Profiling of the secretome of human cancer cells: Preparation of supernatant for proteomic analysis

Secretomic analysis requires removal of serum proteins from cell-culture media. We evaluate the proteins washed from cells prepared in bovine serum-supplemented medium. PBS and serum-free-medium (SFM) were the washing solutions. A Bradford assay was used for total protein concentration and a 1D gel and LC-MS/MS, to assign the protein to human or bovine origin. For both wash solutions, all bovine protein had been removed by the third wash, without compromising the number of living cells. Further washes reduced the number of living cells, especially when using PBS. Proteomic analysis of wash supernatant showed that SFM induced greater lysis of dead cells. Three washes were sufficient to minimize the effects on cell viability, while still removing serum proteins. Washing in SFM resulted in contamination of the wash supernatant with lysed dead cell proteins. Washed cells were incubated in SFM and exposed to ionizing radiation. Analysis of the supernatant showed an increase in human cytoplasmic, plasma membrane, and nuclear protein following irradiation. Secreted proteins were also detected, but in smaller quantities. The significance of these findings extend to in vitro studies of bystander phenomena, since the proteins of lysed dead cells may participate in driving bystander responses

Theories of behaviour change synthesised into a set of theoretical groupings: Introducing a thematic series on the Theoretical Domains Framework

Behaviour change is key to increasing the uptake of evidence into healthcare practice. Designing behaviour-change interventions first requires problem analysis, ideally informed by theory. Yet the large number of partly overlapping theories of behaviour makes it difficult to select the most appropriate theory. The need for an overarching theoretical framework of behaviour change was addressed in research in which 128 explanatory constructs from 33 theories of behaviour were identified and grouped. The resulting Theoretical Domains Framework (TDF) appears to be a helpful basis for investigating implementation problems. Research groups in several countries have conducted TDF-based studies. It seems timely to bring together the experience of these teams in a thematic series to demonstrate further applications and to report key developments. This overview article describes the TDF, provides a brief critique of the framework, and introduces this thematic series. In a brief review to assess the extent of TDF-based research, we identified 133 papers that cite the framework. Of these, 17 used the TDF as the basis for empirical studies to explore health professionals’ behaviour. The identified papers provide evidence of the impact of the TDF on implementation research. Two major strengths of the framework are its theoretical coverage and its capacity to elicit beliefs that could signify key mediators of behaviour change. The TDF provides a useful conceptual basis for assessing implementation problems, designing interventions to enhance healthcare practice, and understanding behaviour-change processes. We discuss limitations and research challenges and introduce papers in this series

The spatial and temporal patterns of falciparum and vivax malaria in Perú: 1994–2006

<p>Abstract</p> <p>Background</p> <p>Malaria is the direct cause of approximately one million deaths worldwide each year, though it is both preventable and curable. Increasing the understanding of the transmission dynamics of falciparum and vivax malaria and their relationship could suggest improvements for malaria control efforts. Here the weekly number of malaria cases due to <it>Plasmodium falciparum </it>(1994–2006) and <it>Plasmodium vivax </it>(1999–2006) in Perú at different spatial scales in conjunction with associated demographic, geographic and climatological data are analysed.</p> <p>Methods</p> <p>Malaria periodicity patterns were analysed through wavelet spectral analysis, studied patterns of persistence as a function of community size and assessed spatial heterogeneity via the Lorenz curve and the summary Gini index.</p> <p>Results</p> <p>Wavelet time series analyses identified annual cycles in the incidence of both malaria species as the dominant pattern. However, significant spatial heterogeneity was observed across jungle, mountain and coastal regions with slightly higher levels of spatial heterogeneity for <it>P. vivax </it>than <it>P. falciparum</it>. While the incidence of <it>P. falciparum </it>has been declining in recent years across geographic regions, <it>P. vivax </it>incidence has remained relatively steady in jungle and mountain regions with a slight decline in coastal regions. Factors that may be contributing to this decline are discussed. The time series of both malaria species were significantly synchronized in coastal (ρ = 0.9, P < 0.0001) and jungle regions (ρ = 0.76, P < 0.0001) but not in mountain regions. Community size was significantly associated with malaria persistence due to both species in jungle regions, but not in coastal and mountain regions.</p> <p>Conclusion</p> <p>Overall, findings highlight the importance of highly refined spatial and temporal data on malaria incidence together with demographic and geographic information in improving the understanding of malaria persistence patterns associated with multiple malaria species in human populations, impact of interventions, detection of heterogeneity and generation of hypotheses.</p

Heel raises versus prefabricated orthoses in the treatment of posterior heel pain associated with calcaneal apophysitis (Sever's Disease): study protocol for a randomised controlled trial

<p>Abstract</p> <p>Background</p> <p>Posterior Heel pain can present in children of 8 to 14 years, associated with or clinically diagnosed as Sever's disease, or calcaneal apophysitis. Presently, there are no comparative randomised studies evaluating treatment options for posterior heel pain in children with the clinical diagnosis of calcaneal apophysitis or Sever's disease. This study seeks to compare the clinical efficacy of some currently employed treatment options for the relief of disability and pain associated with posterior heel pain in children.</p> <p>Method</p> <p>Design: Factorial 2 × 2 randomised controlled trial with monthly follow-up for 3 months.</p> <p>Participants: Children with clinically diagnosed posterior heel pain possibly associated with calcaneal apophysitis/Sever's disease (n = 124).</p> <p>Interventions: Treatment factor 1 will be two types of shoe orthoses: a heel raise or prefabricated orthoses. Both of these interventions are widely available, mutually exclusive treatment approaches that are relatively low in cost. Treatment factor 2 will be a footwear prescription/replacement intervention involving a shoe with a firm heel counter, dual density EVA midsole and rear foot control. The alternate condition in this factor is no footwear prescription/replacement, with the participant wearing their current footwear.</p> <p>Outcomes: Oxford Foot and Ankle Questionnaire and the Faces pain scale.</p> <p>Discussion</p> <p>This will be a randomised trial to compare the efficacy of various treatment options for posterior heel pain in children that may be associated with calcaneal apophysitis also known as Sever's disease.</p> <p>Trial Registration</p> <p>Trial Number: ACTRN12609000696291</p> <p>Ethics Approval Southern Health: HREC Ref: 09271B</p

Animal-related factors associated with moderate-to-severe diarrhea in children younger than five years in western Kenya: A matched case-control study

Background Diarrheal disease remains among the leading causes of global mortality in children younger than 5 years. Exposure to domestic animals may be a risk factor for diarrheal disease. The objectives of this study were to identify animal-related exposures associated with cases of moderate-to-severe diarrhea (MSD) in children in rural western Kenya, and to identify the major zoonotic enteric pathogens present in domestic animals residing in the homesteads of case and control children. Methodology/Principal findings We characterized animal-related exposures in a subset of case and control children (n = 73 pairs matched on age, sex and location) with reported animal presence at home enrolled in the Global Enteric Multicenter Study in western Kenya, and analysed these for an association with MSD. We identified potentially zoonotic enteric pathogens in pooled fecal specimens collected from domestic animals resident at children’s homesteads. Variables that were associated with decreased risk of MSD were washing hands after animal contact (matched odds ratio [MOR] = 0.2; 95% CI 0.08–0.7), and presence of adult sheep that were not confined in a pen overnight (MOR = 0.1; 0.02–0.5). Variables that were associated with increased risk of MSD were increasing number of sheep owned (MOR = 1.2; 1.0–1.5), frequent observation of fresh rodent excreta (feces/urine) outside the house (MOR = 7.5; 1.5–37.2), and participation of the child in providing water to chickens (MOR = 3.8; 1.2–12.2). Of 691 pooled specimens collected from 2,174 domestic animals, 159 pools (23%) tested positive for one or more potentially zoonotic enteric pathogens (Campylobacter jejuni, C. coli, non-typhoidal Salmonella, diarrheagenic E. coli, Giardia, Cryptosporidium, or rotavirus). We did not find any association between the presence of particular pathogens in household animals, and MSD in children. Conclusions and significance Public health agencies should continue to promote frequent hand washing, including after animal contact, to reduce the risk of MSD. Future studies should address specific causal relations of MSD with sheep and chicken husbandry practices, and with the presence of rodents

Solar Radiation and Tidal Exposure as Environmental Drivers of Enhalus acoroides Dominated Seagrass Meadows

There is strong evidence of a global long-term decline in seagrass meadows that is widely attributed to anthropogenic activity. Yet in many regions, attributing these changes to actual activities is difficult, as there exists limited understanding of the natural processes that can influence these valuable ecosystem service providers. Being able to separate natural from anthropogenic causes of seagrass change is important for developing strategies that effectively mitigate and manage anthropogenic impacts on seagrass, and promote coastal ecosystems resilient to future environmental change. The present study investigated the influence of environmental and climate related factors on seagrass biomass in a large ≈250 ha meadow in tropical north east Australia. Annual monitoring of the intertidal Enhalus acoroides (L.f.) Royle seagrass meadow over eleven years revealed a declining trend in above-ground biomass (54% significant overall reduction from 2000 to 2010). Partial Least Squares Regression found this reduction to be significantly and negatively correlated with tidal exposure, and significantly and negatively correlated with the amount of solar radiation. This study documents how natural long-term tidal variability can influence long-term seagrass dynamics. Exposure to desiccation, high UV, and daytime temperature regimes are discussed as the likely mechanisms for the action of these factors in causing this decline. The results emphasise the importance of understanding and assessing natural environmentally-driven change when interpreting the results of seagrass monitoring programs

Lack of association between estrogen receptor β dinucleotide repeat polymorphism and autoimmune thyroid diseases in Japanese patients

BACKGROUND: The autoimmune thyroid diseases (AITDs), such as Graves' disease (GD) and Hashimoto's thyroiditis (HT), appear to develop as a result of complex interactions between predisposing genes and environmental triggers. Susceptibility to AITDs is conferred by genes in the human leukocyte antigen (HLA) and genes unlinked to HLA, including the CTLA-4 gene. Recently, estrogen receptor (ER) β, located at human chromosome 14q23-24.1, was identifed. We analyzed a dinucleotide (CA)n repeat polymorphism located in the flanking region of ERβ gene in patients with AITDs and in normal subjects. High heterozygosity makes this polymorphism a useful marker in the genetic study of disorders affecting female endocrine systems. We also correlated a ERβ gene microsatellite polymorphism with bone mineral density (BMD) in the distal radius and biochemical markers of bone turnover in patients with GD in remission. RESULTS: Fourteen different alleles were found in 133 patients with GD, 114 patients with HT, and 179 controls subjects. The various alleles were designated as allele(*)1 through allele(*)14 according to the number of the repeats, from 18 to 30. There was no significant difference in the distributions of ERβ alleles between patient groups and controls. Although recent study demonstrated a significant relation between a allele(*)9 in the ERβ gene and BMD in postmenopausal Japanese women, there were no statistically significant interaction between this allele and BMD in the distal radius, nor biochemical markers in patients with GD in remission. CONCLUSIONS: The present results do not support an association between the ERβ microsatellite marker and AITD in the Japanese population. We also suggest that the ERβ microsatellite polymorphism has at most a minor pathogenic importance in predicting the risk of osteoporosis as a complication of GD