Prognostic gene expression signature for high-grade serous ovarian cancer.

BACKGROUND: Median overall survival (OS) for women with high-grade serous ovarian cancer (HGSOC) is ∼4 years, yet survival varies widely between patients. There are no well-established, gene expression signatures associated with prognosis. The aim of this study was to develop a robust prognostic signature for OS in patients with HGSOC. PATIENTS AND METHODS: Expression of 513 genes, selected from a meta-analysis of 1455 tumours and other candidates, was measured using NanoString technology from formalin-fixed paraffin-embedded tumour tissue collected from 3769 women with HGSOC from multiple studies. Elastic net regularization for survival analysis was applied to develop a prognostic model for 5-year OS, trained on 2702 tumours from 15 studies and evaluated on an independent set of 1067 tumours from six studies. RESULTS: Expression levels of 276 genes were associated with OS (false discovery rate < 0.05) in covariate-adjusted single-gene analyses. The top five genes were TAP1, ZFHX4, CXCL9, FBN1 and PTGER3 (P < 0.001). The best performing prognostic signature included 101 genes enriched in pathways with treatment implications. Each gain of one standard deviation in the gene expression score conferred a greater than twofold increase in risk of death [hazard ratio (HR) 2.35, 95% confidence interval (CI) 2.02-2.71; P < 0.001]. Median survival [HR (95% CI)] by gene expression score quintile was 9.5 (8.3 to -), 5.4 (4.6-7.0), 3.8 (3.3-4.6), 3.2 (2.9-3.7) and 2.3 (2.1-2.6) years. CONCLUSION: The OTTA-SPOT (Ovarian Tumor Tissue Analysis consortium - Stratified Prognosis of Ovarian Tumours) gene expression signature may improve risk stratification in clinical trials by identifying patients who are least likely to achieve 5-year survival. The identified novel genes associated with the outcome may also yield opportunities for the development of targeted therapeutic approaches.

Highly photostable wide-dynamic-range pH sensitive semiconducting polymer dots enabled by dendronizing the near-IR emitters.

One constraint of semiconducting polymer dots (Pdots), especially those with near-IR emission, is their low effective emitter ratio (∼1.5 mole percent), which limits their pH sensing performance. The other critical issue of existing Pdot-based pH sensors is their poor photostability. To address these issues, we developed a series of Pdots by dendronizing the squaraine-based pH responsive near-IR emitter, which is covalently incorporated into the polyfluorene (PFO) backbone. The fluorescence self-quenching of the NIR squaraine emitter was effectively suppressed at a high emitter concentration of 5 mole percent. Through controlling the individually incomplete energy transfer from the amorphous PFO donor to the blue ß-phase PFO and NIR squaraine emitter, we obtained a ratiometric pH sensor with simultaneously improved pH sensitivity, brightness, and photostability. The Pdots showed a fast and reversible pH response over the whole biological pH range of 4.7 to 8.5. Intracellular pH mapping was successfully demonstrated using this ultra-bright and photostable Pdot-based pH indicator.

Dynamic intracellular delivery of antibiotics via pH-responsive polymersomes.

Reversible addition-fragmentation chain transfer (RAFT) polymerization was employed to prepare a series of copolymers consisting of 2-hdroxyethyl methacrylate (HEMA) and poly(ethylene glycol) methyl ether methacrylate (FWavg ~ 950 Da) (O950) with variable comonomer compositions and molecular weights for use as polymeric scaffolds. Reactivity ratios for the monomer pair were determined to be 1.37 and 0.290 respectively. To these scaffolds trithiocarbonate-based RAFT chain transfer agents (CTAs) were grafted using carbodiimide chemistry. The resultant graft chain transfer agents (gCTA) were subsequently employed to polymerize dimethylaminoethyl methacrylate (DMAEMA) and (HPMA) between degrees of polymerization (DP) of 25 and 200. Kinetic analysis for the polymerization of DMAEMA targeting a DP of 100 from a 34 arm graft gCTA show linear Mn conversion and pseudo first order rate plots with narrow molecular weight distributions that move toward lower elution volumes with monomer conversion. D values for these polymerizations remain low at around 1.20 at monomer conversions as high as 70 %. pH-responsive endosomalytic brushes capable of spontaneously self-assembling into polymersomes were synthesized and a combination of dynamic light scattering (DLS), cryoTEM, and red blood cell hemolysis were employed to evaluate the aqueous solution properties of the polymeric brush as a function of pH. Successful encapsulation of ceftazidime and pH-dependent drug release properties were confirmed by HPLC. Intracellular antibiotic activity of the drug-loaded polymersomes was confirmed in a macrophage coculture model of infection with B. thailandensis and RAW 264.7 cells.

Well-defined single polymer nanoparticles for the antibody-targeted delivery of chemotherapeutic agents.

Aqueous reversible addition-fragmentation chain transfer (RAFT) polymerization was employed to prepare a series of linear copolymers of N,N-dimethylacrylamide (DMA) and 2-hydroxyethylacrylamide (HEAm) with narrow D values over a molecular weight range spanning three orders of magnitude (103 to 106 Da). Trithiocarbonate-based RAFT chain transfer agents (CTAs) were grafted onto these scaffolds using carbodiimide chemistry catalyzed with DMAP. The resultant graft chain transfer agent (gCTA) was subsequently employed to synthesize polymeric brushes with a number of important vinyl monomer classes including acrylamido, methacrylamido, and methacrylate. Brush polymerization kinetics were evaluated for the aqueous RAFT polymerization of DMA from a 10 arm gCTA. Polymeric brushes containing hydroxyl functionality were further functionalized in order to prepare 2nd generation gCTAs which were subsequently employed to prepare polymers with a brushed-brush architecture with molecular weights in excess of 106 Da. These resultant single particle nanoparticles (SNPs) were employed as drug delivery vehicles for the anthracycline-based drug doxorubicin via copolymerization of DMA with a protected carbazate monomer (bocSMA). Cell-specific targeting functionality was also introduced via copolymerization with a biotin-functional monomer (bioHEMA). Drug release of the hydrazone linked doxorubicin was evaluated as function of pH and serum and chemotherapeutic activity was evaluated in SKOV3 ovarian cancer cells.

What childhood obesity prevention programmes work? A systematic review and meta-analysis.

Previous reviews of childhood obesity prevention have focused largely on schools and findings have been inconsistent. Funded by the US Agency for Healthcare Research and Quality (AHRQ) and the National Institutes of Health, we systematically evaluated the effectiveness of childhood obesity prevention programmes conducted in high-income countries and implemented in various settings. We searched MEDLINE®, Embase, PsycINFO, CINAHL®, ClinicalTrials.gov and the Cochrane Library from inception through 22 April 2013 for relevant studies, including randomized controlled trials, quasi-experimental studies and natural experiments, targeting diet, physical activity or both, and conducted in children aged 2-18 in high-income countries. Two reviewers independently abstracted the data. The strength of evidence (SOE) supporting interventions was graded for each study setting (e.g. home, school). Meta-analyses were performed on studies judged sufficiently similar and appropriate to pool using random effect models. This paper reported our findings on various adiposity-related outcomes. We identified 147 articles (139 intervention studies) of which 115 studies were primarily school based, although other settings could have been involved. Most were conducted in the United States and within the past decade. SOE was high for physical activity-only interventions delivered in schools with home involvement or combined diet-physical activity interventions delivered in schools with both home and community components. SOE was moderate for school-based interventions targeting either diet or physical activity, combined interventions delivered in schools with home or community components or combined interventions delivered in the community with a school component. SOE was low for combined interventions in childcare or home settings. Evidence was insufficient for other interventions. In conclusion, at least moderately strong evidence supports the effectiveness of school-based interventions for preventing childhood obesity. More research is needed to evaluate programmes in other settings or of other design types, especially environmental, policy and consumer health informatics-oriented interventions.

Microfluidics for single-cell genetic analysis.

The ability to correlate single-cell genetic information to cellular phenotypes will provide the kind of detailed insight into human physiology and disease pathways that is not possible to infer from bulk cell analysis. Microfluidic technologies are attractive for single-cell manipulation due to precise handling and low risk of contamination. Additionally, microfluidic single-cell techniques can allow for high-throughput and detailed genetic analyses that increase accuracy and decrease reagent cost compared to bulk techniques. Incorporating these microfluidic platforms into research and clinical laboratory workflows can fill an unmet need in biology, delivering the highly accurate, highly informative data necessary to develop new therapies and monitor patient outcomes. In this perspective, we describe the current and potential future uses of microfluidics at all stages of single-cell genetic analysis, including cell enrichment and capture, single-cell compartmentalization and manipulation, and detection and analyses.

Glucose-6-phosphate dehydrogenase--beyond the realm of red cell biology.

Glucose-6-phosphate dehydrogenase (G6PD) is critical to the maintenance of NADPH pool and redox homeostasis. Conventionally, G6PD deficiency has been associated with hemolytic disorders. Most biochemical variants were identified and characterized at molecular level. Recently, a number of studies have shone light on the roles of G6PD in aspects of physiology other than erythrocytic pathophysiology. G6PD deficiency alters the redox homeostasis, and affects dysfunctional cell growth and signaling, anomalous embryonic development, and altered susceptibility to infection. The present article gives a brief review of basic science and clinical findings about G6PD, and covers the latest development in the field. Moreover, how G6PD status alters the susceptibility of the affected individuals to certain degenerative diseases is also discussed.

Potential for biomarkers of chronic kidney disease-mineral bone disorder to improve patient care.

Chronic kidney disease (CKD) is a growing public health problem. Cardiovascular disease is common in CKD, but standard risk assessment tools perform poorly in this population. Equally, despite CKD being associated with an increased risk for death and dialysis, standard biochemical measurements have limited prognostic value. Novel serum biomarkers may aid risk assessment; however, studies have shown varying clinical utility in relation to progression of CKD, incident cardiovascular disease and death. This inconsistency may relate to limitations in our understanding of the biological actions and interactions of these biomarkers. This review discusses a range of biomarkers in relation to these clinical endpoints in CKD-mineral bone disorder. We consider where biomarkers may enhance risk stratification and improve clinical management, but also highlight where they fall short of achieving this objective.

Variation in the heritability of body mass index based on diverse twin studies: a systematic review.

OBJECTIVES: Over the past three decades, twin studies have shown variation in the heritability of obesity. This study examined the difference of body mass index (BMI) heritability (BMI-H) by population characteristics, such as sex, age, time period of observation and average BMI, as well as by broad social-environmental factors as indicated by country-level gross domestic product (GDP) per capita and GDP growth rate. METHODS: Twin studies that reported BMI-H and were published in English from January 1990 to February 2011 after excluding those with disease, special occupations or combined heritability estimates for country/ethnic groups were searched in PubMed. 32 studies were identified from Finland (7), the United Kingdom (6), the United States (3), Denmark (3), China (3), Netherlands (2), South Korea (2), Sweden (2) and four from other countries. Meta-regression models with random effects were used to assess variation in BMI-H. RESULTS: Heterogeneity of BMI-H is significantly attributable to variations in age (<20, 20-55 and ≥56 years), time period of observation (i.e. year of data collection), average BMI and GDP (≤$20,000, $20,001-26,000 and >$26,000). BMI-H was higher in adolescents (<20 years), in studies done in past years, and in populations with higher average BMIs or higher GDP per capita (≥$26,000) than their counterparts. Consistent lowering effects of high GDP growth rate (>median) on BMI-H were shown through stratified analyses by GDP. BMI-H was lower in countries of mid-level GDP, particularly those experiencing rapid economic growth. CONCLUSIONS: BMI-H is sensitive to age, time period of observation, average BMI, GDP and rapid economic growth.

Effective NET formation in neutrophils from individuals with G6PD Taiwan-Hakka is associated with enhanced NADP(+) biosynthesis.

In response to infection, neutrophils employ various strategies to defend against the invading microbes. One of such defense mechanisms is the formation of neutrophil extracellular traps (NETs). Recent studies suggest that reactive oxygen species is a signal critical to NET formation. This prompts us to examine whether neutrophils from individuals with glucose-6-phosphate dehydrogenase (G6PD) Taiwan-Hakka variant, which are prone to oxidative stress generation, have altered ability to form NET. We adopted an image-based method to study the NET formation potential in neutrophils from G6PD-deficient patients. Neutrophils from either normal or G6PD-deficient individuals underwent NETosis in response to phorbol 12-myristate 13-acetate (PMA). The extent of NETosis in the former did not significantly differ from that of the latter. Diphenyleneiodonium sulfate (DPI) and 3-methyladenine (MA) inhibited PMA-stimulated NET formation in these cells, suggesting the involvement of NADPH oxidase and autophagy in the process. Glucose oxidase (GO) and xanthine oxidase/xanthine (XO/X) could induce a similar extent of NET formation in normal and G6PD-deficient neutrophils. GO- or XO-induced NETosis was not inhibitable by MA, implying that reactive oxygen species (ROS) can act as an independent signal for activation of NETosis. Mechanistically, enhanced superoxide production in neutrophils was associated with increases in levels of NAD(+) and NADP(+), as well as activation of NAD(+) kinase. Taken together, these findings suggest that G6PD-deficient neutrophils are as equally efficient as normal cells in NET formation, and their deficiency in G6PD-associated NADPH regeneration capacity is largely compensated for by nicotinamide nucleotide biosynthesis.

Glucose 6-phosphate dehydrogenase deficiency enhances germ cell apoptosis and causes defective embryogenesis in Caenorhabditis elegans.

Glucose 6-phosphate dehydrogenase (G6PD) deficiency, known as favism, is classically manifested by hemolytic anemia in human. More recently, it has been shown that mild G6PD deficiency moderately affects cardiac function, whereas severe G6PD deficiency leads to embryonic lethality in mice. How G6PD deficiency affects organisms has not been fully elucidated due to the lack of a suitable animal model. In this study, G6PD-deficient Caenorhabditis elegans was established by RNA interference (RNAi) knockdown to delineate the role of G6PD in animal physiology. Upon G6PD RNAi knockdown, G6PD activity was significantly hampered in C. elegans in parallel with increased oxidative stress and DNA oxidative damage. Phenotypically, G6PD-knockdown enhanced germ cell apoptosis (2-fold increase), reduced egg production (65% of mock), and hatching (10% of mock). To determine whether oxidative stress is associated with G6PD knockdown-induced reproduction defects, C. elegans was challenged with a short-term hydrogen peroxide (H2O2). The early phase egg production of both mock and G6PD-knockdown C. elegans were significantly affected by H2O2. However, H2O2-induced germ cell apoptosis was more dramatic in mock than that in G6PD-deficient C. elegans. To investigate the signaling pathways involved in defective oogenesis and embryogenesis caused by G6PD knockdown, mutants of p53 and mitogen-activated protein kinase (MAPK) pathways were examined. Despite the upregulation of CEP-1 (p53), cep-1 mutation did not affect egg production and hatching in G6PD-deficient C. elegans. Neither pmk-1 nor mek-1 mutation significantly affected egg production, whereas sek-1 mutation further decreased egg production in G6PD-deficient C. elegans. Intriguingly, loss of function of sek-1 or mek-1 dramatically rescued defective hatching (8.3- and 9.6-fold increase, respectively) induced by G6PD knockdown. Taken together, these findings show that G6PD knockdown reduces egg production and hatching in C. elegans, which are possibly associated with enhanced oxidative stress and altered MAPK pathways, respectively.

Effects of aliasing on the fidelity of a two dimensional array of foci generated with a kinoform.

This paper investigates, through simulation and experiment, the behavior of two dimensional foci arrays generated via phase-only holography where an iterative algorithm was used to produce the kinoforms. Specifically, we studied how aliasing of the signal on a spatial light modulator affects the quality of the foci array as the density and size of the array are varied. This study provides a reference for applications where it is important to understand how the fidelity and overall quality of the foci array changes as the number of foci increases and as the spacing between foci decreases.

Residence time and contact volume in sloped compost and compost/vegetated filter beds.

Little is known about transport mechanisms in sloped dormant vegetated and compost only filters for roadway runoff. Residence time experiments were carried out in triplicate in 0.254 m wide × 0.65 m long by 0.10 m deep beds using a bromide tracer. Bed slope was 12°. Only at the lowest flow rate tested (0.276 l/min per m of filter width) were mean residence times in compost beds with and without dormant grasses different. Pools formed ahead of beds at higher flow rate, and pool depth reached bed depth at 3.54 l/min/m. The ideal model of a well-mixed pool in series with a plug flow porous bed was a good predictor of effluent concentration data at flows ≥2.66 l/min/m. Theoretical contact volume within the beds increased with flow rate to reach approx. 30% of available pore space, while free drainage volume declined. Data shows that designs for sloped compost filter beds must consider flow, bed depth and length, and whether or not areas for pooling are needed.

Accelerated total dose infusion of low molecular weight iron dextran is safe and efficacious in chronic kidney disease patients.

BACKGROUND: Low molecular weight iron dextran (LMWID) is licensed for use as a total dose infusion (TDI) over 4-6 h. In order to improve patient convenience and cost-effectiveness of therapy, we investigated the safety and efficacy of adopting accelerated dosing regimens and compared this with a standard rate LMWID infusion. METHODS: A retrospective study of patients undergoing accelerated and standard rate TDI of LMWID was conducted across three centres. A total of 1904 doses of LMWID were administered at an accelerated rate of 1 g over 1 h 40 min. This was compared with 395 patients who had standard rate infusion of 1 g LMWID over 3-4 h. RESULTS: There were eight minor adverse events in patients receiving accelerated dose LMWID (8/1904, 0.42%) in comparison to one adverse event in patients receiving a standard regimen (1/395, 0.25%). No serious adverse events occurred. Serum haemoglobin and ferritin significantly improved in both groups. CONCLUSION: TDI LMWID is a safe and efficacious method of iron replacement. Accelerated infusion regimen is safe and compares well with standard rate infusion regimen. Furthermore, accelerated TDI of LMWID enables greater numbers of patients to be treated and consequently there appear to be advantages for both patient and health resources.

Endogenous endophthalmitis: 10-year experience at a tertiary referral centre.

PURPOSE: Endogenous endophthalmitis (EE) is a sight-threatening emergency and the aetiology is often multifactorial. Delayed diagnosis may exacerbate the poor visual prognosis. We describe the management and visual outcomes of EE presenting to a tertiary referral centre. PATIENTS AND METHODS: A prospective consecutive case series of 64 patients presenting with presumed EE from 1997 to 2007 to the Royal Victorian Eye and Ear Hospital were included. All data were collected in a standardized manner. Outcome measures included: visual acuity, microbial profiles, and vitrectomy rate. RESULTS: In total, 64 cases of EE were identified over the study period with a mean age of 57.5 years, and 53.5% were male. Presenting acuities ranged from Snellen 6/6 to no perception of light (NPL). Identifiable risk factors were present in 78.1%, with the majority related to intravenous drug abuse. A 64.1% culture positivity rate was recorded. A vitrectomy rate of 57, 56, and 21% was recorded in documented bacterial, fungal, and no growth cases, respectively. Final Snellen acuities ranged from 6/6 to NPL. A total of 5 out of 64 eyes were enucleated, of which 3 identified Klebsiella species. Better visual outcome was documented in fungal cases. CONCLUSION: EE is a serious ocular condition and has a varied aetiology. Visual outcomes are often poor, irrespective of the method of management. Fungal aetiology often confers a better prognosis, and vitrectomy is advocated for bacterial proven cases.

Activation and retrograde transport of protein kinase G in rat nociceptive neurons after nerve injury and inflammation.

Nerve injury elicits both universal and limited responses. Among the former is regenerative growth, which occurs in most peripheral neurons, and among the latter is the long-term hyperexcitability that appears selectively in nociceptive sensory neurons. Since positive injury signals communicate information from the site of an injury to the cell body, we hypothesize that a nerve injury activates both universal and limited positive injury signals. Studies in Aplysia indicate that protein kinase G is a limited signal that is responsible for the induction of long-term hyperexcitability. Given that long-term hyperexcitability contributes to chronic pain after axotomy in rodent neuropathic pain models, we investigated its underlying basis in the rat peripheral nervous system. Using biochemical assays, Western blots, and immunocytochemistry we found that the Type 1alpha protein kinase G is the predominant isoform in the rat periphery. It is present primarily in axons and cell bodies of nociceptive neurons, including populations that are isolectin B4-positive, isolectin B4-negative, and those that express transient receptor potential vanilloid receptor-1. Surprisingly, protein kinase G is not present in the facial nerve, which overwhelmingly contains axons of motor neurons. Crushing the sciatic nerve or a cutaneous sensory nerve activates protein kinase G in axons and results in its retrograde transport to the neuronal somata in the DRG. Preventing the activation of protein kinase G by injecting Rp-8-pCPT-cGMPS into the crush site abolished the transport. The protein kinase A inhibitor Rp-8-pCPT-cAMPS had no effect. Extracellular signal-related kinases 42/44 are also activated and transported after nerve crush, but in both motor and sensory axons. Chronic pain has been linked to long-term hyperexcitability following a nerve inflammation in several rodent models. We therefore injected complete Freund's adjuvant into the hindpaw to induce an inflammation and found that protein kinase G was activated in the sural nerve and transported to the DRG. In contrast, the extracellular signal-related kinases in the sensory axons were not activated by the complete Freund's adjuvant. These studies support the idea that the extracellular signal-related kinases are universal positive axonal signals and that protein kinase G is a limited positive axonal signal. They also establish the association between protein kinase G, the induction of long-term hyperexcitability, and chronic pain in rodents.

A Java application for tissue section image analysis.

The medical industry has taken advantage of Java and Java technologies over the past few years, in large part due to the language's platform-independence and object-oriented structure. As such, Java provides powerful and effective tools for developing tissue section analysis software. The background and execution of this development are discussed in this publication. Object-oriented structure allows for the creation of "Slide", "Unit", and "Cell" objects to simulate the corresponding real-world objects. Different functions may then be created to perform various tasks on these objects, thus facilitating the development of the software package as a whole. At the current time, substantial parts of the initially planned functionality have been implemented. Getafics 1.0 is fully operational and currently supports a variety of research projects; however, there are certain features of the software that currently introduce unnecessary complexity and inefficiency. In the future, we hope to include features that obviate these problems.

Human chromosome 21-specific DNA markers are useful in prenatal detection of Down syndrome.

Trisomy 21 is the most common chromosomal aberration in live births. In this study we employed human chromosome 21-specific short tandem repeat (STR) DNA markers to determine the numbers of chromosome 21 present in fetal cells. Forty amniotic fluid samples from pregnancies complicated with fetal Down syndrome and 98 samples from euploid pregnancies were analyzed for D21S11 and interferon-alpha receptor (IFNAR) gene intervening sequence. Fluorescent dye-labeled primers were used in PCR amplification of these 2 markers. The PCR amplicon was analyzed with an automatic DNA sequence analyzer. The results showed that 35 of 40 fetal Down syndrome samples analyzed for IFNAR showed 3 distinct peaks, while 24 of 30 cases analyzed for D21S11 showed 3 distinct peaks. Two Down syndrome samples showed two uneven peaks. By analyzing 98 euploid pregnancies as controls, the ratios of area under the peaks were determined to be 1.31 +/- 0.22 and 1.96 +/- 0.18 (mean +/- SD) for the euploid pregnancies and pregnancies complicated by fetal Down syndrome with 2 peaks, respectively. Our data showed that altogether 39 of 40 (97.5%) Down syndrome cases were correctly identified based on either the 3-peak pattern in one or more of the DNA markers or the relative peak area ratio calculation. In conclusion, polymorphic STR DNA markers are useful for determining the numbers of chromosome 21 in fetal cells. The high sensitivity and automation of the procedures suggest a good prospect for use of this method in prenatal detection of fetal Down syndrome. However, this is a preliminary investigation and a large-scale study is necessary to validate the clinical application of this protocol.

Supportive care management of brain metastases: what is known and what we need to know. Conference proceedings of the National Cancer Institute of Canada (NCIC) Workshop on Symptom Control in Radiation Oncology.

OBJECTIVES: To review the results of published randomised controlled trials in the treatment of brain metastases and, from the knowledge gained from these trials, to identify potential study questions. MATERIALS AND METHODS: The literature was searched for randomised controlled trials that dealt with the management of brain metastases. Potential research questions were identified on the basis of the results of the literature review. RESULTS: A number of research questions were identified. In the context of the NCIC Symptom Control Group, a trial of supportive care alone vs supportive care and whole-brain radiotherapy (WBRT) in a subset of patients with the diagnosis of brain metastases was deemed to be of highest priority. We discussed a number of issues relating to the feasibility of such a trial. CONCLUSIONS: The optimal management of brain metastases remains elusive. Despite the results of numerous randomised controlled trials, many questions remain unanswered. The magnitude of benefit using WBRT above supportive care alone is uncertain. A trial of supportive care alone vs supportive care and WBRT may be successful once target population, feasibility and methodological issues are thoroughly solved.