Assessment of somaclonal variation in somatic embryo-derived plants of yacon [ Smallanthus sonchifolius (Poepp. and Endl.) H. Robinson] using inter simple sequence repeat analysis and flow cytometry

Background: Yacon ( Smallanthus sonchifolius ) is a root crop native to the Andean region. Low sexual reproductive capacity is amajor constraint facing the genetic breeding of this crop. Biotechnological techniques offer alternative ways to widen genetic variability. We investigated somaclonal variation in regenerants of yacon derived from in vitro somatic embryogenesis using simple sequence repeat (ISSR) analysis and flow cytometry. Results: Twenty tested ISSR primers provided a total of 7848 bands in 60 in vitro regenerants and control plant. The number of bands for each primer varied from3 to 10, and an average of 6.95 bands was obtained per ISSR primer. Eight primers were polymorphic and generated 10 polymorphic bands with 7.19% mean polymorphism. ISSR analysis revealed genetic variability in 6 plants under study. These regenerants had Jaccard's distances 0.104, 0.020, 0.040, 0.106, 0.163 and 0.040. Flow cytometric analysis did not reveal changes of relative nuclear DNA content in regenerants suggesting that the plants obtained via somatic embryogenesis had maintained stable octoploid levels. Conclusions: Our findings show that indirect somatic embryogenesis could be used in yacon improvement to widen genetic variability, especially when low sexual reproductive capacity hinders classical ways of breedin

Patient reported outcomes and recruitment rates following the introduction of principled patient information leaflets (PrinciPILs): Protocol for a meta-analysis

Background The way potential benefits and harms of trial interventions are shared within patient information leaflets (PILs) varies widely and may cause unnecessary harms (“nocebo effects”). The aim of this meta-analysis will be to evaluate the influence on recruitment rates and early effects on patient reported adverse events of principled patient information leaflets (PrinciPILs) compared with standard PILs. Methods Eligible studies will include those that report the effects on recruitment and patient reported adverse events of PrinciPILs compared to standard PILs. We will include in this meta-analysis all the standard PILs in studies within trials (SWATs) of PrinciPILs that were developed as part of the Medical Research Council (MRC) funded PrinciPIL project. By publishing this as a living meta-analysis, we will allow the meta-analysis to be updated with future SWATs of PrinciPILs. We will use the Cochrane Risk of Bias tool to evaluate the risk of bias for each outcome. We will report the total number of studies and participants analysed and the characteristics of included studies (including details of intervention, comparators, outcomes). For dichotomous data, we will calculate the risk difference and the risk ratio (RR) and 95% confidence intervals (CIs). For continuous outcomes we will use weighted mean differences with 95% CIs or standardized mean differences with 95% CIs. We will investigate heterogeneity by visually inspecting the forest plot and by considering the I2 test result. We will assess the certainty warranted for each outcome using the Grading of Recommendations Assessment Development and Evaluation (GRADE). Ethics approval is not applicable since no original data will be collected. The results will be disseminated through peer-reviewed publication and conference presentations. Discussion We will discuss the limitations of the meta-analysis including study risk of bias, inconsistency, heterogeneity, and imprecision. A general interpretation of the results and important implications will be provided

Co-production of guidance and resources to implement principled participant information leaflets (PrinciPILs) [version 1; peer review: 2 approved with reservations]

Background: The way information about potential benefits and harms of trial is presented within participant information leaflets (PILs) varies widely and may cause unnecessary ‘nocebo’ effects. The Medical Research Council (MRC) funded a project that developed seven principles to reduce this variation. However, guidance has not been produced to facilitate the implementation of the principles. Stakeholder involvement is recommended to optimise the way these principles are disseminated and explained. To co-produce recommendations for developing: (1) user-friendly guidance for users of the principles; and (2) resources that support the implementation of the principles. Methods: We held a co-production workshop with representation from the following professional groups: the Health Research Authority (HRA), research ethics committee members, and trial managers. Two rounds of discussions focused on generating recommendations for guidance and resources that support the implementation of the seven principles. Extensive low inference style ethnographic notes were taken, and the data were analysed thematically using deductive codes. The data was collected on October 14, 2022. Results: 25 participants attended a hybrid workshop. Participants recommended that both researchers designing PILs and research ethics committee members should use the principles, and that that they should be simple, mention both benefits and harms explicitly, include examples of visual representations, and provide the evidence base for the principles. Conclusions: We were able to co-produce recommendations for developing and implementing the seven principles within PILs. These recommendations can now be implemented to reduce unexplained variation in the way potential benefits and harms are shared within PILs

Developing core principles for sharing Information about potential intervention benefits and harms in patient information leaflets using a modified Delphi Survey

Patients need to be informed about potential risks of taking part in clinical trials. A problem is that there are no standards telling researchers or ethics committees how these risks need to be communicated. Our background research suggests that the way harms are communicated can actually increase the risks of the harms occurring. Over half of our sample of 250,000 patients who took placebo pills (like sugar pills) in clinical trials reported some negative side effect (like pain or nausea but also more serious things). The aim of this study is to understand what information about trial harms and benefits stakeholders consider to be important for ‘principled participant information leaflets’ or ‘PrinciPILs’ to contain. The stakeholders will include patients, ethics committee members, industry representatives, medico-legal experts, psychologists, and trial managers

Palliative radiotherapy after oesophageal cancer stenting (ROCS): a multicentre, open-label, phase 3 randomised controlled trial

Background: patients with advanced oesophageal cancer have a median survival of 3-6 months, and most require intervention for dysphagia. Self-expanding metal stent (SEMS) insertion is the most typical form of palliation in these patients, but dysphagia deterioration and re-intervention are common. This study examined the efficacy of adjuvant external beam radiotherapy (EBRT) compared with usual care alone in preventing dysphagia deterioration and reducing service use after SEMS insertion.Methods: this was a multicentre, open-label, phase 3 randomised controlled trial based at cancer centres and acute care hospitals in England, Scotland, and Wales. Patients (aged ≥16 years) with incurable oesophageal carcinoma receiving stent insertion for primary management of dysphagia were randomly assigned (1:1) to receive usual care alone or EBRT (20 Gy in five fractions or 30 Gy in ten fractions) plus usual care after stent insertion. Usual care was implemented according to need as identified by the local multidisciplinary team (MDT). Randomisation was via the method of minimisation stratified by treating centre, stage at diagnosis (I-III vs IV), histology (squamous or non-squamous), and MDT intent to give chemotherapy (yes vs no). The primary outcome was difference in proportions of participants with dysphagia deterioration (&gt;11 point decrease on patient-reported European Organisation for Research and Treatment of Cancer quality of life questionnaire-oesophagogastric module [QLQ-OG25], or a dysphagia-related event consistent with such a deterioration) or death by 12 weeks in a modified intention-to-treat (ITT) population, which excluded patients who did not have a stent inserted and those without a baseline QLQ-OG25 assessment. Secondary outcomes included survival, quality of life (QoL), morbidities (including time to first bleeding event or hospital admission for bleeding event and first dysphagia-related stent complications or re-intervention), and cost-effectiveness. Safety analysis was undertaken in the modified ITT population. The study is registered with the International Standard Randomised Controlled Trial registry, ISRCTN12376468, and ClinicalTrials.gov, NCT01915693, and is completed.Findings: 220 patients were randomly assigned between Dec 16, 2013, and Aug 24, 2018, from 23 UK centres. The modified ITT population (n=199) comprised 102 patients in the usual care group and 97 patients in the EBRT group. Radiotherapy did not reduce dysphagia deterioration, which was reported in 36 (49%) of 74 patients receiving usual care versus 34 (45%) of 75 receiving EBRT (adjusted odds ratio 0·82 [95% CI 0·40-1·68], p=0·59) in those with complete data for the primary endpoint. No significant difference was observed in overall survival: median overall survival was 19·7 weeks (95% CI 14·4-27·7) with usual care and 18·9 weeks (14·7-25·6) with EBRT (adjusted hazard ratio 1·06 [95% CI 0·78-1·45], p=0·70; n=199). Median time to first bleeding event or hospital admission for a bleeding event was 49·0 weeks (95% CI 33·3-not reached) with EBRT versus 65·9 weeks (52·7-not reached) with usual care (adjusted subhazard ratio 0·52 [95% CI 0·28-0·97], p=0·038; n=199). No time versus treatment interaction was observed for prespecified QoL outcomes. We found no evidence of differences between trial group in time to first stent complication or re-intervention event. The most common (grade 3-4) adverse event was fatigue, reported in 19 (19%) of 102 patients receiving usual care alone and 22 (23%) of 97 receiving EBRT. On cost-utility analysis, EBRT was more expensive and less efficacious than usual care.Interpretation: patients with advanced oesophageal cancer having SEMS insertion for the primary management of their dysphagia did not gain additional benefit from concurrent palliative radiotherapy and it should not be routinely offered. For a minority of patients clinically considered to be at high risk of tumour bleeding, concurrent palliative radiotherapy might reduce bleeding risk and the need for associated interventions.Funding: National Institute for Health Research Health Technology Assessment Programme.</p

Developing principles for sharing information about potential trial intervention benefits and harms with patients: report of a modified Delphi survey

Background The way information about potential harms of trial intervention is shared within participant information leaflets (PILs) varies widely and can cause subjective ‘nocebo’ harms. This study aimed to develop principles to improve the composition of information about potential trial intervention benefits and harms within PILs so that variability and avoidable harms are reduced. Methods We conducted a two-round modified online Delphi survey, followed by a consensus meeting. For the first round of the survey, 27 statements were developed based on previous research and relevant guidance from the UK, the USA and the World Health Organization. Participants included members from each of the following stakeholder groups: patient and public representatives, research ethics committee members, industry representatives, medico-legal experts, psychologists and trial managers. Each participant was asked to rate their degree of agreement or disagreement with each statement on a 9-point Likert scale. In the second round, participants were invited to reappraise their ratings after reviewing the results of the first round. Finally, two members from each stakeholder group participated in a meeting to confirm those statements for which there was agreement. Results Two hundred and fifty participants completed round 1, and 201 participants completed round 2. In round 1, consensus was reached for 16 statements. In round 2, consensus was reached for an additional three statements. The consensus meeting confirmed the survey results and consolidated the statements. This process resulted in seven principles: (1) all potential harms of a given intervention should be listed, (2) all potential harms should be separated into serious and less serious, (3) it must be made explicit that not all potential harms are known, (4) all potential benefits should be listed, (5) all potential benefits and harms need to be compared with what would happen if the participant did not take part in the trial, (6) suitable visual representations should be added where appropriate and (7) information regarding potential benefits and harms should not be presented apart by one or more pages. Conclusions Our modified Delphi process successfully generated seven principles that can and should be used to guide how information is conveyed to patients in information leaflets regarding potential trial benefits and harms

Monoclonal anti-endoglin antibody TRC105 (carotuximab) prevents hypercholesterolemia and hyperglycemia-induced endothelial dysfunction in human aortic endothelial cells

15 p.-11 fig.Endoglin (Eng) is a co-receptor of the transforming growth factor β superfamily playing an important role in endothelial dysfunction. TRC105 (carotuximab) is a monoclonal antibody that blocks Eng and its downstream Smad signaling pathway. Here we have investigated for the first time the effects of TRC105 treatment on the development of endothelial dysfunction induced by 7-ketocholesterol (7K) or high glucose (HG), focusing on Eng expression, signaling, and function. In the hypercholesterolemia study, human aortic endothelial cells (HAoECs) were treated with TRC105 (300 μg/ml) for 1 h, followed by the addition of 7K (10 μg/ml) for another 12 h. In the hyperglycemia study, HAoECs were exposed to HG (45 mM) for 60 h, followed by the addition of TRC105 for another 12 h, and cells treated with 5mM glucose and 40 mM mannitol served as control. Protein levels, adhesion, and transmigration of monocytes were assessed by flow cytometry, mRNA expression was measured by qRT-PCR. 7K and HG treatment increased protein levels of NF-κB and Eng and adhesion and transmigration of monocytes through HAoECs monolayer. TRC105 pretreatment reduced the 7K- or HG-induced Eng protein levels and pSmad1/5 and pSmad2/3 signaling. Despite increased protein levels of P-selectin and VCAM-1, TRC105 mediated blockage of Eng prevented 7K- and HG-induced adhesion and transmigration of monocytes through endothelial monolayers. These results suggest that TRC105-mediated Eng blockage can counteract the hypercholesterolemia- and hyperglycemia-induced endothelial dysfunction in HAoECs, suggesting that Eng might be a potential therapeutic target in disorders associated with elevated cholesterol and glucose levels.This research was supported by grants from the Grant Agency of Charles University (GAUK No. 1130120), Czech Science Foundation (GACR 22-14961S), Specific University Research (SVV 260 549), efficiency and safety improvement of current drugs and nutraceuticals: advanced methods new challenge grant (EFSA-CDN; No. CZ.02.1.01/0.0/0.0/16_019/0000841), and by Consejo Superior de Investigaciones Científicas (CSIC), grant number: 201920E022.Peer reviewe

Patterns in recent and Holocene pollen accumulation rates across Europe - the Pollen Monitoring Programme Database as a tool for vegetation reconstruction

The collection of modern, spatially extensive pollen data is important for the interpretation of fossil pollen assemblages and the reconstruction of past vegetation communities in space and time. Modern datasets are readily available for percentage data but lacking for pollen accumulation rates (PARs). Filling this gap has been the motivation of the pollen monitoring network, whose contributors monitored pollen deposition in modified Tauber traps for several years or decades across Europe. Here we present this monitoring dataset consisting of 351 trap locations with a total of 2742 annual samples covering the period from 1981 to 2017. This dataset shows that total PAR is influenced by forest cover and climate parameters, which determine pollen productivity and correlate with latitude. Treeless vegetation produced PAR values of at least 140 grains cm−2 yr−1. Tree PAR increased by at least 400 grains cm−2 yr−1 with each 10 % increase in forest cover. Pollen traps situated beyond 200 km of the distribution of a given tree species still collect occasional pollen grains of that species. The threshold of this long-distance transport differs for individual species and is generally below 60 grains cm−2 yr−1. Comparisons between modern and fossil PAR from the same regions show similar values. For temperate taxa, modern analogues for fossil PARs are generally found downslope or southward of the fossil sites. While we do not find modern situations comparable to fossil PAR values of some taxa (e.g. Corylus), CO2 fertilization and land use may cause high modern PARs that are not documented in the fossil record. The modern data are now publicly available in the Neotoma Paleoecology Database and aid interpretations of fossil PAR data.publishedVersio

Genetics of Host Response to Leishmania tropica in Mice – Different Control of Skin Pathology, Chemokine Reaction, and Invasion into Spleen and Liver

Several hundred million people are exposed to the risk of leishmaniasis, a disease caused by intracellular protozoan parasites of several Leishmania species and transmitted by phlebotomine sand flies. In humans, L. tropica causes cutaneous form of leishmaniasis with painful and long-persisting lesions in the site of the insect bite, but the parasites can also penetrate to internal organs. The relationship between the host genes and development of the disease was demonstrated for numerous infectious diseases. However, the search for susceptibility genes in the human population could be a difficult task. In such cases, animal models may help to discover the role of different genes in interactions between the parasite and the host. Unfortunately, the literature contains only a few publications about the use of animals for L. tropica studies. Here, we report an animal model suitable for genetic, pathological and drug studies in L. tropica infection. We show how the host genotype influences different disease symptoms: skin lesions, parasite dissemination to the lymph nodes, spleen and liver, and increase of levels of chemokines CCL2, CCL3 and CCL5 in serum