Leishmania genus. There is no human vaccine available and it is considered by many studies as apotential effective tool for disease control. To discover novel antigens, computational programs have been used in reverse vaccinology strategies. In this work, we developed a validation antigen approach that integrates prediction of B and T cell epitopes, analysis of Protein-Protein Interaction (PPI) networks and metabolic pathways. We selected twenty candidate proteins from Leishmania tested in murine model, with experimental outcome published in the literature. The predictions for CD4+ and CD8+ T cell epitopes were correlated with protection in experimental outcomes. We also mapped immunogenic proteins on PPI networks in order to find Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways associated with them. Our results suggest that non-protective antigens have lowest frequency of predicted T CD4+ and T CD8+ epitopes, compared with protective ones. T CD4+ and T CD8+ cells are more related to leishmaniasis protection in experimental outcomes than B cell predicted epitopes. Considering KEGG analysis, the proteins considered protective are connected to nodes with few pathways, including those associated with ribosome biosynthesis and purine metabolism.Leishmaniasis is a wide-spectrum disease caused by parasites from  Leishmania genus. It is prevalent in Americas, Europe, Africa and Asia. Overall, human infection is caused by at least 20 species whose vectors are phlebotomine sandflies [Leishmaniasis is a wide-spectrum disease caused by parasites from andflies . Althoug+ cells and T CD8+ cells epitopes and subcellular localization. They are valuable in a pre-screening evaluation for vaccine targets and searching for diagnostic markers.In the advent of reverse vaccinology, in the latest years, a great effort has been made by bioinformaticians in order to provide epitopes predictors programs. Currently, it is possible to scan entire genomes searching for immunogenic epitopes and then select promising proteins for vaccine development. The bottleneck in this workflow analysis is the validation of predictions for protozoan parasites. Many predictors are available for B cells, T CD4The building of protein-protein interaction (PPI) networks may give some insights to understand the biological role of these targets, and so might be a valuable asset in vaccine development. These networks are constituted by nodes that correspond to proteins, connected by edges, representing the interactions between two connected proteins. With PPI networks, we can have an overview of protein relationships and notice those with high connections . Hub proteins tend to have essential role in the parasite metabolism and might be good candidates to vaccinal and drug target ,4.Leishmania vaccine research, we developed an approach that integrates prediction of B and T cell epitopes, analysis of PPI networks and metabolic pathways. With the aim of validating this methodology, we selected Leishmania proteins tested as vaccine candidates in murine model, with experimental outcome (EO) published in the literature. After predicting epitopes in the selected proteins using specific computational programs, we correlated the predictions for T CD4+ and T CD8+ cells with protection in EO. Finally, we mapped the immunogenic proteins on PPI networks in order to find Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways associated with them.To support Leishmania species that were used in studies aiming the vaccine development against these parasites.Through the use of text mining technics from Pubmed website that included, but was not restricted to, categorization and entity extraction, we were able to identify and select 20 proteins from six different It is important to highlight that, for each one of those proteins, a specific MySQL ID was assigned to link GI accession number and TriTrypDB specific ID. Based on the results published, the EO was categorized into: (a) “no protection” (nine proteins); (b) “partial protection” (five proteins); and (c) “protection” (six proteins). The accession numbers of these proteins are depicted in With the purpose of selecting potential immunogenic epitopes in the selected experimental dataset, Structured Query Language (SQL) statements were used. The results obtained in terms of number of predicted binding Major Histocompatibility Complex (MHC) class I and II epitopes, B cell epitopes and subcellular locations are detailed in Specifically regarding the epitopes capacity to bind MHC class I, MHC class II and epitopes for B cell activation, considering a path from “no protection” to “protection” groups, a gradual increase of the number of predicted epitopes for T cells and B cells was observed.+ epitopes obtained from NetMHC and NetCTL; and (b) prediction for B cell epitopes obtained from AAP12, BCPred12 and BepiPred. The consensus predictions were obtained overlapping identical predictions made by different methodologies.To evaluate the possible association between the number of predicted epitopes (NPE) for B and T cells, and the EO of selected proteins, the following consensus predictions were produced: (a) prediction for T CD8Leishmania that cause CL and VL were correlated with NPE (+ T cells (p < 0.05) (data not shown). On the other hand, for CL analyses, it was observed significant correlation between EO and NPE for T CD4+, T CD8+ and B cells, as shown in +, T CD8+ and B cells), a significant correlation exists between NPE specific to CD4+ and CD8+ T cells with r = 0.752/p < 0.05 and r = 0.793/p < 0.05. In addition, a weak association with B cell predicted epitopes (r = 0.515/p < 0.05) was observed. In other words, non-protective antigens have lowest frequency of predicted T CD4+ and T CD8+ epitopes, compared with protective ones.In regards to CM analysis  approaches were applied to visualize the potential associations determined through Spearman r and Chi-square distance, respectively. Firstly, the disease (leishmaniasis) was stratified into cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL) and the EO of antigens from with NPE a. Regardanalysis c, consid+ and T CD8+, p < 0.05  and haplotype b binders  identified in the “protection” group were superior to those ones identified in the “no protection” group. In summary, our results indicate that epitopes for MHC class I and II haplotype b and d are more frequent in the success antigens tested for vaccine development. In addition, a detailed analysis in which MHC class I and II haplotypes were individually investigated revealed a strong association between NPE from MHC class I haplotype d and EO (p < 0.05 and r = 0.855) that is not observed for MHC class II haplotype d and b.To hypothesize possible reasons linked with vaccine success or failure, an evaluation of allele-specific affinity (MHC I and II) was investigated. As illustrated in r = −8.55) between the number of connected pathways and EO of the selected proteins (We chose Cytoscape to model PPI networks using data from STRING v.10. proteins c.Leishmania species), there are not strong and validated platforms to identify promising antigens for Leishmania vaccines [Nowadays, many computational methodologies have been described for epitope predictions of bacterial, fungal and others microorganisms. For protozoan , so there is no standardization to appoint if a vaccine indicates protection or not . We trie+ and T CD8+ cells could be associated with protection in EO. In this regard, our results revealed that there are strong correlation and association between predicted epitopes and the EO. The inertia values show the powerful association between the variables. The CM dimension 1 represents the highest inertia allowing the interpretation of the results in the first dimension. T CD4+ and T CD8+ predicted epitopes versus EO show higher inertia values when compared to B cells predicted epitopes emphasizing that epitopes for CD4+ and CD8+ T cells are crucial for Leishmania vaccines success. We used all available human and mouse alleles to restrict the epitopes allowing enhance the assertive prediction. This analysis is useful to identify conserved epitopes that can bind various alleles of MHC appointing for rare and promising epitopes. The in silico analyses and in vivo validation of epitopes demonstrates that some algorithms may be important tools for the identification of epitopes, and consequently of immunogenic proteins. The algorithm NetCTL version 1.2 makes prediction of peptide–MHC class I binding, proteasomal C terminal cleavage, both using artificial neural networks, and TAP transport efficiency using weight matrix. The tree predictions are then integrated [Regarding the platform validation, we performed the epitope mapping using ad-hoc algorithms. Our results suggest that antigens with more predicted epitopes for T CD4tegrated . Anothertegrated . To perftegrated . Then wetegrated ,30,31. Ftegrated . The intL. major, obtaining 26 potential epitopes through prediction consensus. Fourteen of them revealed to be immunogenic epitopes that were capable to stimulate T cells to produce IFN-γ. Other studies employing computational predictions in specific Leishmania proteins have shown that it is quite possible to use combined algorithms in epitopes searching that could be validated by in vivo experiments [+ epitopes in 63 L. braziliensis proteins, demonstrating a cytotoxic activity of some predicted epitopes in Leishmania infected mice. Recently, in silico methods for linear epitope predictions  were combined with molecular modeling to identify potential epitopes in the whole L. braziliensis proteome. Therefore, the pipeline was validated based on stimulation of human peripheral blood mononuclear cells (PBMCs) proliferation. The results obtained after the in vitro assays showed that six of ten selected epitopes could be classified as potentially immunogenic [Herrera-Najera et al.  performeeriments . Duarte eriments  developeunogenic .Leishmania proteins are more immunogenic and considered better targets for vaccine development [L. amazonensis used in this work) and the large number of linear epitopes. Nevertheless, our results of epitope prediction indicate a higher assertive and successful prediction, so it can be a useful approach for vaccine development against leishmaniasis.The role of B cell epitopes still hasroom for discussion since there is no consensus if immunoglobulins could be associated with resistance or susceptibility in leishmaniasis ,38. On telopment ,40. ThisLeishmania spp. [Leishmania parasites. Our analyses showed that many of the selected antigens do not have any KEGG pathway associated to them, but, instead, are connected to proteins that are part of some pathway. Pathways associated with ribosome biosynthesis, purine metabolism and metabolic processes are present in “protection” group networks. Ribosome related proteins were considered relevant molecules during infection, since in some circumstances they can modulate cell activities and cytokine release. Many works associated these pathways to immune response [Leishmania major gene considered to be homologous to the mammalian ribosomal protein S3a. This ribosomal protein can be found in many other Leishmania species such as L. infantum, L. amazonensis, and L. mexicana. The article authors suggested that this protein could participate in the Th1/Th2 immune response balance during leishmaniasis [Leishmania acidic ribosomal proteins, also called P-type proteins [L. donovani, where the majority of virulent proteins (secreted proteins) belong mainly to metabolic and biosynthesis processes [Leishmania knocked-out for protein kinases and phosphatases possible involved in parasite metabolism regulation were generated. After this process, in many cases highly attenuated or completely avirulent parasites could be observed [Leishmania major mutant lacking the regulatory subunit of the Ca2+/calmodulin-dependent serine/threonine-specific phosphatase. This modified Leishmania grew normally at 27 °C. However, this parasite lost viability when exposed to 34 °C [Leishmania major parasites. These knocked-out parasites exhibited slower growth than wild-type parasites and were unable to survive or replicate in macrophages in vitro. These parasites were not capable of inducing disease or establish infection in mice in vivo [Leishmania virulence, since down regulation of metabolic genes causes latency of many Leishmania species. One possible application of these attenuated or avirulent parasites could be in whole parasites vaccines. Carter et al. [Leishmania survival. Surprisingly, our findings show that proteins associated to protection are connected to few pathways when compared to proteins that are not protective.To better understand the biological importance of vaccine candidates, we proposed the use of PPI networks enriched with KEGG pathways information. It is well known that some proteins are essential for specific biological processes of nia spp. ,42. In tresponse . Cordeirmaniasis . Soto anproteins . Anotherproteins . Our finrocesses . To checobserved . Nadererto 34 °C . Target  in vivo . In addi in vivo  have shor et al.  have notIn summary, in this work, we proposed and validated a computational approach regarding epitope prediction, topological structure and pathway analyses to drive a rational vaccine design against leishmaniasis.Leishmania proteins tested in murine model, with EO published in the literature, were selected. Studies describing vaccine effectiveness after challenge with Leishmania spp. were preferentially chosen. Bearing in mind that there is no standardization of the protection concept and that there is variation of results in the literature, it was necessary to create three categories as already described. According to our categorization, “no protection” group includes antigens that promote no or slight reduction of parasite burden or lesions after Leishmania challenge. The “protection” group includes antigens that promoted significantly strong reduction of the parasite burden or lesions showing strong immune response to Leishmania antigens. Thus, the term partial protection was used to classify antigens that are between a potential protection and no protection at all. The “partial protection” group comprises proteins that could slightly reduce the parasite burden and/or lesions more than “no protection” group. In addition, these proteins can elicit some immune response which results in ineffective protection. It is important to highlight that the influence of adjuvants was not taken into account for the categorization, thus only the response of antigens tested alone was chosen to categorize the groups. Twenty candidates from Old and New World Leishmania species were categorized according with their experimental results in the following groups: (a) “no protection”; (b) “partial protection”; and (c) “protection”. The selected experimentally validated data included information from proteins of L. amazonensis, L. braziliensis, L. major, L. mexicana, L. donovani and L. infantum that were subsequently used in this study to corroborate the in silico bioinformatics predictions. The selected antigens are described in Leishmania species were obtained from TriTrypDB (Kinetoplastid Genomics Resource) and the sequence of L. amazonensis was downloaded from http://bioinfo08.ibi.unicamp.br/leishmania/ [The predicted proteome sequences of dermatotropic and viscerotropic shmania/ . Detaile+ and T CD8+ epitopes, B cell epitopes and subcellular localization of proteins.All proteomic data used in this work were screened in order to predict T CD4+ epitope prediction (MHC class I binding epitopes), algorithms NetCTL [+ epitopes, NetMHCII [http://www.uniprot.org/) and IEDB  (http://www.iedb.org/). The analytical workflow used in this study is presented in For T CD8s NetCTL ,53,54 ans NetCTL ,56,57 weNetMHCII ,58 was uNetMHCII  and BCPRNetMHCII ,30,31 weNetMHCII ,55,56,57NetMHCII  describiTo carry out the predictions, the algorithms were parameterized for eukaryotic genomes. For MHC class I binding epitopes, predictions for 12 human supertypes and seven mice alleles were performed. The following alleles were used: A1, A2, A3, A24, A26, B7, B8, B27, B39, B44, B58, B62, H2-Db, Dk-H2, H2-Dd, H2-Kb, H2-Kd, Kk-H2, and H2-Ld. Concerning MHC class II binding epitopes, we used 14 human alleles and three mice alleles bringing the total of different alleles to 17, as follows: HLA-DRB1*01:01, HLA-DRB1*03:01, HLA-DRB1*04:01, HLA-DRB1*04:04, HLA-DRB1*04:05, HLA-DRB1*07:01, HLA-DRB1*08:02, HLA-DRB1*09:01, HLA-DRB1*11:01, HLA-DRB1*13:02, HLA-DRB1*15:01, HLA-DRB3*01:01, HLA-DRB4*01:01, HLA-DRB5*01:01, H2-IAs, H2-IAd and H2-IAb. http://www.mysql.com). The use of a database system in this work represents a crucial step that allows the integration of the results from all predictors and a way of getting a data receptacle or conceptual repository from which is possible to extract data correlation, helping in the identification of target proteins. The MySQL GUI Tools (http://dev.mysql.com/downloads/gui-tools/5.0.html) were used as a graphical user interface for our MySQL database. The relational model was built in MySQL Workbench (http://wb.mysql.com). To extract, parse and load data into database, specific Perl scripts were developed using DBI (the Perl interface to databases) and BioPerl modules. The relational schema is presented in Taking into account the great amount of data generated by the algorithms, we constructed a relational database using MySQL as Relational Database Management System (RDBMS) . The networks were built using the target proteins as central nodes and expanding them with the first neighbors, using 500 as maximum value of interactions. From STRING, we obtained the PPI network data and KEGG [p < 0.05) of proteins involved in these networks. Then we built the protein networks using Cytoscape [In this topic, two groups were created: P (protection), aggregating proteins classified as “partial protection” and “protection”, and NP (no protection) for proteins classified as “no protection”. We entered these proteins into STRING v.10  looking and KEGG  pathway ytoscape , adding p < 0.05.The analyses were performed using SPSS version 20 . Association coefficients were determined using Spearman two tailed test and correspondence analysis were determined by Chi-squared, statistical significance was considered when + and CD8+ T cells are more related to leishmaniasis protection in EO than B cells. For a deeper analysis, we also used PPI networks enriched with KEGG pathways information. According to our results, proteins associated to protection are connected to few pathways when compared to proteins classified as “no protection”. In addition, analysis of PPIs and KEGG pathways associated to proteins from “protection” group corroborate the idea already published in the literature that reverse vaccinology approaches are able to identify proteins related to pathogenicity of infectious agents, helping researchers to understand virulence mechanisms and how immune responses from hosts are able to fight them. Obtained results may be helpful in discovering new potential antigens using computational approaches.In this work, we validated a computational approach regarding epitope prediction, topological structure and pathway analyses to drive a rational vaccine design against leishmaniasis, using antigens tested in murine model described in literature. Our results suggest that CD4


The association of fasting glucose with the risk of death according to sex and age remains unclear, and insufficient information is available on sex- and age-specific glucose concentrations within ethnic groups. This study analyzed a sample of 12,455,361 Korean adults who participated in health examinations during 2001–2004, and were followed up until 2013. Men had 3.0 mg/dL (0.167 mmol/L) higher mean glucose concentrations than women (94.7 vs. 91.7 mg/dL), although women over 73 years had higher levels. For glucose levels of 100–199 mg/dL, each 18 mg/dL (1 mmol/L) increase in fasting glucose increased mortality by 13% . In individuals with fasting glucose levels of 100–125 mg/dL, each 18 mg/dL increase in fasting glucose was associated with a 30% increase in the risk for mortality  in those aged 18–34 years, a 32% increase  in those aged 35–44 years, and a 10% increase  in those aged 75–99 years. The fasting glucose levels associated with the lowest mortality were 80–94 mg/dL regardless of sex and age. Prediabetes (100–125 mg/dL) was associated with higher mortality. The associations of hyperglycemia with mortality were stronger at younger ages. However, the current cut-points, for example, 100 or 110 mg/dL for prediabetes, are somewhat arbitrarily chosen due to inconsistent results in prior studies6. Identifying proper cut-points for fasting glucose and overall mortality would help inform public and clinical actions to prevent premature death from health issues related to hyperglycemia and hypoglycemia.Diabetes is a worldwide epidemic. Fasting glucose measurement remains the main test for the diagnosis of diabetes and prediabetes, and it is a key indicator of future diabetes and cardiovascular disease9 and may differ by sex10, the sex- and age-specific associations of fasting glucose with all-cause mortality have rarely been examined. It is unclear whether variation according to age and sex is present in the association between fasting glucose and the risk of death and/or the range of fasting glucose associated with a minimal risk of death12. Additionally, age- and sex-specific fasting glucose concentrations may differ across ethnic and regional groups13, meaning that the association of fasting glucose levels with diseases and mortality may also vary by ethnicity14. Furthermore, detailed sex- and age-specific fasting glucose concentrations in the Korean population have not been reported.Although fasting glucose levels are generally known to increase with advancing ageThrough a large prospective cohort study that included 12.8 million participants, we set out to elucidate whether the association between fasting glucose levels and all-cause mortality varies by sex and age, and to estimate sex- and age-specific levels of fasting glucose associated with minimal mortality. Precise estimates could help determine cut points for the management of hyperglycemia and hypoglycemia. Additionally, detailed estimates of mean (and median) concentrations of fasting glucose according to sex and age could help in the management of sex- and age- specific aspects of glucose metabolism and related metabolic disorders.15. Individuals with missing information on serum glucose, blood pressure, total cholesterol, and body-mass index (BMI) were excluded , as were 3,665 persons with extremely low weight (<30 kg), high BMI (≥50 kg/m2), or short stature , and another 210 missing date of the health examination. Persons with self-reported diabetes  were also excluded. The final study population included 12,455,361 participants, followed up until the 31st December 2013 through the NHIS database of beneficiary status, with participants’ deaths being ascertained from the Resident Register of Korea15. All data are collected and maintained by the NHIS according to several Korean laws. For the research in accord with the conditions documented at Korean laws, health examination data can be provided to the researchers by the NHIS without specific informed consents from the participants. This study was approved by Institutional Review Board of Catholic Kwandong University, Republic of Korea. Anonymized data were provided to the authors by the NHIS, and they were only available through a specific computer within the NHIS headquarters.The National Health Insurance Service (NHIS) provides compulsory health insurance that covers 97% of the Korean population. The Korean Metabolic Risk factor (KOMERIT) study was designed primarily to assess the risk of death associated with metabolic risk factors, and includes 12,845,017 NHIS beneficiaries aged 18–99 years who participated in routine health examinations during 2001–200415. BMI was calculated by weight in kilograms divided by the square of height in meters (kg/m2). Smoking history, alcohol use, and known diabetes status were self-reported via questionnaire. Health examination and data collection followed a standard protocol, the Health Examination Practice Guide, officially registered by the Ministry of Health and Welfare . Eight (or seven) groups , were used in additional analysis to show the difference in hazard ratios (HRs) for sex- and age-specific groups. The glucose category with the lowest mortality in sex and age-adjusted analysis was used as the referent. Fasting glucose level was also analyzed as a continuous variable and HRs per 18 mg/dL (1 mmol/L), rather than 10 mg/dL, increase in fasting glucose were calculated for between-study comparisonHRs for various fasting glucose categories with respect to the reference group were calculated using Cox proportional hazards models stratified by age (years) at baseline  after adjustment for age at baseline , sex (when applicable), smoking status , alcohol use , physical activity , body-mass index , systolic blood pressure (continuous variable), serum total cholesterol (continuous variable). Age at baseline was stratified for analyses into six groups . The HRs for a restricted cubic spline transformation of fasting glucose with 5 knots  with mortality in participants having fasting glucose ≤300 mg/dL, were also plotted. Subgroup analyses with varying categories of fasting glucose served as sensitivity analyses.15. All p-values were two-sided. All analyses used SAS version 9.4 .Sex- and age-standardized death rates per 100,000 person-years  was calculated for each fasting glucose groupDuring 134.9 million person-years of follow-up of 12,455,361 people (43.1% women), 416,557 men and 215,832 women died. At baseline the mean (standard deviation) age was 44.0(14.1) years and the mean fasting glucose level was 93.4(27.5) mg/dL Table –S3. HighA unimodal distribution of fasting glucose was observed with a peak at 85–89 mg/dL Figure . Men hadJ-curve-like associations between fasting glucose levels and mortality, with a nadir at around 80–94 mg/dL in both men and women Fig.  and in einteraction = 0.190), while women had stronger association than men in the age group of 65–74 years  in the sex- and age-specific analysis. For glucose levels below 100 mg/dL, the inverse associations between fasting glucose and mortality were stronger in men than in women (Pinteraction = 0.014), especially in the age groups of 55–64 and 75–99 years.For glucose levels of 100–199 mg/dL assuming a log-linear relationship, the HRs per 18 mg/dL (1 mmol/L) increase in fasting glucose were 1.13  in men and 1.12 (1.11 to 1.13) in women (Table interaction between age groups < 0.001 in both the 100–125 and 100–199 mg/dL ranges). The inverse associations in the range below 100 mg/dL were similar across age groups (Pinteraction = 0.198).In the age-specific analysis for glucose levels of 100–199 mg/dL, the HRs per 18 mg/dL (1 mmol/L) increase in fasting glucose were 1.17  in the age group of 35–44 years and 1.09  in the age group of 75–99 years  is concordant with no increase in mean fasting glucose. Furthermore, a well-known sharp decline in estrogen around the time of menopause corresponds with a sharp increase in fasting glucose in women around 50 years in the present study.Fasting glucose levels generally increased with age and were higher in men compared with women, consistent with previous research26. In this systematic review, IFG-ADA was associated with higher mortality in younger persons with an average age <55 years , and in studies in which individuals with baseline cardiovascular disease were not excluded , whereas such an association was not found in older persons (age ≥55 years)  or in studies that excluded individuals with baseline cardiovascular disease . In our study, the multivariate, smoking status-adjusted HRs associated with IFG-ADA were 1.12 (1.11 to 1.13) compared to normoglycemia (70–99 mg/dL)  was associated with higher mortality by 13%  in all eligible studies, and by 9%  in smoking-adjusted studiesL) Table . AlthougConsidering the fairly strong relative risk associated with prediabetes (100–125 mg/dL), younger individuals with prediabetic glucose levels (particularly 110–125 mg/dL), should be carefully managed to prevent premature death. Individuals whose glucose is below 80 mg/dL  should be followed clinically.30, whereas the Diabetes Control and Complications Trial (DCCT)/the Epidemiology of Diabetes Interventions and Complications (EDIC) study, and the follow-up study in the United Kingdom Prospective Diabetes Study (UKPDS) demonstrated that better glycemic control reduced vascular complications and all-cause mortality32. The participants in the DCCT/EDIC and UKPDS were younger and had less pre-existing cardiovascular complications than trials that showed no beneficial effect of intensive glycemic control on mortality. Our analysis showed that the estimated relative risk associated with increasing fasting glucose levels generally weakened with advancing age. These results suggest that glucose lowering interventions may be more beneficial for younger persons with fewer vascular complications33. In this regard, glycemic management in younger adults with prediabetic hyperglycemia (as well as those with diabetes) may be beneficial for preventing premature death.Three large clinical trials in diabetic populations, failed to show a beneficial effect of intensive glycemic control on overall mortality and major vascular complications34; <4th decile, 4th-7th decile, ≥8th decile) in the subpopulation analysis, however, showed no change in the findings. Nevertheless, unadjusted and residual confounding might affect the study results. Information on fasting glucose and other variables might vary in quality depending on the time and the hospitals. However, since the potential variability in data quality by hospitals would most likely be non-differential according to mortality, it would be unlikely to overestimate the associations. Nonetheless, this is a limitation of the study. Relative risk based on a single measurement of fasting glucose may underestimate the true association, due to a regression dilution effect35.The very large number of people and the complete follow-up for death are clear strengths of the study; however, its limitations are to be considered. Causal inference may be limited because of the observational nature of the study. Information on cause-specific mortality was not available due to privacy issues. Other measures of glucose tolerance such as postprandial glucose and HbA1c were also unavailable. Some potential confounders including socioeconomic status and dietary factors were not adjusted for in the main analysis. Including a socioeconomic status variable  were generally the same regardless of sex and age, while the relative risk associated with increasing fasting glucose above those levels was greater at younger ages than at older ages. Prediabetic hyperglycemia was associated with excess mortality, especially in the range of 110–125 mg/dL.Supplementary information


CYP2C19 can result in important interindividual variations regarding therapeutic efficacy. Therefore, we aimed to evaluate the impact of the CYP2C19⁎2 variant (rs4244285) on in-stent restenosis occurrence in Chilean patients who underwent PCI and received clopidogrel. A total of 77 cases with stenosis >50% in the angioplasty site  and 86 controls  were studied. The polymorphism was genotyped using TaqMan® Drug Metabolism Genotyping Assays. Overall, CYP2C19⁎2 allele frequency was 8.3%. Diabetes, chronic lesions, and bare metal stents (BMS) were observed more often in cases than in controls . Genotypic frequencies did not differ significantly between the groups (p = 0.15). Nonetheless, the mutated allele was observed in a greater proportion in patients without in-stent restenosis (p = 0.055). There was no significant association between the rs4244285 variant and the occurrence of in-stent restenosis after PCI . In summary, no association was identified between the CYP2C19⁎2 variant and the development of coronary in-stent restenosis. Clopidogrel is an antiplatelet drug especially used in patients undergoing percutaneous coronary interventions (PCI). Polymorphisms within Clopidogrel is an antiplatelet drug widely used to reduce the risk of acute myocardial infarction and stroke, as well as for secondary prevention of atherosclerotic events in patients at high risk , 2. ThisCYP2C19 polymorphisms have been previously associated with large interindividual variations in terms of therapeutic efficacy in patients treated with clopidogrel [∗2 corresponds to a change of guanine by adenine at position 681 in exon 5 and results in a loss-of-function allele associated with decreased conversion of clopidogrel to its active metabolite, increased platelet reactivity during therapy, and poor clinical outcome, with a potential increase of in-stent thrombosis [∗17 variant is associated with increased antiplatelet therapeutic response and a better clinical outcome [pidogrel –8. Differombosis . On the  outcome –13. Addi outcome , 15, whi outcome .∗2/∗2, ∗2/∗3, and ∗3/∗3) are advised to use drugs such as prasugrel and ticagrelor [∗2 variant was determined in Chilean patients with coronary disease, evaluating its role in the occurrence of in-stent restenosis.Currently in the USA, patients with a poor metabolizer phenotype  was determined in 163 unrelated Chilean patients with coronary heart disease (CHD) admitted to the respective cardiology service of Dr. Hernán Henríquez Aravena Hospital (Temuco) or Dr. Guillermo Grant Benavente Hospital (Concepción), both in Chile. Patients underwent a successful PTCA and a coronary angiography control after 6 months, and all received DAPT. Patients with stenosis >50% were defined as cases and those with stenosis <50% were defined as controls by two experienced cardiologists. In total, both groups consisted of 77 cases and 86 controls. A record of demographic and anthropometric data, occurrence of myocardial infarction, presence of diabetes, biochemical tests , and smoking was elaborated, in addition to angiographic variables of the affected vessel, lesion characteristics, and stent type. The study protocol was approved by 2 independent ethics committees: Scientific Ethics Committee of Universidad de La Frontera and the Scientific Ethics Committee of the Health Service in Concepción. All individuals signed informed consent.This work corresponds to an incident unpaired case-control investigation. The presence of the CYP2C19Blood samples used for biochemical determination and DNA extraction were obtained by venipuncture following 10–12 hours of fasting. Genomic DNA was extracted from leukocytes by a method optimized by Salazar et al. . DNA quat-test for unpaired samples when normal distribution of data was observed, according to D'Agostino testing, or by the nonparametric Mann–Whitney U test. To determine differences between noncontinuous variables, genotypic distribution, allele frequencies, and Hardy-Weinberg equilibrium, chi-square test (χ2) was used. The association between restenosis and the CYP2C19∗2 variant was estimated by calculating the odds ratio (OR) with a confidence interval (CI) of 95%. Additionally, a stratified Mantel-Haenszel analysis was performed to identify potentially confounding variables. A p value less than 0.05 was considered statistically significant.Data are presented as percentages and means ± standard deviation (SD). The differences between the means of two continuous variables were compared using two-tailed Student's p = 0.08). A majority of male patients were observed in both groups . No significant differences regarding demographic parameters, smoking, body mass index (BMI), and blood pressure were observed. On the other hand, diabetes was more frequent in cases than in controls   .p = 0.92) and diameter  values were comparable between the two groups. On the other hand, a higher proportion of subjects with chronic lesions developed restenosis than patients with acute lesions (p = 0.042). Moreover, when comparing stent types (BMS versus DES), the presence of BMS in the case group was significantly higher than in controls .∗2 variant was 8.3% (data not shown). Genotypic distribution and allelic frequencies for each group are shown in χ2 = 0.23, p = NS) and controls . The minor allele was present in a greater proportion in the control group versus cases . No significant differences were observed in genotype frequencies between patients with restenosis and controls (p = 0.15). A higher minor allele frequency (MAF) was observed in patients who did not develop in-stent restenosis, though not significant (p = 0.055), with an odds ratio of 0.44 . Since no patients from the case group carried the homozygous variant, and just one individual presented this genotype in the control group, we were unable to study more profoundly the isolated effect of the AA genotype on restenosis development. However, we observed that individuals from the control group carrying the mutated allele and receiving clopidogrel developed significantly less in-stent restenosis during the study period, discarding diabetes as a possible confounding variable by a stratified and adjusted Mantel-Haenszel analysis.Considering the total cohort, allele frequency for the CYP2C19 CYP2C19 have been widely associated with therapeutic response variability to multiple drugs, as published by the Clinical Pharmacogenetics Implementation Consortium (CPIC) and reported as a warning by the FDA within Plavix (clopidogrel) indications [ CYP2C19 genotypes in patients undergoing percutaneous coronary intervention (PCI), mainly due to the presence of loss-of-function alleles, which can result in subjects with a poor metabolizer phenotype who, therefore, should receive an alternative to clopidogrel therapy. This work constitutes the first study evaluating the association between the loss-of-function CYP2C19∗2 allele and the presence of in-stent restenosis in Chilean CAD patients. Our results show a MAF of 8.3% for the CYP2C19∗2 variant in the entire study group and 5% and 10% allelic distribution in cases and controls, respectively. Allelic frequency for this variant was previously reported to reach 12% for Chileans in a study including the general population [∗2 MAF obtained is comparable to different African populations such as Egyptians (10.9%) [Sequence alterations inications . It has pulation ; howeverpulation , Colombipulation , and Mexpulation  as well pulation . Moreove (10.9%) , Tanzani (10.9%) , and Tun (10.9%) ; Europea (10.9%) ; and als (10.9%)  and Pale (10.9%) .∗/2∗, and in intermediate metabolizers compared to extensive metabolizers (1∗/1∗) (p = 0.008). Sawada et al. [∗ allele, it did not reach statistical significance (p = 0.06), which did occur when comparing the presence of subclinical thrombi, more frequent in the first group (p = 0.0002). Our data indicate a higher proportion of the mutated allele in patients who did not develop restenosis . Even though these results are similar to those reported by Nozari et al. [∗2 allele and in-stent restenosis occurrence, the elevated frequency of the mutated allele in controls rather than cases represents a controversial and unexpected finding, mainly due to the fact that several studies identify this polymorphism as a risk variant. However, various interethnic studies have also demonstrated the highly polymorphic nature of CYP2C19, which could partly explain the differences observed between different populations, especially for the higher MAF found in control subjects from our study. In line with this, a previous 2012 case-control investigation performed in the Korean population described that patients with essential hypertension (EH) had a lower frequency of the CYP2C19∗3 variant allele than did normal controls [On the other hand, our results indicate a lack of association between the loss-of-function variant and restenosis development in the studied subjects. The relationship between low clopidogrel response and restenosis has been previously discarded in a study reporting that low platelet response to clopidogrel had no significant impact on restenosis after DES implantation . Howevera et al.  describei et al. , as theycontrols , which i CYP2C19 genotypes  represent risk factors for restenosis in patients with stenting in the vertebral artery (p = 0.001) [∗2 phenotype carriers or both, required a 2–4-fold higher clopidogrel dose to obtain similar platelet reactivity to patients medicated with 75 mg having neither diabetes nor the loss-of-function variant [Contrasting our results, a Chinese investigation established that loss-of-function= 0.001) . Another= 0.001) –39, it h= 0.001) , demonst= 0.001) . In a re variant . With re variant –45. CYP2C19 variants not included in this study are present, such as CYP2C19∗17, which in combination with a loss-of-function variant results in an intermediate phenotype and not in poor drug metabolism.Among the limitations of our work, we can find the restricted sample size, creating the need to replicate this study in a larger cohort. Additionally, we cannot exclude that different∗2 variant and in-stent restenosis development. Although our results provide valuable information regarding the prevalence of this variant in the Chilean population, the greater proportion of mutated alleles in controls versus cases is to some extent contradictory. However, this outcome should be interpreted with caution, since additional clinical and angiographic factors found in the case group may predispose importantly to restenosis development, particularly in the case of diabetics.In summary, no association was found between the loss-of-function CYP2C19


In patients with one to three brain metastases who undergo resection, options for post-operative treatments include whole-brain radiotherapy (WBRT) or stereotactic radiosurgery (SRS) of the resection cavity. In this meta-analysis, we sought to compare the efficacy of each post-operative radiation modality with respect to tumor recurrence and survival.Pubmed, Embase and Cochrane databases were searched through June 2016 for cohort studies reporting outcomes of SRS or WBRT after metastasis resection. Pooled effect estimates were calculated using fixed-effect and random-effect models for local recurrence, distant recurrence, and overall survival.2: 3.35%, P-heterogeneity = 0.36, 3 studies), 1.09 for distant recurrence , and 2.99 for leptomeningeal disease . For the same comparison, the risk ratio for median overall survival was 0.47  in a fixed-effect model, but was no longer significant  in a random-effect model. SRS was associated with a lower risk of leukoencephalopathy , yet with a higher risk of radiation-necrosis .Eight retrospective cohort studies with 646 patients (238 with SRS versus 408 with WBRT) were included in the analysis. Comparing SRS to WBRT, the overall crude risk ratio using the fixed-effect model was 0.59 for local recurrence (95%-CI: 0.32–1.09, IBased on retrospective cohort studies, the results of this study suggest that SRS of the resection cavity may offer comparable survival and similar local and distant control as adjuvant WBRT, yet may be associated with a higher risk for developing leptomeningeal disease. Future research on SRS should focus on achieving a better understanding of the various factors that may favor SRS over WBRT. Brain metastases are an increasingly common complication of systemic cancers and represent a significant source of morbidity and mortality in cancer patients –5. ApproTraditionally, the standard of care for patients with solitary brain metastasis has been resection plus whole-brain radiotherapy (WBRT) , 7. ReseFurthermore, WBRT is associated with both short and long-term neurological complications, such as radiation-induced edema, leukoencephalopathy, and cognitive deficits , 7, 10. In this study, we used the current published literature to compare local recurrence, distant recurrence, leptomeningeal disease, and overall patient survival amongst patients who underwent resection plus SRS to patients who underwent resection plus WBRT for treatment of one to three intracranial metastases. Given the available data and abovementioned RCT, we hypothesize that SRS may lead to similar rates of local control as compared to WBRT following resection, but, based upon the targeted scope of SRS, may lead to poorer rates of distant control and overall patient survival.This meta-analysis was done in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement . PubMed,Inclusion criteria consisted of any study that described adult patients (≥18 years of age) with a diagnosis of systemic cancer who underwent neurosurgery followed by whole-brain radiotherapy (WBRT) or by stereotactic radiosurgery (SRS) for treatment of up to three metastases to the brain. SRS was defined as a single or few fractions of high dose radiation to a small intracranial target that was encompassed by at least 50% of the prescribed dose . BecauseThe following data were extracted from each study whenever possible: author and year of published articles, number of patients in the study, patient characteristics , number of intracranial lesions at presentation, extent of extracranial disease), duration of follow-up, and primary and secondary outcomes of the study. When available, relative risks (RR) comparing SRS to WBRT were extracted for local recurrence risk, distant recurrence, leptomeningeal disease, and overall survival. Duration of overall survival (OS) and progression free survival (PFS) for both local and distant recurrence was also noted. Furthermore, data on the number of patients that died from neurological causes and on observed neurotoxicity were extracted. In one study reporting outcomes of multiple metastases per patient, the reported percent recurrence by number of cavities was considered as equivalent to the percent recurrence by number of patients, as the number of metastases per patient approximated 1:1 .Study quality was assessed by the New-Castle Ottawa Scale, and the quality of our recommendation was assessed using the Grading of Recommendations, Assessment, Development and Evaluations (GRADE) criteria. Both assessments were made by two authors (NL and IM) independently , 22. DisTo minimize heterogeneity amongst studies being compared, only cohort studies were included in the meta-analysis. Data analysis was performed using Comprehensive Meta-Analysis (CMA) Version 3 . Overall fixed and random risk ratios were calculated for the same outcomes, depending on the available data in the articles. I-squared values were calculated to assess heterogeneity. For median overall survival, relative risks were calculated based on group size and median overall survival both for SRS and WBRT . Meta-reThe search strategy resulted in 9908 articles after removal of duplicates. After screening for titles and abstracts, 410 full-texts were screened. Eight retrospective cohort studies provided outcomes for 646 patients treated with WBRT (n = 408) or SRS n = 238) following resection  that disfavored SRS over WBRT by a 68% higher rate of distant recurrence [p-value comparing just these two treatment arms did find that 5 patients treated with adjuvant SRS were nearly four times as likely to experience local recurrence as compared to 14 patients treated with adjuvant WBRT [Data regarding local recurrence was available in five out of the eight studies included in our review , 26, 27.currence . One Itaant WBRT , 26. Onl2 = 49.44%, p-heterogeneity = 0.10, 3 studies). This analysis was based on 163 patients receiving post-operative WBRT and 138 patients receiving postoperative SRS.The pooled risk ratio comparing local recurrence incidence between SRS vs. WBRT groups was 0.79  and demonstrated no significant difference in rates of local recurrence  and continent (p = 0.04) were identified as a source of heterogeneity. With increasing age, the RR for local recurrence was in favor of SRS as compared to WBRT. Furthermore, Europe, in comparison to North America was associated with a RR for local recurrence in favor of WBRT. Journal impact factor was not identified as a source of heterogeneity (p = 0.97).Univariate meta-regression on age , age (p = 0.12), and journal impact factor (p = 0.52) were not identified as sources of heterogeneity. This analysis was based on 163 patients receiving postoperative WBRT and 138 patients receiving postoperative SRS.Data regarding distant progression was available in three out of the eight studies included in our study Fig.  5, 20, , 20, 26.Two studies reported on the development of leptomeningeal disease (LMD) following radiotherapy , 20. Ret2: 14.4% p-heterogeneity: 0.28, 2 studies) between the included studies. This remained significant in the random effects model . The analysis is based on 192 patients receiving resection followed by WBRT and 152 patients receiving resection followed by SRS.The pooled relative risk for the development of LMD following postoperative SRS to postoperative WBRT was 2.99 (95% CI 1.55–5.76), indicating that resection followed by SRS increases the risk for developing LMD as compared to resection followed by WBRT Fig. . There wp = 0.02). In contrast, they reported a slightly higher risk of death (HR = 1.04) in 11 patients treated with WBRT after resection as compared to SRS alone [p = 0.06) [Only one study out of the 8 included in this review found a significant difference in survival between patients treated with SRS following resection as opposed to WBRT . In theiRS alone . While tRS alone , 26 whilRS alone , 27, 28  = 0.06) .2: 86.6%, p-heterogeneity < 0.01; 4 studies), suggesting a favorable outcome for WBRT with considerable heterogeneity  and journal impact factor (p < 0.01) were identified as sources of heterogeneity. Europe and Asia, in comparison to North America, were associated with a RR for overall survival in favor of SRS. A lower impact factor journal showed a RR in favor of WBRT as compared to SRS. Age was not identified as a source of heterogeneity (p = 0.14).The pooled relative risk for median overall survival comparing patients treated with SRS after resection to those treated with WBRT was 0.51 . After 1 year, the rate of radiographic leukoencephalopathy was 7% following SRS vs. 47% following WBRT (p < 0.01). The relative risk of leukoencephalopathy for SRS compared to WBRT in this study was 0.15 (95% CI: 0.11–0.21).Radiation necrosis and leukoencephalopathy were each reported in one study. Patel et al. found that symptomatic radiation necrosis, requiring the use of steroid or other interventions, was 27% in the SRS group and 0% in the WBRT group  . PrimaryThis meta-analysis suggested that post-operative, localized SRS may offer local protection of a similar degree as post-operative WBRT in patients with one to three lesions. WBRT as compared to SRS, however, seemed to offer better protection from LMD and may offer better protection from distant progression of intracranial metastases with improved survival, albeit not reaching statistical significance in this meta-analysis. However, because WBRT has been consistently associated with post-operative development of physical and cognitive symptoms, SRS is a valid alternative to consider, and can be used in multiple sessions if distant metastases eventually do develop. Notably, the long-term risks of radiation from SRS must also be considered and weighed against the long-term risks of WBRT.


Female gender has been associated with musculoskeletal complaints (MSCs), but there are limited studies on how other factors may influence women and men differently. The aim of this prospective cohort study was to explore possible predictors of MSCs in women and men free of MSCs at baseline.The present study included participants from the population-based Tromsø study, with baseline data from 1994–1995 and follow-up data from 2007–2008. MSCs were defined as having pain and/or stiffness in muscles and joints for 3 consecutive months during the past year. Predictors of MSCs were examined through binary logistic regression analyses and presented as odds ratios with 95% confidence intervals.2  and smoking . Age and physical activity level were not significantly associated with MSCs. Gender-stratified analyses revealed that mental health complaints  predicted MSCs in men , but not in women. Current smoking  and poor self-perceived health  showed slightly higher odds ratios among women than men, but the gender differences were not significant.At baseline 4,496 participants reported no MSCs and among these 2,015 (44.8%) and 441 (9.8%) participants reported mild or severe MSCs, respectively, at follow-up. Female gender predicted MSCs in multivariable logistic regression analyses . Educational level of primary/secondary school  was the strongest predictor of MSCs, followed by poor self-perceived health . Other predictors were BMI ≥30 kg/mThe present study demonstrates that several negative health determinants are predicting subsequent MSCs. However, the examined risk factors could not explain the higher prevalence of MSCs in women. Chronic musculoskeletal complaints (MSCs) are recognised as a major and costly health problem in Norway  and otheMany studies have reported gender differences in the prevalence of MSCs , 20–22, Using data from 13 years of follow-up, the aim of the present study was to determine which factors predicted MSCs in a prospective cohort of Northern Norwegian adult women and men with no MSCs at baseline.The Tromsø study is a longitudinal, population-based, multi-purpose health study carried out in the municipality of Tromsø, Northern Norway. The study currently consists of seven health surveys (Tromsø 1–7); Tromsø 1 was performed in 1974 and Tromsø 7 was completed at the end of 2016. The participation rate for all six surveys has ranged from 65% to 77% . All parThe present analyses used data from Tromsø 4 (1994–1995) and Tromsø 6, as information on MSCs in these surveys was comparable. Tromsø 4 included men and women aged 25–97 years. All inhabitants of Tromsø, Northern Norway, older than 25 years  were invited and 27,158 attended . After excluding those who had moved away or died, the participation rate for Tromsø 4 was 77%. Tromsø 6 included men and women aged 30–87 years; of the 19,762 invited, 12,984 (66%) attended [Chronic MSCs were defined as pain and/or stiffness in muscles and joints lasting for at least 3 consecutive months during the previous year (hereafter referred to as simply MSCs). Information on MSCs in Tromsø 4 (baseline) was collected through the screening question, “Have you during the last year suffered from pain and/or stiffness in muscles and joints that have lasted continuously for at least 3 months?” (yes/no). Respondents who answered “no” were categorised as not having MSCs. The reliability of this screening question has been evaluated to be good . The Tro2, 25–29.9 kg/m2 and ≥30 kg/m2). Participants were categorised as married if they had answered either “married” or “registered partnership”. The question on self-perceived health had four alternatives: very good, good, not so good and poor, merged into two categories: “poor” or “good”. Those reporting current cigarette, cigar, or pipe smoking at baseline were categorised as current smokers. Information on mental health was taken from the CONOR-Mental Health Index (MHI), a 7-item questionnaire. Questions were answered using a 4-level scale, ranging from “no” (1) to “very” (4). Thus, the total score for all seven questions ranged from 7–28. The CONOR-MHI average score was calculated by dividing the total score by the number of items, i.e., seven. Based on a validation study, the cut-off was set to 2.15, creating two groups: CONOR-MHI <2.15 and CONOR-MHI ≥2.15, i.e. indicating mental health complaints [https://en.uit.no/prosjekter/prosjekt?p_document_id=80172) [The baseline risk factors included age, gender, marital status, current smoking, self-perceived health, mental health complaints, education level, body mass index (BMI), and leisure time physical activity. Information on these factors was collected through questionnaires, except for BMI, which was calculated from data on height and weight measured in light clothing and without shoes. Three categories of BMI were created (BMI ≤24.9 kg/mmplaints . This inmplaints . The qued=80172) .In total, 10,326 individuals attended both baseline and follow-up. In order to create valid information on MSCs, 3,902 individuals who reported MSCs or had missing information on MSCs (n = 9) at baseline were excluded, as were those with missing answers on one or more of the six MSCs variables at follow-up . Because of low participation rates in the age groups 80–84 years (42.4%) and 85–87 years (30.5%) at follow-up, we excluded these age groups (n = 134). Thus, 4,496 participants reporting absence of MSCs at baseline were included in this study .Differences in the baseline characteristics of participants with no complaints, mild complaints, and severe complaints were analysed using ANOVA for continuous variables, and the Pearson chi-square test for categorical variables. Bonferroni corrections were conducted to adjust for multiple comparisons. The influence of independent variables on MSCs was evaluated using binary logistic regression to estimate odds ratios (OR) with 95% confidence intervals (CI) and p-values for trend analyses. Logistic regression analyses were first performed using the dichotomised dependent variable on overall MSCs (mild/severe MSCs versus no MSCs). Logistic regression was then performed on the dichotomised dependent variable on only severe MSCs. Predictors of multiregional MSCs were identified in a regression model using MSCs in ≥3 regions versus <3 regions as the dependent variable. Finally, we evaluated predictors of MSCs for each body region separately. Crude logistic regression analyses were first performed and then adjusted for age and gender. Covariates that showed a significant relationship in the age- and gender-adjusted models were included in multivariable models. Finally, logistic regression analyses including interaction terms were performed to examine gender differences in predictors of MSCs and to assess the significance level.Missing values on independent variables at baseline were: marital status (n = 6), smoking status (n = 7), BMI (n = 5), self-perceived health (n = 2), mental health (n = 67), educational level (n = 8), and physical activity level (n = 40). Thus of the 4,496 participants in this study, 4,373 were included in the multivariable regression models. The statistical analyses were performed by SPSS version 21. All tests were performed two-sided and a p-value <0.05 was considered significant.The Regional Committee for Medical and Health Research Ethics and the Norwegian Data Inspectorate approved the Tromsø study, and written informed consent was obtained from all participants.2 and 8.0% were obese (BMI ≥30 kg/m2). Nearly one out of four (23.4%) reported primary/secondary school as their highest level of education , with odds ratios that were slightly higher in women. Mental health complaints were not significantly predicting MSCs in this multivariable analysis of the total population. However, after stratification for gender, mental health complaints were shown to predict MSCs in men , but not in women . The interaction term analysis with gender and mental health complaints did almost reach the significance level of 5% (p = 0.051). Furthermore, BMI ≥30 kg/m2 increased the risk of MSCs in men , but not in women . However, the interaction term analysis with gender and BMI ≥30 kg/m2 was not significant (p = 0.537), as were the interaction terms involving the other covariates analysed in this multivariable regression model. Finally, reporting a sedentary physical activity level did not predict MSCs in this multivariable analysis.The associations were similar in the multivariable regression analysis of mild/severe versus no MSCs . Female The results of the multivariable analysis on severe (model 2) and multiregional MSCs (model 3) compared to severe and mild together (model 1) are shown in 2 at baseline strongly predicted MSCs of the hip/leg/feet . Physical activity level was not significantly associated with MSCs in the upper back, lumbar back, hip/leg/feet, and other regions.Female gender, poor self-perceived health, and an educational level of primary/secondary or technical school increased the risk of reporting MSCs in all six body regions . Mental 2 , primary/secondary school as their highest level of education , poorer self-perceived health , and to be current smokers . Furthermore, the excluded population was more likely to report a sedentary physical activity level than the study sample . Marital status and high physical activity level were similar in the two populations. A sensitivity analysis was performed using logistic regression models in which all 6,415 participants reporting no MSCs at baseline were included. Those who had reported MSCs in any region at follow-up were included among participants with overall MSCs and those who did not report MSCs at follow-up were coded as no MSCs  and the excluded population . Excluded participants were more likely to be female , have a BMI ≥30 kg/mThe effects of demographic, socioeconomic, self-reported health, and lifestyle factors were examined in this prospective cohort study on self-reported MSCs after 13 years of follow-up in the population of Tromsø, Northern Norway. The odds of having MSCs in any body region were 46% higher in women than men. The strong association between female gender and MSCs is broadly supported by other studies , 34, incLow educational level, followed by poor self-perceived health were the strongest predictors in the regression analyses of overall MSCs (mild/severe MSCs versus no MSCs), and they were also important predictors in the analyses of individual body regions. This emphasises findings of previous studies examining how socioeconomic status and poor self-perceived health associates to MSCs , 25, 35.The results of our gender-stratified analyses of physical activity showed an association with overall MSCs, but only among women in age-adjusted models. Moreover, these models showed a dose-response relationship with a significant trend. However, after adjustment for other covariates, the significant association did not persist in multivariable analyses for any level of physical activity. Neither did region-specific analyses add convincing evidence of a relationship between physical activity level and MSCs. Thus, we were not able to unambiguously show that low physical activity level increases the risk of MSCs, which is in accordance with the cross-sectional study based on the Tromsø study (2007–08) , but in Mundal et al used the hospital anxiety and depression scale (HADS) to assess mental health complaints, and found that it predicted the onset of chronic widespread MSCs in a study with a similar design . HADS anIt is broadly accepted that smoking is associated with MSCs , 19, 43.Both overweight and obesity were significantly predicting MSCs in our study. The strongest association with obesity was found in the lower extremities, increasing the odds of MSCs by 74%. The suggested underlying mechanisms of this are multi-factorial –48. A crKvalheim et al explored the relationship between female reproductive health and chronic widespread MSCs in their study on a general population . They foThe major strengths of this study are the prospective design and the inclusion of MSCs-free subjects at baseline. Thus, we can be certain that the risk factors developed before, and not as a consequence of the outcome. The baseline characteristics included in this study were chosen based on previous findings from the Tromsø study . AlthougThe prevalence of possible risk factors in our study sample changed as we chose to exclude a relatively large proportion of participants. This was due to incomplete follow-up data and difficulties in building an overall MSCs end-point variable. In sum, our study sample had lower levels of negative health determinants than did the excluded population. This may have biased our results as we included only those who fully understood the questionnaire and provided complete answers. It is likely that the complete-case approach to self-reported MSCs makes our results more conservative. However, the sensitivity analysis did not reveal any changes in the associations.In conclusion, the results of this study helps clinicians outline individuals at risk of developing musculoskeletal complaints as it has demonstrated that several factors are predicting the onset of both mild and severe MSCs. With the prospective design, large study population, and the assumption of temporality, the present study indicates that, although not large, there are some differences between men and women regarding the effects of risk factors on the development of MSCs. However, no specific risk factors that could explain the higher prevalence of MSCs in women were revealed. This warrants further research to focus on other factors that could influence gender differences in the development of MSCs.All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.S1 AppendixThose reported MSCs at any body region were included as having MSCs, and those who did not report MSCs were coded as no MSCs.1Musculoskeletal complaints (Mild/severe pain and/or stiffness in muscles and joints lasting at least 3 months during the past year), 2The Tromsø Study; a prospective study of an arctic general population consisting of 4,496 adult men and women free of MSCs at baseline, 3Binary regression, adjusted for age and gender (bold text = significant result), 4Cohort of Norway Mental Health Index ≥2.15, OR: odds ratio,*significant interaction with gender; p<0.05.(DOCX)Click here for additional data file.S2 AppendixThose reported MSCs at any body region were included as having MSCs, and those who did not report MSCs were coded as no MSCs.1Musculoskeletal complaints (Mild/severe pain and/or stiffness in muscles and joints lasting at least 3 months during the past year), 2The Tromsø Study; a prospective study of an arctic general population consisting of 6,415 adult men and women free of MSCs at baseline, 3Multivariable logistic regression analyses, the models included all variables listed (bold text = significant result), 4Cohort of Norway Mental Health Index ≥2.15, OR: odds ratio. Interaction term analyses did not reveal significant gender interactions in this multivariable logistic regression analysis, p<0.05.(DOCX)Click here for additional data file.


This study retrospectively evaluated overall survival (OS) by treatment of recurrent or metastatic uterine adenosarcoma including surgery, radiation, chemotherapy, and hormonal therapy and evaluated OS and progression-free survival (PFS) after 1st line systemic chemotherapy.  78 patients with recurrent or metastatic adenosarcoma comprised the study population. The Kaplan-Meier method was used to estimate OS and PFS. The log-rank test was performed to test the difference in survival between groups. p=0.035. Median OS differed by surgery for 1st recurrence 26.3 months versus 15.1 months. OS was not influenced by chemotherapy, p=0.58, palliative radiation, p=0.58, or hormonal therapy, p=0.15. The response rate (CR + PR) per RECIST 1.1 for chemotherapy was 31.2% for doxorubicin-based regimens and 14.3% for gemcitabine/docetaxel. OS since 1st line chemotherapy was not significantly different among chemotherapy regimens. However, the median PFS was superior for doxorubicin/ifosfamide (15.4 months) compared to gemcitabine/docetaxel (5.0 months), platinum-based regimens (5.7 mo), or other doxorubicin-based regimens (6.5 months).  Median OS from diagnosis of recurrent or metastatic disease was 1.8 yrs. OS was influenced by pathology on recurrence,  These results suggest that surgery is an important treatment modality for recurrent or metastatic uterine adenosarcoma, and the most effective chemotherapeutics are doxorubicin/ifosfamide and gemcitabine/docetaxel. Uterine adenosarcoma is an extremely rare subtype of uterine sarcoma, which represents only 5.5 to 9% of all uterine sarcomas , 2. UterThere is a specific FIGO uterine adenosarcoma staging system which divides between stage Ia, Ib, and Ic by the presence and extent of myometrial invasion . The majUnfortunately, given the rarity of adenosarcoma, there are limited data to guide treatment decisions in the recurrent or metastatic setting. There is no standard treatment for recurrent or metastatic disease, though surgery is often preformed . A recenThe purpose of this study was to examine the role of surgery, radiation, and hormonal therapy in the recurrent or metastatic setting. Furthermore, this is the first study to report response rates, overall survival, and progression-free survival of adenosarcoma patients treated with chemotherapy for recurrent or metastatic disease.With approval by the MD Anderson Cancer Center Institutional Review Board, a search was conducted of the Institutional Tumor Registry. Seventy-eight patients treated at the MD Anderson Cancer Center from August 1982 to December 2014 with recurrent or metastatic uterine or extrauterine adenosarcoma were identified. The diagnosis of uterine adenosarcoma was confirmed by MD Anderson sarcoma or gynecologic pathologists. Demographic, clinicopathologic, and treatment characteristics were abstracted from the patients' medical records. Then, a deidentified database was constructed of all adenosarcoma patients. The stage was assigned using the International Federation of Gynecology and Obstetrics 2009 uterine adenosarcoma staging system .Patients' demographic and clinical characteristics were analyzed, with categorical variables summarized in frequency tables while continuous variables summarized using mean (±S.D.) and median (range). The product limit method of Kaplan and Meier was used to estimate overall survival (OS) and progression-free survival (PFS) . OS was Seventy-eight patients with recurrent or metastatic uterine or extrauterine adenosarcoma who received treatment at the MD Anderson Cancer Center were identified from a retrospective review of the Institutional Tumor Registry and were included in this study. The median follow-up time since recurrent or metastatic disease was 8.2 years. The demographic and clinical characteristics for all patients included in this study are summarized in Treatment on recurrence varied greatly and was influenced by the location and extent of recurrence. p=0.035, HR = 0.47 (95% CI 0.23–0.96)  .Forty-five patients underwent surgery for recurrence or metastatic disease.p=0.54  or platinum-based regimens (0.49). The median overall survival for doxorubicin-based regimens, other than AI, was 13.1 months. Additionally, patients who received AI had improved PFS (15.4 months) compared to other doxorubicin-based regimens (6.5 months), gemcitabine/docetaxel (5.0 mo), and platinum-based regimens (5.7 months)  . This diPatients were analyzed for other factors which may have influenced the effectiveness of their chemotherapeutic regimens, such as chemotherapy dosing, # of chemotherapy cycles, presence of sarcomatous overgrowth (SO), point in their disease course when they received chemotherapy, and time to recurrence prior to initiation of chemotherapy, a potential indicator of the aggressiveness of their disease. Patients did not significantly differ in terms of chemotherapy dosing, # of chemotherapy cycles, presence of SO, or point in their disease course when they received chemotherapy. The median time to recurrence prior to initiation of 1st line chemotherapy was 10.1 months for all doxorubicin-based regimens, 12.9 months for AI, 9.6 months for other doxorubicin-based regimens, 8.1 months for platinum-based regimens, and 21.7 months for gemcitabine/docetaxel, indicating a possible physician bias for treating patients with doxorubicin-based regimens in patients with quicker relapses, so more aggressive disease.p=0.15.Twenty-eight patients received hormonal therapy at some point during their treatment course. Seven patients received more than 1 line of hormonal therapy. Initial hormonal therapies included GnRH agonists , progesterones , SERMs , and aromatase inhibitors (anastrozole 3 pts and letrozole 1 pt). The medial overall survival for patients with recurrent disease who received hormonal therapy was 34.7 months compared to 17.6 months, a trend toward improved outcomes that was not statistically significant, Out of these twenty-eight patients, there were four that derived several years of benefit from hormonal therapy. Two patients treated with leuprolide had disease control for >2 years. Their tumors were not stained for the estrogen receptor (ER) or progesterone receptor (PR). A third patient was placed on leuprolide after surgical resection with the development of progression after two years. This patient was then placed on megestrol acetate without response and then on anastrozole with a complete response lasting for eight years. This patient developed a 2nd malignancy. She was taken off anastrozole at the time of surgery for her cholangiocarcinoma. Shortly after completing adjuvant chemotherapy for her cholangiocarcinoma, she developed recurrence of her adenosarcoma, biopsy proven. She was placed back on anastrozole with response lasting for another five years. More recently, she developed progression and is now on systemic chemotherapy with trabectedin. This patient's tumor stained for ER 80% and PR 40%. The mesenchymal portion of her initial tumor was described as endometrial stromal sarcoma, though she did have sarcomatous overgrowth noted as well. A fourth elderly patient with locally advanced unresectable disease was treated with leuprolide and carboplatin for seven cycles leading to a partial response. Her leuprolide was continued after chemotherapy resulting in a complete response after five years with resolution of her pelvic mass and remaining subcentimeter pelvic lymphadenopathy. She continued leuprolide for 14 years, at which point her leuprolide was stopped. She then developed radiographic recurrence within the abdomen and was restarted on leuprolide with stable disease until her death at age 95. The ER/PR status and SO status of her tumor are unknown.In this study, we report the largest single-institution experience with recurrent or metastatic uterine and extrauterine adenosarcoma. There is no standard treatment for patients with local recurrence or metastatic uterine adenosarcoma. Treatment options include surgery, radiation, chemotherapy, and hormonal therapy. This study is the first to show that the pathology on recurrence influences outcomes. Specifically, those patients that present with a recurrence that is purely high-grade sarcoma have significantly worse outcomes than those patients who present with a recurrent tumor with mixed epithelial and mesenchymal components. This may represent selective clonal evolution of these tumors such that patients with a recurrence of pure high-grade sarcoma have a more clinically aggressive course.The majority of uterine adenosarcomas recur locally, suggesting that resection of a local recurrence may improve overall survival and time to next recurrence. Previous reports have indicated a benefit for secondary cytoreduction of recurrent adenosarcoma in terms of overall survival and time to next recurrence , 21. Our2 of gemcitabine and 75 to 100 mg/m2 of docetaxel; the patients that received doxorubicin-based regimens received 60 to 75 mg/m2 of doxorubicin, 7.5 to 10 gm/m2 of ifosfamide, and 750 to 1000 mg/m2 of dacarbazine. This is standard sarcoma chemotherapy dosing.This is the first retrospective report to examine the use of systemic chemotherapy in a large population of recurrent uterine adenosarcoma. Case reports and case series have described responses in adenosarcoma with the use of doxorubicin-based regimens –26, gemcThe OS after 1st line chemotherapy for recurrent or metastatic disease was not statistically different between AI, gem/doc, or platinum-based regimens. However, OS may be affected by subsequent therapies such as surgery, hormonal therapy, or further chemotherapy. Additionally, there was a trend toward worse survival for patients that received other doxorubicin-based regimens excluding AI, suggesting that ifosfamide is required to obtain the higher response rate and clinical benefit. However, whether this effective is limited to ifosfamide alone or a result of synergy between doxorubicin/ifosfamide, as noted in soft tissue sarcomas, is undetermined .PFS may be a better measure of benefit from systemic chemotherapy. Patients who received AI had the longest PFS (15.4 months), with a trend toward statistical significance when compared to patients who received doxorubicin alone or in combination with a 2nd agent or platinum-based regimens. If the goal of therapy is to produce reduction in tumor size prior to surgical resection of recurrent disease, AI chemotherapy may have advantages over gemcitabine/docetaxel. If patients are older with poor function status and multiple medical comorbidities, they are not a doxorubicin/ifosfamide candidate; then gemcitabine/docetaxel may be the preferred regimen. Cisplatin- and carboplatin-based regimens appear to be the least effective in uterine adenosarcomas and should not be recommended for treatment of recurrent or metastatic disease. Further study is required to evaluate the role of trabectedin in the treatment of uterine adenosarcomas.It should be noted that there was likely a physician bias in choosing treatment with doxorubicin-based regimens over gemcitabine/docetaxel. There was a shorter median time to recurrence prior to the start of 1st line chemotherapy with AI (12.9 months) or other doxorubicin-based regimens (9.6 months) compared to gemcitabine/docetaxel (21.7 months), indicating that physicians were more likely to treat with doxorubicin-based regimens than with gemcitabine/docetaxel for patients that had quicker relapses, or more aggressive disease. This suggests a benefit of doxorubicin/ifosfamide over gemcitabine/docetaxel in the treatment of uterine adenosarcomas, despite similar median OS, as patients with more aggressive disease would be expected to have worse survival.Evidence for the use of hormonal therapy in uterine adenosarcoma is even more limited than evidence for the use of chemotherapy. Case reports or case series have noted responses to hormonal therapy lasting 10 months to 7 years , 30–33. This study is limited by its retrospective nature and small sample size, though this is the largest single-institution recurrent or metastatic uterine or extrauterine adenosarcoma series to date. Overall, uterine adenosarcoma is a rare disease with limited evidence-based data to determine treatment recommendations. Treatment of recurrence or metastatic disease can consist of surgery, chemotherapy, preferable with doxorubicin/ifosfamide or gemcitabine/docetaxel, or hormonal therapy in select patients.


Inflammatory arthritis leads to work disability, absenteeism and presenteeism (i.e. at-work productivity loss) at high cost to individuals, employers and society. A trial of job retention vocational rehabilitation (VR) in the United States identified this helped people keep working. The effectiveness of this VR in countries with different socioeconomic policies and conditions, and its impact on absenteeism, presenteeism and health, are unknown. This feasibility study tested the acceptability of this VR, modified for the United Kingdom, compared to written advice about managing work problems. To help plan a randomized controlled trial, we tested screening, recruitment, intervention delivery, response rates, applicability of the control intervention and identified the relevant primary outcome.A feasibility randomized controlled trial with rheumatoid, psoriatic or inflammatory arthritis patients randomized to receive either job retention VR or written information only . Following three days VR training, rheumatology occupational therapists provided individualised VR on a one to one basis. VR included work assessment, activity diaries and action planning, and (as applicable) arthritis self-management in the workplace, ergonomics, fatigue and stress management, orthoses, employment rights and support services, assistive technology, work modifications, psychological and disclosure support, workplace visits and employer liaison.Fifty five (10%) people were recruited from 539 screened. Follow-up response rates were acceptable at 80%. VR was delivered with fidelity. VR was more acceptable than written advice only (7.8 versus 6.7). VR took on average 4 h at a cost of £135 per person. Outcome assessment indicated VR was better than written advice in reducing presenteeism (Work Limitations Questionnaire (WLQ) change score mean: VR = −12.4 (SD 13.2); control = −2.5 (SD 15.9), absenteeism, perceived risk of job loss and improving pain and health status, indicating proof of concept. The preferred primary outcome measure was the WLQ, a presenteeism measure.This brief job retention VR is a credible and acceptable intervention for people with inflammatory arthritis with concerns about continuing to work due to arthritis.ISRCTN 76777720. Registered 21.9.12.The online version of this article (doi:10.1186/s12891-017-1671-5) contains supplementary material, which is available to authorized users. Work problems are common in people with inflammatory arthritis. A third of employed people with rheumatoid arthritis (RA) stop working within three years of diagnosis due to arthritis (i.e. become work disabled) and 50% within 10 years . This isn = 242)  minutes work advice (without a structured work assessment), provision of work advice booklets and signposting to other services , 13. ConIntervention group participants also received written work self-help information and usual care. Treating occupational therapists received three days VR training [12]. Participants were seen within four weeks of referral. Direct VR consisted of up to 4.5 h of 1:1 meetings, starting with a structured work interview and ending with a telephone review. Up to 1.5 h extra contact was also possible. We planned VR to be longer than in Allaire et al.’s trial  as: therMeetings were at mutually agreed times (often early or late in the day to fit around participants’ work commitments) and locations  spread over two to four months. The intervention started with a structured work interview and job discussion  and detailed assessment of work barriers. This was conducted using the UK Work Experience Survey-Rheumatic Conditions (WES-RC) –19. ThisThe WORK-IA VR Resource Manual was provided to support VR delivery. :Demographic and work data: age, gender, condition duration, marital and living status (at baseline only); medication regimen; current job(s); years in current main job; whether disclosed arthritis to employer and/or work colleagues; and number of normal working hours.Employment status: whether in full- or part-time work, on long term sick leave, or stopped working (with date when stopped and reason).i)RA- Work Instability Scale (RA-WIS): 23 true/false items measuring, at present, the degree of mismatch between functional abilities and workplace demands [ii)Work Activities Limitations Scale (WALS): 12 items (time frame unspecified) indicating degree of difficulty performing physical work activities, time, mental and output demands , with additional items for whether help or equipment are used for each .iii)The Work Limitations Questionnaire (WLQ): 25 items, indicating the amount of time, in the last two weeks, a person was limited in: physical work demands, time demands, mental-interpersonal demands and output demands. The four sub-scale scores and Summed score  range from 0 to 100%. A percentage Productivity Loss score can also be calculated .Presenteeism: these measures evaluate the effect of a condition on the quality or quantity of work productivity . Three mWork self-efficacy: confidence in ability to continue working with arthritis; and ability to manage arthritis at work ).Satisfaction with work rehabilitation advice received (0–10 NRS at six months only).SF-12v2 Health Survey: 12 items, assessed over the last four weeks, scored as physical and mental health sub-scales .Multi-dimensional Health Assessment Questionnaire (HAQ): the modified HAQ (eight activity limitation items); psychological status HAQ (four items); pain, fatigue and a global rating of health measured using 100 mm Visual Analogue Scales (VAS), assessed over the last week .Hand/ wrist pain: pain in the last week during moderate activity (100 mm VAS).Euroqol five dimensions questionnaire (EQ5D): measuring quality of life .A self-report measure was resource use questionnaire .In addition to the questionnaire booklet, participants filled out a monthly tear-off calendar, to record their work status each day . This was modified from Part 1 of the Health and Labour Questionnaire . ParticiTime to temporary or permanent job loss (days): recording on which date their contract ended, if they stopped working and, if a new job was obtained, their contract started.Absenteeism: over nine months, the number of days sickness absence attributable to either arthritis or other causes (e.g. common ailments), not including days not normally worked.Participants were not telephoned to obtain missing data from returned questionnaires or minimal datasets from non-responders, apart from for the absenteeism measure.Therapists completed a VR Treatment Record of: duration of treatment contact for direct VR (i.e. with participant) and indirect VR ; treatment location; and travel mileage.A statistically based sample size estimate was inapplicable for this feasibility study. Randomising 100 participants would permit estimation of the percentage of overall drop-outs to +/− 10% points at most, with 95% confidence. Drop-outs were considered those who did not attend VR or did not return follow-up questionnaires.Analyses were mainly descriptive in order to determine if a definitive RCT is feasible. Recruitment and retention rates were summarised descriptively. For each outcome measure and trial arm, the proportion of missing data was described and the outcome at follow-up summarised . Where, applicable, change from baseline was also summarised and effect sizes were calculated) as mean change/standard deviation at baseline) to compare the internal responsiveness of outcomes. An effect size of 0.2 is interpreted as small, 0.5 medium and 0.8 large effect sizes . All anaEthics approvals were obtained to schedule. Sites recruited varyingly between November 2011 and September 2012 and actively recruited for 6.5 months (1QR 5–7) . This was because Local NHS Trust Research and Development department (R&D) approvals were delayed and took until month 14 at five sites and month 17 at one site. Five sites were re-scheduled to open for ten and one for seven months. Average time to recruit the first patient was 14 weeks (IQR 10.5–17.25) as, for the first three to four months, there was little or no research facilitator availability at four sites. At six months, two sites closed: research facilitator support was withdrawn at one and the only occupational therapist went on long-term sick leave at the other.n = 199) were eligible. Of these, 28% of those eligible  consented. At screening, most had RA-WIS scores ≥10 (i.e. moderate to high risk of work disability). Only six (11%) had scores <10 but answered “yes” to concerns about continuing working with arthritis. Their average RA-WIS score at screening was 5.5 (SD 2.7) but rose to 7.8 (SD3.7) at baseline, as three now reported RA-WIS scores ≥10. The two intervention group participants with scores <10 at screening, had multiple problems identified in the WES-RC, indicating VR was applicable. Given most people with concerns had RA-WIS scores ≥10, and those scoring <10 at screening had scores close to or ≥10 at baseline, using the concerns criterion is applicable for a future trial. Most participants completed baseline questionnaires at home, with only 10 choosing to do so in clinic. Within the two groups, 1/26 in the control group was incorrectly referred to occupational therapy by the CTU and received VR; and 4/29 in the intervention group did not complete VR . By six months, one person in the intervention group withdrew.The flow of participants is shown in Fig. n = 144; or 27% overall) of those eligible did not consent. Almost half  were recorded as “no reason given” for non-participation. The trial manager identified these were often people needing time to consider participation after screening but some research facilitators were unable to contact patients by telephone to complete consent. Research facilitators worked during the day (when most participants were at work) and not evenings, when patients could be more readily contactable. The other half of patients (n = 73) not consenting gave understandable reasons  thought VR would interfere too much with their work/ daily life and 23% (n = 33) did not want VR/work advice. During the trial, the trial manager identified some research facilitators were unclear about flexibility of the VR and employer contact being unnecessary. To help research facilitators allay patients concerns, further site training was provided.However, 72% (n = 340) were ineligible. Of these, 8% (n = 41) of patients had incomplete screening. The trial manager identified these were patients sent invitation letters prior to clinics, added to the screening list but some research facilitators were then unable to attend clinic to complete screening in person or contact patients later by telephone.Overall, 63%  and Driving to work, n = 13); Physical Demands ; Work/mental time demands . Matching solutions included, respectively: advance planning, daily activities and driving advice; pacing and moving and handling advice/ training; and fatigue management.The monitoring visits confirmed occupational therapists conducted assessments appropriately. The WES-RC review indicated therapists identified a wide range of problems and provided appropriate treatment matching problems. The commonest problems were: Preparing for/getting to work  participants in both groups correctly completed all questionnaire items, apart from two (one in each group) having missing data in either the output demands or time management demands scales of the WLQ. At six and nine months, 44/55 and 43/55 respectively returned questionnaires  in the RA-WIS, WLQ Summed and Productivity Loss scores, confidence in managing arthritis at work, physical function (SF-12v2), pain, hand pain and perceived health status Tables  and 4. In = 25 participants attending VR). Direct VR lasted on average 3.92 (SD 1.61) hours and indirect VR was an additional 1.68 (SD 1.46) hours. The average cost per participant of VR was £135.18 (SD £81.80) for occupational therapy staff time and mileage costs.Participants in the intervention group attended on average 2.86 (SD 1.55) VR sessions  reported no difficulties completing presenteeism or absenteeism measures and expressed no preference for the most important to use. Completion rates were better for presenteeism than absenteeism measures. The three presenteeism measures  had similar psychometric properties, correlated similarly with other health and work measures and had similar completion rates. Larger effect sizes were seen for the WLQ Summed score and RA-WIS , suggesting time to job loss and employment status are not applicable outcomes unless a trial has a long follow- up (e.g. four to five years).Participants interviewed about the outcome measures  provided detailed data but completion rates were lower than for presenteeism and had too high a responder burden. Three monthly self-report of sick leave days  would be less onerous and could lead to better response rates. The WLQ and RA-WIS presenteeism measures are considered the best options. The Outcome Measurement in RA Clinical Trials (OMERACT) Work Group previously had identified the WLQ, WALS and RA-WIS as presenteeism measures , 38. HowOnly 20% of people with RA consider they receive sufficient information and practical support about employment issues from their Rheumatology service . Many emDuring the study, some research facilitators highlighted patients, particularly in areas with higher social deprivation and in precarious employment, expressed concerns about taking time out of work and/ or did not want VR, influencing their non-participation. The British Society of Rheumatology is now supporting training for Rheumatology teams to have “work conversations” with patients to promote the importance of staying working and obtaining work advice. In a future trial, ensuring teams have training in, and implement work conversations, could prime patients to perceive VR as important to receive.There were limitations in this study. The two groups differed in occupational groupings and, in a definitive RCT, trial stratification by occupational group (levels 1 to 4) would reduce this disparity. The recruitment time was nearly half that planned, partly due to the time obtaining R&D approvals. The NHS Health Research Authority has since streamlined approvals processes, with targets for completions and recruitment. Our recruitment strategy was screening in two Rheumatology clinics per week. However, limited research facilitator availability caused delays in sites starting, periods of no screening and early site closure. Only 10% of patients screened were randomised into the study. This was however, consistent with pre-trial predictions. A two-week audit before the trial at one site identified 25% of Rheumatology out-patients were employed people with RA, PsA or IA. Studies evaluating RA-WIS scores in employed people with RA identified 34 to 69% had scores ≥10%, i.e. moderate to high risk of work disability , 46–49. Research facilitators had problems contacting up to 20% of those identified to complete screening or consent. Research facilitators worked only during the day and often only had patients’ home landline telephone numbers and not work/ mobile telephone numbers and e-mail addresses. This prevented contacting employed patients in the evenings. It is clearly important to ensure a range of contact details are obtained at the point of first contact with potential participants. In consequence, in a future trial, employing a research fellow, working flexibly into some evenings, would assist contacting patients outside normal work hours. The research fellow can then complete screening and consent by telephone with patients, ensure baseline questionnaires are sent out/returned and patients randomised (subject to patients’ permission for contact details to be forwarded). When research facilitators are unavailable to complete screening and recruitment, a research fellow can also complete study procedures. To increase numbers of people with RA, PsA or IA referred for screening, working age/ employed patients could be identified by healthcare team members/ research facilitators from department databases, medical and therapy notes and in therapy departments. Such patients could be mailed invitation packs by research facilitators, with interested patients returning a reply form to the research fellow who completes study procedures.Whilst an 80% retention rate is suggested as near the limit of validity for RCTs , our 20%The findings from this study suggest job retention VR provided by NHS occupational therapists with VR training is credible, acceptable, deliverable and promising for employed people with arthritis with concerns about continuing to work in future because of their condition. The finding regarding screening and recruitment suggest recruitment could be challenging and requires additional research fellow support. Consideration needs to be given to the number needing to be screened (around 1350) to achieve the recruitment target, the number of sites and length of recruitment period required to achieve the target. There is a need for a definitive trial of this brief job retention VR in the UK, with the caveat of ensuring sufficient funding for out-of-hours and remote screening, recruitment and retention support to ensure timely recruitment and maximised response rates.