Disorders of gut-brain interaction (DGBI), formerly termed functional gastrointestinal disorders (FGID), are highly prevalent although exact pathophysiological mechanisms remain unclear. Intestinal immune activation has been recognized, but increasing evidence supports a pivotal role for an active inflammatory state in these disorders. In functional dyspepsia (FD), marked eosinophil and mast cell infiltration has been repeatedly demonstrated and associations with symptoms emphasize the relevance of an eosinophil-mast cell axis in FD pathophysiology. In this Review, we highlight the importance of immune activation in DGBI with a focus on FD. We summarize eosinophil biology in both homeostasis and inflammatory processes. The evidence for immune activation in FD is outlined with attention to alterations on both cellular and molecular level, and how these may contribute to FD symptomatology. As DGBI are complex and multifactorial conditions, we shed light on factors associated to, and potentially influencing immune activation, including bidirectional gut-brain interaction, allergy and the microbiota. Crucial studies reveal a therapeutic benefit of treatments targeting immune activation, suggesting that specific anti-inflammatory therapies could offer renewed hope for at least a subset of DGBI patients. Lastly, we explore the future directions for DGBI research that could advance the field. Taken together, emerging evidence supports the recognition of FD as an immune-mediated organic-based disorder, challenging the paradigm of a strictly functional nature. Functional dyspepsia (FD) is a chronic gastrointestinal (GI) disorder affecting nearly 10% of the population as surveyed by internet questionnaires and personal interviews worldwide . FD can Cardinal symptoms of FD include postprandial fullness and early satiation, both meal-related symptoms that identify the subgroup of postprandial distress syndrome (PDS). Patients suffering from painful symptoms such as epigastric pain and burning are classified as epigastric pain syndrome (EPS) subtype . AccordiAlthough the presence of major structural alterations is not in line with the current Rome IV diagnostic criteria, microscopic duodenal alterations have become increasingly recognized as a part of FD pathophysiology. Low-grade immune activation in the duodenum, manifested by increased mucosal eosinophil and mast cell numbers, was associated with increased permeability of the duodenal epithelium , challenKnown risk factors for FD are female gender, anxiety and use of non-steroidal anti-inflammatory drugs or antibiotics . Other fIn this Review, we provide a narrative overview on immune activation in DGBI with a focus on FD and particular attention for the role of eosinophils in homeostasis and inflammation. We also discuss various factors associated to immune activation and how they can contribute to FD pathophysiology, before we address overlapping symptomatology and inflammatory differences with other DGBI. Finally, we discuss the evidence for anti-inflammatory therapies in FD and explore future directions in the field.4β7 in order to migrate into the small intestine and colon 6-dependent mechanism  , inflammin vitro  as well in vitro . Th2-celin vitro . Eosinopistamine . Moreoveistamine . Alternaistamine . Eosinop FD patients, implicating other inflammatory cell types in the observed immune activation  lymphocytes which were associated to various GI symptoms and gastric emptying (GE)    . This suRemarkably, only few studies reported alterations in the immune cell profile of the stomach and none were related to FD symptoms , therebyp = 0.03] and mast cell  counts in the stomach were indeed increased in FD patients compared to controls, with even more convincing evidence for duodenal eosinophil  and mast cell  infiltration, but not for other cell types  = 0.36, ll types . Conflicvice versa remains to be determined. These two pathophysiological events most likely amplify each other, leading to a vicious circle, which complicates untangling these mechanisms even more.Besides increased mucosal infiltration of inflammatory cells and subsequent release of their granular components, altered presence of soluble factors in the mucosal environment further substantiates the evidence for immune activation in FD . A recenEvidence for neuro-immunological mechanisms on the molecular level include increased inducible nitric oxide synthase (iNOS) in the duodenum, which was linked to mast cell degranulation as well as upper abdominal distention scores in PDS patients . In addi1R)-mediated sensitization of TRPV1 contributed to the development of visceral hypersensitivity -α and IL-1β act as potent activators of the HPA axis, confirming the relation between stress and immune activation. These cytokines can exert stimulatory effects on CRH release in the brain, thereby inducing a stress response . In an IAs eosinophils and mast cells are classically involved in allergic conditions, allergy and atopy are indeed associated with DGBI. Duodenal eosinophilia was common in allergic patients {odds ratio (OR) [95% confidence interval (CI)] 5.04 [2.12–11.95]} without an association to specific upper GI symptoms . Asthma In spite of multiple studies linking allergies to DGBI, these patients do not experience more IgE-mediated food responses compared to the general population. The prevalence of food hypersensitivity in DGBI patients was estimated to be 4% (8/200) , which iBesides, the relatively new clinical entity of non-celiac gluten sensitivity (NCGS), defined as both GI and extra-intestinal reactions in response to gluten intake in the absence of wheat allergy and celiac disease, shares a highly similar symptom pattern to DGBI . Indeed,A study in IBS found that 84% of 197 included patients experienced symptoms after intake of specific foods, and multiple food triggers were linked to more severe symptoms and lower quality of life . Indeed,Taken together, atopy, allergies and untoward food reactions are linked to various DGBI although more mechanistic studies are needed to untangle the immunological maze of factors contributing to overlap between these conditions, and mechanisms primarily driving GI symptom generation, respectively. In addition, studies focusing on the immune-related pathophysiology in DGBI should acknowledge the presence of atopy and allergy as potential confounders in order to isolate the drivers of DGBI symptoms . Only byex vivo findings were corroborated in vivo using CLE  on intestinal inflammation were suggested by oral butyrate administration resulting in reduced colonic infiltration of neutrophils and eosinophils in mice during colitis, but not in healthy mice . These bFrequently overlapping symptom patterns with other DGBI complicate the diagnosis and management of FD. FD and IBS are the most prevalent DGBI  and overWhereas FD presents with symptoms thought to originate from the gastroduodenal region, IBS presents with lower GI complaints such as constipation, diarrhea and abdominal pain. Moreover, the inflammatory infiltrate in IBS predominantly consists of mast cells, initially reported in the terminal ileum  and cecu+ macrophage infiltration in the myenteric plexus of the gastric wall   or non-iA better understanding of cellular and molecular changes underlying the pathophysiology in different DGBI is essential to fulfill the diagnostic and therapeutic unmet needs in the field. Eventually, implementing objective measures, of which the intestinal immune activation profile appears to be a prime candidate, in addition to symptomatologic evaluation as part of the diagnostic work-up could promote faster and more accurate diagnosis, and allow targeted therapies to be developed.Helicobacter pylori-positive patients  antagonists suppress gastric acid secretion, albeit less powerful but with similar efficacy compared to PPI , IL-25 and IL-33, which activate eosinophils as part of a Type 2 allergic response, often in a cooperative manner and involving ILC2 induction , 2018. AResearch in the field of DGBI is rapidly evolving, and although the presence of intestinal immune activation in these disorders has widely been recognized, increasing evidence supports the concept of an elevated immune response as key pathophysiological event in DGBI. Increased infiltration and degranulation of both eosinophils and mast cells is frequently linked with typical GI symptoms, while treatments reducing local immune activation also yield therapeutic benefit. Immune activation should therefore be regarded as an important contributor to DGBI pathophysiology, and both direct anti-inflammatory therapies, as well as preventive or therapeutic approaches indirectly leading to reduced immune cell infiltration, should be studied further.Furthermore, the separate events in the triad of mucosal alterations, DGBI symptomatology and psychosocial comorbidities all have been demonstrated independently, while cross-links have been made by associations between those factors. Nevertheless, describing the cause-consequence relationship between the three aforementioned manifestations continues to be one of the major challenges in DGBI research and should have the highest priority in future studies.MC wrote the original draft. All authors contributed to the manuscript, revised and approved the final version of the review.LW reports financial support for research from Danone and MyHealth; has served on the advisory board of Naturex; has served on the Speaker bureau for Falk Pharma, Takeda and MyHealth. TV reports financial support for research from Danone, MyHealth, Takeda and VectivBio; has served on the Speaker bureau for Abbott, Falk Pharma, Fresenius Kabi, Menarini, Remedus, Takeda, Truvion and VectivBio; reports consultancy fees from Baxter, Falk Pharma, Takeda, VectivBio and Zealand Pharma. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Global plastic production has increased exponentially since the 1960s, with more than 6300 million metric tons of plastic waste generated to date. Studies have found a range of human health outcomes associated with exposure to plastic chemicals. However, only a fraction of plastic chemicals used have been studied in vivo, and then often in animals, for acute toxicological effects. With many questions still unanswered about how long-term exposure to plastic impacts human health, there is an urgent need to map human in vivo research conducted to date, casting a broad net by searching terms for a comprehensive suite of plastic chemical exposures and the widest range of health domains.2017 Guidance for the Conduct of Joanna Briggs Institute (JBI) Scoping Reviews, to be reported in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews (PRISMA-ScR) Checklist. A literature search of primary clinical studies in English from 1960 onwards will be conducted in MEDLINE (Ovid) and EMBASE (Ovid) databases. References eligible for inclusion will be identified through a quality-controlled, multi-level screening process. Extracted data will be presented in diagrammatic and tabular form, with a narrative summary addressing the review questions.This protocol describes a scoping review that will follow the recommended framework outlined in the This scoping review will comprehensively map the primary research undertaken to date on plastic exposure and human health. Secondary outputs will include extensive databases on plastic chemicals and human health outcomes/impacts.https://archive.org/details/osf-registrations-gbxps-v1, 10.17605/OSF.IO/GBXPSOpen Science Framework (OSF)-Standard Pre-Data Collection Registration, The online version contains supplementary material available at 10.1186/s13643-022-02010-6. Continuous human exposure to plastic chemicals has become inevitable due to its ubiquity in the environment , 2. ‘PlaAs of 2017, approximately 8300 million metric tons (Mt) of virgin plastic chemicals had been produced, and as of 2015, 6300 million Mt of plastic waste generated –11 and mPlastic materials are used in every aspect of our lives  and commOf the thousands of plastic chemicals in use globally today, more than 20% have been identified as substances of concern by the European Union on the basis of persistence, bioaccumulation and/or toxicity, while 39% remain unclassified in terms of hazard . Some plWe are not aware of any sources or publications comprehensively summarising research conducted to date on how plastic chemicals and plastic particles are impacting human health, or which populations, demographically and geographically, have been studied in this respect. Preliminary searches conducted in Prospero, Epistimonikos, Cochrane, MEDLINE (Ovid) and Embase (Ovid) databases in May 2020 did not identify any previous or current scoping review on this broad topic. A number of systematic and literature reviews were identified, focusing on a small number of classes of plastic chemicals  and bisphenol A –55 and cTo demonstrate the scope of research conducted to date, and to inform future research efforts, we will conduct a scoping review to chart published primary human in vivo studies that investigated the relationship between exposure to plastic materials and human health since 1960, when global plastic production began to increase exponentially . The scoIn primary human in vivo research published since 1960 on the effects of plastic materials on health, which plastic chemicals/particles and which health domains have (and have not) been investigated?Which populations  have been examined?Which study designs were adopted?When were these studies conducted over time (Jan 1960–Jan 2021)?In relation to our main question, the following questions will inform sub-analyses:The Population, Exposure and Outcome (PEO) mnemonic Table  was creaThe review will consider studies that investigate any population globally, and any age group, including the developing foetus. Animal studies and in vitro human studies will be excluded.Only in vivo directly measured exposures  of plastic materials that meet inclusion criteria will be included. All avenues of exposure are considered relevant, including environmental contact via activities of everyday living, occupational exposure, accidental ingestion through contaminated food or water, cross-placental exposure and ingestion through breast milk.additive — plasticiser, additive — flame retardant, per- and polyfluoroalkyl substances (PFAS), individual polymer or polymer class and bisphenols will be included, unless commonly used in biomedical applications or as food additives . Additional plastic chemicals identified during the screening process, or missing plastic chemicals that are structurally and systematically related to included substances , will undergo the same selection criteria and subsequently included or excluded. Polymer metabolites, monomers other than bisphenols and plastic additives other than plasticisers and flame retardants are excluded from this review.This scoping review focuses on plastic materials to which humans are generally exposed, other than at the point of synthesis or product manufacturing. A systematic decision-making process Fig.  will be A comprehensive and searchable database of included plastic chemicals will be developed, indexing the multiple names, synonyms and commonly used terminology describing these chemicals.Only physiological and psychological changes in human health that are directly measured in vivo are included.Measures of direct impact on human health as a diagnosis of illness or disease, for example hypothyroidism;Measures of physiological or biochemical change associated with a disease/disorder where no illness/disease has been diagnosed, for example oestrogen levels as a measure of change in ovarian function; andMeasures of physiological or biochemical change within an organ/body system that are not associated with a specific disorder, for example genetic and epigenetic changes.Health outcome measures include any assessment of change in structure or function of the human body, whether undertaken at the clinical, functional, physiological or cellular level. From preliminary searches, we anticipate encountering three types of health outcome measures in relation to plastic material exposures:International Classification of Disease 11th Revision (ICD-11) [Health outcome measures listed above will be classified using the hierarchical structure of the (ICD-11) , with anThis scoping review considers published peer-reviewed journal articles presenting primary human research, inclusive of all study designs. Excluded sources are literature/systematic reviews, meta-analyses, editorials, book reviews/chapters, perspectives, opinion/commentary pieces, conference abstracts and proceedings and grey literature.Updated methodological guidance for the conduct of scoping reviews [Preferred reporting items for systematic reviews and meta-analyses extension for scoping reviews (PRISMA-ScR) Checklist [Conducting high quality scoping reviews – challenges and solutions [The scoping review uses the methodological framework as described in the  reviews  and repohecklist   and Embase (Ovid) databases and examining the hierarchy of controlled vocabulary in these databases (MeSH and Emtree respectively). Indexed terms were selected representing plastic materials  and a broad range of health outcome categories informed by preliminary searches and expert input from author CS, a consultant paediatrician with relevant research expertise . MeSH and Emtree terms, corresponding text words, text words contained in the titles and abstracts of key articles, and terms and synonyms for all specific plastic chemicals included in our database, will comprise the final search strategy, being developed in consultation with research librarian, author TS. A large number of exclusion terms such as ‘plastic surgery’, plasticity, etc., will be utilised to minimise irrelevant results.In summary, the search strategy will include the following: Health outcomes AND plastic material exposures NOT exclusions, limited to English language and dates ranging from 1960 to current. A search filter will be used in each database  to identify human-only studies and exclude animal studies –81. The Scopus and Web of Science databases were investigated as sources; however, due to the lack of controlled vocabulary and no filters for human-only studies, these will not be searched.in preparation).In anticipation of a very large number of results from this search, the reference lists of retrieved papers will not be reviewed. We will, however, review the reference lists associated with a related ‘Umbrella Review’ of systematic reviews with meta-analyses on this topic, being conducted by the Minderoo Foundation and reported separately  systematic review software application for screening and data extraction and deduplicated. Relevant articles will then be selected via two screening stages, each preceded by an in-depth training process for the team of reviewers, who have postgraduate and doctoral degrees in chemistry, biochemistry, environmental and health sciences. Training will involve the provision of guidelines, including inclusion/exclusion criteria and practical examples of their application, education on classifying study design, access to our database of included plastic chemicals and demonstration of the screening process using a predetermined set of references to be included for the scoping review, ensuring consistency and reliability of assessment decisions.References will first be screened for relevance by title, abstract and index terms. Reviewers will initially screen several sets of the same 100 references and discuss their screening decisions, proceeding to single-screening after achieving an 85% level of agreement with subsequently agreed (resolved) group decisions over three sets. References without abstracts but potentially relevant to the scoping review will be included at the first screening stage. Auditing will be performed on a random selection of screened references to evaluate screening accuracy, and we will use the inbuilt artificial intelligence (DistillerAI) tool to identify possible false exclusions, re-screening references as necessary and thereby minimising the number of relevant references being discarded.During the second screening stage, full-text articles of each reference will be reviewed for eligibility. Training will again involve the screening of several sets of references, with discussions after each set about screening decisions. Reviewers will proceed to single-screening only after achieving an 85% level of agreement with subsequently resolved group decisions and erroneously excluding no more than two per 100 eligible references. Eligible references will proceed to data extraction, whereas references excluded once will require a second reviewer to confirm the exclusion. Any disagreements will be resolved by a third reviewer with relevant experience. The selection process will be reported via a PRISMA flowchart Fig. .The data to be extracted will include specific publication details, study characteristics, relevant plastic material exposures detected in the body and the health outcome measures analysed in relation to exposures. In relation to study design, we anticipate a large number of epidemiological studies involving multiple populations and/or time points of assessment. A simple classification of study design will therefore be implemented, with studies classified as ‘Experimental’ , ‘Longitudinal’ , ‘Cross-sectional’  or ‘Case Study/Series’ . Data extraction forms designed in DistillerSR , this scoping review aims to identify a wider suite of health outcome measures that should be taken into consideration for plastic material exposure research. This database could also benefit future research endeavours.In summary, this scoping review will provide an overview of the research so far undertaken on plastic material exposure and human health. This can serve as a basis to identify research clusters, by topic and geographical location of studied populations, as well as knowledge gaps, helping to prioritise specific areas in need of more in-depth research. Additionally, alongside the primary outputs, findings may be used to strengthen efforts to better regulate the production, importation and use of plastic materials that human populations are ubiquitously exposed to.Additional file 1. Completed PRISMA-P checklist.Additional file 2. Draft data extraction forms.


Adolescents and young adults (AYA) in low to middle income countries (LMIC) have poorer outcomes along each step in the HIV continuum of prevention and care compared to younger children or older adults. The use of mHealth technology provides a potentially promising implementation strategy for interventions to remedy these disparities. We therefore conducted a systematic review of the English literature and conference proceedings from January 1, 2000 to April 1, 2021 evaluating mHealth interventions targeting AYA along each step of the HIV continuum of care in LMIC. We identified 27 mHealth interventions across the HIV continuum, with no interventions addressing transition from pediatric to adult care. The majority of studies were single arm, uncontrolled or underpowered, with few randomized trials resulting in mixed and inconclusive outcomes. mHealth interventions have potential to remedy disparities along the HIV continuum of care for AYA in LMIC but larger, powered randomized trials are needed.The online version contains supplementary material available at 10.1007/s10461-022-03840-0. In 2020, there were an estimated 1.75 million adolescents between the ages of 10 and 19 living with HIV globally, with only about half receiving antiretroviral treatment –5. AdoleAYA in particular experience poorer HIV outcomes at each step of the care continuum compared to younger children or older adults. Despite the scale up of global antiretroviral therapy (ART), including international guidelines supporting early initiation of ART with simple, potent, and well-tolerated regimens , adolescmHealth, the use of mobile wireless technologies for health, may improve the disparities along the AYA HIV continuum of care. Greater than 80% of the population in LMIC access health information and services through mHealth that can address healthcare delivery gaps across the care continuum –23. mHeaOur systematic review evaluates mHealth interventions addressing the HIV continuum of care, including HIV prevention, HIV testing, linkage to care, adherence/retention in care, viral suppression and transition to adult care that have been designed for and evaluated among AYA in LMIC.Appendix 1 for search terms. We conducted six independent searches for each step in the HIV continuum of care. To be eligible, studies had to meet the following inclusion criteria: (1) original peer-reviewed studies published in English; (2) involve mHealth interventions addressing HIV prevention and the AYA HIV continuum of care that have been piloted, implemented, or evaluated in LMIC (as defined by the World Bank Group); and (3) include interventions where the median age of participants was between 10 and 25 years. For inclusion in our review, we defined interventions as having been purposefully designed and prospectively assigned to influence bio/behavioral outcomes related to HIV. We defined mHealth as clinical and public health practices supported by the provision of health services and information via mobile technologies, such as mobile phones, tablet computers and personal digital assistants (PDA) [We searched PubMed and Google Scholar from January 1, 2000, to April 1, 2021 and online conference proceedings from the International AIDS Society (IAS), International Workshop on Pediatric HIV/AIDS, the International AIDS Conference (AIDS), the Conference on Retrovirology and Opportunistic Infections (CROI), and HIV Research 4 Prevention (R4P) from January 1, 2019, to April 1, 2021. Key words and medical subject headings relevant to age  were cross-referenced with terms associated with mHealth  and each step in the HIV continuum of care . See ts (PDA) . We exclFollowing the search, all identified references were imported into Covidence, a systematic review management system, and were de-duplicated. Eight authors  screened the remaining citations by title, abstract, and full-text against pre-determined, aforementioned inclusion and exclusion criteria by independent reviewers for each step in the HIV continuum of care, with discrepancies resolved by consensus.After meeting our inclusion and exclusion criteria, we then extracted data from the full text articles/abstracts and created summarized tables organized by each step in the HIV continuum of care. We created a template for study extraction including study characteristics and findings. The template included the following: authors, title, year, country, age range, number of subjects, intervention description, and results of the intervention. For studies that included samples from multiple age groups, we extracted only the data that pertained to adolescents and young adults.Fig. Overall, we reviewed 912 articles and 90 conference abstracts in our initial screening  related and non-PrEP prevention interventions. Independent searches for PrEP related and non-PrEP HIV prevention interventions were conducted to individually highlight the intervention strategies and outcomes for both the biomedical and behavioral approaches to HIV prevention.We identified nine studies in which mHealth was used as or with an intervention to promote PrEP adherence among AYA in LMIC Fig. a. These Four randomized controlled trials (RCTs) have been published to date, none of which found a benefit from mHealth , 40, 51.Our search of abstracts revealed five additional studies. In a conference abstract about the POWER demonstration project, Celum et al. reported high PrEP initiation but low persistence when PrEP was delivered in multiple youth-friendly models in South Africa and Kenya; SMS, WhatsApp, and phone calls were utilized for PrEP refill reminders . Pintye Our search found four interventions that have been published addressing mHealth interventions for non-PrEP related HIV prevention among AYA in LMICs Fig. b. These Several studies were published on the development, pilot testing and outcomes of Tumaini, an interactive narrative-based game aimed to prevent HIV through improving health-related knowledge and skills. The preliminary studies examining the acceptability, feasibility, and mechanisms of effect of the mHealth intervention –60 were The three remaining HIV prevention studies targeted increased HIV prevention knowledge and decreasing HIV risk behaviors. Ybarra et al. conducted a beta test of In This toGether, a text-messaging based HIV prevention program aimed to increase condom use and STI/HIV testing, among 34 older adolescents in Uganda . They foWe identified seven mHealth interventions addressing HIV diagnosis and linkage to care among AYA in LMIC Fig. c. These The technology approaches to promote HIV testing and linkage to care included tablet-based education and service linkage facilitated by field workers; smartphone and web-based psychoeducational and HIV testing facilitation apps; SMS-based educational messages; and social media and geosocial outreach. These approaches were very often facilitated by peers .In terms of the tablet-based, facilitated approaches, an interactive tablet-based psychosocial app tailored to men, EPIC-HIV, was used by fieldworkers in rural KwaZulu-Natal, South Africa, to promote uptake of rapid HIV testing and linkage to care by targeting intrinsic motivation and self-determination. The fieldworkers facilitated the intervention immediately prior to an invitation to complete testing as part of an annual HIV surveillance program . SimilarMulakat sought to increase HIV testing among MSM via peer mobilization in virtual spaces, including social and geosocial media  [Mulakat was evaluated in a single arm post-test only design, finding relatively low seroprevalence of 3.2%, with a high percentage of first-time testers (99%), but only 50% linked to care. The virtual mentors program used a prospective case-control design, with each mentee matched to two control patients with similar HIV testing and counseling dates. In the intervention group 80% linked to ART versus 43% in the control group, although the time to linkage was much longer in the intervention group.Two interventions used existing social media apps to promote HIV testing and linkage to care among youth. In Mumbai, India, Project  Grindr) . Initial Grindr) . ProjectTwo interventions used smartphone or web apps to promote HIV testing. As described above, Garg and colleagues developed an Android-based app, RumahSELA, to promote HIV testing among key populations including MSM, TGW and PWUD in several provinces of Indonesia, facilitated by peers . In addiFinally, in Kambu County, Central Kenya, an intervention targeted to young college women promoted HIV testing via SMS text messages focused on sexual and reproductive health by increasing knowledge, risk perception and promoting risk reduction . Weekly Our search yielded seven peer-reviewed articles and one conference abstract reporting effect of mHealth interventions on ART adherence or retention in care in LMIC youth Fig. d. Of theFour studies reported SMS-based interventions –48, 50, Of the 8 studies included, one reported a significant association between the intervention and ART adherence and one reported significant improvement in viral suppression , 50. SánOur search identified three peer-reviewed articles evaluating mHealth interventions on viral load or viral suppression among AYA in LMICs Fig. e 36, 4444, 46. TThe three studies which met inclusion criteria used varied platforms , all included the option for SMS messaging. The mHealth interventions were delivered by peers (youth living with HIV) in one study, while the others included SMS-based messages from research staff and healthcare providers. Two studies focused on non-adherent/non-suppressed youth and one on youth newly initiating ART; only one study was an RCT. Abiodun et al.’s RCT (as described above in adherence/retention) evaluated an SMS intervention among youth in Nigeria who were non-suppressed and found a significantly higher rate of viral suppression at 20 weeks follow-up in the intervention arm . The twoFig. Our systematic review found did not find any relevant articles in the published literature of mHealth interventions addressing healthcare transition for AYA living with HIV in LMICs as indicated in AYA comprise a growing number of individuals living with HIV in LMIC; however, the percentage of AYA who are aware of their diagnosis, retained in care and virally suppressed remains low , 11–14. Many of the studies used mHealth as an implementation strategy as a component of a more complex intervention; however, few studies used implementation science frameworks or measured implementation outcomes such as cost, fidelity, and/or sustainability, thus, limiting the ability to measure the effectiveness of mHealth in isolation. In studies that did not find benefit when using mHealth, it is unclear whether the intervention itself, the implementation strategy, study design or lack of power, contributed to ineffectiveness. Future studies involving mHealth should be conceptualized within an implementation science framework to adequately measure effectiveness of mHealth as an implementation strategy.Despite the popularity of mobile communications among youth generally and mHealth within the HIV field, a relatively small number of mHealth interventions have been developed to promote PrEP adherence among adolescents in resource-limited settings –42, 51. Case finding and diagnosis is the critical first step in the HIV care continuum and AYA in LMIC are at high risk of going undiagnosed . We idenThere was a shortage of adequately powered rigorously designed evaluations of mHealth interventions for ART adherence and retention in care in YLWH. We found only four randomized evaluations that were conducted with the primary aim of assessing clinical effect , 47, 50.There is a lack of mHealth interventions targeting AYA living with HIV transitioning to adult-oriented care despite an expected oncoming wave of youth living with HIV aging into adulthood and requiring adult-oriented services in the next 5–10 years –72. DespDespite the overall increase in mobile phone ownership and use in recent years worldwide, AYA remain a challenging population to effectively engage in mHealth, especially in LMIC. In particular, mHealth interventions have difficulty engaging individuals of lower socioeconomic status, especially AYA who may not have their own phone or the ability to pay for data plans. Previous studies have shown that while most AYA in LMICs have access to a mobile phone, it can be either through owning or through borrowing one from someone else, including parents, siblings, friends, or other family members . SeveralWhile there is increasing use of mHealth interventions to improve health outcomes along the HIV continuum of prevention and care for AYA in LMIC, study outcomes are mixed. This is in part due to intervention design and difficulty evaluating individual intervention components, in addition to a lack of large-scale randomized-controlled trials powered to detect group differences. Although several studies comment on acceptability and/or feasibility of the intervention, there is a lack of rigorous implementation design in order to assess key components of the intervention and its implementation process. Future studies can utilize an implementation framework including using mHealth as an implementation strategy in combination with other interventions  to measure intervention effectiveness, costing, fidelity and sustainability.This review had several limitations. We included mHealth interventions that were specifically targeting AYA in LMIC and did not include larger studies with broader age ranges where some youth were included. Additionally, mHealth is a broad term and comprises a variety of interventions of varying intensities, from simple text messages to more complex interventions with interactive games and peer interaction, making an evaluation of mHealth interventions challenging. Many studies described in this review were one-off or short term-interventions. Studies of long-term engagement in care, adherence, and viral suppression are required. Many of the mHealth intervention were smaller pilot studies that have not been scaled widely, despite their promise. Despite these weaknesses, the results from most studies largely support the potential efficacy of mHealth approaches among populations of youth in LMICs, particularly key populations.The use of mHealth technology is a promising implementation strategy to address the disparities along the HIV continuum of prevention and care for AYA in LMIC. There is a high degree of acceptability and feasibility of mHealth among AYA; however, results of studies are conflicting due to predominance of uncontrolled-single arm studies with only a few adequately powered randomized trials. There is a need to utilize the principles of implementation science to better design, evaluate, and eventually scale, delivery of effective mHealth interventions for AYA in LMIC. While none of the studies evaluated mHealth interventions during the COVID-19 pandemic, the ability of mHealth to be delivered without face-to-face contact is an attractive service delivery model during a pandemic and should be further evaluated.Below is the link to the electronic supplementary material.Supplementary Material 1


Literature reports superior performance when focusing one’s attention during a movement on environmental effects of that movement  compared to focusing on the moving body . Nevertheless, IF instructions still play an important role in the daily practice of coaches, trainers, and therapists. The current review compiles evidence for focus-of-attention concepts on movement form corrections and technique training. Reviews on the topic and selected additional papers addressing the effect of attentional focus on movement form or on kinetic, kinematic or muscle activity data were included. Both EF and IF instructions affect movement form. The reviews revealed that IF instructions seem to be better applicable to direct movement form changes than EF instructions. In contrast, EF instructions better facilitate optimization within the whole-body coordination, often resulting in better performance outcomes not directly linked to movement pattern changes. Several studies discuss focus-of-attention effects in the context of the optimal feedback control theory expanding on the constrained action hypothesis. In summary, EF and IF instructions both affect form and performance of movements, however, their relative efficacy is situation dependent. The often-purported superiority of EF over IF instructions cannot be generalized to all application contexts. Instruction and feedback on how a movement is performed are important interactions between coaches and athletes and play an important role in the development of highly skilled movement performances. How exactly these instructions are worded might draw athletes’ attention on how their body performs the movement ; or might direct their attention towards an external goal within the environment  . For morThe observed difference in movement performance outcomes due to attentional focus has been explained by the constrained action hypothesis , saying However, in contrast to what experimental results and theoretical considerations suggest, IF still plays an important role in the daily practice of coaches, athletes, and movement therapists. In some application domains, IF is even the dominant form of instruction and feedback ,17,18. FThe number of studies investigating focus of attention has continuously increased over the last years, as has the number of systematic reviews and meta-analyses on the topic. While in 2013 Wulf reviewed a set of 53 studies, in 2021 Chua and colleagues provided a meta-analysis of 143 studies ,11. Withidentify topic areas for which evidence regarding focus of attention effects is provided and discussed in the available scientific literature;specifically consider EF and IF instructions towards movement form and movement technique corrections.The current paper intends to appraise evidence of focus-of-attention concepts in movement form corrections to support or decline a paradigm change to EF usage within sport-technical and therapeutic teaching. Thereto it critically assesses and summarizes results of systematic reviews and selected additional recent studies to:The discussion of these points is intended to provide a theoretical framework beyond the constrained action hypothesis for the focus of attention effects.The current study is a narrative review of the up-to-date knowledge on focus instructions applicable to physiotherapy or movement technique training. PubMed, PSYNDEXplus and Web of Science were searched for reviews using the terms “focus of attention”, “motor skill”, or “motor performance”. Only papers written in English were considered. Reviews published since 2011, thus representing the last 10 years of research, were scrutinized. To assess the magnitude of differences, effects sizes were compiled from systematic reviews and are presented here with Cohen’s d , or with Hedge’s g (with the same ranges). Supplemental articles were added if they addressed the effect of attentional focus on movement form or movement technique and reported on kinetic, kinematic or muscle activity differences. Some articles included in the reviewed reviews were selected for further discussion if they helped clarify specific focus effects.Thirteen reviews were found, summarizing more than 250 studies investigating the focus of attention effects in instructions see . Five diMotor balance: In particular balance tasks seem to benefit from EF conditions , providiExplosive skills: Similar to the benefit of EF in balance tasks, performance output of explosive jumps and muscular maximum strength increased when EF conditions were applied ,22. EF iYouth sport: Concerning youth sport, two reviews indicate inconsistent results. Barillas et al. reported 35 studies with contradictory outcomes, concluding with preference of EF instructions while respecting person-specific responses . AnotherTraining with patients: Consideration of focused instructions on physio-therapy and training with patients was summarized by four reviews, two of them criticizing insufficient evidence for an endorsement of EF ,29. HoweMotor coordination and skill technique: Out of 258 studies gathered in the different reviews, only 43 addressed changes in movement form or movement patterns due to focused instructions. Effect sizes compiled in the extensive review of Chua and colleagues  on motorTaken together, the results corroborate the way instructions are worded and thus the way attention is focused on aspects of the movement does affect movement performance and does modify movement execution. A large proportion of the scientific literature indicates advantages in EF instructions over IF instructions with regard to movement outcome and learning. However, a closer examination specifically at adaptations in movement-coordination reveals some uncertainty about the expected effects. To classify focus effects on movement technique in more detail, three main questions will be discussed. First, what is the impact on movement form when using EF or IF? Second, how can we apply this knowledge for instructing technique training? And third, what kind of explanation and motor control model could fit to observed behaviors?Comparing the effect of instructions used for jumps, typically like “push the floor behind” (EF) versus “open your knee angle explosively” (IF), led to higher jump length or jump height in EF condition ,33. ResuComparable observations were made in continuous movement patterns, like running. Movement technique measures in sprinting revealed that expert athletes showed no difference in sprint time or in push-off forces when using IF or EF, whereas collegiate athletes produced slower sprint times with IF, but again without differences in the push-off forces . In anotSuperiority of EF instructions due to enhanced performance outcome is commonly attributed to the assumption of executing better suiting movement patterns. In contrast, studies show inconsistent results. There is evidence in the literature that specific instructions (EF as well as IF) induce movement changes exactly at the addressed movement part. For example, Wulf et al.  implemenThe intense search for studies investigating movement form when using EF or IF instructions showed that only a few studies on this specific topic have been conducted so far. This mirrors critical differences in the measurement complexity of motor skills observed in focus studies. The relative sparsity of such studies might have two distinct reasons. First, performance outcome measures like jumping height or distance are well defined and straightforward to assess, in contrast to movement technique analysis, which requires rather complex procedures and the selection of representative kinematic and kinetic parameter subsets. Second, whereas performance outcome can be easily addressed with an EF instruction, technique training often requires instructions aimed at the motion of specific body parts, provoking the difficulty to directly address these parts by EF instructions. Some authors automatically define the naming of body parts in the wording as IF instructions . In ordeWithin that context, Kons  could shSingh and Wulf  conducteLohse et al.  observedObserving muscle activity during motor performance confirmed that for IF conditions, higher EMG amplitudes occur ,24. FocuThe study of Coker  specificThe impact of the frequency of instructions might be interesting information for practitioners. Most of the studies provided instructions immediately before each movement execution or block of trials with reminders to make sure that attention is adequately directed ,40,45. NEven though the body of literature referring to the focus effect is large, only a relatively small number of studies observed movement form corrections and technique adaptation, showing divergent results. IF and EF instructions differed in their impact on movement form and movement technique. However, this effect turned out to be small. Therefore, a general superiority of EF instructions in technique training is not sufficiently legitimate.Literature indicates that EF instructions facilitate optimization of existing co-ordination patterns within movement execution, whereas IF is able to adjust movement trajectories in a desired direction, thus potentially being more capable of movement technique corrections. Results support the implementation of imagining techniques to adapt EF on partial movement form corrections where possible.Study-results of focus effects seem to be in line with ideas of the Optimal Feedback Control Theory. Instructions directing attention to specific movement sequences or specific joints, or segments, should be expected to act on this segment. However, such a focus might interfere with the integration of movement elements into the whole movement, which can lead to a worse movement outcome. External focused instructions then seem to facilitate optimization of movement pattern integration and, as a result, movement execution.Summing up the main findings of the reviews and important focus-of-attention literature addressing movement form and technique training, we can state:The huge body of focus studies clearly demonstrates that shifts in attentional focus and subsequent changed success in movement outcome happens by a small change of wording within the movement instruction. This underpins the importance of sophisticated strategies for creating instructions to direct the focus on athletes by trainers and coaches. Some of the systematic reviews promote a positive EF effect, but this notion does not hold for movement form corrections and technique training, where still only insufficient evidence is available. If coaches head for optimization in the whole-body co-ordination, then EF instructions with imagined movement effects seem more suitable. In contrast, IF instructions should be favored to provoke movement form changes and technique corrections. The use of both implicit focused instructions—to become aware of movement patterns—and explicit focused instructions—to provide optimization of whole-body-movement execution, may lead to more success in movement technique training.


Emergency physicians and trauma surgeons are increasingly confronted with pre-injury direct oral anticoagulants (DOACs). The objective of this study was to assess if pre-injury DOACs, compared to vitamin K antagonists (VKA), or no oral anticoagulants is independently associated with differences in treatment, mortality and inpatient rehabilitation requirement.We performed a review of the prospectively maintained institutional trauma registry at an urban academic level 1 trauma center. We included all geriatric patients (aged ≥ 65 years) with tICH after a fall, admitted between January 2011 and December 2018. Multivariable logistic regression analysis controlling for demographics, comorbidities, vital signs, and tICH types were performed to identify the association between pre-injury anticoagulants and reversal agent use, neurosurgical interventions, inhospital mortality, 3-day mortality, and discharge to inpatient rehabilitation.A total of 1453 tICH patients were included . DOAC use was independently associated with lower odds of receiving specific reversal agents  than VKA patients. DOAC use was independently associated with requiring neurosurgical intervention . VKA use, but not DOAC use, was independently associated with inhospital mortality, or discharge to hospice care  compared to controls. VKA use was independently associated with higher odds of discharge to inpatient rehabilitation  compared to controls.Despite the higher neurosurgical intervention rates, patients with pre-injury DOAC use were associated with comparable rates of mortality and discharge to inpatient rehabilitation as patients without anticoagulation exposure. Future research should focus on risk assessment and stratification of DOAC-exposed trauma patients. Direct oral anticoagulants (DOACs) were first introduced in 2008 for the prevention of thromboembolism after elective arthroplasty, and in 2010 for non-valvular atrial fibrillation (AF) patients . DOACs aAt the same time, DOAC use may add considerable risks to the setting of acute major hemorrhaging. . EmergenDespite that DOAC use is prevalent in the geriatric trauma population, many aspects of the clinical presentation, management and prognosis are still unknown. This unfamiliarity potentially influences the treatment of tICH in DOAC-anticoagulated trauma patients. For instance, where reversal strategies for VKA-anticoagulated patients are widely established, reversal protocols for DOACs are still in the development phase, and based on less robust clinical evidence. Two DOAC reversal agents are currently available: idarucizumab to reverse dabigatran and andexanet alfa for reversal of factor Xa inhibitors, but clinical hemostatic effectiveness of both agents is yet to be confirmed in randomized controlled trials –16. FurtThe primary objective of this study was therefore to establish if pre-injury DOAC use in geriatric patients sustaining traumatic intracranial hemorrhage after falls is associated with differences in the rate of reversal agent use and of neurosurgical intervention, and with increased mortality and inpatient rehabilitation requirement.This study is reported on in accordance with the STrengthening the Reporting of OBservational studies in Epidemiology (STROBE) statement . A retroThe outcomes of interest were use of anticoagulant-specific reversal agents ; number and types of neurosurgical interventions; mortality, and discharge to inpatient rehabilitation. The definition of inhospital mortality includes patients discharged to hospice care. Similarly, 3-day mortality includes patients discharged to hospice care on or before hospital day 3. Discharge to inpatient rehabilitation was analyzed only in patients surviving until discharge.2-analysis. Missingness of each variable was summarized. Missingness was assumed to be not-at-random, and missing data were imputed using multiple imputation with chained equations. Multivariable logistic regression was performed on the dataset with imputed missing data to assess the independent associated between pre-injury anticoagulant exposure and the use of anticoagulant-specific reversal agents, neurosurgical interventions, discharge to inpatient rehabilitation, 3-day mortality, and inhospital mortality. All variables with a p value of less than 0.100 on univariate analysis were included as covariates, with the exception of INR on admission, as it does not reflect the degree of anticoagulation for DOAC-anticoagulated patients, thereby biasing the results toward favorable effects for VKA patients. Only anticoagulated patients were included in the baseline comparison and regression analysis of use of anticoagulant-specific reversal agents.All normally distributed continuous variables are displayed as means with standard deviations. Median and interquartile range notation was used for nonparametric data. Normally distributed continuous variables were compared using one-way ANOVA test and using Kruskall–Wallis test for skewed distributions. Categorical variables are summarized as proportions and compared using ChiConsidering the large number of covariables studied, collinearity analysis was performed. A mean variance inflation factor (VIF) over five was considered an indicator of collinearity. New multivariable logistic regression models were constructed after sequentially omitting the variable with the strongest collinearity until the mean VIF was below five.p < 0.001). The majority of patients were transferred from an outside hospital. No clinically relevant differences were found between groups on presentation, except for median INR, which was highest in the VKA group . No differences between the groups were found for platelet count, hemoglobin and blood alcohol concentration.A total of 1453 patients meeting all inclusion criteria were identified in the trauma registry. Of these, 52 used pre-injury DOACs, 376 used pre-injury VKAs, and 1025 used neither DOAC nor VKA (Table p = 0.079). The majority of patients sustained subdural and/or subarachnoid hemorrhages. Hemorrhagic contusions and intraparenchymal hemorrhage were more frequent in the DOAC group . The median Rotterdam CT score did not differ significantly between the groups . Skull fractures were most prevalent in the control group .ISS did not differ between the three groups . More patients in the DOAC group underwent decompressive craniotomy, but this difference was not statistically significant . No difference was seen in the rate of burr holes and intracranial pressure monitor placement. Extraventricular drain placement was performed significantly more frequently in the DOAC group . Fewer patients in the DOAC group were admitted to the ICU , with no difference in duration of ICU stay . There was no statistically significant difference in mechanical ventilation rates, ventilation days or hospital length of stay. In-hospital mortality or discharge to hospice care was highest in the VKA group and lower in the DOAC and control groups . This pattern was also seen for mortality or discharge to hospice care at day 3, albeit not statistically significant on univariate analysis . Fewer patients in the DOAC group were discharged to inpatient rehabilitation than in the VKA and control groups .DOAC patients were significantly less likely to receive a specific reversal agent than VKA patients  are presented in Table Treatment of anticoagulated patients and prognosis of intracranial hemorrhage differed significantly depending on pre-injury anticoagulant exposure in geriatric trauma patients with traumatic intracranial hemorrhage after a fall. Pre-injury DOAC users were less likely to receive reversal therapy and more likely to undergo a neurosurgical intervention than patients using VKA or no anticoagulation medication. Pre-injury DOAC use was not associated with mortality or discharge to hospice care, whereas pre-injury use of VKAs was an independent risk factor for mortality.Clinical outcomes for anticoagulated blunt tICH patients have been the subject of substantial interest: numerous observational cohort studies have compared presentation, management, and outcomes of tICH patients on pre-injury DOACs and VKAs –31. The First, the rate of DOAC reversal varied, with individual studies reporting between 12 and 40% use of predominantly nonspecific reversal agent use , 30, 31.Second, rates of neurosurgical intervention in smaller cohorts showed widely differing rates at times favoring pre-injury DOACs and at other times favoring VKAs, while larger, more contemporaneous cohorts reported similar to potentially rates of neurosurgery in for pre-injury DOAC patients –28, 31. Owing to these differences in patient management, the true association between pre-injury anticoagulation and clinical outcomes is difficult to determine. The most reliable information on outcomes of anticoagulated tICH patients could come from subset analyses from randomized controlled trials comparing efficacy and safety of chronic oral anticoagulant use. In two studies comparing DOACs to VKAs (RE-LY and ROCKET-AF), however, sample sizes were too small to determine a survival benefit of either type in the setting of tICH , 33.Alternatively, studies using national databases may diminish the impact of heterogeneity between hospitals, as well as sample size issues. While the association between pre-injury anticoagulation, treatment and outcomes has been studied in the general trauma population, analysis of the tICH subpopulation may add further information .This study is limited by several factors arising from its retrospective design. First, we were not able to report ICH progression rate at a fixed timepoint. While ICH progression is sometimes preferred over mortality as an outcome, especially in studies of treatment effects in the early phase of tICH management, this study aimed to describe management and prognosis, and therefore discharge to inpatient rehabilitation and mortality were deemed appropriate outcomes. Secondly, retrospective data collection impeded collection of the modified Rankin scale, which is commonly used in studies to assess general neurological functioning after stroke and intracranial hemorrhage. We did also not account for the degree of anticoagulation, as anti-FXa assays were not used, nor were such assays validated during the study period. Moreover, the use of non-anticoagulant-specific reversal agents, e.g. 3 or 4 factor prothrombin complex concentrate, was not collected in this study. The DOAC group was smaller than the VKA cohort. Imbalance in group sizes reduces the statistical power of tests performed, resulting in lower chances of detecting significant differences between groups. Despite this limitation, reported differences in reversal agent use, neurosurgical interventions, and mortality or hospice care were statistically significant. Sensitivity analysis demonstrated a limited impact of collinearity, as evidenced by the effect sizes not changing significantly pre- and post-collinearity correction.In patients with tICH after falls, pre-injury DOAC was associated with similar rates of mortality and discharge to inpatient rehabilitation, whereas pre-injury VKA exposure was associated with higher inhospital mortality and more frequent discharge to inpatient rehabilitation. Vigilance remains warranted in the treatment of DOAC-anticoagulated trauma patients, as differences in treatment allude to various risk profiles. Future research should focus on risk assessment and stratification of DOAC-exposed trauma patients.


PLOS Biology, Kreher and colleagues show in a mouse model that in vivo, neurons and not only myelinating glia are primary effectors of disease progression in Krabbe disease. The neuron-specific model generated allows the unprecedented capacity to investigate the neuronal autonomous component of this disorder.In this issue of  This Primer explores the implications of a PLOS Biology showing that in vivo, neurons (not only myelinating glia) are primary effectors of disease progression in Krabbe disease; the neuron-specific animal model described allows an unprecedented opportunity to investigate the neuronal-autonomous component of this disorder. Galc expression, psychosine accumulates in myelinating glia and neurons [In Krabbe disease, also known as globoid cell leukodystrophy, mutations in β-galactosylceramidase (GALC), a lysosomal enzyme responsible for the catabolism of myelin galactolipids, lead to the accumulation of its substrates galactocerebroside and psychosine . Whereas neurons . As a po neurons  and axon neurons  occur in neurons , and the neurons , furtherPLOS Biology, Kreher and colleagues use the recently generated conditional Galc floxed mouse [Galc heterozygosity , neuronal ablation was achieved by Synapsin 1–mediated expression of cre recombinase. The homozygous loss of Galc in neurons  is shown to be sufficient to induce a neurological phenotype with growth and motor-coordination defects. The analysis of Syn1Cre:Galcf/− mice revealed that psychosine accumulates in the nervous system where neuronal and axon degeneration, neuroinflammation (astrocytosis and microgliosis), and a mild reduction in myelination occur , in which all murine cells only have one functional Galc allele. Although Galc+/− mice do not develop an overt phenotype or pathology, and express normal levels of several myelin components, Galc heterozygosity causes a reduction in Galc activity [Galcf/f mice in a wild-type (WT) background would serve to fully decouple effects of loss of Galc activity in neurons from possible mild consequences of Galc heterozygosity.In this issue of ed mouse  to elimion occur . Thus, tactivity , impaireactivity . Thus, sThe findings of Kreher and colleagues underscore the need to further understand the specific cellular and molecular mechanisms of GALC function in neurons. A substantial body of data already shows the cell-specific deleterious effects of psychosine in this cell type. Psychosine can intercalate in membrane lipid rafts disrupting their structure and, as a consequence, impairing lipid-raft mediated signaling, including critical pathways for neuronal development and function such as PKC, Erk, and AKT-GSK3β ,9. In adPLOS Biology, the study of Kreher and colleagues opens new windows of action in Krabbe disease by highlighting that in vivo neurons are an important primary cellular target in this disorder. Although originally classified as a myelin disorder—leukodystrophy—in which the involvement of CNS and peripheral nervous system (PNS) myelin mediated a secondary neurodegeneration, there is now mounting evidence of an additional neuron-autonomous effect behind Galc deficiency. It is possible that similar cell-autonomous mechanisms unrelated to myelinating glia operate in other leukodystrophies, especially for those in which the disease-causing gene shows generalized expression in multiple cell types. Thus, not only new therapeutic strategies focused on targeting and correcting primary neuronal defects but also further knowledge on cell-specific primary effects caused by the accumulation of toxic substrates in different cell types of the nervous system are needed in Krabbe disease and also in other leukodystrophies.In summary, in this issue of


In 2016, the European Hematology Association (EHA) published the EHA Roadmap for European Hematology Research1aiming to highlight achievements in the diagnostics and treatment of blood disorders, and to better inform European policy makers and other stakeholders about the urgent clinical and scientific needs and priorities in the field of hematology. Each section was coordinated by one to two section editors who were leading international experts in the field. In the five years that have followed, advances in the field of hematology have been plentiful. As such, EHA is pleased to present an updated Research Roadmap, now including eleven sections, each of which will be published separately. The updated EHA Research Roadmap identifies the most urgent priorities in hematology research and clinical science, therefore supporting a more informed, focused, and ideally a more funded future for European hematology research. The eleven EHA Research Roadmap sections include Normal Hematopoiesis; Malignant Lymphoid Diseases; Malignant Myeloid Diseases; Anemias and Related Diseases; Platelet Disorders; Blood Coagulation and Hemostatic Disorders; Transfusion Medicine; Infections in Hematology; Hematopoietic Stem Cell Transplantation; CAR-T and Other Cell-based Immune Therapies; and Gene Therapy.3Relapse following allogeneic hematopoietic stem cell transplantation (HSCT) has remained unchanged over the past 3 decades, as reviewed in the second workshop convened by the National Institutes of Health (NIH).5 have a high risk of relapse post-transplant. Thus, the first measure is to reduce tumor burden pretransplant.The strongest predictor of relapse is phase of the disease. Patients not in hematologic remission at the time of transplantation, or patients with minimal residual disease (MRD),6 or chimeric antigen receptor T cells (CART)7: the role of allogeneic HSCT following CART CD19 therapy should be explored, to maintain the achievement of MRD negativity.A molecular remission pretransplant can be achieved with targeted therapy in acute lymphoid leukemia (ALL), using bispecific antibodies such as blinatumomab8 Several Flt3 inhibitors are now available and should be tested before allogeneic HSCT to induce MRD negativity. For patients with Flt3-negative AML, CART cell therapy needs to be expanded: a phase 1 trial with CART CD44v6 cells has been opened . Further support needs to be given to the control of MRD in AML before HSCT.In patients with Flt3-positive acute myeloid leukemia (AML), midostaurin has shown encouraging results when used in induction.9 A prospective randomized trial has been designed to confirm this finding (NCT04217278).The intensity of the conditioning regimen is another major player and should be further explored. In children with ALL, conventional total body irradiation (TBI) has been shown to be superior to chemotherapy (NCT01423747). The problem is the use of TBI in adults, and total marrow irradiation should be explored. In AML, thiotepa, busulfan, and fludarabine may reduce post-transplant relapse as compared with busulfan fludarabine.Whether patients with myelodysplastic syndrome should be treated with azacytidine before an allogeneic HSCT remains uncertain: a randomized trial comparing upfront HSCT to azacytidine followed by HSCT has been activated (NCT04184505).10 and needs to be assessed in prospective trials: a previous small, randomized study has shown improved leukemia control with prophylaxis as compared with MRD based, pre-emptive therapy.11 For patients with Flt3-positive AML, 2 recent randomized studies have shown significantly improved survival with post-HSCT prophylactic sorafenib,13 although this drug has currently no European Medicines Agency (EMA) approval in AML. Other Flt3 inhibitors such as gilteritinib need to be tested post-HSCT. A randomized trial with azacytidine to prevent leukemia relapse in AML has recently failed.14The role of post-HSCT prophylactic tyrosine kinase inhibitors for Philadelphia-positive ALL is still being discussedSelection of optimal donor for leukemia control is an additional relevant issue, which is being addressed by 3 prospective randomized trials . These studies are designed to enroll almost 1000 patients, and we should then know whether a family haploidentical or an unrelated donor is best, for patients with acute leukemia.15 However, there is no proof that prophylactic infusions of NK cells will reduce leukemia relapse in acute leukemia. Prospective randomized trials testing the role of NK cell infusions in AML patients with active disease at transplant should be pursued (NCT 04166929).The use of cellular therapy as prophylaxis should be considered: natural killer (NK) cells have shown to possess antitumor activity and may be given without the risk of graft versus host disease (GvHD).2 The use of bispecific antibodies and CART cells are expanding the therapeutic options for relapsed ALL patients and should be tested further.17 New treatment modalities such as the combination of azacytidine and venetoclax, for AML, should also be prospectively explored.The outcome of patients following a hematologic relapse will depend on the pace of the underlying disease. Patients with acute leukemia, relapsing within 6 months are at very high risk of early death, whereas delayed relapses can be successfully treated in a proportion of cases with a second transplant or chemotherapy and DLI.Clinical trials need to be designed addressing (1) pre-transplant consolidation for MRD eradication and (2) the impact of conditioning regimen intensity on MRD clearance post HSCT. The selection of optimal donors (3) and conditioning regimens (4) should also be assessed prospectively. Post-transplant prophylaxis with drugs, cellular therapy, as well as monitoring of MRD must be improved, particularly in patients with acute leukemia, in whom time is crucial.Hematopoiesis is critically regulated by nonhematopoietic cells that support the production of blood and immune cells according to the organismal demands. Mesenchymal stromal cells (MSCs) are key components of the hematopoietic microenvironment. Recent technological developments (most notably single-cell studies) have made it possible to describe different MSC subpopulations, several of which seem to show organ-specific properties and gene expression profiles. However, functional studies will be critical in the near future for fulfilling the perceived potential of stromal cells in immune modulation and tissue regeneration.18 Computational methods have been developed to infer candidate gene networks in HSC-supporting MSCs19 and potential ligand–receptor interactions.20 Future studies will be essential to reveal the specific functions and potential of these MSC subpopulations.HSCs and their microenvironment represent probably the best-characterized hierarchical stem cell system in vertebrates. They have paved the path to understanding how cell lineages are organized, their fate determined, and how they contribute to function in other organs. Recent European research has revealed the cellular and spatial organization of different bone marrow MSC subsets.21 Extracellular vesicles from lymphoma B cells render MSCs protumorigenic.22 Notch2 signaling on MSCs triggers Wnt-dependent survival of chronic lymphocytic leukemia.23 Particularly, Wnt5A from MSCs is required for the engraftment of normal and leukemic HSCs.24 MSCs remodel the extracellular matrix to support the progression of B-cell ALL,25 support the growth of myeloproliferative neoplasms26 or differentiate into myofibroblasts in myelofibrosis.27 Finally, mitochondrial transfer from MSCs protects ALL and AML cells from oxidative stress induced by chemotherapy.29 Therefore, elucidating and targeting niche-dependent mechanisms of tumorigenesis and resistance will be key to eradicate cancer.Recent European research has demonstrated a key role for MSCs in the initiation, progression, and drug resistance of a variety of hematological neoplasms through paracrine or cell-contact–dependent mechanisms.30 Targeting MSCs could help mitigate the damage to normal hematopoiesis by aging, AML, infection, or iron overload in beta-thalassemia.34 Additionally, MSC coinfusion could increase the efficiency of HSCT35 and minimize the risks of graft failure in gene therapy applications associated with low conditioning regimens and infusion of limited numbers of gene-edited HSCs.36European research over the past 5 years has shown that MSCs enhance HSC mobilization for subsequent apheresis from peripheral blood and transplantation (HSCT).MSCs have a profound impact on different immune cells, having therapeutic benefits in sepsis, autoimmune disorders or graft-versus-host disease (GVHD), and may influence the outcome of immunotherapies. Additionally, research on MSCs could influence the outcome and treatment of steadily increasing systemic conditions, such as diabetes, obesity, and aging.Increasing research in the hematopoietic microenvironment and particularly MSCs will, scientifically, feed into other stem cell systems and, clinically, provide new ways to modulate and treat hematopoietic diseases, immune responses, and regenerative processes. As a result of this productive research, Europe continues to be the world region with profound understanding of MSC contributions to controlling the hematopoietic niche and the composition of immune cell populations of each individual in health and disease.37 If HSCs could be generated from a patient’s own hPSCs, genetic engineering could easily be used to correct genetic defects before differentiation into transplantable HSCs, which would overcome some caveats of conventional HSCT therapies.The present availability of human pluripotent stem cells  raises promise for a universal resource for cell based therapies in regenerative medicine. Rapid progress has been made in generating hPSCs amenable for clinical applications, culminating in reprogramming of adult somatic cells to autologous hPSCs that can be indefinitely expanded in vitro. However, a major challenge remains how to differentiate hPSCs to specific lineages efficiently and how to select for cells that will function normally upon transplantation in adults. Another major challenge, which until now has not been solved, is how to generate true HSCs that can be transplanted, engraft and maintain all (blood) stem cell properties. The hematopoietic lineage has been particularly refractory compared with other cell types and derivation of adult-type HSCs has not yet been possible even though various other blood lineages can be obtained.38Skin fibroblasts were the first human somatic cells to be reprogrammed into hiPSC but blood cells are increasingly used because of the availability of banked  blood samples covering a range of disease, age, gender, and ethnic backgrounds. Complementing hiPSC generation are attempts at “direct reprogramming” using sets of lineage specific transcription factors that act as master regulators and convert cells to a new differentiation state without intermediate pluripotency. Generating hiPSC from mature B cells requires silencing of lineage-specific factors such as PAX5, so that direct reprogramming into blood cells where this would not be necessary, may therefore be quite feasible.Cdx/Hox), certain cell nonautonomous effects and exposure to physical stimuli (flow) that occur at specific stages of development. hPSC derivatives thus remain immature and, to date, all hPSC-derived HSCs have been deficient in their developmental potential, ability to self-renew and to engraft upon transplantation in mice, even though animal studies have shown that this is possible in the context of a developing embryo . Normal HSC development during embryogenesis occurs in several distinct temporal/spatial waves, each characterized by its own set of hematopoietic progenitor cells (HPCs). Only those produced from the latter, or “definitive,” wave give rise to mature, functional HSCs but during in vitro differentiation, it is believed that the HSC-like cells produced are from the more primitive waves. These give rise to HSC-like cells biased toward myeloid lineages at the expense of lymphoid potential. These are unable to self-renew in culture and lack long-term engraftment capacity. Key to overcoming this HSC bottleneck will be understanding: (1) the ontogeny of human HSCs, (2) how they progress at the molecular level to become mature, adult HSCs, and (3) intrinsic and extrinsic factors that govern HSC behavior and function.The first steps in directed differentiation of hPSCs are guided by morphogens  which are important in development Figure . hPSCs pSeveral prominent European hematologists are using genomic approaches  to examine the molecular mechanisms that regulate cell fate decisions  by HSCs or HPCs and their more differentiated progeny. This focus on genetic modification of stem cells, especially using gamma-retroviral and lentiviral vectors, has brought Europe in the global forefront of genetically modified stem cell therapy for treatment of various types of severe combined immunodeficiency (SCID). This has been in part through EU FP7 funded projects that included PERSIST, EuroStemCell, and NOVEXPAND, among others. This momentum is poised to combine HSC expansion with gene editing approaches that will move the field forwards toward innovative therapies in the coming decade. Several European consortia are currently applying for funding in the Horizon Europe scheme that has started 2021.40 Recent study has established the potential for site specific gene editing to repair disease mutations in the human genome by targeting the integration of a corrective cDNA into the IL2RG locus of HSCs from a SCID-X1 patient.41 For homologous recombination-based gene editing approaches to be successful, HSC need to proliferate and template DNA harboring the corrective sequences needs to be efficiently introduced into the target cells. This can be readily achieved in hiPSC so that these cells are prime targets for gene editing techniques. However, as gene editing can also be done in primary HSCs, it is crucial to find conditions that allow HSC to proliferate without loss of phenotype. Therefore, HSC expansion protocols  are directly relevant for gene repair described here. Niche signals, expansion, directed differentiation toward induced-HSCs (iHSCs), and genetic modification of HSCs and their progeny are related research topics that need an integrated approach for optimal exploitation in the clinical context.An exciting new development in the gene therapy field is the use of designer nucleases, most recently CRISPR/Cas9 based, that are proving exceptionally efficient in engineering the genome.ERG, HOXA5, HOXA9, HOXA10, LCOR, RUNX1, and SPI.42 However, the overexpression of such factors, in particular HOX genes, is associated with acute leukemogenesis. This indicates that more research is needed, not only for efficacy but also safety of hiPSC-derived iHSC. Careful single cell RNAseq, ATAC-seq and other -omics approaches comparing bona fide HSCs with iHSC may help in defining the relevant conditions to generate transplantable iHSC. Finally, in all of these approaches, residual undifferentiated hiPSC may also present a risk for clinical application of these cells.Ex vivo generation of HSCs is currently difficult and requires ectopic expression of a number of transcription factors including A major bottleneck to curing genetic diseases originating from mutations in HSCs is that even though efficient methods for gene targeting are now available, HSCs from neither adult nor hPSC sources can be expanded sufficiently for this to become routine practice. The combined projects in this section seek to address this, bringing safe gene therapy to the clinic via HSCs. New inroads into the generation of iHSC are needed to realize the true potential of these cells for clinical use in blood-mediated diseases.Reduction of leukemia relapse remains a priority for future multimodal approach, including optimization of conditioning regimens and targeted drug therapy.An immunologic control of leukemia with cellular and/or antibody-based therapy needs to be expanded.Expansion of HSCs from either adult or hPSC sources in culture: even though efficient methods for gene targeting are now available, clinical use of these cells will require scaled cell production for the approach to become routine practice.More insight into the generation of iHSC (either by direct reprogramming or from hiPSCs) is needed to realize the full potential of these cells for clinical use in blood-mediated diseases.iHSC need to be engrafting upon transplantation, maintain self-renewal and multilineage differentiation.Costs of hPSC generation under good manufacturing practice (GMP) conditions need to be significantly reduced.Regulator engagement is essential early in the clinical development phase to guide products efficiently to the clinic.WF is a chairman DSMB in the Glycostem Therapeutics and advisor for Starfish Innovations. DK receives research funding from Merck. CM received research funding from Sartorius. AB receives speaker’s fees from SANOFI, MSD, Therakos, Novartis, JAZZ, ADIENNE. All the other authors have no conflicts of interest to disclose.