The function and mechanisms of action of ghrelin and obestatin in ovarian cancer

In this study, we have demonstrated that the preproghrelin derived hormones, ghrelin and obestatin, may play a role in ovarian cancer. Ghrelin and obestatin stimulated an increase in cell migration in ovarian cancer cell lines and may play a role in cancer progression. Ovarian cancer is the leading cause of death among gynaecological cancers and is the sixth most common cause of cancer-related deaths in women in developed countries. As ovarian cancer is difficult to diagnose at a low tumour grade, two thirds of ovarian cancers are not diagnosed until the late stages of cancer development resulting in a poor prognosis for the patient. As a result, current treatment methods are limited and not ideal. There is an urgent need for improved diagnostic markers, as well better therapeutic approaches and adjunctive therapies for this disease. Ghrelin has a number of important physiological effects, including roles in appetite regulation and the stimulation of growth hormone release. It is also involved in regulating the immune, cardiovascular and reproductive systems and regulates sleep, memory and anxiety, and energy metabolism. Over the last decade, the ghrelin axis, (which includes the hormones ghrelin and obestatin and their receptors), has been implicated in the pathogenesis of many human diseases and it may t may also play an important role in the development of cancer. Ghrelin is a 28 amino acid peptide hormone that exists in two forms. Acyl ghrelin (usually referred to as ghrelin), has a unique n-octanoic acid post-translational modification (which is catalysed by ghrelin O-acyltransferase, GOAT), and desacyl ghrelin, which is a non-octanoylated form. Octanoylated ghrelin acts through the growth hormone secretagogue receptor type 1a (GHSR1a). GHSR1b, an alternatively spliced isoform of GHSR, is C-terminally truncated and does not bind ghrelin. Ghrelin has been implicated in the pathophysiology of a number of diseases Obestatin is a 23 amino acid, C-terminally amidated peptide which is derived from preproghrelin. Although GPR39 was originally thought to be the obestatin receptor this has been disproven, and its receptor remains unknown. Obestatin may have as diverse range of roles as ghrelin. Obestatin improves memory, inhibits thirst and anxiety, increases pancreatic juice secretion and has cardioprotective effects. Obestatin also has been shown to regulate cell proliferation, differentiation and apoptosis in some cell types. Prior to this study, little was known regarding the functions and mechanisms of action ghrelin and obestatin in ovarian cancer. In this study it was demonstrated that the full length ghrelin, GHSR1b and GOAT mRNA transcripts were expressed in all of the ovarian-derived cell lines examined (SKOV3, OV-MZ-6 and hOSE 17.1), however, these cell lines did not express GHSR1a. Ovarian cancer tissue of varying stages and normal ovarian tissue expressed the coding region for ghrelin, obestatin, and GOAT, but not GHSR1a, or GHSR1b. No correlations between cancer grade and the level of expression of these transcripts were observed. This study demonstrated for the first time that both ghrelin and obestatin increase cell migration in ovarian cancer cell lines. Treatment with ghrelin (for 72 hours) significantly increased cell migration in the SKOV3 and OV-MZ-6 ovarian cancer cell lines. Ghrelin (100 nM) stimulated cell migration in the SKOV3 (2.64 +/- 1.08 fold, p <0.05) and OV-MZ-6 (1.65 +/- 0.31 fold, p <0.05) ovarian cancer cell lines, but not in the representative normal cell line hOSE 17.1. This increase in migration was not accompanied by an increase in cell invasion through Matrigel. In contrast to other cancer types, ghrelin had no effect on proliferation. Ghrelin treatment (10nM) significantly decreased attachment of the SKOV3 ovarian cancer cell line to collagen IV (24.7 +/- 10.0 %, p <0.05), however, there were no changes in attachment to the other extracellular matrix molecules (ECM) tested (fibronectin, vitronectin and collagen I), and there were no changes in attachment to any of the ECM molecules in the OV-MZ-6 or hOSE 17.1 cell lines. It is, therefore, unclear if ghrelin plays a role in cell attachment in ovarian cancer. As ghrelin has previously been demonstrated to signal through the ERK1/2 pathway in cancer, we investigated ERK1/2 signalling in ovarian cancer cell lines. In the SKOV3 ovarian cancer cell line, a reduction in ERK1/2 phosphorylation (0.58 fold +/- 0.23, p <0.05) in response to 100 nM ghrelin treatment was observed, while no significant change in ERK1/2 signalling was seen in the OV-MZ-6 cell line with treatment. This suggests that this pathway is unlikely to be involved in mediating the increased migration seen in the ovarian cancer cell lines with ghrelin treatment. In this study ovarian cancer tissue of varying stages and normal ovarian tissue expressed the coding region for obestatin, however, no correlation between cancer grade and level of obestatin transcript expression was observed. In the ovarian-derived cell lines studied (SKOV3, OV-MZ-6 and hOSE 17.1) it was demonstrated that the full length preproghrelin mRNA transcripts were expressed in all cell lines, suggesting they have the ability to produce mature obestatin. This is the first study to demonstrate that obestatin stimulates cell migration and cell invasion. Obestatin induced a significant increase in migration in the SKOV3 ovarian cancer cell line with 10 nM (2.80 +/- 0.52 fold, p <0.05) and 100 nM treatments (3.12 +/- 0.68 fold, p <0.05) and in the OV-MZ-6 cancer cell line with 10 nM (2.04 +/- 0.10 fold, p <0.01) and 100 nM treatments (2.00 +/- 0.37 fold, p <0.05). Obestatin treatment did no affect cell migration in the hOSE 17.1normal ovarian epithelial cell line. Obestatin treatment (100 nM) also stimulated a significant increase in cell invasion in the OV-MZ-6 ovarian cancer cell line (1.45 fold +/- 0.13, p <0.05) and in the hOSE17.1 normal ovarian cell line cells (1.40 fold +/- 0.04 and 1.55 fold +/- 0.05 respectively, p <0.01) with 10 nM and 100 nM treatments. Obestatin treatment did not stimulate cell invasion in the SKOV3 ovarian cancer cell line. This lack of obestatin-stimulated invasion in the SKOV3 cell line may be a cell line specific result. In this study, obestatin did not stimulate cell proliferation in the ovarian cell lines and it has previously been shown to have no effect on cell proliferation in the BON-1 pancreatic neuroendocrine and GC rat somatotroph tumour cell lines. In contrast, obestatin has been shown to affect cell proliferation in gastric and thyroid cancer cell lines, and in some normal cell lines. Obestatin also had no effect on attachment of any of the cell lines to any of the ECM components tested (fibronectin, vitronectin, collagen I and collagen IV). The mechanism of action of obestatin was investigated further using a two dimensional-difference in gel electrophoresis (2D-DIGE) proteomic approach. After treatment with obestating (0, 10 and 100 nM), SKOV3 ovarian cancer and hOSE 17.1 normal ovarian cell lines were collected and 2D-DIGE analysis and mass spectrometry were performed to identify proteins that were differentially expressed in response to treatment. Twenty-six differentially expressed proteins were identified and analysed using Ingenuity Pathway Analysis (IPA). This linked 16 of these proteins in a network. The analysis suggested that the ERK1/2 MAPK pathway was a major mediator of obestatin action. ERK1/2 has previously been shown to be associated with obestatin-stimulated cell proliferation and with the anti-apoptotic effects of obestatin. Activation of the ERK1/2 signalling pathway by obestatin was, therefore, investigated in the SKOV3 and OV-MZ-6 ovarian cancer cell lines using anti-active antibodies and Western immunoblots. Obestatin treatment significantly decreased ERK1/2 phosphorylation at higher obestatin concentrations in both the SKOV3 (100 nM and 1000 nM) and OV-MZ-6 (1000 nM) cell lines compared to the untreated controls. Currently, very little is known about obestatin signalling in cancer. This thesis has demonstrated for the first time that the ghrelin axis may play a role in ovarian cancer migration. Ghrelin and obestatin increased cell migration in ovarian cancer cell lines, indicating that they may be a useful target for therapies that reduce ovarian cancer progression. Further studies investigating the role of the ghrelin axis using in vivo ovarian cancer metastasis models are warranted.

Clinical dyslipidaemia is associated with changes in the lipid composition and inflammatory properties of apolipoprotein-B-containing lipoproteins from women with type 2 diabetes

The aim of this study was to use lipidomics to determine if the lipid composition of apolipoprotein-B-containing lipoproteins is modified by dyslipidaemia in type 2 diabetes and if any of the identified changes potentially have biological relevance in the pathophysiology of type 2 diabetes. VLDL and LDL from normolipidaemic and dyslipidaemic type 2 diabetic women and controls were isolated and quantified with HPLC and mass spectrometry. A detailed molecular characterisation of VLDL triacylglycerols (TAG) was also performed using the novel ozone-induced dissociation method, which allowed us to distinguish vaccenic acid (C18:1 n-7) from oleic acid (C18:1 n-9) in specific TAG species. Lipid class composition was very similar in VLDL and LDL from normolipidaemic type 2 diabetic and control participants. By contrast, dyslipidaemia was associated with significant changes in both lipid classes (e.g. increased diacylglycerols) and lipid species (e.g. increased C16:1 and C20:3 in phosphatidylcholine and cholesteryl ester and increased C16:0 [palmitic acid] and vaccenic acid in TAG). Levels of palmitic acid in VLDL and LDL TAG correlated with insulin resistance, and VLDL TAG enriched in palmitic acid promoted increased secretion of proinflammatory mediators from human smooth muscle cells. We showed that dyslipidaemia is associated with major changes in both lipid class and lipid species composition in VLDL and LDL from women with type 2 diabetes. In addition, we identified specific molecular lipid species that both correlate with clinical variables and are proinflammatory. Our study thus shows the potential of advanced lipidomic methods to further understand the pathophysiology of type 2 diabetes.

The OnTrack Diabetes web-based program for type 2 diabetes and dysphoria self-management: A randomized controlled trial protocol

Background: The prevalence of type 2 diabetes is rising with the majority of patients practicing inadequate disease self-management. Depression, anxiety, and diabetes-specific distress present motivational challenges to adequate self-care. Health systems globally struggle to deliver routine services that are accessible to the entire population, in particular in rural areas. Web-based diabetes self-management interventions can provide frequent, accessible support regardless of time and location Objective: This paper describes the protocol of an Australian national randomized controlled trial (RCT) of the OnTrack Diabetes program, an automated, interactive, self-guided Web program aimed to improve glycemic control, diabetes self-care, and dysphoria symptoms in type 2 diabetes patients. Methods: A small pilot trial is conducted that primarily tests program functionality, efficacy, and user acceptability and satisfaction. This is followed by the main RCT, which compares 3 treatments: (1) delayed program access: usual diabetes care for 3 months postbaseline followed by access to the full OnTrack Diabetes program; (2) immediate program: full access to the self-guided program from baseline onward; and (3) immediate program plus therapist support via Functional Imagery Training (FIT). Measures are administered at baseline and at 3, 6, and 12 months postbaseline. Primary outcomes are diabetes self-care behaviors (physical activity participation, diet, medication adherence, and blood glucose monitoring), glycated hemoglobin A1c (HbA1c) level, and diabetes-specific distress. Secondary outcomes are depression, anxiety, self-efficacy and adherence, and quality of life. Exposure data in terms of program uptake, use, time on each page, and program completion, as well as implementation feasibility will be conducted. Results: This trial is currently underway with funding support from the Wesley Research Institute in Brisbane, Australia. Conclusions: This is the first known trial of an automated, self-guided, Web-based support program that uses a holistic approach in targeting both type 2 diabetes self-management and dysphoria. Findings will inform the feasibility of implementing such a program on an ongoing basis, including in rural and regional locations.

Medical Data Access Accountability in EHR Systems, A Practical Perspective

The world has experienced a large increase in the amount of available data. Therefore, it requires better and more specialized tools for data storage and retrieval and information privacy. Recently Electronic Health Record (EHR) Systems have emerged to fulfill this need in health systems. They play an important role in medicine by granting access to information that can be used in medical diagnosis. Traditional systems have a focus on the storage and retrieval of this information, usually leaving issues related to privacy in the background. Doctors and patients may have different objectives when using an EHR system: patients try to restrict sensible information in their medical records to avoid misuse information while doctors want to see as much information as possible to ensure a correct diagnosis. One solution to this dilemma is the Accountable e-Health model, an access protocol model based in the Information Accountability Protocol. In this model patients are warned when doctors access their restricted data. They also enable a non-restrictive access for authenticated doctors. In this work we use FluxMED, an EHR system, and augment it with aspects of the Information Accountability Protocol to address these issues. The Implementation of the Information Accountability Framework (IAF) in FluxMED provides ways for both patients and physicians to have their privacy and access needs achieved. Issues related to storage and data security are secured by FluxMED, which contains mechanisms to ensure security and data integrity. The effort required to develop a platform for the management of medical information is mitigated by the FluxMED's workflow-based architecture: the system is flexible enough to allow the type and amount of information being altered without the need to change in your source code.

Identifying the knowledge management structures in enterprise systems projects

Enterprise System (ES) implementation and management are knowledge intensive tasks that inevitably draw upon the experience of a wide range of people with diverse knowledge capabilities. Knowledge Management (KM) has been identified as a critical success factor in ES projects. Despite the recognized importance of managing knowledge for ES benefits realization, systematic attempts to conceptualize KM-structures have been few. Where the adequacy of KM-structures is assessed, the process and measures are typically idiosyncratic and lack credibility. Using the ‘KM-process’, itself based in sociology of knowledge, this paper conceptualizes four main constructs to measure the adequacy of KM-structures. The SEM model is tested using 310 responses gathered from 27 ES installations that had implemented SAP R/3. The findings reveal six constructs for KM-structure. Furthermore, the paper demonstrates the application of KM-structures in the context of ES using the Adaptive Structuration Theory. The results demonstrate that having adequate KM-structures in place, while necessary, is not sufficient. These rules and resources must be appropriated to have greater positive influence on the Enterprise System. Furthermore, the study provides empirical support for knowledge-based theory by illustrating the importance of knowledge use/re-use (vs. knowledge creation) as the most important driver in the process of KM.

Open Source Enterprise Systems: Towards a Viable Alternative

Enterprise systems are located within the antinomy of appearing as generic product, while being means of multiple integrations for the user through configuration and customisation. Technological and organisational integrations are defined by architectures and standardised interfaces. Until recently, technological integration of enterprise systems has been supported largely by monolithic architectures that were designed, and maintained by the respective developers. From a technical perspective, this approach had been challenged by the suggestion of component-based enterprise systems that would allow for a more user-focused system through strict modularisation. Lately, the product nature of software as proprietary item has been questioned through the rapid increase of open source programs that are being used in business computing in general, and also within the overall portfolio that makes up enterprise systems. This suggests the potential for altered technological and commercial constellations for the design of enterprise systems, which are presented in different scenarios. The technological and commercial decomposition of enterprise software and systems may also address some concerns emerging from the users’ experience of those systems, and which may have arisen from their proprietary or product nature.