A novel method for sample preparation of fresh lung cancer tissue for proteomics analysis by tumor cell enrichment and removal of blood contaminants

<p>Abstract</p> <p>Background</p> <p>In-depth proteomics analyses of tumors are frequently biased by the presence of blood components and stromal contamination, which leads to large experimental variation and decreases the proteome coverage. We have established a reproducible method to prepare freshly collected lung tumors for proteomics analysis, aiming at tumor cell enrichment and reduction of plasma protein contamination. We obtained enriched tumor-cell suspensions (ETS) from six lung cancer cases (two adenocarcinomas, two squamous-cell carcinomas, two large-cell carcinomas) and from two normal lung samples. The cell content of resulting ETS was evaluated with immunocytological stainings and compared with the histologic pattern of the original specimens. By means of a quantitative mass spectrometry-based method we evaluated the reproducibility of the sample preparation protocol and we assessed the proteome coverage by comparing lysates from ETS samples with the direct lysate of corresponding fresh-frozen samples.</p> <p>Results</p> <p>Cytological analyses on cytospin specimens showed that the percentage of tumoral cells in the ETS samples ranged from 20% to 70%. In the normal lung samples the percentage of epithelial cells was less then 10%. The reproducibility of the sample preparation protocol was very good, with coefficient of variation at the peptide level and at the protein level of 13% and 7%, respectively. Proteomics analysis led to the identification of a significantly higher number of proteins in the ETS samples than in the FF samples (244 vs 109, respectively). Albumin and hemoglobin were among the top 5 most abundant proteins identified in the FF samples, showing a high contamination with blood and plasma proteins, whereas ubiquitin and the mitochondrial ATP synthase 5A1 where among the top 5 most abundant proteins in the ETS samples.</p> <p>Conclusion</p> <p>The method is feasible and reproducible. We could obtain a fair enrichment of cells but the major benefit of the method was an effective removal of contaminants from red blood cells and plasma proteins resulting in larger proteome coverage compared to the direct lysis of frozen samples. This sample preparation method may be successfully implemented for the discovery of lung cancer biomarkers on tissue samples using mass spectrometry-based proteomics.</p

Fetal Exposure to PCBs and Their Hydroxylated Metabolites in a Dutch Cohort

Polychlorinated biphenyls (PCBs) are still the most abundant pollutants in wildlife and humans. Hydroxylated PCB metabolites (OH-PCBs) are known to be formed in humans and wildlife. Studies in animals show that these metabolites cause endocrine-related toxicity. The health effects in humans have not yet been evaluated, especially the effect on the fetus and newborn. The aim of this study is to measure the levels of PCBs and OH-PCBs in maternal and cord blood samples in a population with background levels of PCBs. We analyzed 51 maternal and corresponding cord blood samples in the northern part of the Netherlands. The PCB concentrations in maternal plasma ranged from 2 to 293 ng/g lipid, and OH-PCB concentrations from nondetectable (ND) to 0.62 ng/g fresh weight. In cord plasma, PCB concentrations were 1–277 ng/g lipid, and OH-PCB concentrations, ND to 0.47 ng/g fresh weight. The cord versus maternal blood calculated ratio was 1.28 ± 0.56 for PCBs and 2.11 ± 1.33 for OH-PCBs, expressed per gram of lipid. When expressed per gram fresh weight, the ratios are 0.32 ± 0.15 and 0.53 ± 0.23 for PCBs and OH-PCBs, respectively. A significant correlation between the respective maternal and cord levels for both PCBs and OH-PCBs was found. Our results indicate that OH-PCBs and PCBs are transferred across the placenta to the fetus in concentrations resulting in levels of approximately 50 and 30%, respectively, of those in maternal plasma. More research in humans is needed to evaluate potential negative effects of these endocrine disruptors on the fetus

Balancing extensive ambition and a context overflowing with opportunities and demands: A grounded theory on stress and recovery among highly educated working young women entering male-dominated occupational areas

Several factors underline the issue of stress-related health among young highly educated women. Major societal changes might provide more new challenges with considerably changed and expanded roles than were expected by earlier generations, especially among women. The quantity of young women with higher education has also increased threefold in Sweden in less than two decades and there are a growing number of young women that hereby break with traditional gender positions and enter new occupational areas traditionally dominated by men. The research questions in the present study were: “What is the main concern, regarding stress and recovery, among young highly educated working women breaking with traditional gender positions and entering male-dominated occupational areas?” and “How do they handle this concern?” We conducted open-ended interviews with 20 informants, aged 23-29 years. The results showed that the synergy between highly ambitious individuals and a context overflowing with opportunities and demands ended up in the informants’ constantly striving to find a balance in daily life (main concern). This concern refers to the respondents experiencing a constant overload of ambiguity and that they easily became entangled in a loop of stress and dysfunctional coping behavior, threatening the balance between stress and sufficient recovery. In order to handle this concern, the respondents used different strategies in balancing extensive ambition and a context overflowing with opportunities and demands (core category). This preliminary theoretical model deepens our understanding of how the increasing numbers of highly educated young women face complex living conditions endangering their possibility of maintaining health and work ability

Ancestral SARS-CoV-2-specific T cells cross-recognize the Omicron variant

The emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron (B.1.1.529) variant of concern (VOC) has destabilized global efforts to control the impact of coronavirus disease 2019 (COVID-19). Recent data have suggested that B.1.1.529 can readily infect people with naturally acquired or vaccine-induced immunity, facilitated in some cases by viral escape from antibodies that neutralize ancestral SARS-CoV-2. However, severe disease appears to be relatively uncommon in such individuals, highlighting a potential role for other components of the adaptive immune system. We report here that SARS-CoV-2 spike-specific CD4+ and CD8+ T cells induced by prior infection or BNT162b2 vaccination provide extensive immune coverage against B.1.1.529. The median relative frequencies of SARS-CoV-2 spike-specific CD4+ T cells that cross-recognized B.1.1.529 in previously infected or BNT162b2-vaccinated individuals were 84% and 91%, respectively, and the corresponding median relative frequencies for SARS-CoV-2 spike-specific CD8+ T cells were 70% and 92%, respectively. Pairwise comparisons across groups further revealed that SARS-CoV-2 spike-reactive CD4+ and CD8+ T cells were functionally and phenotypically similar in response to the ancestral strain or B.1.1.529. Collectively, our data indicate that established SARS-CoV-2 spike-specific CD4+ and CD8+ T cell responses, especially after BNT162b2 vaccination, remain largely intact against B.1.1.529

Insulin resistance:Genetic associations with depression and cognition in population based cohorts

We are grateful to the families who took part in GS:SFHS, general practitioners and the Scottish School of Primary Care for their help in recruitment, and the whole GS:SFHS team that includes academic researchers, clinic staff, laboratory technicians, clerical workers, IT staff, statisticians and research managers. The research reported here, and the genotyping of GS:SFHS samples was funded by the Wellcome Trust, (Wellcome Trust Strategic Award ‘STratifying Resilience and Depression Longitudinally’ (STRADL) Reference 104036/Z/14/Z) and by the Medical Research Council. SF acknowledges support from the National Institute of Mental Health, USA (R01MH113619; R01MH116147) and the consortium for Psychopathology and Allostatic load across the Life Span (PALS; https://www.pals-network.org) AMM acknowledges the financial support received from the Dr. Mortimer and Theresa Sackler Foundation. IJD and AMM are members of The University of Edinburgh Centre for Cognitive Ageing and Cognitive Epidemiology, part of the cross council Lifelong Health and Wellbeing Initiative (MR/K026992/1). Generation Scotland received core support from the Chief Scientist Office of the Scottish Government Health Directorates (CZD/16/6) and the Scottish Funding Council (HR03006). Funding from the Biotechnology and Biological Sciences Research Council and Medical Research Council is gratefully acknowledged.Peer reviewedPublisher PD

Children’s and adolescents’ rising animal-source food intakes in 1990–2018 were impacted by age, region, parental education and urbanicity

Animal-source foods (ASF) provide nutrition for children and adolescents’ physical and cognitive development. Here, we use data from the Global Dietary Database and Bayesian hierarchical models to quantify global, regional and national ASF intakes between 1990 and 2018 by age group across 185 countries, representing 93% of the world’s child population. Mean ASF intake was 1.9 servings per day, representing 16% of children consuming at least three daily servings. Intake was similar between boys and girls, but higher among urban children with educated parents. Consumption varied by age from 0.6 at <1 year to 2.5 servings per day at 15–19 years. Between 1990 and 2018, mean ASF intake increased by 0.5 servings per week, with increases in all regions except sub-Saharan Africa. In 2018, total ASF consumption was highest in Russia, Brazil, Mexico and Turkey, and lowest in Uganda, India, Kenya and Bangladesh. These findings can inform policy to address malnutrition through targeted ASF consumption programmes.publishedVersio

Incident type 2 diabetes attributable to suboptimal diet in 184 countries

The global burden of diet-attributable type 2 diabetes (T2D) is not well established. This risk assessment model estimated T2D incidence among adults attributable to direct and body weight-mediated effects of 11 dietary factors in 184 countries in 1990 and 2018. In 2018, suboptimal intake of these dietary factors was estimated to be attributable to 14.1 million (95% uncertainty interval (UI), 13.8–14.4 million) incident T2D cases, representing 70.3% (68.8–71.8%) of new cases globally. Largest T2D burdens were attributable to insufficient whole-grain intake (26.1% (25.0–27.1%)), excess refined rice and wheat intake (24.6% (22.3–27.2%)) and excess processed meat intake (20.3% (18.3–23.5%)). Across regions, highest proportional burdens were in central and eastern Europe and central Asia (85.6% (83.4–87.7%)) and Latin America and the Caribbean (81.8% (80.1–83.4%)); and lowest proportional burdens were in South Asia (55.4% (52.1–60.7%)). Proportions of diet-attributable T2D were generally larger in men than in women and were inversely correlated with age. Diet-attributable T2D was generally larger among urban versus rural residents and higher versus lower educated individuals, except in high-income countries, central and eastern Europe and central Asia, where burdens were larger in rural residents and in lower educated individuals. Compared with 1990, global diet-attributable T2D increased by 2.6 absolute percentage points (8.6 million more cases) in 2018, with variation in these trends by world region and dietary factor. These findings inform nutritional priorities and clinical and public health planning to improve dietary quality and reduce T2D globally.publishedVersio