Enhancement of DNA repair using topical T4 endonuclease V does not inhibit melanoma formation in Cdk4R24C/R24C/Tyr-NrasQ61K mice following neonatal UVR

To further investigate the use of DNA repair-enhancing agents for skin cancer prevention, we treated Cdk4R24C/R24C/NrasQ61K mice topically with the T4 endonuclease V DNA repair enzyme (known as Dimericine) immediately prior to neonatal ultraviolet radiation (UVR) exposure, which has a powerful effect in exacerbating melanoma development in the mouse model. Dimericine has been shown to reduce the incidence of basal-cell and squamous cell carcinoma. Unexpectedly, we saw no difference in penetrance or age of onset of melanoma after neonatal UVR between Dimericine-treated and control animals, although the drug reduced DNA damage and cellular proliferation in the skin. Interestingly, epidermal melanocytes removed cyclobutane pyrimidine dimers (CPDs) more efficiently than surrounding keratinocytes. Our study indicates that neonatal UVR-initiated melanomas may be driven by mechanisms other than solely that of a large CPD load and/or their inefficient repair. This is further suggestive of different mechanisms by which UVR may enhance the transformation of keratinocytes and melanocytes.

Differential roles of the pRb and Arf/p53 pathways in murine naevus and melanoma genesis

We report on a systematic analysis of genotype-specific melanocyte (MC) UVR responses in transgenic mouse melanoma models along with tumour penetrance and comparative histopathology. pRb or p53 pathway mutations cooperated with NrasQ61K to transform MCs. We previously reported that MCs migrate from the follicular outer root sheath into the epidermis after neonatal UVR. Here, we found that Arf or p53 loss markedly diminished this response. Despite this, mice carrying these mutations developed melanoma with very early age of onset after neonatal UVR. Cdk4R24C did not affect the MC migration. Instead, independent of UVR exposure, interfollicular dermal MCs were more prevalent in Cdk4R24C mice. Subsequently, in adulthood, these mutants developed dermal MC proliferations reminiscent of superficial congenital naevi. Two types of melanoma were observed in this model. The location and growth pattern of the first was consistent with derivation from the naevi, while the second appeared to be of deep dermal origin. In animals carrying the Arf or p53 defects, no naevi were detected, with all tumours ostensibly skipping the benign precursor stage in progression.

The association between MC1R genotype and BRAF mutation status in cutaneous melanoma : findings from an Australian population

There is increasing epidemiological and molecular evidence that cutaneous melanomas arise through multiple causal pathways. The purpose of this study was to explore the relationship between germline and somatic mutations in a population-based series of melanoma patients to reshape and refine the divergent pathway model for melanoma. Melanomas collected from 123 Australian patients were analyzed for melanocortin-1 receptor (MC1R) variants and mutations in the BRAF and NRAS genes. Detailed phenotypic and sun exposure data were systematically collected from all patients. We found that BRAF-mutant melanomas were significantly more likely from younger patients and those with high nevus counts, and were more likely in melanomas with adjacent neval remnants. Conversely, BRAF-mutant melanomas were significantly less likely in people with high levels of lifetime sun exposure. We observed no association between germline MC1R status and somatic BRAF mutations in melanomas from this population. BRAF-mutant melanomas have different origins from other cutaneous melanomas. These data support the divergent pathways hypothesis for melanoma, which may require a reappraisal of targeted cancer prevention activities.

Melanocyte homeostasis in vivo tolerates Rb1 loss in a developmentally independent fashion

There has been uncertainty regarding the precise role that the pocket protein Rb1 plays in murine melanocyte homeostasis. It has been reported that the TAT-Cre mediated loss of exon 19 from a floxed Rb1 allele causes melanocyte apoptosis in vivo and in vitro. This is at variance with other findings showing, either directly or indirectly, that Rb1 loss in melanocytes has no noticeable effect in vivo, but in vitro leads to a semi-transformed phenotype. In this study, we show that Rb1-null melanocytes lacking exon 19 do not undergo apoptosis and survive both in vitro and in vivo, irrespective of the developmental stage at which Cre-mediated ablation of the exon occurs. Further, Rb1 loss has no serious long-term ramifications on melanocyte homeostasis in vivo, with Rb1-null melanocytes being detected in the skin after numerous hair cycles, inferring that the melanocyte stem cell population carrying the Cre-mediated deletion is maintained. Consequently, whilst Rb1 loss in the melanocyte is able to alter cellular behaviour in vitro, it appears inconsequential with respect to melanocyte homeostasis in the mouse skin.

Inverse expression states of the BRN2 and MITF transcription factors in melanoma spheres and tumour xenografts regulate the NOTCH pathway

The use of adherent monolayer cultures have produced many insights into melanoma cell growth and differentiation, but often novel therapeutics demonstrated to act on these cells are not active in vivo. It is imperative that new methods of growing melanoma cells that reflect growth in vivo are investigated. To this end, a range of human melanoma cell lines passaged as adherent cultures or induced to form melanoma spheres (melanospheres) in stem cell media have been studied to compare cellular characteristics and protein expression. Melanoma spheres and tumours grown from cell lines as mouse xenografts had increased heterogeneity when compared with adherent cells and 3D-spheroids in agar (aggregates). Furthermore, cells within the melanoma spheres and mouse xenografts each displayed a high level of reciprocal BRN2 or MITF expression, which matched more closely the pattern seen in human melanoma tumours in situ, rather than the propensity for co-expression of these important melanocytic transcription factors seen in adherent cells and 3D-spheroids. Notably, when the levels of the BRN2 and MITF proteins were each independently repressed using siRNA treatment of adherent melanoma cells, members of the NOTCH pathway responded by decreasing or increasing expression, respectively. This links BRN2 as an activator, and conversely, MITF as a repressor of the NOTCH pathway in melanoma cells. Loss of the BRN2-MITF axis in antisense-ablated cell lines decreased the melanoma sphere-forming capability, cell adhesion during 3D-spheroid formation and invasion through a collagen matrix. Combined, this evidence suggests that the melanoma sphere-culture system induces subpopulations of cells that may more accurately portray the in vivo disease, than the growth as adherent melanoma cells.

Melanoma cell invasiveness is regulated by miR-211 suppression of the BRN2 transcription factor

To identify microRNAs potentially involved in melanomagenesis, we compared microRNA expression profiles between melanoma cell lines and cultured melanocytes. The most differentially expressed microRNA between the normal and tumor cell lines was miR-211. We focused on this pigment-cell-enriched miRNA as it is derived from the microphthalmia-associated transcription factor (MITF)-regulated gene, TRPM1 (melastatin). We find that miR-211 expression is greatly decreased in melanoma cells and melanoblasts compared to melanocytes. Bioinformatic analysis identified a large number of potential targets of miR-211, including POU3F2 (BRN2). Inhibition of miR-211 in normal melanocytes resulted in increased BRN2 protein, indicating that endogenous miR-211 represses BRN2 in differentiated cells. Over-expression of miR-211 in melanoma cell lines changed the invasive potential of the cells in vitro through directly targeting BRN2 translation. We propose a model for the apparent non-overlapping expression levels of BRN2 and MITF in melanoma, mediated by miR-211 expression.

The effect of MC1R variants and sunscreen on the response of human melanocytes in vivo to ultraviolet radiation and implications for melanoma.

We conducted a clinical trial to compare the molecular and cellular responses of human melanocytes and keratinocytes in vivo to solar-simulated ultraviolet radiation (SSUVR) in 57 Caucasian participants grouped according to MC1R genotype. We found that, on average, the density of epidermal melanocytes 14 days after exposure to 2 minimal erythemal dose (MED) SSUVR was twofold higher than baseline (unirradiated) skin. However, the change in epidermal melanocyte counts among people carrying germline MC1R variants (97% increase) was significantly less than those with wild-type MC1R (164% increase; P = 0.01). We also found that sunscreen applied to the skin before exposure to 2 MED SSUVR completely blocked the effects of DNA damage, p53 induction, and cellular proliferation in both melanocytes and keratinocytes.

Exposure to solar ultraviolet radiation is associated with a decreased folate status in women of childbearing age

In vitro studies indicate that folate in collected human blood is vulnerable to degradation after exposure to ultraviolet (UV) radiation. This has raised concerns about folate depletion in individuals with high sun exposure. Here, we investigate the association between personal solar UV radiation exposure and serum folate concentration, using a three-week prospective study that was undertaken in females aged 18–47 years in Brisbane, Australia (153 E, 27 S). Following two weeks of supplementation with 500 μg of folic acid daily, the change in serum folate status was assessed over a 7-day period of measured personal sun exposure. Compared to participants with personal UV exposures of <200 Joules per day, participants with personal UV exposures of 200–599 and >600 Joules per day had significantly higher depletion of serum folate (p = 0.015). Multivariable analysis revealed personal UV exposure as the strongest predictor accounting for 20% of the overall change in serum folate (Standardised B = −0.49; t = −3.75; p = <0.01). These data show that increasing solar UV radiation exposures reduces the effectiveness of folic acid supplementation. The consequences of this association may be most pronounced for vulnerable individuals, such as women who are pregnant or of childbearing age with high sun exposures.

A place to make, hack, and learn: makerspaces in Australian public libraries

Content-creation spaces, or ‘makerspaces’, are an emerging phenomenon in public libraries worldwide. This study investigated the current state of makerspaces in Australian public libraries. Qualitative interviews with three information professionals formed the data collection. Thematic analysis of interviews addressed two research questions: What are the issues and challenges of creating makerspaces within Australian public libraries? How can they be addressed? Findings revealed the substantive benefits of these spaces, including enhanced community engagement, development of a new form of library as ‘third place’, and transforming the library's image from that of a place where works are consumed to that of a place where works are created. Additionally the study highlighted significant challenges to creating these spaces, including budgetary constraints, resistance to change within organisations and proving the relevance of such spaces within a library context. The study provides suggestions for overcoming these obstacles and provides areas for further research in the area, including larger studies across a broader geographic area and further investigation and follow-up into upcoming programs within existing makerspaces.

The glycosphingolipid P1 is an ovarian cancer-associated carbohydrate antigen involved in migration

Background The level of plasma-derived naturally circulating anti-glycan antibodies (AGA) to P1 trisaccharide has previously been shown to significantly discriminate between ovarian cancer patients and healthy women. Here we aim to identify the Ig class that causes this discrimination, to identify on cancer cells the corresponding P1 antigen recognised by circulating anti-P1 antibodies and to shed light into the possible function of this glycosphingolipid. Method An independent Australian cohort was assessed for the presence of anti-P1 IgG and IgM class antibodies using suspension array. Monoclonal and human derived anti-glycan antibodies were verified using three independent glycan-based immunoassays and flow cytometry-based inhibition assay. The P1 antigen was detected by LC-MS/MS and flow cytometry. FACS-sorted cell lines were studied on the cellular migration by colorimetric assay and real-time measurement using xCELLigence system. Results Here we show in a second independent cohort (n=155) that the discrimination of cancer patients is mediated by the IgM class of anti-P1 antibodies (P=0.0002). The presence of corresponding antigen P1 and structurally related epitopes in fresh tissue specimens and cultured cancer cells is demonstrated. We further link the antibody and antigen (P1) by showing that human naturally circulating and affinity-purified anti-P1 IgM isolated from patients ascites can bind to naturally expressed P1 on the cell surface of ovarian cancer cells. Cell-sorted IGROV1 was used to obtain two study subpopulations (P1-high, 66.1%; and P1-low, 33.3%) and observed that cells expressing high P1-levels migrate significantly faster than those with low P1-levels. Conclusions This is the first report showing that P1 antigen, known to be expressed on erythrocytes only, is also present on ovarian cancer cells. This suggests that P1 is a novel tumour-associated carbohydrate antigen recognised by the immune system in patients and may have a role in cell migration. The clinical value of our data may be both diagnostic and prognostic; patients with low anti-P1 IgM antibodies present with a more aggressive phenotype and earlier relapse.

Government grants - an abrogation or management of risks?

New public management (NPFM), with its hands-on, private sector-style performance measurement, output control, parsimonious use of resources, disaggreation of public sector units and greater competition in the public sector, has significantly affected charitable and nonprofit organisations delivering community services (Hood, 1991; Dunleavy, 1994; George & Wilding, 2002). The literature indicates that nonprofit organisations under NPM believe they are doing more for less: while administration is increasing, core costs are not being met; their dependence on government funding comes at the expense of other funding strategies; and there are concerns about proportionality and power asymmetries in the relationship (Kerr & Savelsberg, 2001; Powell & Dalton, 2011; Smith, 2002, p. 175; Morris, 1999, 2000a). Government agencies are under increased pressure to do more with less, demonstrate value for money, measure social outcomes, not merely outputs and minimise political risk (Grant, 2008; McGreogor-Lowndes, 2008). Government-community service organisation relationships are often viewed as 'uneasy alliances' characterised by the pressures that come with the parties' differing roles and expectations and the pressures that come with the parties' differing roles and expectations and the pressurs of funding and security (Productivity Commission, 2010, p. 308; McGregor-Lowndes, 2008, p. 45; Morris, 200a). Significant community services are now delivered to citizens through such relationships, often to the most disadvantaged in the community, and it is important for this to be achieved with equity, efficiently and effectively. On one level, the welfare state was seen as a 'risk management system' for the poor, with the state mitigating the risks of sickness, job loss and old age (Giddens, 1999) with the subsequent neoliberalist outlook shifting this risk back to households (Hacker, 2006). At the core of this risk shift are written contracts. Vincent-Jones (1999,2006) has mapped how NPM is characterised by the use of written contracts for all manner of relations; e.g., relgulation of dealings between government agencies, between individual citizens and the state, and the creation of quais-markets of service providers and infrastructure partners. We take this lens of contracts to examine where risk falls in relation to the outsourcing of community services. First we examine the concept of risk. We consider how risk might be managed and apportioned between governments and community serivce organisations (CSOs) in grant agreements, which are quasiy-market transactions at best. This is informed by insights from the law and economics literature. Then, standard grant agreements covering several years in two jurisdictions - Australia and the United Kingdom - are analysed, to establish the risk allocation between government and CSOs. This is placed in the context of the reform agenda in both jurisdictions. In Australia this context is th enonprofit reforms built around the creation of a national charities regulator, and red tape reduction. In the United Kingdom, the backdrop is the THird Way agenda with its compacts, succeed by Big Society in a climate of austerity. These 'case studies' inform a discussion about who is best placed to bear and manage the risks of community service provision on behalf of government. We conclude by identifying the lessons to be learned from our analysis and possible pathways for further scholarship.

Prognostic value of BRAF mutations in localized cutaneous melanoma

BACKGROUND: BRAF mutations are frequent in melanoma but their prognostic significance remains unclear. OBJECTIVE: We sought to further evaluate the prognostic value of BRAF mutations in localized cutaneous melanoma. METHODS: We undertook an observational retrospective study of 147 patients with localized invasive (stages I and II) cutaneous melanomas to determine the prognostic value of BRAF mutation status. RESULTS: After a median follow-up of 48 months, patients with localized melanomas with BRAF-mutant melanomas exhibited poorer disease-free survival than those with BRAF-wt genotype (hazard ratio 2.2, 95% confidence interval 1.1-4.3) even after adjustment for Breslow thickness, tumor ulceration, location, age, sex, and tumor mitotic rate. LIMITATIONS: The retrospective design and the small number of events are limitations. CONCLUSIONS: Our findings suggest that reappraisal of clinical treatment approaches for patients with localized melanoma harboring tumors with BRAF mutation might be warranted

Action regulation across the adult lifespan (ARAL): A metatheory of work and aging

The goals of this article are to integrate action regulation theory (ART) with the lifespan developmental perspective and to outline tenets of a new metatheory of work and aging. The action regulation across the adult lifespan (ARAL) theory explains how workers influence, and are influenced by, their environment across different time spans. First, the basic concepts of ART are described, including the sequential and hierarchical structure of actions, complete tasks and actions, foci of action regulation, and the action-regulating mental model. Second, principles of the lifespan developmental perspective are delineated, including development as a lifelong and multidirectional process, the joint occurrence of gains and losses, intraindividual plasticity, historical embeddedness, and contextualism. Third, propositions of ARAL theory are derived by analyzing workers’ action regulation from a lifespan developmental perspective (i.e., effects of aging on action regulation), and by analyzing aging and development in the work context from an ART perspective (i.e., effects of action regulation on age-related changes in cognition and personality). Fourth, we develop further propositions to integrate ART with lifespan theories of motivation and socioemotional experience. Finally, we discuss implications for future research and practice based on ARAL theory.