Determining conditions for successful culture of multi-cellular 3D tumour spheroids to investigate the effect of mesenchymal stem cells on breast cancer cell invasiveness

Mesenchymal stem cells have been widely implicated in tumour development and metastases. Moving from the use of two-dimensional (2D) models to three-dimensional (3D) to investigate this relationship is critical to facilitate more applicable and relevant research on the tumour microenvironment. We investigated the effects of altering glucose concentration and the source of foetal bovine serum (FBS) on the growth of two breast cancer cell lines (T47D and MDA-MB-231) and human bone marrow-derived mesenchymal stem cells (hBM-MSCs) to determine successful conditions to enable their co-culture in 3D tumour spheroid models. Subsequently, these 3D multicellular tumour spheroids were used to investigate the effect of hBM-MSCs on breast cancer cell invasiveness. Findings presented herein show that serum source had a statistically significant effect on two thirds of the growth parameters measured across all three cell lines, whereas glucose only had a statistically significant effect on 6%. It was determined that the optimum growth media composition for the co-culture of 3D hBM-MSCs and breast cancer cell line spheroids was 1 g/L glucose DMEM supplemented with 10% FBS from source A. Subsequent results demonstrated that co-culture of hBMMSCs and MDA-MB-231 cells dramatically reduced invasiveness of both cell lines (F(1,4) = 71.465, p = 0.001) when embedded into a matrix comprising of growth-factor reduced base membrane extract (BME) and collagen

Defining the genetic susceptibility to cervical neoplasia - a genome-wide association study

Funding: MAB was funded by a National Health and Medical Research Council (Australia) Senior Principal Research Fellowship. Support was also received from the Australian Cancer Research Foundation. JL holds a Tier 1 Canada Research Chair in Human Genome Epidemiology. The Seattle study was supported by the following grants: NIH, National Cancer Institute grants P01CA042792 and R01CA112512. Cervical Health Study (from which the NSW component was obtained) was funded by NHMRC Grant 387701, and CCNSW core grant. The Montreal study was funded by the Canadian Institutes of Health Research (grant MOP-42532) and sample processing was funded by the Reseau FRQS SIDA-MI. The Swedish Research Council, the Swedish Foundation for Strategic Research, the ALF/LUA research grant in Gothenburg and Umeå, the Lundberg Foundation, the Torsten and Ragnar Soderberg’s Foundation, the Novo Nordisk Foundation, and the European Commission grant HEALTH-F2-2008-201865-GEFOS, BBMRI.se, the Swedish Society of Medicine, the KempeFoundation (JCK-1021), the Medical Faculty of Umeå University, the County Council of Vasterbotten (Spjutspetsanslag VLL:159:33-2007). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscriptPeer reviewedPublisher PDFPublisher PD

Setting sodium targets for pre-packaged foods in China — an exploratory study

IntroductionSetting sodium targets for pre-packaged food has been a priority strategy for reducing population sodium intake. This study aims to explore the attitudes and considerations of researchers and key stakeholders toward implementing such policy in China.MethodsAn exploratory study comprising a survey and a focus group discussion was conducted among 27 purposively selected participants including 12 researchers, 5 consumers, 4 administrators, 3 industry association representatives and 3 food producers. The survey/discussion covered the key questions considered when developing/promoting sodium targets. Free-text responses were manually classified and summarized using thematic analysis.ResultsTwo-thirds of the participants supported target-setting policy. Researchers and administrators were most supportive, and food producers and associations were least supportive. Adapted WHO food categorization framework was well accepted to underpin target-setting to ensure international comparability and applicability for Chinese products. Maximum values were the most agreed target type. The WHO benchmarks were thought to be too ambitious to be feasible given the current food supply in China but can be regarded as long-term goals. Initially, a reduction of sodium content by 20% was mostly accepted to guide the development of maximum targets. Other recommendations included implementing a comprehensive strategy, strengthening research, engaging social resources, establishing a systematic monitoring/incentive system, maintaining a fair competitive environment, and developing a supportive information system. Target-setting policy was acceptable by most stakeholders and should be implemented alongside strategies to reduce discretionary salt use.DiscussionOur findings provide detailed guidance for the Chinese government when developing a target-setting strategy. The methods and results of this study also provide meaningful references for other countries to set sodium targets for pre-packaged foods and implement other salt reduction strategies simultaneously

The role of deadenylation in the degradation of unstable mRNAs in trypanosomes

Removal of the poly(A) tail is the first step in the degradation of many eukaryotic mRNAs. In metazoans and yeast, the Ccr4/Caf1/Not complex has the predominant deadenylase activity, while the Pan2/Pan3 complex may trim poly(A) tails to the correct size, or initiate deadenylation. In trypanosomes, turnover of several constitutively-expressed or long-lived mRNAs is not affected by depletion of the 5′–3′ exoribonuclease XRNA, but is almost completely inhibited by depletion of the deadenylase CAF1. In contrast, two highly unstable mRNAs, encoding EP procyclin and a phosphoglycerate kinase, PGKB, accumulate when XRNA levels are reduced. We here show that degradation of EP mRNA was partially inhibited after CAF1 depletion. RNAi-targeting trypanosome PAN2 had a mild effect on global deadenylation, and on degradation of a few mRNAs including EP. By amplifying and sequencing degradation intermediates, we demonstrated that a reduction in XRNA had no effect on degradation of a stable mRNA encoding a ribosomal protein, but caused accumulation of EP mRNA fragments that had lost substantial portions of the 5′ and 3′ ends. The results support a model in which trypanosome mRNAs can be degraded by at least two different, partially independent, cytoplasmic degradation pathways attacking both ends of the mRNA

Natural experiments for the evaluation of place-based public health interventions:a methodology scoping review

Place-based public health evaluations are increasingly making use of natural experiments. This scoping review aimed to provide an overview of the design and use of natural experiment evaluations (NEEs), and an assessment of the plausibility of the as-if randomisation assumption. A systematic search of three bibliographic databases (Pubmed, Web of Science and Ovid-Medline) was conducted in January 2020 to capture publications that reported a natural experiment of a place-based public health intervention or outcome. For each, study design elements were extracted. An additional evaluation of as-if randomisation was conducted by twelve 12 of this paper’s authors who evaluated the same set of 20 randomly selected studies and assessed ‘as-if’ randomisation for each. 366 NEE studies of place-based public health interventions were identified. The most commonly used NEE approach was a Difference-in-Differences study design (25%), followed by before-after comparisons studies (23%) and regression analysis studies. 42% of NEEs had likely or probably as-if randomisation of exposure (the intervention), while for 25% this was implausible. An inter-rater agreement exercise indicated poor reliability of as-if randomisation assignment. Only about half of NEEs reported some form of sensitivity or falsification analysis to support inferences. NEEs are conducted using many different designs and statistical methods and encompass various definitions of a natural experiment, while it is questionable whether all evaluations reported as natural experiments should be considered as such. The likelihood of as-if randomisation should be specifically reported, and primary analyses should be supported by sensitivity analyses and/or falsification tests. Transparent reporting of NEE designs and evaluation methods will contribute to the optimum use of place-based NEEs

A rare mutation in SMAD9 associated with high bone mass identifies the SMAD-dependent BMP signalling pathway as a potential anabolic target for osteoporosis

Novel anabolic drug targets are needed to treat osteoporosis. Having established a large national cohort with unexplained high bone mass (HBM), we aimed to identify a novel monogenic cause of HBM and provide insight into a regulatory pathway potentially amenable to therapeutic intervention. We investigated a pedigree with unexplained HBM in whom previous sequencing had excluded known causes of monogenic HBM. Whole exome sequencing identified a rare (minor allele frequency 0.0023), highly evolutionarily conserved missense mutation in SMAD9 (c.65T>C, p.Leu22Pro) segregating with HBM in this autosomal dominant family. The same mutation was identified in another two unrelated individuals both with HBM. In silico protein modeling predicts the mutation severely disrupts the MH1 DNA-binding domain of SMAD9. Affected individuals have bone mineral density (BMD) Z-scores +3 to +5, mandible enlargement, a broad frame, torus palatinus/mandibularis, pes planus, increased shoe size, and a tendency to sink when swimming. Peripheral quantitative computed tomography (pQCT) measurement demonstrates increased trabecular volumetric BMD and increased cortical thickness conferring greater predicted bone strength; bone turnover markers are low/normal. Notably, fractures and nerve compression are not found. Both genome-wide and gene-based association testing involving estimated BMD measured at the heel in 362,924 white British subjects from the UK Biobank Study showed strong associations with SMAD9 (P-GWAS = 6 x 10(-16); P-GENE = 8 x 10(-17)). Furthermore, we found Smad9 to be highly expressed in both murine cortical bone-derived osteocytes and skeletal elements of zebrafish larvae. Our findings support SMAD9 as a novel HBM gene and a potential novel osteoanabolic target for osteoporosis therapeutics. SMAD9 is thought to inhibit bone morphogenetic protein (BMP)-dependent target gene transcription to reduce osteoblast activity. Thus, we hypothesize SMAD9 c.65T>C is a loss-of-function mutation reducing BMP inhibition. Lowering SMAD9 as a potential novel anabolic mechanism for osteoporosis therapeutics warrants further investigation. (c) 2019 The Authors. Journal of Bone and Mineral Research published by American Society for Bone and Mineral Research

Beyond NIMBYs and NOOMBYs:what can wind farm controversies teach us about public involvement in hospital closures?

Background Many policymakers, researchers and commentators argue that hospital closures are necessary as health systems adapt to new technological and financial contexts, and as population health needs in developed countries shift. However closures are often unpopular with local communities. Previous research has characterised public opposition as an obstacle to change. Public opposition to the siting of wind farms, often described as NIMBYism (Not In My Back Yard), is a useful comparator issue to the perceived NOOMBYism (Not Out Of My Back Yard) of hospital closure protestors. Discussion The analysis of public attitudes to wind farms has moved from a fairly crude characterisation of the ‘attitude-behaviour gap’ between publics who support the idea of wind energy, but oppose local wind farms, to empirical, often qualitative, studies of public perspectives. These have emphasised the complexity of public attitudes, and revealed some of the ‘rational’ concerns which lie beneath protests. Research has also explored processes of community engagement within the wind farm decision-making process, and the crucial role of trust between communities, authorities, and developers. Summary Drawing on what has been learnt from studies of opposition to wind farms, we suggest a range of questions and approaches to explore public perspectives on hospital closure more thoroughly. Understanding the range of public responses to service change is an important first step in resolving the practical dilemma of effecting health system transformation in a democratic fashion

Investigating the Effects of Indirect Coculture of Human Mesenchymal Stem Cells on the Migration of Breast Cancer Cells: A Systematic Review and Meta-Analysis

Purpose: Breast cancer is the most diagnosed cancer and the leading cause of cancer death in women globally, and mesenchymal stem cells have been widely implicated in tumour progression. This systematic review and meta-analysis seeks to identify and summarise existing literature on the effects of human mesenchymal stem cells (hMSCs) on the migration of breast cancer cells (BCCs) in vitro, to determine the direction of this relationship according to existing research and to identify the directions for future research. Methods: A systematic literature search was conducting using a collection of databases, using the following search terms: in vitro AND mesenchymal stem cells AND breast cancer. Only studies that investigated the effects of human, unmodified MSCs on the migration of human, unmodified BCCs in vitro were included. Standardised mean differences (SMDs) were calculated to determine pooled effect sizes. Results: This meta-analysis demonstrates that hMSCs (different sources combined) increase the migration of both MDA-MB-231 and MCF-7 cell lines in vitro (SMD = 1.84, P  = .03 and SMD = 2.69, P  < .00001, respectively). Importantly, the individual effects of hMSCs from different sources were also analysed and demonstrated that MSCs derived from human adipose tissue increase BCC migration (SMD = 1.34, P  = .0002) and those derived from umbilical cord increased both MDA-MB-231 and MCF-7 migration (SMD = 3.93, P  < .00001 and SMD = 3.01, P  < .00001, respectively). Conclusions: To our knowledge, this is the first systematic review and meta-analysis investigating and summarising the effects of hMSCs from different sources on the migration of BCCs, in vitro

An exploration of the role of exercise in modulating breast cancer progression in vitro (a systematic review and meta-analysis)

Background: Breast cancer is the most prevalent cancer in women worldwide. In the UK, approximately 5% of all breast cancers are already metastatic at the time of diagnosis. An abundance of literature shows exercise can have beneficial effects on the outcome and prognosis of breast cancer patients, yet the molecular mechanisms remain poorly understood. There are several in vitro models that aim to recapitulate the response of breast cancer to exercise in vivo: this systematic review and meta-analysis summarises the existing literature. Methods: The following search terms were used to conduct a systematic literature search using a collection of databases (last search performed May 2020): 'in vitro' and 'exercise' and 'breast cancer'. Only studies that investigated the effects of exercise on breast cancer in vitro were included. Standardised mean differences (SMD) were calculated to determine pooled effect sizes. Results: This meta-analysis has successfully demonstrated that various identified exercise interventions on breast cancer cells in vitro significantly reduced breast cancer cell viability, proliferation, and tumourigenic potential (SMD: -1.76, p = 0.004, SMD: -2.85, p = 0.003 and SMD: -3.15, p = 0.0008, respectively). A clear direction of effect was found with exercise on breast cancer cell migration in vitro, however this effect was not significant (SMD: -0.62, p = 0.317). Conclusion: To our knowledge, this is the first meta-analysis and systematic review investigating and summarising literature on exercise and breast cancer in vitro, highlighting models used and priority areas for future research focus