifts. A promising insurgent strategy is to attack states in ways that lead angry publics and leaders to discount the historically established risks and take flawed but popular decisions to use repressive measures. Such decisions may be enabled by a visceral belief in the power of coercion and selective use of examples of where robust measures have indeed suppressed resistance. To avoid such counterproductive behaviours the cases of apparent 'successful repression' must be understood. This thesis tests whether robust state action is correlated with reduced support for insurgents, analyses the causal mechanisms of such shifts and examines whether such reduction is because of compulsion or coercion? The approach is founded on prior research by the RAND Corporation which analysed the 30 insurgencies most recently resolved worldwide to determine factors of counterinsurgent success. This new study first re-analyses their data at a finer resolution with new queries that investigate the relationship between repression and insurgent active support. Having determined that, in general, repression does not correlate with decreased insurgent support, this study then analyses two cases in which the data suggests repression seems likely to be reducing insurgent support: the PKK in Turkey and the insurgency against the Vietnamese-sponsored regime after their ousting of the Khmer Rouge. It applies 'structured-focused' case analysis with questions partly built from the insurgency model of Leites and Wolf, who are associated with the advocacy of US robust means in Vietnam. This is thus a test of 'most difficult' cases using a 'least likely' test model. Nevertheless, the findings refute the deterrence argument of 'iron fist' advocates. Robust approaches may physically prevent effective support of insurgents but they do not coercively deter people from being willing to actively support the insurgency.

The roles of the preproghrelin-derived peptides - ghrelin, desacyl ghrelin and obestatin - in prostate cancer

Prostate cancer is the second most common cause of cancer related deaths in Western men. Despite the significant improvements in current treatment techniques, there is no cure for advanced metastatic, castrate-resistant disease. Early detection and prevention of progression to a castrate-resistant state may provide new strategies to improve survival. A number of growth factors have been shown to act in an autocrine/paracrine manner to modulate prostate cancer tumour growth. Our laboratory has previously shown that ghrelin and its receptors (the functional GHS-R1a and the non-functional GHS-R1b) are expressed in prostate cancer specimens and cell lines. We have shown that ghrelin increases cell proliferation in the PC3 and LNCaP prostate cancer cell lines through activation of ERK1/2, suggesting that ghrelin could regulate prostate cancer cell growth and play a role in the progression of the disease. Ghrelin is a 28 amino-acid peptide hormone, identified to be the natural ligand of the growth hormone secretagogue receptor (GHS-R1a). It is well characterised as a growth hormone releasing and as an orexigenic peptide that stimulates appetite and feeding and regulates energy expenditure and bodyweight. In addition to its orexigenic properties, ghrelin has been shown to play a regulatory role in a number of systems, including the reproductive, immune and cardiovascular systems and may play a role in a number of pathological conditions such as chronic heart failure, anorexia, cachexia, obesity, diabetes and cancer. In cancer, ghrelin and its receptor are expressed in a range of tumours and cancer cell lines and ghrelin has been demonstrated to modulate cell proliferation, apoptosis, migration and invasion in some cell types. The ghrelin gene (GHRL) encodes preproghrelin peptide, which is processed to produce three currently known functional peptides - ghrelin, desacyl ghrelin and obestatin. Prohormone convertases (PCs) have been shown to cleave the preproghrelin peptide into two primary products - the 28 amino acid peptide, ghrelin, and the remaining 117 amino acid C-terminal peptide, C-ghrelin. C-ghrelin can then be further processed to produce the 23 amino acid peptide, obestatin. Ghrelin circulates in two different forms - an octanoylated form (known as ghrelin) and a non-octanoylated form, desacyl ghrelin. The unique post-translational addition of octanoic acid to the serine 3 residue of the propeptide chain to form acylated ghrelin is catalysed by ghrelin O-acyltransferase (GOAT). This modification is necessary for binding of ghrelin to its only known functional receptor, the GHS-R1a. As desacyl ghrelin cannot bind and activate the GHS-R1a, it was initially thought to be an inactive peptide, despite the fact that it circulates at much higher levels than ghrelin. Further research has demonstrated that desacyl ghrelin is biologically active and shares some of the actions of ghrelin, as well as having some opposing and distinct roles. Interestingly, both ghrelin and desacyl ghrelin have been shown to modulate apoptosis, cell differentiation and proliferation in some cell types, and to stimulate cell proliferation through activation of ERK1/2 and PI3K/Akt pathways. The third known peptide product of the ghrelin preprohormone, obestatin, was initially thought to oppose the actions of ghrelin in appetite regulation and food intake and to mediate its effects through the G protein-coupled receptor 39 (GPR39). Subsequent research failed to reproduce the initial findings, however, and the possible anorexigenic effects of obestatin, as well as the identity of its receptor, remain unclear. Obestatin plays some important physiological roles, including roles in improving memory, the inhibition of thirst and anxiety, increased secretion of pancreatic juice, and regulation of cell proliferation, survival, apoptosis and differentiation. Preliminary studies have also shown that obestatin stimulates cell proliferation in some cell types through activation of ERK1/2, Akt and PKC pathways. Overall, however, at the commencement of this PhD project, relatively little was known regarding the functions and mechanisms of action of the preproghrelin-derived functional peptides in modulating prostate cancer cell proliferation. The roles of obestatin, and desacyl ghrelin as potential growth factors had not previously been investigated, and the potential expression and regulation of the preproghrelin processing enzymes, GOAT and prohormone convertases was unknown in prostate cancer cell lines. Therefore, the overall objectives of this study were to: 1. investigate the effects of obestatin on cell proliferation and signaling in prostate cancer cell lines 2. compare the effects of desacyl ghrelin and ghrelin on cell proliferation and signaling in prostate cancer cell lines 3. investigate whether prostate cancer cell lines possess the necessary enzymatic machinery to produce ghrelin and desacyl ghrelin and if these peptides can regulate GOAT expression Our laboratory has previously shown that ghrelin stimulates cell proliferation in the PC3 and LNCaP prostate cancer cell line through activation of the ERK1/2 pathway. In this study it has been demonstrated that treatments with either ghrelin, desacyl ghrelin or obestatin over 72 hours significantly increased cell proliferation in the PC3 prostate cancer cell line but had no significant effect in the RWPE-1 transformed normal prostate cell line. Ghrelin (1000nM) stimulated cell proliferation in the PC3 prostate cancer cell line by 31.66 6.68% (p<0.01) with the WST-1 method, and 13.55 5.68% (p<0.05) with the CyQUANT assay. Desacyl ghrelin (1000nM) increased cell proliferation in PC3 cells by 21.73 2.62% (p<0.01) (WST-1), and 15.46 7.05% (p<0.05) (CyQUANT) above untreated control. Obestatin (1000nM) induced a 28.37 7.47% (p<0.01) (WST-1) and 12.14 7.47% (p<0.05) (CyQUANT) significant increase in cell proliferation in the PC3 prostate cancer cell line. Ghrelin and desacyl ghrelin treatments stimulated Akt and ERK phosphorylation across a range of concentrations (p<0.01). Obestatin treatment significantly stimulated Akt, ERK and PKC phosphorylation (p<0.05). Through the use of specific inhibitors, the MAPK inhibitor U0126 and the Akt1/2 kinase inhibitor, it was demonstrated that ghrelin- and obestatin-induced cell proliferation in the PC3 prostate cancer cell line is mediated through activation of ERK1/2 and Akt pathways. Although desacyl ghrelin significantly stimulated Akt and ERK phosphorylation, U0126 failed to prevent desacyl ghrelin-induced cell proliferation suggesting ghrelin and desacyl ghrelin might act through different mechanisms to increase cell proliferation. Ghrelin and desacyl ghrelin have shown a proliferative effect in osteoblasts, pancreatic -cells and cardiomyocytes through activation of ERK1/2 and PI3K/Akt pathways. Here it has been shown that ghrelin and its non-acylated form exert the same function and stimulate cell proliferation in the PC3 prostate cancer cell line through activation of the Akt pathway. Ghrelin-induced proliferation was also mediated through activation of the ERK1/2 pathway, however, desacyl ghrelin seems to stimulate cell proliferation in an ERK1/2-independent manner. As desacyl ghrelin does not bind and activate GHSR1a, the only known functional ghrelin receptor, the finding that both ghrelin and desacyl ghrelin stimulate cell proliferation in the PC3 cell line suggests that these peptides could be acting through the yet unidentified alternative ghrelin receptor in this cell type. Obestatin treatment also stimulated PKC phosphorylation, however, a direct role for this pathway in stimulating cell proliferation could not be proven using available PKC pathway inhibitors, as they caused significant cell death over the extended timeframe of the cell proliferation assays. Obestatin has been shown to stimulate cell proliferation through activation of PKC isoforms in human retinal epithelial cells and in the human gastric cancer cell line KATO-III. We have demonstrated that all of the prostate-derived cell lines examined (PC3, LNCaP, DU145, 22Rv1, RWPE-1 and RWPE-2) expressed GOAT and at least one of the prohormone convertases, which are known to cleave the proghrelin peptide, PC1/3, PC2 and furin, at the mRNA level. These cells, therefore, are likely to possess the necessary machinery to cleave the preproghrelin protein and to produce the mature ghrelin and desacyl ghrelin peptides. In addition to prohormone convertases, the presence of octanoic acid is essential for acylated ghrelin production. In this study octanoic acid supplementation significantly increased cell proliferation in the PC3 prostate cancer cell line by over 20% compared to untreated controls (p<0.01), but surprisingly, not in the DU145, LNCaP or 22Rv1 prostate cancer cell lines or in the RWPE-1 and RWPE-2 prostate-derived cell lines. In addition, we demonstrated that exogenous ghrelin induced a statistically significant two-fold decrease in GOAT mRNA expression in the PC3 cell line (p<0.05), suggesting that ghrelin could pontentially downregulate its own acylation and, therefore, regulate the balance between ghrelin and desacyl ghrelin. This was not observed, however, in the DU145 and LNCaP prostate cancer cell lines. The GOAT-ghrelin system represents a direct link between ingested nutrients and regulation of ghrelin production and the ghrelin/desacyl ghrelin ratio. Regulation of ghrelin acylation is a potentially attractive and desirable tool for the development of better therapies for a number of pathological conditions where ghrelin has been shown to play a key role. The finding that desacyl ghrelin stimulates cell proliferation in the PC3 prostate cancer cell line, and responds to ghrelin in the same way, suggests that this cell line expresses an alternative ghrelin receptor. Although all the cell lines examined expressed both GHS-R1a and GHS-R1b mRNA, it remains uncertain whether these cell lines express the unidentified alternative ghrelin receptor. It is possible that the varied responses seen could be due to the expression of different ghrelin receptors in different cell lines. In addition to GOAT, prohormone convertases and octanoic acid availability may regulate the production of different peptides from the ghrelin preprohormone. The studies presented in this thesis provide significant new information regarding the roles and mechanisms of action of the preproghrelin-derived peptides, ghrelin, desacyl ghrelin and obestatin, in modulating prostate cancer cell line proliferation. A number of key questions remain to be resolved, however, including the identification of the alternative ghrelin/desacyl ghrelin receptor, the identification of the obestatin receptor, a clarification of the signaling mechanisms which mediate cell proliferation in response to obestatin treatment and a better understanding of the regulation at both the gene and post-translational levels of functional peptide generation. Further studies investigating the role of the ghrelin axis using in vivo prostate cancer models may be warranted. Until these issues are determined, the potential for the ghrelin axis, to be recognised as a novel useful target for therapy for cancer or other pathologies will be uncertain.