Gossypium spp. L., Malvaceae) is the world’s largest source of natural fibers. Virus outbreaks are fast and economically devasting regarding cotton. Identifying new viruses is challenging as virus symptoms usually mimic nutrient deficiency, insect damage, and auxin herbicide injury. Traditional viral identification methods are costly and time-consuming. Developing new resistant cotton lines to face viral threats has been slow until the recent use of molecular virology, genomics, new breeding techniques (NBT), remote sensing, and artificial intelligence (AI). This perspective article demonstrates rapid, sensitive, and cheap technologies to identify viral diseases and propose their use for virus resistance breeding.Cotton ( Gossypium spp. L.  is the largest source of natural fibers in the world. The widely cultivated cotton cultivars are allotetraploid species. More than 90% of the annual cotton crop worldwide is Gossypium hirsutum (AD1), Upland or American cotton, and less than 10% is Gossypium barbadense (AD2), extra-long-staple, or Pima cotton , digital disease assessment and phenotyping (DDAP), artificial intelligence (AI), and biotechnological tools may accelerate and enhance the breeding of virus-resistant cotton and help sustain the cotton industry.Solemoviridae , a positive sense (+) ssRNA virus, causes the Cotton Blue Disease (CBD) that affects cotton crops in South America and Africa , one of the most polyphagous aphid species and worldwide-distributed, hosts over 40 viruses , a “cryptic species complex,” which is one of the most agriculturally important pests worldwide  or cotton leaf curl Kokhran virus (CLCuKoV), with the cotton leaf curl Multan betasatellite  as B. tabaci is present in their territories.The 1992–1993 CLCuD epidemic in Pakistan caused economic losses calculated in one billion US dollars  and is aBegomovirus that mimics nutrition deficiency, is hard to diagnose, and proliferates through the Brazillian cotton belt in the Midwest and Northeast regions  and cotton leaf curl Gezira virus associated with cotton leaf curl Gezira betasatellite and cotton leaf curl Gezira alphasatellite, and African cotton mosaic disease virus is spreading at a quick rate from Western to Central Africa and Indo-Pak subcontinent .The cotton bunchy top (CBT), a polerovirus from the Oceania continent, is cotton’s most viral severe disease in Australia. CBT often occurs in patches or on crop edges and is associated with the highest aphid activity areas .Ilavirus, family Bromoviridae), transmitted by thrips, was reported to infect cotton in specific regions of India and Pakistan  transmitted by aphids are coinfecting cotton plants  are the safest way to confirm a virus species and/or strain and a resistant cotton line. After confirming viral resistance, breeders can plan the crosses and breeding schemes. Moreover, molecular and discovery breeders can initiate molecular marker discovery for resistant genes and molecular breeding strategies with the knowledge of contrasting resistant and susceptible cotton lines.Nowadays, molecular virus detecting techniques, such as high-throughput sequencing (HTS), allow the detection of all viruses present in a plant (virome), including undescribed new viruses . HoweverVirome information is essential for breeding and molecular breeding schemes to understand the availability and type of viral-resistant genes needed for elite line introgression. Developing a new resistant elite variety can be fast if a known virus causes the symptoms and resistant germplasm and molecular markers are available. When dealing with a new virus or coinfection, developing a new resistant elite variety will demand germplasm and gene prospection, resources, and time.After completing the virus sequencing step, it is possible to develop new sensitive virus detection techniques and replace costly qRT-PCR-based diagnosis testing systems . Some acin situ screening of human viruses using paper strips. The development of such POCT tools for plant viruses will enable a large-scale, fast, and robust detection of virus on-site, aiding in selecting virus-resistant cotton lines without waiting for symptoms.Among CRISPR-NAT tools, CAS9 has shown to be an ultrasensitive pathogen detection method. Converting the developing NAT to point-of-care testing (POCT) will drastically reduce costs and accelerate viral identification due to on-site identification and easy handling . CRISPR/MVDT are so fast and sensitive that CLRDV is detectable in susceptible cotton plants 24 h after inoculation and 2–5 days in systemic leaves after aphid infection, respectively . MVDT coi) and selection accuracy (r), parameters of the “breeder’s equation” Several available technologies permit digital disease assessment and phenotyping (DDAP) and may reduce operational time, costs, breeding score subjectivity, and increases the genetic gain (ΔG) by increasing selection intensity  and lower reflectance values in the red region  can detect viral diseases with decreased dependence on weather conditions . Swarm r2 of data and can capture healthy vegetation’s spectral signature and anomalies caused by viruses , labeling training data, and high processing time, Deep Learning (DL) enables an automatic and hierarchical learning process . DL has Pathogen-resistant cotton lines appear to have relatively low yields, low fiber quality, and high linkage drag, complicating the development of elite resistant varieties. Implementing integrated cotton molecular breeding strategies , such asMoreover, new breeding techniques (NBT) may be the quickest path to developing elite resistant varieties without the worries of linkage drag . Among tAV2) and antisense coat protein (ACP) genes of CLCuV provide resistance to CLCuD  plants expressing the antisense movement protein  project E-26/010.001716/2019.The 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.


For many therapeutic areas in modern medicine, precision medicine is the current treatment paradigm. Interestingly, or probably disappointingly, the concept of precision medicine often only encompasses selecting the right drug for the right patient. Subsequent selection of the right dose for the right patient is at least equally important and is only scarcely considered. Therapeutic Drug Monitoring (TDM) can be defined as assessing the adequacy of the drug plasma concentrations in relation to a target concentration or concentration window at a specific time in a dosing interval. This evaluation, following appropriate clinical interpretation and according to the drug pharmacokinetic/pharmacodynamic (PK/PD) properties, can guide dosing. However, finding the optimal dosing in order to guarantee a therapeutic exposure remains complicated. Sources of PK variability, including age, genetic heritage, and disease conditions, all influence the chances of achieving therapeutic outcomes. This aspect is particularly evident in children and neonates where physiological changes, associated with growth and maturation, dramatically influence drug PK properties. Similarly, pediatric patients subjected to medical procedures, including dialysis or extracorporeal membrane oxygenation (ECMO), may require special dose considerations since these procedures could significantly affect PK parameters. In pediatrics, drug-administration issues are more prominent than in adults. To adjust the dose and to secure intake in pediatrics, adult formulations may need to be crushed, dissolved, or extemporaneously prepared as liquid. In the absence of reliable data on these manipulations, TDM may contribute to safe and effective therapy in pediatric practice.The utility of TDM is often underestimated or exclusively applied to molecules where monitoring of drug concentrations is mandatory due to safety concerns. However, TDM is less commonly used for optimization towards effective drug exposure. Although the multiple benefits of TDM are well known and recognized by clinicians, this knowledge seems to be only theoretical and its introduction into clinical practice remains far away from the application. TDM should rely on analytical methods such as high-performance liquid chromatography coupled to UV (HPLC-UV) or to mass spectrometry (LC-MS/MS) characterized by fast detection, and high accuracy and precision. These instruments are not always available for every drug and in every analytical laboratory, making TDM often unfeasible. Another issue that limits the application of TDM in pediatric patients is the invasiveness of blood sampling. The impact of blood sampling in children should not be underestimated. Even with professional psychological support, many children describe blood sampling as a dismal experience, and -naturally- TDM should therefore only be applied if clinically relevant. Low sampling volume limits in neonates and children make the bioanalytical procedures not always applicable, thereby limiting the use of TDM. To overcome this issue, several microsampling methods have been proposed including volumetric absorptive microsampling (VAMS) and dried blood spots (DBS) sampling. However, exhaustive analytical and clinical validation of these methods for conducting PK studies and TDM interventions is still rather limited.off-label in neonates and children. These studies could be particularly useful to overcome the physiological and ethical issues connected to the realization of PK clinical studies on these subjects.The aim of this Research Topic was to show the utility of TDM applied to the routine clinical practice in pediatric patients. To this scope, we have collected reviews, research articles, case reports, and both pre-clinical and clinical study protocols that investigate the utility and the application of TDM in different pediatric settings and disease conditions. Moreover, an important contribution has been provided by population pharmacokinetic (popPK) studies that demonstrate how modelling and simulation approaches could provide important dosing guidance for drugs used Xu et al. These authors report two cases of critically ill pediatric patients with acute kidney injury requiring CRRT and receiving polymyxin B treatment due to carbapenem-resistant organism bloodstream infections. TDM of polymyxin B revealed an increased clearance of polymyxin B through CRRT that required supplanted dosing of the drug Xu et al. Similarly, a popPK analysis, aimed at describing primary PK/PD parameters of vancomycin and meropenem in pediatric patients undergoing ECMO, showed that the PK/PD target for vancomycin was achieved partially with conventional doses meanwhile higher dosing with extended infusion was needed in the case of meropenem Zylbersztajn et al. Even pathological conditions including hematological malignancies can affect PK properties of many administered drugs. An augmented renal clearance (ARC) is increasingly recognized in pediatric oncologic patients. In these immunocompromised subjects, broad-spectrum beta-lactams are commonly prescribed for empirical or selective treatment of bacterial infections. André et al. compared trough concentrations of meropenem and piperacillin in a cohort of unselected pediatric hematology-oncology patients stratified according to their estimated renal function as decreased, normal or with ARC, and to their neutrophil count. The results obtained by this retrospective evaluation showed that intermittent administration of meropenem and piperacillin often fails to ensure sufficient exposure in treated patients even at maximal recommended daily dosage, perhaps due to an increased drug clearance. Therefore, the authors suggest systematic TDM alongside assessment of renal function, as valid support for dosage adjustment in order to guarantee an appropriate antibiotic therapy in pediatric patients affected by malignancies André et al. These articles confirm that TDM may represent a valuable tool for assisting clinicians in optimizing antimicrobial exposure. A proof of concept in this direction has been provided by Gatti et al. The authors report their experience as clinical pharmacological advice (CPA) and conclude that a TDM-based real-time CPA may be particularly useful to optimize antimicrobial therapies in different challenging pediatric settings Gatti et al. The prospective EXPAT Kids study aims to evaluate PK/PD target attainment within the first 36 h after initiation of beta-lactam dosing in critically ill children. Schouwenburg et al. will also investigate the association between PK/PD target attainment and patient characteristics and clinical outcomes. Aiming to reduce the workload for medical staff, they will validate the use of residual material from heparinized astrup syringes for TDM. The results of the EXPAT Kids study are eagerly awaited and are expected to provide important new insights to improve clinical outcomes through better beta-lactam dosing.Infections represent one of the main complications among hospitalized neonatal and pediatric patients, especially during long-standing periods in intensive care units (ICUs) where the presence of central catheters for parental nutrition and ventilators for respiratory support are often sources of bacterial colonization and require an appropriate shunt lock therapy to avoid systemic infections . Therefovan der Stoep et al. provided an overview of the possible relationships between PK parameters and clinical outcomes or toxicities for the most commonly used conditioning agents in pediatric HSCT van der Stoep et al.Brooks et al. developed a popPK model for tacrolimus intravenous continuous infusion in pediatric and young adult HCT patients Brooks et al. The model was implemented in a Bayesian dosing tool, thereby making it available at the point-of-care and allowing a clinical impact.Hematopoietic stem cell transplantation (HSCT) remains the only curative treatment in several pediatric hematological pathologies. Although this therapeutic approach has significantly improved clinical outcomes of pediatric patients with malignant and non-malignant disorders, graft-versus-host disease (GVHD) is an important cause of morbidity and mortality in patients receiving HSCT . One of Gao et al. in Chinese pediatric patients with ALL with the noteworthy aim to establish a personalized dosage regimen. In particular, potential covariates such as age, body weight, and biochemical measurements  on MTX PK disposition were investigated. The results obtained from this study revealed that body weight and serum creatinine levels (SCr) were significant covariates on the disposition of MTX. Therefore, this popPK model combined with an a posteriori Bayesian approach can be used to estimate individual PK parameters and optimize personalized MTX therapy for pediatric patients with ALL Gao et al.High-dose methotrexate (HD-MTX) is widely used in pediatric acute lymphoblastic leukemia (ALL) treatment regimens. A popPK model of HD-MTX has been developed by Xia et al. have analyzed plasma valproic acid (VPA) concentrations in 711 pediatric patients with epilepsy and compared a routine Enzyme Multiplied Immunoassay Technique (EMIT) with a validated in-house LC-ESI-MS/MS method on the same samples. Consistency between the two assays was evaluated using linear regression and Bland-Altman analysis. Results revealed that both methods were closely correlated although EMIT assay overestimated VPA levels in human plasma compared with LC-ESI-MS/MS method. Therefore, the authors conclude that switching from immunoassays to LC-based techniques for TDM of VPA deserves close attention and a therapeutic range of 35.0–75.0 μg/ml could be more feasible. However, further studies are required to evaluate the eligibility of this alternative range in clinical practice Xia et al.Different types of bioanalytical techniques are commonlyused in clinical laboratories for TDM. Historically, gas chromatography (GC), HPLC-UV, and LC-MS/MS are the most frequently used technologies to measure drug concentrations in different biological matrices. Immunoassays, including the enzyme-linked immunosorbent assay (ELISA), are also used for the quantification of several drugs. Usually, these assays are easily accessible for many laboratories and do not require highly specialized personnel. However, immunoassays are not available for all drugs monitored in clinical laboratories and their sensitivity, specificity, and accuracy toward the target compound is lower than GC, HPLC-UV, or LC-MS/MS. Therefore, the latter techniques are usually preferred to perform TDM although not always accessible to all laboratories. In a recent study, ABL)-ELISA assay has been developed and validated by Montecchini et al. to investigate ABL1 in vitro activity in four immortalized leukemic cell lines. The assay was further validated on blasts derived from an adult affected by chronic myeloid leukemia (BCR-ABL1 positive) and a child affected by ALL (BCR-ABL1 negative). Phosphorylation of PABL was inhibited after incubating BCR-ABL1 positive cell lysates with imatinib, but not with ruxolitinib. In conclusion, the authors suggest that the PABL-based ELISA assay provides a novel in vitro tool for screening both the aberrant ABL1 activity in BCR-ABL1 like ALL leukemic cells and their potential response to tyrosine kinase (TK) inhibitors Montecchini et al.Similarly, a novel peptide biosensor (PLong et al. have evaluated the PK profile of caffeine in preterm infants alongside the safety and efficacy of different doses of caffeine, therapeutic concentration ranges, and the impact of genetic variability on caffeine therapy. Although safety and efficacy of standard-dose caffeine have been already assessed, evidence for the safety of higher administered doses is not yet explored. Therefore, the authors suggest the utility of TDM when dose optimization is required for preterm infants who lack clinical response to standard-dose caffeine. In fact, even polymorphisms in PD-related genes have a significant impact on the inter-individual variability in pharmacological response to caffeine, therefore an individualized therapy based on TDM data could be particularly useful during routine clinical practice Long et al.Neonatal pharmacology deserves particular attention due to the complexity of maturational and physiological changes that not only characterize neonates but also affect the response of these patients to pharmacological treatments. Although pre-terms are generally considered part of the neonatal population, they are physiologically and pharmacologically different from full-term neonates and, therefore, require specific considerations . For exaIn conclusion, TDM in pediatrics represents, similarly to adults, a useful tool that allows a more tailored drug administration and, therefore, the individualization of therapy to avoid risks of therapeutic failures or drug toxicity. The contributions to this issue of Frontiers in Pediatrics clearly demonstrate the value of TDM for tailoring the administration of various drugs in pediatric patients, mainly during antimicrobial and immunosuppressive therapies. However, various challenges remain, preventing a wide uptake in routine clinical care. Overcoming the bioanalytical issues and providing a software tool for model-informed dosing support may be key for TDM to grow into a user-friendly and reliable tool for improving treatments in children and neonates.


Expanding availability to naloxone is a core harm reduction strategy in efforts to address the opioid epidemic. In the US, state-level legislation is a prominent mechanism to expand naloxone availability through various venues, such as community pharmacies. This qualitative study aimed to identify and summarize the views of experts on state-level naloxone access laws.We conducted a three-round modified-Delphi process using the online ExpertLens platform. Participants included 46 key stakeholders representing various groups  with expertise naloxone access laws. Participants commented on the effectiveness and implementability of 15 state-level naloxone access laws (NALs). We thematically analyzed participant comments to summarize views on NALs overall and specific types of NAL.Participants commented that the effectiveness of NALs in reducing opioid-related mortality depends on their ability to make sustained, significant impacts on population-level naloxone availability. Participants generally believed that increased naloxone availability does not have appreciable negative impacts on the prevalence of opioid misuse, opioid use disorder (OUD), and non-fatal opioid overdoses. Implementation barriers include stigma among the general public, affordability of naloxone, and reliance on an inequitable healthcare system.Experts believe NALs that significantly increase naloxone access are associated with less overdose mortality without risking substantial unintended public health outcomes. To maximize impacts, high-value NALs should explicitly counter existing healthcare system inequities, address stigmatization of opioid use and naloxone, maintain reasonable prices for purchasing naloxone, and target settings beyond community pharmacies to distribute naloxone. Mortality from opioid-related overdoses remains a significant public health issue , with a Despite the priority of addressing the opioid crisis and empirical evidence supporting naloxone’s efficacy in reversing overdoses, there is substantial variation in state adoption of NALs , mixed rhttps://osf.io/f4hk8/). A separate manuscript reports quantitative findings from the online modified-Delphi process [The study was deemed exempt by the RAND Human Subjects Protection Committee ID-2018–0506). We prospectively registered the study on the Open Science Framework (18–0506.  process .Our study was guided by the GRADE Evidence-to-Decision (EtD) Framework, which facilitates structured and transparent use of evidence to inform public health recommendations and decisions . Our quaWe recruited participants identifying with one of several stakeholder groups . We first developed a recruitment list from published research related to NALs, project advisory board suggestions, and relevant organizations’ member lists. We also used a “snowball sampling” approach allowing approached stakeholders to nominate further participants. We then emailed a recruitment survey to potential participants to indicate whether they would be willing to participate in the online modified-Delphi process, offering a $300 gift card or prepaid debit card for study completion. Individuals indicating interest provided demographic data and stated their preference to participate in the panel focused on either (a) effectiveness or (b) implementability of NALs. Participants electronically provided informed consent, which included information about sharing de-identified information and assurances of protecting confidentiality of responses and discussion comments. We aimed to recruit 40–80 participants (20–40 participants per panel), based on guidance for online expert panels that aim to engage large, diverse, and geographically-distributed groups of stakeholders in consensus exploration .We conducted two online modified-Delphi panels concurrently during summer 2020 using RAND’s ExpertLens system: one on NAL effectiveness and another on their implementability. We explicitly instructed participants that, while there are several critically important channels for naloxone distribution, we specifically were interested in understanding policies that target naloxone access and distribution through pharmacies . In Round One, participants rated each policy on four outcomes (Effectiveness Panel) or implementability criteria (Implementation Panel). In Round Two, participants explored areas of agreement and disagreement by discussing Round One results in an anonymous, asynchronous online forum. In Round Three, participants re-rated Round One results following Round Two discussion . Each roWe constructed a list of 15 NALs through an iterative process of feedback from the project’s advisory board and cognitive testing on the ExpertLens platform  agreed to participate: 24 in the panel on NAL effectiveness (Panel A) and 22 in the panel on NAL implementability (Panel B). Most panelists were female (54%), White Non-Hispanic (78%), identified as researchers (61%), and resided in the Northeast 41%). We did not identify any discernable differences in final themes by participant demographics. Overall, participants provided 2,658 comments: 1,479 in Round One , 319 in Round 2 , and 860 in Round Three . Across all comments, participants cited several studies as warrant for their claims , 35–41.%. We didWe organized overall themes by the domains of the GRADE EtD Framework: effectiveness, acceptability, feasibility, affordability, and equity . As with acceptability, participants noted stigma substantially influences stakeholder support for NALs and consequently NAL feasibility across a range of organizations and settings. Specific stakeholders included first responders, healthcare organizations, healthcare and social service providers, insurance companies, law enforcement, lawyers and legal experts, licensing boards, naloxone manufacturers, pharmacists and pharmacy chains, prescribers, professional associations, regulators, and state legislatures. Organizations and settings included including clinics, healthcare systems, hospitals, and pharmacies.Variability in ratings on the societal affordability of NALs was largely a factor of the cost of naloxone itself, who pays for naloxone, and effectiveness of an NAL in reducing overdoses. The cost of naloxone significantly influenced views on affordability because naloxone is the most significant direct and ongoing cost of NALs, and the cost of naloxone can vary due to numerous factors . Consequently, as part of their rationale for affordability ratings, participants frequently commented on naloxone-related cost burdens for specific stakeholders . From a societal perspective, many participants rating NALs as highly affordable noted cost savings associated with decreased overdose mortality.Variability in equitability ratings largely related to inequities and disparities of society and the systems in which NALs are implemented, rather than the equitability of NALs per se. Namely, participants frequently commented on structural, systemic, and institutional oppression many populations face in the USA due to their race, ethnicity, socioeconomic status, and place of residence—and how the resultant unjust stratification of health opportunities and outcomes are likely to replicate in implementation of NALs that rely on pharmacies and the existing healthcare system. In addition, participants often identified discrimination and interpersonal biases held by healthcare providers and pharmacists as another potential source of inequitable implementation. Participants also noted pharmacies are a suboptimal mechanism for distributing naloxone to people most likely to experience or witness an overdose, as pharmacies are often less prevalent in specific areas  and less accessible to specific populations . Consequently, even NALs that increase the number of places and ways that people can access naloxone still could widen disparities in naloxone access if implemented solely through pharmacies. Lastly, participants generally posited that lowering out-of-pocket naloxone cost increases an NAL’s equitability, particularly for low-income and under- or uninsured populations.We organized themes for specific NALs by policy groupings: liability policies, education and training requirements, co-prescribing naloxone, layperson accessibility, expanded pharmacy access, and cost subsidization . For example, one participant noted that all communications and education tools from the federal Rural Health Opioid Program  are freely available for replication. Putting aside concerns about stigma of substance use and naloxone generally, perceived acceptability of these laws specifically stems from agreement among the general public to provide information about proper use for all medications. However, concerns about equitability arose by the interaction of the onerous aspects of this mandate with the structural oppression and interpersonal discrimination faced by many people and communities affected by these policies. Consequently, panel consensus reflected views that these policies likely disproportionately negatively impact marginalized communities, namely people of color, people with mental health disorders, people who are homeless, rural residents, and individuals with lower socioeconomic status or communities with less resources. That said, conditional on receiving training, these policies could improve equity among those who previously have been underserved in terms of medical education.Naloxone co-prescribing requirements were generally viewed as effective in substantially increasing naloxone pharmacy distribution, but only modestly effective in reducing overdose mortality. While participants noted evidence that these policies expand access to naloxone through pharmacies –37, thisParticipants often showed support for the principle of using risk indicators beyond opioid overdose in order to cover overdose risk more broadly. However, participants noted numerous issues of this policy in-practice: patient perceptions of additional questions as invasive, provider burdens due to increased complexity and difficulty in obtaining this information , uncertainty about the indicators to use (and subsequent waste in resources if the wrong indicators are used), and greater subjectivity (compared to the more objective criteria of using only prescribed opioid dosages) increasing the opportunity for interpersonal biases to yield further inequities.Participants generally viewed NALs facilitating greater accessibility of naloxone to anyone (regardless of opioid use status) as effective in removing barriers for laypersons and thereby increasing naloxone pharmacy distribution. Namely, over-the-counter (OTC) pharmacy supply was one of the NALs that participants believed had the best chance to destigmatize acquiring naloxone, because many third parties  are likely to acquire naloxone via a pharmacy . These ratings translated into perceptions of meaningful reductions in fatal overdoses from authorizing OTC naloxone pharmacy supply, but not from third party prescription due to the latter’s reliance on physician prescriptions and narrower focus on laypersons who may not actually witness overdoses. Conversely, participants viewed third party prescriptions as slightly more equitable than OTC pharmacy supply due to concerns about prohibitory retail costs of OTC naloxone for low-income persons. While both OTC supply and third-party prescribing had consensus ratings of “high” affordability, participants raised financial concerns for both , but viewed both as generally acceptable to the public and feasible changes to prescriber and dispenser practices.Participants also viewed expanded pharmacy access laws as effective in facilitating naloxone availability by removing patient barriers to acquiring naloxone  and enabling pharmacist autonomy. As evidence for their acceptability, participants frequently cited successful adoption of these NALs without much public pushback (likely due to their commensurability with obtaining other types of medication from pharmacists). Participants explained the “high feasibility” consensus was conditional on pharmacist willingness to dispense naloxone without physician involvement and lack of opposition from prescribers concerned about professional scope creep. “High affordability” was generally due to eliminating costs associated with prescriber office visits. Statewide standing/protocol orders and pharmacist prescriptive authority had the added benefit of lesser administrative costs compared to collaborative practice agreements . Removing the need to access prescribers increased participant views of equitability, though participants noted the remaining possibilities of pharmacist bias and limited access to pharmacies.Participants viewed cost-subsidization NALs as effective in significantly facilitating naloxone pharmacy distribution because they address the significant barrier of out-of-pocket costs to individuals. Participants particularly viewed statewide free naloxone as the most equitable NAL in effectively facilitating naloxone pharmacy distribution and reducing overdose mortality, given its elimination of out-of-pocket costs and potential for destigmatizing naloxone. However, they also considered it the least affordable policy for states—and least acceptable as a result. While insurance coverage was not seen as equitable and effective in reducing overdose mortality due to many high-risk patients not having insurance, participants viewed insurance coverage as the most implementable cost-subsidization policy due to public support of shifting burdens to insurance companies covering the medication. Participants similarly viewed state subsidies for naloxone purchase through insurance as less effective due to its reliance on high-risk patients having insurance, although they also saw it as less implementable than insurance coverage as it only provides assistance with co-pays (rather than full coverage) and costs fall on the state (rather than insurance companies).Experts believed that the effectiveness of NALs in reducing opioid-related mortality requires sustained, significant impacts on population-level naloxone availability. This necessitates addressing implementation barriers that apply broadly across NALs, including affordability of naloxone itself, reliance on an inequitable healthcare system, and stigma—which can mitigate the effectiveness of all types of policies considered , 47. ExpRegarding specific NALs, expanded pharmacy access laws appeared to be the most valuable set of policies—with statewide standing or protocol orders considered particularly “high-value”—as these policies remove the most barriers to naloxone distribution and access. This does not imply these policies are absent implementation barriers, such as failure of pharmacies to stock naloxone and high out-of-pocket costs to purchasers ; thus, tThe following limitations should be considered when interpreting the study’s results. Our method required participants to have stable Internet access, proficiency with online survey systems, and several hours of availability over the three rounds. While our sample size is commensurate with recommendations for online modified-Delphi processes , we did Experts believe NALs that significantly increase naloxone access are associated with less overdose mortality without risking substantial unintended public health outcomes. To maximize impact, “high-value policies” explicitly counter existing inequities in the healthcare system, address stigmatization of opioid use and naloxone, maintain reasonable prices for purchasing naloxone, and target settings beyond community pharmacies to distribute naloxone.


WHO pneumonia guidelines recommend that children (aged 2-59 months) with chest indrawing pneumonia and without any “general danger sign” can be treated with oral amoxicillin without hospital admission. This recommendation was based on trial data from limited contexts whose generalisability is unclear. This review aimed to identify which children with chest-indrawing pneumonia in low- and middle-income countries can be safely treated at home, and under what conditions is it safe to do so.We searched MEDLINE, EMBASE, and PubMed for observational and interventional studies of home-based management of children (aged 28 days to four years) with chest-indrawing pneumonia in low- or middle-income countries.We included 14 studies, including seven randomised trials, from a variety of urban and rural contexts in 11 countries. Two community-based and two hospital-based trials in Pakistan and India found that home treatment of chest-indrawing pneumonia was associated with similar or superior treatment outcomes to hospital admission. Evidence from trials (n = 3) and observational (n = 6) studies in these and other countries confirms the acceptability and feasibility of home management of chest-indrawing pneumonia in low-risk cases, so long as safeguards are in place. Risk assessment includes clinical danger signs, oxygen saturation, and the presence of comorbidities such as undernutrition, anaemia, or HIV. Pulse oximetry is a critical risk-assessment tool that is currently not widely available and can identify severely ill patients with hypoxaemia otherwise possibly missed by clinical assessment alone. Additional safeguards include caregiver understanding and ability to return for review.Home treatment of chest-indrawing pneumonia can be safe but should only be recommended for children confirmed to be low-risk and in contexts where appropriate care and safety measures are in place. World Health Organization (WHO) clinical guidelines for children are intended to provide evidence-based guidance to health workers in diverse clinical settings, with a focus on low-resource and smaller health facilities -3. The gThe 2013 revisions were informed by evidence that oral antibiotics are equivalent to intravenous antibiotics for most children with pneumonia -12 and sHowever, the studies which influenced this guideline change were conducted in limited contexts and under controlled trial conditions. The observed pneumonia mortality rates were very low compared to most low- and middle-income country (LMIC) settings, the rates of wheezing and viral aetiology were high, and large numbers of patients were excluded. This raised concern that they may not be representative of the populations for whom WHO guidelines are intended – particularly in higher mortality contexts with higher rates of bacterial pneumonia and in the absence of the level of monitoring usually associated with clinical trials . While tA subsequent study from Kenya sought to evaluate the appropriateness of this change in guidelines, adding to the evidence showing that chest indrawing in sub-Saharan Africa hospital contexts, along with other factors not currently incorporated in the WHO classification of severity of pneumonia , were significant risk factors for mortality ,21.This review aimed to examine all published studies from LMICs which analysed home treatment of chest indrawing pneumonia, in comparison to referral or admission for inpatient hospital treatment, to establish the degree to which we can be confident in recommending such management, and under what conditions.We conducted a systematic search of Medline, Embase, and PubMed (for articles not yet indexed in Medline) for all relevant articles published since January 1, 2000 . We mapped search terms to medical subject headings where possible, using Boolean operators to combine searches into our final systematic search query. We used synonyms of “pneumonia”, “chest-indrawing”, “home treatment” and “child” to target the search strategy, with oversight from an experienced medical librarian to ensure all relevant papers were identified. We also searched reference lists of all included references for eligible studies. The specific search terms used for our Medline search and further details of the search strategy, information sources, and data collection processes are included in Appendix S1 in the We included studies published since 2000 evaluating outcomes for children (aged 28 days to 4 years) with WHO-defined pneumonia with chest-indrawing (“chest-indrawing pneumonia”) treated at home (We used a standardised data extraction form to extract data relevant to our review. Two reviewers (CW and PW) independently extracted data from each eligible study and entered it into an Excel spreadsheet . We resolved disagreements by discussion and contacted study authors where appropriate to resolve any uncertainties. We did not attempt a meta-analysis of extracted data as our primary goal was to understand treatment outcomes with respect to population and context. We categorised context, population and outcome data, then qualitatively synthesised results to determine whether children with WHO-defined chest-indrawing pneumonia could be safely managed at home, and in what contexts.We assessed the quality and risk of bias of all included studies by using the Effective Public Health Practice Project (EPHPP) Quality Assessment Tool ,23. UsinEthical approval for this study was not required.We retrieved 1521 references from database searches, screened 1131 unique articles and identified 26 articles for full-text screening. Of these, 12 were excluded because it was impossible to extract data specific to patients with chest indrawing pneumonia (n = 8) -31 or spCharacteristics of included papers can be found in The studies took place in 9 lower-middle income countries and 2 low-income countries. Seven studies were predominately in urban settings ,37,39,41All studies included children 59 months of age or younger, except one  which inFour studies reported vaccination coverage, with generally high coverage ranging from 74% to 97% receiving all age-appropriate vaccines according to local guidelines ,18,39,43Many patients were excluded following screening in most of these studies, with numbers of enrolled patients ranging from 5%  to 45%  of the tAll studies either compared management of pneumonia with chest indrawing at home vs hospital management, compared management algorithms that mandated hospital management in one arm and allowed home treatment in another, or were observational studies of home care.Oxygen saturation was documented as having been measured in patients in eight of the 14 studies -41,43, aTen of the 14 studies described the training procedures for study staff, which ranged from one day  to one wThe primary outcome of eight studies ,39,41,42Other studies’ primary outcomes were appropriateness of treatment compared to guidelines , successOf the seven included randomised or cluster-randomised trials, four -15,17 diOne other randomised trial from urban Bangladesh  found noThese trials reported low treatment failure rates for those treated in the community , and mortality was very low .These trials included several important safety measures in selecting patients for community-based treatment and monitoring them and responding to deterioration  and very low mortality (range = 0%-0.6%) among children with chest-indrawing pneumonia treated in the community  would not have been referred compared to 8% using the existing guideline  [The inclusion of pulse oximetry was found to improve pneumonia diagnosis and identification of hypoxaemia in a cluster randomised trial among rural primary care facilities in Ethiopia , an obsedrawing) . The Tandrawing) . The Ethdrawing) . The Papdrawing) .While community health workers’ assessments and that of more highly trained health professionals were generally concordant ,42, chilStudies found mixed effects of community-based treatment guidelines on the number and proportion of patients presenting, being referred, and adhering to treatment recommendations. An observational study in Bangladesh that introduced community-based management of chest-indrawing pneumonia after observing low referral completion rates found increased numbers of children presenting with chest-indrawing pneumonia to primary care facilities, lower referral initiation rates, and minimal change in the number or proportion following through with referral recommendations . Other sThe economic cost of management of chest indrawing pneumonia in the community, where examined, was significantly lower than the cost of inpatient management ,36.Evidence from four trials suggests similar or lower treatment failure rates among children with chest-indrawing pneumonia treated at home compared to referral or inpatient management. However, these trials were all conducted in Pakistan or India and excluded patients with significant comorbidities, making generalisability limited – particularly in populations where malnutrition or other comorbidities are common and settings where the aetiology and associated features of pneumonia differ.Evidence from other interventional and observational studies supports the feasibility of home management of chest-indrawing pneumonia, if safeguards are in place, with relatively low rates of treatment failure and mortality. These studies included settings with higher HIV prevalence and likely higher rates of bacterial pneumonia than peri-urban Pakistan and India, where the RCTs were conducted. They also represented care models based in the community, primary care facilities, secondary and tertiary hospitals, and “day clinics”. In all these studies, the conditions for safe outpatient management of chest-indrawing pneumonia were carefully defined. Children with hypoxaemia or danger signs and those with HIV, malnutrition, anaemia, or other comorbidities, were all managed as inpatients. A strong emphasis was placed on appropriate training of the health care staff, education of caregivers, and appropriate and timely follow-up of all patients.Pulse oximetry was a core part of risk assessment in most included studies , identifying severely ill children with hypoxaemia who otherwise may not have been appropriately referred or treated with oxygen. This finding concurs with other studies on pulse oximetry that have found pulse oximetry is an objective measure that improves hypoxaemia detection and risk stratification and provides health care workers and patients/families with greater confidence in treatment plans ,44-49.Aside from pulse oximetry, there is insufficient evidence supporting the inclusion of other point-of-care tests  as part of routine triage of children with chest-indrawing pneumonia ,50.These findings are supported by a recent study on the management of chest-indrawing pneumonia by community health workers in Bangladesh, India, Ethiopia, and Malawi, which found equivalent treatment outcomes compared to facility-based care . This stBased on existing evidence, we suggest that home treatment of chest-indrawing pneumonia may be appropriate for low-risk patients with adequate safety and care structures  from 8 This review was limited by the number and quality of studies included. We identified few randomised trials, from narrow geographical and epidemiological settings, and their trial conditions may not reflect what is possible in actual practice. To address these issues, we included interventional and observational studies that could provide more ‘real-life’ data on the conditions required to provide safe care in the community. Future studies exploring the implementation of home management of chest-indrawing pneumonia in more routine practice settings and in diverse contexts will give greater clarity and confidence about how to safely manage these children without admission.Home treatment of chest-indrawing pneumonia can produce treatment outcomes comparable to hospital-based care for carefully selected populations in certain contexts. However, home treatment should only be recommended in children who are low risk and in contexts where adequate care and safety provisions are available. Prospective operational research into home treatment of chest indrawing pneumonia in settings outside of South Asia, particularly Sub-Saharan Africa, would help with determining the generalisability of the findings of existing trials.


Palliative care is an interdisciplinary care to optimize physical, psychosocial, and spiritual symptoms of patients and their families whose quality of life is impaired by serious, life-limiting illness. In 2021, the importance of providing palliative care in the intensive care unit (ICU) is well recognized by various studies to alleviate physical symptoms due to invasive treatments, to set patient-centered goals of care, and to provide end-of-life care. This paper summarizes the evidence known to date on primary palliative care delivered in the ICU settings. We will then discuss the potential benefits and harms of primary palliative care so that critical care clinicians are better equipped to decide what services might best improve the palliative care needs in their ICUs. Palliative care is an interdisciplinary care to optimize physical, psychosocial, and spiritual symptoms of patients and their families whose quality of life is impaired by serious, life-limiting illness. Up to 75% of patients admitted to the intensive care units (ICU) experience distressful symptoms   who introduces the concept of organ donation. Occasionally, a palliative care physician will continue this conversation with the family of a patient who has been declared brain dead about what happens next. Some data exist that communication training may be helpful for palliative care physicians to discuss this topic with the families of brain-dead patients .Many palliative care interventions in the ICU have focused on supporting families of ICU patients, specifically targeting the domains of patient-/family-centered decision-making, emotional and practical support, and communication . ResearcOverall, effects of these primary palliative care interventions on family satisfaction and anxiety/depression scores have been mixed, with some studies showing modest improvements and others showing no difference between groups , 29–32. In contrast to the mixed results observed in family satisfaction and psychological distress, multiple studies have found that family-focused, primary palliative care interventions are consistently associated with decreased critical care use at the end of life , 32–36. Taken together, these studies suggest that while family-focused primary palliative care interventions in the ICU have variable effects on family satisfaction and psychological distress, they are a cost-effective intervention for reducing potentially undesired critical care at the end of life and do not seem to increase overall mortality. Future primary palliative care research should investigate why some family support interventions seemed to improve family satisfaction and psychological distress, and to elucidate which interventions accomplish both improving the family experience and reducing intensity of care at the end of life. As discussed below, it may be that complementary clinician-focused or systems-based interventions improve these outcomes as well.The impact of interventions on clinicians has also been increasingly explored and studied over the past several decades . These iIn a 2014 national study of 89 hospitals and 71 ICU training programs, quality of education in primary palliative care skills and presence of evidence-based bedside tools were both associated with reduced ICU length of stay . SeveralRecently, the importance of communication skills training for critical care clinicians has been attracting attention. Oppenheim et al. reported that in a simulated conversation between an ICU health care provider and a patient's family regarding the patient's prognosis, family members interpret physicians’ indirect response to questions about prognosis as more optimistic than direct responses . TherefoCritical care clinicians receive family-centered communication skills training as part of their ICU training, which can improve healthcare professionals' self-efficacy and family satisfaction . In the Attempts to implement multifaceted bundles to improve critical care clinicians’ ability to provide palliative care have had mixed results. For example, the largest such project published to date showed a positive outcome in the initial single site study, but no difference in the subsequent multisite study , 49.Finally, relatively little has been published about the impact of palliative care interventions on critical care clinician wellness. One single site study showed that trainees felt palliative care involvement was supportive for them and helped them be better clinicians . “Death In addition to interventions targeted individual critical care clinicians, there have been some interventions aimed at designing systems to help support the reliable delivery of palliative care in the ICU. The idea of system design as a lever to facilitate best-care, has been well documented in the ICU literature . The intIn summary, although several exploratory or quality improvement studies have investigated the impact of system interventions to drive best-care, and some suggest promise, none have demonstrated a system solution that yet warrants wide-scale adoption.Multi-level ICU palliative care interventions are interventions that include components from across several domains  and frequently target multiple outcomes. As detailed above, family-, clinician-, and system-level interventions individually have yielded some promising results to improve the overall quality of care. However, the impact on patient/surrogate-centered outcomes has been limited in many trials. The motivation for multi-level interventions is to leverage the benefits found in single-level interventions and to extend the scope or reach of those benefits by addressing theoretical barriers to improvement. Multi-level ICU interventions are variably designed, with outcomes often set to test a combination of surrogates’ psychological distress, ICU utilization metrics, and quality of communication measures. At their core, most focus on addressing communication and information exchange , 62.Nearly all multi-level palliative care ICU trials have shown significant reductions in ICU length of stay without increasing mortality , 62, 63.Taken together, multi-level trials hold promising results, particularly in improving surrogate psychological symptoms as compared to single-level interventions. However, even here the scope and impact of palliative ICU interventions could be improved. Several theoretical limitations may explain the variable or limited impact of multi-level interventions. First, few interventions have included components that target the post-ICU period, when bereavement and other challenges may pose a substantial burden for surrogates . Second,We have described the potential benefits of primary palliative care interventions that focus on the families, clinicians, and system levels. Many demonstrated modest effects on the overall quality of care yet often lacked patient-centered benefits. What about the potential harms of implementing these interventions in the ICU settings? The main costs of implementations are clinician training, clinical time to deliver primary palliative care services, and system-level tools . The potential barriers –69 and rIn summary, we described the modest benefits of primary palliative care interventions in the ICU settings. Overall improvement in quality of care can be expected despite the mixed level of evidence in patient-centered outcomes. Given the low potential harm and cost of implementing these primary palliative care interventions, healthcare organizations may consider initial investment to disseminate some forms of primary palliative care interventions in the ICU settings.


Ischemia with nonobstructive coronary artery disease (INOCA) is increasingly recognized as a significant cause of angina, myocardial remodeling, and eventually heart failure (HF). Coronary microvascular dysfunction (CMD) is a major endotype of INOCA, and it is caused by structural and functional alterations of the coronary microcirculation. At the same time, atrial cardiomyopathy (ACM) defined by structural, functional, and electrical atrial remodeling has a major clinical impact due to its manifestations: atrial fibrillation (AF), atrial thrombosis, stroke, and HF symptoms. Both these pathologies share similar risk factors and have a high comorbidity burden. CMD causing INOCA and ACM frequently coexist. Thus, questions arise whether there is a potential link between these pathologies. Does CMD promote AF or the reverse? Which are the mechanisms that ultimately lead to CMD and ACM? Are both part of a systemic disease characterized by endothelial dysfunction? Lastly, which are the therapeutic strategies that can target endothelial dysfunction and improve the prognosis of patients with CMD and ACM? This review aims to address these questions by analyzing the existing body of evidence, offering further insight into the mechanisms of CMD and ACM, and discussing potential therapeutic strategies. Ischemic heart disease and atrial fibrillation (AF) are major cardiovascular diseases that cause a significant burden on the health system, with increased morbidity and mortality, not only in the elderly but also among the younger population. While obstructive coronary artery disease (CAD) has been extensively characterized with well-defined therapeutic options, ischemia with nonobstructive coronary artery disease (INOCA) still faces many unanswered questions regarding pathophysiology and optimal therapy, despite being increasingly recognized as a significant cause of angina, myocardial remodeling, and eventually heart failure (HF) . Up to 7At the same time, AF is more than just an arrhythmia, as it might reflect an underlying diseased atrium with structural, functional, and electrical remodeling, all of these processes being incorporated under the term ‘atrial cardiomyopathy’ (ACM). ACM has been recently recognized as a clinically relevant entity in which atrial dysfunction is considered the primordial pathological alteration, leading to cardiovascular consequences such as AF, HFpEF, and embolic stroke due to atrial thrombosis even in the absence of atrial arrhythmias .Despite apparently different cardiovascular pathogenies, coronary vascular dysfunction and ACM share similar risk factors, such as arterial hypertension, diabetes mellitus (DM), obesity, obstructive sleep apnea (OSA), or dysfunctional epicardial adipose tissue (EAT) ,5. MoreoThere are limited data exploring the potential link between these pathologies, whether they share similar pathogenic mechanisms or whether one promotes the other. Additionally, the particular association between ACM and CMD has been poorly characterized in previously published papers. This review aims to describe existing evidence on the relationship between ACM and phenotypes of INOCA with a focus on CMD. We also explore the role of systemic ED as an underlying pathogeny and describe its associations with atrial ED and CMD. In the second part of the manuscript, current knowledge of potential therapeutic strategies targeting the common pathogenic substrate of these diseases is addressed. We emphasize the role of non-pharmacological measures such as weight loss and continuous positive airway pressure (CPAP) therapy in OSA and question whether EAT could be a new therapeutic target to improve cardiovascular outcomes. Finally, we discuss the available evidence on pharmacotherapies with a potential role in modulating systemic ED, ACM manifestations, and CMD.The coronary arterial system includes three major compartments, each with a specific vessel structure and function. The proximal compartment is formed by the epicardial coronary arteries, which are large, conductive vessels with a three-layer wall and a main role in transportation ,8. CoronCMD encompasses structural and functional abnormalities of the coronary microcirculation that alter the myocardial perfusion and ultimately cause ischemia. Structural remodeling implies luminal narrowing of arterioles by media thickening and intimal proliferation, as well as perivascular fibrosis and capillary rarefaction . FunctioCurrently, there are no available instruments to directly visualize the coronary microvasculature. The diagnostic algorithm of INOCA has been already extensively described . In brieAlthough composed of a single row of cells covering the endocardium and the blood vessels, the endothelium plays a major role in the modulation of vascular tone and contributes to processes such as inflammation, hemostasis, or thrombosis. Endothelial cells synthesize NO in response to hemodynamic shear stress, which promotes vasodilatation and inhibits endothelial thrombosis . In addiThe first detailed analysis of ACM was published in 2016, in an Expert Consensus Document of several societies, which defined it as “any complex of structural, architectural, contractile or electrophysiological changes affecting the atria with the potential to produce clinically relevant manifestations”. The pathophysiological processes leading to the development of ACM are frequently intricate and include atrial cardiomyocyte dysfunction, fibrotic changes, and non-fibrotic atrial infiltration with inflammatory cells or adipocytes . These aBoth CMD and ACM share similar risk factors, suggesting a possible common pathophysiological pathway causing these pathogenies. Traditional cardiovascular risk factors and emerging entities with deleterious cardiovascular effects such as OSA and dysfunctional EAT promote ED, which contributes to CMD and ACM development, reinforcing their possible association ,5.Aging is associated with significant atrial remodeling, due to increased atrial fibrosis. Atrial dilatation is more severe in the elderly, and AF incidence increases with age . MoreoveSmoking significantly increases the risk of AF by up to 33%, and it is also associated with INOCA endotypes ,23. The Obesity is a comorbidity with major deleterious systemic effects. Adipose tissue is an active metabolic organ that secretes proinflammatory cytokines, creating a low-grade systemic inflammatory environment. Chronic vascular inflammation promotes ED, which alters the coronary arterial vasomotor response, causing CMD . In addiArterial hypertension is the underlying cause of at least 1 in 5 cases of incident AF, being the most prevalent modifiable risk factor for AF . IncreasDiabetes mellitus is another major risk factor for both CMD and ACM. Persistent hyperglycemic stress causes mitochondrial dysfunction with increased oxidative stress production, and activation of the advanced glycation end product (AGE)–AGE receptor (RAGE) system . These mOSA is a sleep disorder characterized by multiple nocturnal pauses in breathing due to airway collapse, followed by reoxygenation. OSA is associated with multiple cardiovascular consequences, and ED appears to mediate this pathogenic link . After tIn addition, it appears that OSA might contribute to the development of CMD in patients with nonobstructive CAD, most probably due to repeated episodes of hypoxemia–reoxygenation, which resemble the ischemia–reperfusion injury. In line with this, a meta-analysis showed that OSA was significantly associated with lower CFR . AnotherAt the atrial level, OSA triggers a true myopathic process, with electrophysiological changes caused by intermittent hypoxemia and hypercapnia, increases in intrathoracic pressure, and sympathovagal activation, which significantly augment the risk of AF . In factEAT is the visceral fat deposit of the heart and it is located beneath the epicardium and in direct contact with the myocardium. It is typically located in the atrioventricular and interventricular grooves, and it can surround the entire myocardial surface when the amount of fat increases. It is also arranged along the epicardial coronary branches, and it can extend into the myocardium as it follows the coronary adventitia. There is no anatomical structure separating the myocardium from the EAT; thereby, these structures share the same microcirculation, which enables cellular and molecular interactions at this level .EAT is an organ with a higher metabolic activity compared to subcutaneous fat, and under abnormal conditions, it directly exerts multiple cardiac deleterious effects. Metabolic disorders such as obesity and DM induce systemic inflammation which signals epicardial adipocyte proliferation and dysfunction. Subsequently, defective adipocytes secrete proinflammatory cytokines such as IL-6, IL-1β, and IL-8 which promote myocardial inflammation, cardiomyocyte dysfunction, and fibrosis, by paracrine and endocrine signaling . All theA large amount of published data shows a strong association between EAT and AF, which might be explained by the development of an inflammatory atrial myopathy ,37,38. SPatients with AF frequently associate anginal symptoms, with demonstrated ischemia and ventricular remodeling in the absence of obstructive CAD . Such maThis assumption is further implied by the work of Wijesurendra et al. who analyzed MBF with CMR at rest and after adenosine in patients with AF and without DM or obstructive CAD compared to matched controls in sinus rhythm, before and after ablation of AF. In the AF group, before ablation, patients had significantly lower baseline and stress MBF relative to controls, suggesting that AF is associated with significant CMD. In addition, the impaired MBF was proportionally correlated with significantly larger LA volumes, and lower LA emptying fraction, advocating for a proportional link between the severity of CMD and ACM. After ablation, baseline and stress MBF did not ameliorate, implying that CMD is not reversible after sinus rhythm restoration. This finding suggests that CMD is not only a direct effect of the AF but may reflect an underlying vascular pathogenic state such as ED. Similarly, the LA emptying fraction remained unchanged after ablation despite considerable reduction in LA maximal volume, reinforcing the existence of an atrial myopathic process with atrial mechanical dysfunction .The relationship between CMD and AF appears, however, to be bidirectional, as illustrated in A growing body of evidence lately led to the assumption that cardiac alteration causing HFpEF is a consequence of systemic inflammation, in the context of associated comorbidities  3,49,5050,51,52.Coronary microvascular ED is a central mechanistic pathway leading to HFpEF in the context of chronic inflammation  52]. Du. Du52]. Overcoming this limitation, Ozcan et al. directly evaluated the relationship between CMD, AF, and HFpEF in patients undergoing invasive coronary angiography and functional testing with adenosine infusion. CMD was defined as abnormal CFR in the absence of obstructive CAD. The results provided evidence of the clinical association between CMD and AF, as patients with AF were more likely to have abnormal CFR than patients without AF (61% versus 37%). Similarly, patients with CMD tended to have higher rates of AF than patients with normal CFR (32% versus 15%). Interestingly, HFpEF was present in nearly all patients with AF and CMD (91%), and the majority of patients with both AF and HFpEF associated abnormal CFR (71%). What is more, the presence of either CMD, AF, or HFpEF was associated with lower survival free of HF hospitalization at one year .In HFpEF, ACM might result predominantly from high left ventricular filling pressures or might develop synchronously with the LV myopathy secondary to myocardial inflammation and CMD  57,58].,58.57,58Concluding the above, there is a clear association between ACM, CMD, and HFpEF, which might reflect a signature profile of cardiac dysfunction in patients with a high comorbidity burden and systemic inflammation ,56,59.Recently published work led to a new perspective on CMD and ACM, implicating that both are cardiovascular manifestations of a pathogenic systemic state characterized by ED, affecting multiple territories such as the heart, brain, kidney, and peripheral arteries ,21. The The integration of CMD in the bigger picture of systemic ED is suggested by several studies exploring this potential link. Recently, Al-Badri et al. aimed to assess whether coronary endothelial function might be echoed by peripheral endothelial function in the femoral microcirculation. They included 85 patients, in whom they measured endothelium-dependent coronary epicardial and microvascular function with intracoronary ACh infusions, as well as endothelium-independent pathways with adenosine and sodium nitroprusside. Quantitative coronary angiography was performed before and after each vasoactive infusion, and coronary vascular resistance (CVR) and coronary flow reserve (CFR) were measured. Similarly, femoral vasoactive infusions with ACh and sodium nitroprusside were performed, and femoral vascular resistance (FVR) was measured. The results showed that the percentage change in FVR correlated with the percentage change in CVR in response to ACh, indicating a significant correlation between the magnitude of coronary and femoral microvascular vasodilation induced by ACh. Additionally, there was a significant correlation between changes in the epicardial coronary diameter and femoral microvascular dilation in response to ACh, as patients with epicardial coronary vasoconstriction had attenuated femoral vasodilation compared with those with epicardial coronary vasodilation. Similarly, the changes in CFR with adenosine-reflecting coronary microvascular dilation correlated with the changes in FVR with Ach-reflecting femoral microvascular dilation. Conversely, there was no correlation between the endothelium-independent coronary and femoral microvascular vasodilation in response to sodium nitroprusside infusion . These rAnother study found that CMD and epicardial vasospasm, assessed by invasive coronary functional testing with ACh and adenosine in patients with INOCA, were associated with systemic microvascular dysfunction, reflected in the altered function of peripheral small arteries from biopsies of subcutaneous gluteal fat, which had reduced maximum relaxation to ACh and increased vasoconstriction in response to ET-1 and a thromboxane agonist . SimilarThese results support the concept of ED as a systemic underlying pathogenic process and suggest that the detection of peripheral ED might be a marker of coronary endothelial dysfunction .ED is a major contributor to the pathophysiology of the wide spectrum of ACM .First, atrial ED promotes endothelial microthrombi formation with atrial thrombosis even in the absence of atrial arrhythmias, and a wealth of data suggests that ACM might be a potential etiology of embolic strokes of undetermined source (ESUS), which account for up to 30% of ischemic strokes ,19. In tSecondly, atrial ED promotes atrial electrical remodeling and potentiates an arrhythmic atrial substrate, ultimately causing AF. Electron microscopy studies showed that AF is associated with major endothelium ultrastructural changes, such as endothelial cell desquamation and subendothelial edema which further promotes endothelial cell disruption . In addiMultiple studies found an association between AF and circulating biomarkers of ED. VWF, which is produced by endothelial cells in response to endothelial damage and promotes platelet aggregation, has higher levels in patients with AF compared to those in sinus rhythm and correlates with the arrhythmic burden ,71,72,73The integration of AF in the systemic state of ED is supported by considerable reports showing correlations with peripheral ED. In line with this, one study demonstrated that patients with permanent AF had lower values of RHI assessed by PAT and a higher burden of inflammatory cytokines compared to controls in sinus rhythm; they also had more comorbidities, substantiating the notion of systemic ED caused by the comorbidity-related inflammation . SeveralWhile it is difficult to ascertain which is the primary pathogeny, whether AF or ED, respectively, the abovementioned data underlines the existence of a vicious circle in which ED creates an atrial arrhythmic substrate manifesting as AF, and AF promotes ED at the same time .The main studies describing the interplay between CMD, ACM, and ED are summarized in As evidenced by the above summary, ED is a systemic state which integrates both CMD and ACM development. These pathogenies share common risk factors and comorbidities which cause systemic inflammation and oxidative stress, potentiating ED. Currently, there is little information from randomized controlled trials regarding specific strategies to treat and reverse ED. Considering the shared systemic risk factors which promote ED, it appears legitimate that preventive lifestyle interventions such as smoking cessation and weight loss, and controlling comorbidities such as arterial hypertension, DM, and dyslipidemia, could ameliorate ED and the cardiovascular damage associated with it.In the following section, we discuss the impact of weight loss and CPAP therapy for OSA on systemic ED, CMD, and ACM manifestations. We also provide current knowledge on potential actions that target EAT. Finally, we analyze the effect on ED of pharmacotherapies used to treat arterial hypertension, diabetes, and dyslipidemia, and assess whether they might have a role in ameliorating CMD and ACM markers. Obesity is a major contributor to the development of vascular ED, and multiple studies have demonstrated that weight loss could significantly improve it ,88,89.A recent meta-analysis showed that obese patients who had undergone bariatric surgery had significant improvement in endothelial function ascertained by increasing FMD measurements, independent of the weight loss, suggesting that weight-independent mechanisms such as endocrine and incretin-mediated effects and improved inflammatory status might contribute as well to the amelioration of ED .The beneficial effect of weight loss translates into improved cardiovascular outcomes with a decrease in the incidence of AF. In a trial, patients with a weight loss > 10% had a 6-fold higher probability of arrhythmia-free survival compared with those with <10% weight loss . At the Similarly, it appears that weight loss and improved exercise capacity ameliorate CMD, reflected in increases in CFR assessed by Doppler echocardiography of the left anterior descendent artery ,95. ConvED is an early anomaly in OSA, and continuous positive airway pressure (CPAP) therapy could potentially modulate it.A meta-analysis evaluating this effect found a significant improvement in peripheral ED with CPAP use, noted in an important increment in brachial FMD of 3.9%, whereas no significant change was observed in the group with standard therapy or sham CPAP . SimilarThere is little evidence regarding the impact of CPAP therapy in patients with OSA and CMD. A study showed that short-term (2 weeks) discontinuation of therapeutic CPAP in moderate to severe OSA did not affect the coronary microvascular function, assessed with adenosine-induced MBF using PET . AnotherCPAP therapy appears to exert a benefic effect on ACM substrate. This concept is illustrated by the results of the SLEEP–AF study in which patients with AF and at least moderate OSA had improved parameters at electrophysiologic study after CPAP therapy compared to baseline electrophysiologic evaluation . SimilarAs previously stated, EAT creates an inflammatory environment that promotes CMD and AF. In line with this, reducing epicardial fat emerges as a potential therapeutic target to counteract coronary and atrial inflammation, and to decrease the risk of subsequent morbidity .Several studies demonstrated that significant weight loss following bariatric surgery is associated with a reduction in EAT ,109. MorVarious pharmacotherapies with direct or indirect action on adiposity might reduce or improve the function of EAT.Statin therapy appears to act on epicardial fat causing its reduction, independent of the cholesterol-lowering effect, which might suggest that other mechanisms such as their anti-inflammatory properties might be implicated ,112. ConGlucagon-like peptide 1 receptor agonists (GLP- 1RAs) are a class of drugs originally designed for the treatment of type 2 diabetes but are now also used to treat obesity due to their weight loss benefits. This additional effect is achieved secondary to GLP- 1RAs property of reducing the appetite and the sensation of hunger, as well as increasing satiety . Liraglu2 were randomly assigned to receive dapagliflozin in addition to metformin, or metformin alone. The results showed a significant decrease of EAT in the dapagliflozin group by up to 20% from baseline to 24 weeks, whereas in the metformin group the reduction in EAT was smaller. Interestingly, EAT thickness reduction assessed echocardiographically was significantly greater than body weight loss  which might suggest the contribution of alternative pathways independent of weight loss to epicardial fat reduction [Sodium–glucose cotransporter 2 inhibitors (SGLT2-is) are a revolutionary class of drugs due to their strong cardio–renal protective effect. While initially addressed to diabetic patients for their glucose-lowering action, they are currently one of the main pillars in the treatment of heart failure, as they demonstrated in large clinical trials a significant reduction in cardiovascular death and heart failure hospitalization . SGLT2-ieduction . Severaleduction ,122. Coneduction . Howevereduction . Anothereduction .In addition to their lipid-lowering effect, statins have multiple pleiotropic properties that not only slow the atherosclerosis progression but also induce positive changes in the coronary microvasculature. Such mechanisms include reducing inflammation and oxidative stress, silencing profibrotic pathways, and improving endothelial function, which contribute altogether to vascular endothelial health .The effect of statins on ED was suggested by early studies which showed increases in brachial FMD after statin treatment . A potenExperimental and observational studies suggested a potential antiarrhythmic effect of statins, demonstrating that these drugs protect against AF . PossiblInterestingly, the potential benefit of statins on both CMD and ACM, which are pathologies frequently associated with HFpEF, as part of the systemic ED, might explain the beneficial effect of statins on mortality observed in HFpEF patients without CAD, which is not translated in the population of patients with heart failure with reduced ejection fraction (HFrEF) ,139.ED is strongly linked to RAAS activation, which causes vasoconstriction, endothelial cell proliferation, and vascular oxidative stress. Indeed, ample evidence suggests that RAAS blockers can improve ED. A meta-analysis showed that treatment with ACE-Is significantly improved brachial FMD versus placebo or no treatment. Additionally, ACE-Is were more efficacious in improving brachial FMD when compared with other antihypertensive therapies, and there was no significant difference between ACE-I and ARB effect on brachial FMD . SimilarACE-Is are demonstrated to impact coronary microvascular function, most probably by various mechanistic pathways, such as improved systemic hemodynamics and coronary endothelial function, and blockade of the local deleterious effects of angiotensin II. The potential role of ACE-Is in the management of CMD was illustrated in the WISE study, which included women with INOCA and CMD defined by a CFR < 3, randomized to receive quinapril or placebo for 16 weeks. CFR was assessed at baseline and after completion of treatment with invasive intracoronary Doppler measurements, before and after adenosine infusion. The results showed that CFR improvement was higher in the ACE-Is group and that the benefit was greater in patients with lower baseline CFR (≤2.5). In addition, women receiving ACE-Is had significantly improved anginal symptoms compared to placebo . The samRAAS activation has a major contribution to the development of ACM by activating proinflammatory and profibrotic pathways ,145. ExpSacubitril/valsartan is the first drug in the class of the angiotensin receptor–neprilysin inhibitors (ARNI). Its mechanisms of action include both blocking RAAS activation with valsartan, an ARB, and preserving the natriuretic peptides (NPs) by inhibiting their degradation with sacubitril, a neprilysin inhibitor. The NP system includes hormones such as atrial NP (ANP) and B-type NP (BNP), which are secreted by cardiomyocytes in response to myocardial wall stretch, as well as C-type NP (CNP), which is mainly produced by endothelial cells . ANP andRecent data suggest that endothelial CNP contributes to the modulation of distal arteriolar and capillary blood flow by mediating the cross talk between endothelial cells and pericytes . In addiWhile there are no data regarding sacubitril/valsartan effects on CMD, we could assume that it may potentially improve it given its mechanisms of action, which include blocking the deleterious vascular effects of angiotensin II with valsartan and preserving the vascular and endothelial benefits promoted by NPs with sacubitril. A study evaluating this hypothesis could provide interesting results.Apart from promoting positive ventricular remodeling which translates into improved outcomes in patients with HF, emerging evidence suggests that ARNI could be beneficial in ACM as well. Through their antihypertrophic and antifibrotic effects, NPs prevent atrial structural remodeling and mechanical dysfunction. By counteracting atrial fibrosis, NPs contribute to normal electrical impulse generation and conduction, indirectly exerting an antiarrhythmic effect . MoreoveBeyond their metabolic benefits, SGLT2-is determine a myriad of pleiotropic effects such as an increase in hematocrit, lower values of uric acid and urine albumin: creatinine ratio, improvement in the tubuloglomerular function, and a reduction of inflammation and oxidative stress ,162,163.Several studies tested the effectiveness of SGLT2-is on peripheral ED. In the DEFENCE study, dapagliflozin on top of metformin led to a significant improvement of FMD in type 2 DM patients compared to single therapy with metformin. Moreover, dapagliflozin reduced oxidative stress, as suggested by lower urinary levels of oxidative stress biomarkers . Data frThere is little data specifically describing the effect of SGLT2-is on coronary microvasculature. A recent study analyzed the effects of empagliflozin by evaluating heart biopsies from HFpEF patients as well as from rats that were treated in vivo . IncreasExtending the effects of SGLT2-is, it appears that these drugs could potentially target the pathogenic atrial substrate from ACM as illustrated by several experimental studies. In line with this, a study using a diabetic rat model demonstrated that empagliflozin has incremental positive effects on left atrium structure and function. Specifically, empagliflozin administration in diabetic rats was associated with significantly smaller LA diameters and with attenuated LA interstitial fibrosis and atrial myocyte hypertrophy; moreover, it prevented DM-induced atrial electrical remodeling and AF inducibility, and it improved the mitochondrial function . In anotGLP-1RAs demonstrated conflicting results regarding the potential benefit on systemic ED. While improving endothelial function markers in experimental research, liraglutide did not significantly increase endothelium-dependent vasodilation assessed by measuring forearm blood flow using venous occlusion plethysmography after 12 weeks in a cohort of diabetic patients without other cardiovascular diseases . SimilarGLP-1RAs are demonstrated to reduce the rates of cardiovascular mortality and myocardial infarction, and improvements in CMD might be a potential mechanism . In an eSince GLP-1RAs demonstrated to reduce EAT volume and its metabolic activity, we could assume that they ameliorate periatrial inflammation with a positive effect on atrial arrhythmias . An expeOf particular importance for the clinician is the clinical recognition of the profile of the patient with multiple comorbidities, INOCA, and/or manifestations of ACM as it could provide a more accurate and comprehensive estimation of the patient’s prognosis and risk of future cardiovascular events. Both CMD and ACM, most frequently manifesting as AF, may be part of the same underlying disease process represented by endothelial dysfunction and may potentiate each other. Future studies are needed to further evaluate these associations and to systematically investigate the causality relationship between CMD and AF. Controlling cardiovascular risk factors which cause endothelial dysfunction improves these conditions, and their particular contribution to the long-term prognostic benefit remains to be established in future studies. Endothelial dysfunction emerges as a new therapeutic target, and further research is needed to assess to what extent reversing this common pathophysiological mechanism could indeed improve CMD and ACM manifestations. Certain pharmacotherapies such as statins and ACE-Is have been shown to positively influence CMD and ACM. ARNI, SGLT2-is, and GLP-1RAs are newer therapies which may have the potential to improve coronary and atrial endothelial dysfunction as well. Further studies are needed to clarify whether these drugs act on a common pathogenic substrate and whether their administration translates into long-term positive outcomes. This concept merits indeed further exploration as it could provide a new understanding of the treatment of these pathogenies, centered on targeting their common underlying pathogenic process. In the same line, of major significance would be to evaluate whether administration of these therapies in patients with a high comorbidity burden and endothelial dysfunction but without clinically manifesting CMD or ACM could potentially prevent their development, thus decreasing related morbidity and improving patient’s prognosis.CMD causing INOCA and ACM share similar risk factors and are frequent encounters in patients with a high comorbidity burden. CMD and AF are often found in association, and one may promote the other. Both pathologies can lead to HFpEF, and endothelial dysfunction is a common mechanistic pathway. Targeting endothelial dysfunction emerges as a new therapeutic strategy that could improve cardiovascular morbidity. Nonpharmacological measures such as weight loss and CPAP therapy for OSA ameliorate systemic endothelial dysfunction and can reduce atrial and vascular adverse remodeling. Targeting epicardial adipose tissue could reduce periatrial and coronary microvasculature inflammation and dysfunction. Certain therapies such as statins, ACE-Is, ARNI, SGLT2-is, and GLP-1RAs demonstrated a potential beneficial effect on endothelial dysfunction, reversing this pathogenic substrate and improving CMD and ACM manifestations.


As the single cell lining of the heart and all blood vessels, the vascular endothelium serves a critical role in maintaining homeostasis via control of vascular tone, immune cell recruitment, and macromolecular transit. For victims of acute high-dose radiation exposure, damage to the vascular endothelium may exacerbate the pathogenesis of acute and delayed multi-organ radiation toxicities. While commonalities exist between radiation-induced endothelial dysfunction in radiosensitive organs, the vascular endothelium is known to be highly heterogeneous as it is required to serve tissue and organ specific roles. In keeping with its organ and tissue specific functionality, the molecular and cellular response of the endothelium to radiation injury varies by organ. Therefore, in the development of medical countermeasures for multi-organ injury, it is necessary to consider organ and tissue-specific endothelial responses to both injury and candidate mitigators. The purpose of this review is to summarize the pathogenesis of endothelial dysfunction following total or near total body irradiation exposure at the level of individual radiosensitive organs. Victims of whole or near whole body radiation exposure will experience multi-organ acute and late toxicities. This review focuses on the potential role of the vascular endothelium in the progression of organ specific sequelae following the type of high dose exposure that may be likely to occur in a catastrophic nuclear or radiological event . While iAt the cellular level, ionizing radiation is known to induce immediate endothelial damage characterized by apoptotic cell death, the release of pro-inflammatory cytokines, expression of cellular adhesion molecules, and loss of endothelial barrier function ,7. Data In general, ionizing radiation induces direct cellular injury via three mechanisms: induction of DNA damage, damage to the plasma membrane, and generation of reactive oxygen species (ROS) as illustrated in In endothelial cells, radiation-induced endothelial p53 activation typically promotes p21-mediated cell cycle arrest and the development of a senescent cell phenotype . AlthougIn addition to DNA damage, ionizing radiation causes direct damage to the cell membrane causes direct damage to the cellular membrane by lipid peroxidation and fragmentation. Radiation induces hydrolysis of the membrane lipid sphingomyelin to ceramide by the membrane bound lipid enzyme acidic sphingomyelinase (ASMase) ,25. CeraFinally, radiation injury is known to induce the generation of ROS. Although ROS are generated within seconds following radiation injury , only a Endothelial cells which escape both radiation-induced senescence or apoptotic cell death, may have long-term functional impairment as illustrated in The expression of cell adhesion molecules on the endothelium is a key step in the initiation and progression of inflammatory cardiovascular diseases as these adhesion molecules promote leukocyte adhesion and transendothelial migration of neutrophils . Recent Another important characteristic of functional impairment in the irradiated endothelium is a loss of endothelial barrier function. As will be discussed later, endothelial permeability varies in an organic-specific manner. In general, the endothelium serves as a selective barrier which regulates the transport of fluids, solutes, macromolecules, and cells from the blood to the tissue space. Barrier integrity is maintained by endothelial cell-specific junctional proteins of the adherens, tight, and gap junctions . Several2-) and peroxynitrite (ONOO−) instead of the vasodilator nitric oxide (NO). Superoxide dismutase (SOD) mimetics which act to scavenge ROS have shown promise in mitigation of lung and intestinal radiation injury [Finally, dysfunction of the endothelium is generally associated with impaired endothelial nitric oxide synthase (eNOS) activity. Here, “uncoupled” eNOS favors production of the ROS super oxide  microenvironment which includes the endothelial cells (ECs) commonly divided into two vascular niches: the arteriolar and sinusoidal vessels. The arteriolar and sinusoidal endothelial cells defining these distinct vascular niches  are knowPhysically, the sinusoidal endothelium forms a network of permeable capillaries which serve to traffic nutrients and hematopoietic cells to and from the BM microenvironment . Early tBone marrow endothelial cells (BMECs) play a key role in restoring hematopoiesis following radiation injury in part by secretion of hematopoietic cytokines such as G-CSF, EGF, pleiotrophin, jagged-1, CCL5 and E-selection ,61,62,63Collectively, these studies emphasize the fundamental significance of the endothelium in regulating hematopoietic regeneration following radiation injury. Accordingly, several studies have evaluated whether therapeutic delivery of endothelial cells can improve hematopoietic regeneration after injury. Multiple groups have demonstrated that infusion of endothelial cells or endothelial cell-derived extracellular vesicles (EVs) can accelerate hematopoietic recovery and rescues lethally irradiated mice from death due to hematopoietic syndrome ,67,68,69In addition to endothelial cellular therapies, therapies targeting the endogenous bone marrow endothelium have also been evaluated as potential medical countermeasures for hematopoietic acute radiation syndrome. Notably, activated protein C (APC) is a factor which counters radiation-induced loss of endothelial thrombomodulin to reverse pro-thrombotic and pro-fibrogenic remodeling of the endothelium . Among oPersons exposed to whole body doses as low as 1.5 Gy may experience radiation injury to the gastrointestinal (GI) tract as evidenced by early symptoms of nausea, vomiting, and anorexia. Without treatment, doses above 5 Gy may result in GI death due to denudation of the intestinal mucosal barrier and subsequent fluid loss, hemorrhage, and/or sepsis ,76. The In the setting of acute intestinal injury, it has been debated whether GI toxicity results from initiation of apoptotic death in the endothelium or the intestinal crypt ,79. WhilSurvivors of high-dose acute radiation exposure are at increased risk for developing a range of late multi-organ morbidities which are collectively referred to as the delayed effects of acute radiation exposure (DEARE). Sustained vascular dysfunction is a common trait which has been observed in multiple late radiation-sensitive tissues including the lung ,83,84,85In addition to life-threatening acute GI syndrome, survivors of acute radiation exposure may experience radiation enteropathy a late-radiation toxicity characterized by mucosal loss, fibrosis, and sustained vascular dysfunction . The mecRadiation-induced lung injury (RILI) can occur in two phases: an acute inflammatory stage (pneumonitis) 8–16 weeks following exposure and a late fibrotic injury which can take months to years to evolve . These aRadiation pneumonitis is primarily an inflammatory condition characterized by pulmonary edema, immune cell infiltration, and clinical symptoms of labored breathing/cough. Animal models have been instructive in characterizing the acute vascular changes during pneumonitis. At one month following 10 Gy thoracic irradiation, marked changes in the pulmonary vasculature have been observed including a loss of the pulmonary vessels and increased vascular resistance . ConcurrPathologic alterations in vascular permeability during pneumonitis are attributable in part to alteration of lung endothelial sphingolipids . SimilarSchultz-Hector and Trott provided a comprehensive review of the manifestation of cardiovascular diseases in both clinical cohorts exposed to heart irradiation and epidemiologic analysis of the Life Span Study of Japanese atomic bomb survivors . This reSurvivors of acute radiation syndrome are at increased risk for developing late kidney injury or radiation nephropathy. In a cohort analysis of Japanese atomic bomb survivors, radiation dose was significantly associated with chronic kidney disease . The gloRadiation-induced EC injury is a major mediator and regulator of wider tissue damage in the different vascular beds. The data reviewed here highlight the role of the highly heterogeneous endothelium in the progression of multi-organ injury. While the endothelium of radiosensitive organs is highly specialized, there may be common mechanisms which can be targeted to systemically promote normalization of the endothelium post radiation injury. Several aforementioned therapies (summarized in