Grounding Evaluation Capacity Development in Systems Theory

While “systemic thinking” is popular in the context of capacity development and evaluation, there is currently a lack of understanding about the benefits to employing systems theory in evaluation capacity development. Systems theory provides a useful orientation to the work involved in complex systems (e.g. national evaluation systems). This article illustrates how evaluation capacity development practitioners can use systems theory as a conceptual tool to gain a better understanding of the functional aspects and interrelationships present within a given evaluation system. Specifically, the systems theory perspective can help elucidate the reasons for the success or failure of a given evaluation capacity development program or activity. With the goal of motivating evaluation capacity development practitioners to use systems theory in their work, this article presents a systems theory framework for evaluation capacity development and offers practical examples of how it can be adopted

Statin-induced expression of CD59 on vascular endothelium in hypoxia: a potential mechanism for the anti-inflammatory actions of statins in rheumatoid arthritis

Hypoxia, which leads to dysfunctional cell metabolism, and complement activation both play central roles in the pathogenesis of rheumatoid arthritis (RA). Recent studies have reported that mice deficient for the complement-inhibitory protein CD59 show enhanced susceptibility to antigen-induced arthritis and reported that statins have anti-inflammatory effects in RA. We hypothesized that the anti-inflammatory effect of statins in RA relates in part to their ability to increase CD59 expression in hypoxic conditions and therefore to reduce complement activation. Flow-cytometric analysis showed that CD59 expression on endothelial cells (EC) was unaffected by atorvastatin in normoxia (21% O(2)), whereas in hypoxic conditions (1% O(2)) an up to threefold dose-dependent increase in CD59 expression was seen. This effect of hypoxia was confirmed by treatment of EC with chemical mimetics of hypoxia. The upregulation of CD59 protein expression in hypoxia was associated with an increase in steady-state mRNA. L-Mevalonate and geranylgeraniol reversed the response, confirming a role for inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase and geranylgeranylation. Likewise, inhibition by N(G)-monomethyl-L-arginine and N(G)-nitro-L-arginine methyl ester confirmed that CD59 upregulation in hypoxia was nitric oxide dependent. The expression of another complement-inhibitory protein, decay-accelerating factor (DAF), is known to be increased by atorvastatin in normoxia; this response was also significantly enhanced under hypoxic conditions. The upregulation of CD59 and DAF by atorvastatin in hypoxia prevented the deposition of C3, C9 and cell lysis that follows exposure of reoxygenated EC to serum. This cytoprotective effect was abrogated by inhibitory anti-CD59 and anti-DAF mAbs. The modulation of EC CD59 and DAF by statins under hypoxic conditions therefore inhibits both early and late complement activation and may contribute to the anti-inflammatory effects of statins in RA

Inactivation of the von Hippel-Lindau tumour suppressor gene induces Neuromedin U expression in renal cancer cells.

BACKGROUND: 209 000 new cases of renal carcinoma are diagnosed each year worldwide and new therapeutic targets are urgently required. The great majority of clear cell renal cancer involves inactivation of VHL, which acts as a gatekeeper tumour suppressor gene in renal epithelial cells. However how VHL exerts its tumour suppressor function remains unclear. A gene expression microarray comparing RCC10 renal cancer cells expressing either VHL or an empty vector was used to identify novel VHL regulated genes. FINDINGS: NMU (Neuromedin U) is a neuropeptide that has been implicated in energy homeostasis and tumour progression. Here we show for the first time that VHL loss-of-function results in dramatic upregulation of NMU expression in renal cancer cells. The effect of VHL inactivation was found to be mediated via activation of Hypoxia Inducible Factor (HIF). Exposure of VHL expressing RCC cells to either hypoxia or dimethyloxalylglycine resulted in HIF activation and increased NMU expression. Conversely, suppression of HIF in VHL defective RCC cells via siRNA of HIF-α subunits or expression of Type 2C mutant VHLs reduced NMU expression levels. We also show that renal cancer cells express a functional NMU receptor (NMUR1), and that NMU stimulates migration of renal cancer cells. CONCLUSIONS: These findings suggest that NMU may act in an autocrine fashion, promoting progression of kidney cancer. Hypoxia and HIF expression are frequently observed in many non-renal cancers and are associated with a poor prognosis. Our study raises the possibility that HIF may also drive NMU expression in non-renal tumours

HIF-1alpha and HIF-2alpha are differentially activated in distinct cell populations in retinal ischaemia.

BACKGROUND: Hypoxia plays a key role in ischaemic and neovascular disorders of the retina. Cellular responses to oxygen are mediated by hypoxia-inducible transcription factors (HIFs) that are stabilised in hypoxia and induce the expression of a diverse range of genes. The purpose of this study was to define the cellular specificities of HIF-1alpha and HIF-2alpha in retinal ischaemia, and to determine their correlation with the pattern of retinal hypoxia and the expression profiles of induced molecular mediators. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the tissue distribution of retinal hypoxia during oxygen-induced retinopathy (OIR) in mice using the bio-reductive drug pimonidazole. We measured the levels of HIF-1alpha and HIF-2alpha proteins by Western blotting and determined their cellular distribution by immunohistochemistry during the development of OIR. We measured the temporal expression profiles of two downstream mediators, vascular endothelial growth factor (VEGF) and erythropoietin (Epo) by ELISA. Pimonidazole labelling was evident specifically in the inner retina. Labelling peaked at 2 hours after the onset of hypoxia and gradually declined thereafter. Marked binding to Müller glia was evident during the early hypoxic stages of OIR. Both HIF-1alpha and HIF-2alpha protein levels were significantly increased during retinal hypoxia but were evident in distinct cellular distributions; HIF-1alpha stabilisation was evident in neuronal cells throughout the inner retinal layers whereas HIF-2alpha was restricted to Müller glia and astrocytes. Hypoxia and HIF-alpha stabilisation in the retina were closely followed by upregulated expression of the downstream mediators VEGF and EPO. CONCLUSIONS/SIGNIFICANCE: Both HIF-1alpha and HIF-2alpha are activated in close correlation with retinal hypoxia but have contrasting cell specificities, consistent with differential roles in retinal ischaemia. Our findings suggest that HIF-2alpha activation plays a key role in regulating the response of Müller glia to hypoxia

Role of the von Hippel-Lindau tumour suppressor gene in regulating renal epithelial cell characteristics

Inactivation of the von Hippel-Lindau (VHL) tumour suppressor gene underlies both VHL disease, an inherited multi-cancer syndrome, and the majority of sporadic clear cell renal cell carcinomas (CCRCCs). The best established function of pVHL is regulation of HIF (hypoxia inducible factor). However, how VHL functions as a renal tumour suppressor gene remains to be fully elucidated. There is extensive evidence that changes in cell-cell adhesion and loss of epithelial cytoarchitecture are pivotal in the progression of tumours of epithelial origin, allowing cell-cell dissociation and subsequent invasion. The main objective of this thesis has been to identify whether VHL regulates epithelial cell characteristics, and, if so, whether this is mediated via HIF. The major findings are as follows: First, VHL loss-of-function has striking effects on the expression and localisation of several tight junction (TJ) components (occludin, claudin 1 and ZO-l) in both VHL defective CCRCC cells and sporadic CCRCC tissue (compared to adjacent unaffected kidney). Secondly disruption of TJs was detected in the earliest lesions of VHL inactivation in VHL patient kidneys, suggesting a potential role in tumour initiation. Thirdly re-expression the adherens junction (AJ) protein E-cadherin did not rescue TJ formation, showing that the TJ defect occurs independently of AJ breakage. Fourthly, VHL is required for formation of the primary cilium, a luminal hair-like structure which senses renal tubular flow and which is disturbed in most renal cystic diseases. Finally activation of HIF promotes dedifferentiation of renal epithelial cells. Dedifferentiation may allow cells to tolerate further mutations and undergo full malignant transformation. I have also shown that HDAC inhibitors can reverse this phenotype and therefore may be of use therapeutically. In summary, the work presented in this thesis provides significant insights into understanding how the VHL/HIF pathway contributes to tumour development in both VHL defective renal cancers and hypoxic non-renal tumours.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

The metastasis suppressor RARRES3 as an endogenous inhibitor of the immunoproteasome expression in breast cancer cells

In breast cancer metastasis, the dynamic continuum involving pro-and anti-inflammatory regulators can become compromised. Over 600 genes have been implicated in metastasis to bone, lung or brain but how these genes might contribute to perturbation of immune function is poorly understood. To gain insight, we adopted a gene co-expression network approach that draws on the functional parallels between naturally occurring bone marrow-derived mesenchymal stem cells (BM-MSCs) and cancer stem cells (CSCs). Our network analyses indicate a key role for metastasis suppressor RARRES3, including potential to regulate the immunoproteasome (IP), a specialized proteasome induced under inflammatory conditions. Knockdown of RARRES3 in near-normal mammary epithelial and breast cancer cell lines increases overall transcript and protein levels of the IP subunits, but not of their constitutively expressed counterparts. RARRES3 mRNA expression is controlled by interferon regulatory factor IRF1, an inducer of the IP, and is sensitive to depletion of the retinoid-related receptor RORA that regulates various physiological processes including immunity through modulation of gene expression. Collectively, these findings identify a novel regulatory role for RARRES3 as an endogenous inhibitor of IP expression, and contribute to our evolving understanding of potential pathways underlying breast cancer driven immune modulation