Jan Hontelez and co-authors discuss the use of different types of evidence to inform HIV program integration. Sustainable Development Goal 3 aims to “ensure healthy lives and promote well-being for all at all ages” and has set a target of achieving global universal health coverage, representing a major policy shift away from mostly disease-specific “vertical programmes”.While health service integration can be a promising strategy to improve healthcare coverage, health outcomes, and efficiency, the exact impact of integration in different settings is hard to predict, and policy makers need to choose from a large variety of integration strategies and opportunities with varying levels of scientific evidence.Using the case of health service integration for HIV in low- and middle-income countries, we outline implementation strategies for integration opportunities with lacking or scarce high-level causal evidence, based on existing frameworks and methodologies from within and beyond healthcare and implementation science.Proper use of scientific evidence in other contexts requires adequate and systematic assessments of the transportability of an intervention. Several methods exist that allow for judging transferability and comprehensively identifying key context-specific indicators across studies that can affect the reported impact of interventions.When (transferable) evidence is absent, we propose that by drawing on well-established design and implementation methodologies—underpinned by ongoing learning and iterative improvement of local service delivery strategies—countries could substantially improve decision-making even in the absence of scientific evidence.Reaching the goal of making the HIV response an integral part of a larger, universal, people-centred health system that meets the needs and requirements of citizens can be facilitated by applying lessons learned from implementation science and novel design methodologies.The Sustainable Development Goal (SDG) 3 aims to “ensure healthy lives and promote well-being for all at all ages” ,2 and haPolicy makers now face the daunting challenge of proposing, adapting, and implementing healthcare interventions tailored to specific contexts and population needs, often with limited high-quality scientific evidence on the impact of candidate interventions. Although evidence-based medicine remains the gold standard for decision-making in healthcare , randomiA recent comprehensive systematic review that was undertaken as part of the UNAIDS 2021–2025 HIV/AIDS target estimations and resource needs exercise, analysed and synthesised the existing evidence on the impact of integration of HIV services with other health services . The finSuitability of evidence for decision-making depends on both the quality of the evidence and the usefulness for a new context: Results from experiments in one context may or may not be useful for policy decisions in another context. As a context is defined by different determinants, which can be measured and accounted for to some extend , in theTable 1). First of all, transferability should be judged by local policymakers, experts, and other stakeholders. How does the context in which the evidence for a certain intervention is generated differ from the context in which a policy decision need to be made, for instance, in terms of epidemiology and disease burden, target population, financial and human resources, health services in place, and targets for control? Mehrotra and colleagues (2019) propose a “transportability framework” to understand HIV programme effects in different contexts to enable judgement of external validity of interventions and translate this into policy action and testable insight [Different methods could be applied for judging overall transferability of the available evidence .Transporting and adapting context-specific findings certainly enhances the usefulness of context-specific effect studies. Many promising integration opportunities, however, have limited to no quantitative scientific evidence on impact. This is especially true for key populations in the HIV response, such as transgender people, migrants, men who have sex with men, and sex workers; for particular geographic regions, such as Latin America, Southeast Asia, and the Russian Federation; and for specific disease-based opportunities for integration, such as cervical cancer, mental health, or schistosomiasis. By drawing on well-established design and implementation methodologies—underpinned by ongoing learning and iterative improvement of local service delivery strategies—countries could substantially improve decision-making even in the absence of scientific evidence (The scope of claims that require evidence should be judiciously made. Cartwright and Stegenga (2013) propose to approach the issue from the policy makers’ perspective and work backwards; assessing what the likelihood is that a proposed policy will be effective in achieving a predefined target and what type of evidence would be relevant for the evaluation ,23.There are many examples in which integration opportunities are straightforward, while the risks are likely negligible, allowing for implementation even without a solid evidence base. For instance, the integration of health services is often a sensible choice when the services are targeted at the same end users, like integrating HIV services and gender-affirming therapies for transgender men and women  or integWhen scientific evidence is lacking, but the intervention of interest is already implemented in a given or a comparable context, expert evidence surveys can be administered to local experts to gather information about their unpublished observations and case series as a basis for developing recommendations that are as evidence-based as possible at a particular point in time .When scientific evidence is lacking and comparable interventions have yet to be implemented, implementation and testing of understudied integration opportunities can be aided by lessons learned in other sectors, such as business or software design. Applying systems-thinking processes to integrating the HIV response, for instance, by framing problem definitions and applying context analyses, ideation, creative thinking, prototyping, testing, and evaluation, could ensure that the right solutions to context-specific problems are generated and iteratively refined . SimilarFor people-centred approaches to health systems integration to be successful, deliberative inclusive processes —in whichWhile redesigning the HIV response to meet the changing contextual needs, we should also ensure that we accelerate the sharing of knowledge within the public domain. Approaches, such as “A/B testing” ,34, i.e.Health policy makers and funders are tasked with the daunting challenge of redesigning the HIV response to meet the current complex epidemiological, financial, and political challenges. By applying lessons learned and methods from a range of scientific fields—such as innovation models, systems dynamics models, implementation science, design research, and learning systems—policy makers, funders, and practitioners could help accelerate and maximise the transfer of limited, and often context-specific, knowledge on best practices for developing integrated HIV services. Furthermore, by expanding the evidence with results from widely applied methods that are not randomised controlled trials, policy makers could help ensure that the HIV response is an integral part of a larger, universal, people-centred health system that meets the needs and requirements of citizens. Such a shift in decision-making design could prove to be key in shaping the HIV response within the UHC agenda in rapidly changing contexts.


Evidence suggests social isolation and loneliness are prevalent within the population and may potentially be exacerbated due to the pandemic. Social connections have powerful influences on health and longevity, and lacking social connection qualifies as a risk factor for premature mortality. Evidence from the recent National Academy of Science consensus report on social isolation and loneliness among older adults will be summarized, providing the scope of the health effects, potential mechanisms and risk factors, as well as current gaps in the evidence. Importantly, this evidence points to several implications for solutions across sectors, including medical and healthcare practice and policy.


Whilst access to cannabis-based medicinal products (CBMPs) has increased globally subject to relaxation of scheduling laws globally, one of the main barriers to appropriate patient access remains a paucity of high-quality evidence surrounding their clinical effects.Whilst randomised controlled trials (RCTs) remain the gold-standard for clinical evaluation, there are notable barriers to their implementation. Development of CBMPs requires novel approaches of evidence collection to address these challenges. Real world evidence (RWE) presents a solution to not only both provide immediate impact on clinical care, but also inform well-conducted RCTs. RWE is defined as evidence derived from health data sourced from non-interventional studies, registries, electronic health records and insurance data. Currently it is used mostly to monitor post-approval safety requirements allowing for long-term pharmacovigilance. However, RWE has the potential to be used in conjunction or as an extension to RCTs to both broaden and streamline the process of evidence generation.Novel approaches of data collection and analysis will be integral to improving clinical evidence on CBMPs. RWE can be used in conjunction or as an extension to RCTs to increase the speed of evidence generation, as well as reduce costs. Currently, there is an abundance of potential data however, whilst a number of platforms now exist to capture real world data it is important the right tools and analysis are utilised to unlock potential insights from these. There an humans , 3. Limin humans . There in humans . The quan humans .The National Institute for Health and Care Excellence in the UK has only recommended licensed CBMPs for a limited range of indications . ChangesRandomised controlled trials (RCTs) are necessary and should continue to be the standard against which medical evidence is upheld. However, they are expensive, time consuming and subject to their own limitations . Whilst RCTs are not infallible—they are expensive and time consuming. Globally $100 billion USD is spent on biomedical research . In the 9-tetrahydrocannabinol (THC) there are over 140 cannabinoids, as well as flavonoids, terpenes, and other compounds within the flower of different cannabis plants [In addition to cannabidiol (CBD) and (−)-trans-Δs plants . These cs plants . The cons plants , 13.The route of administration further affects the pharmacokinetics of CBMPs and the associated outcome of any trial. CBMPs can be administered sublingually, trans-dermally, via inhalation, or orally . This suAn appropriately blinded assessment against placebo or active therapy is the optimal design for RCTs. It has been difficult to identify a placebo that cannot be distinguished against an active CBMP according to absence of both vasoactive and psychoactive effects, as well as the typical aroma associated with cannabis . This prProduction methods and import costs mean that CBMPs are typically expensive, adding further to high research costs . ResearcRCTs are possible with CBMPs; however, the above issues present legitimate challenges. In many chronic diseases there is a need for novel therapeutics and CBMPs are therefore being utilised based on best available evidence. Due to the challenges in developing CBMPs through a traditional drug development pipeline, the exploration of its utility should not be limited to traditional methods. It is important that we capture a suite of real-world evidence (RWE) to inform prescribing guidelines, regulations, and clinical trials. By leaning on RWE there is an opportunity to improve the quality and design of RCTs and clinical evidence in general, via a top-down approach .evidence derived from health data sourced from non-interventional studies, registries, electronic health records and insurance data as opposed to the highly controlled setting of RCTs [RWE is defined as  of RCTs . There i of RCTs . It is c of RCTs . There i of RCTs .Consistent use of RWE to aid regulatory decision making is yet to be normalised, but the promise is apparent . RecentlNHS England and NHS improvement published a review on the barriers to accessing CBMPs in the UK . Their rRWE is already being incorporated into the scientific literature on cannabis Table . Early eBetween these study designs it is important to be aware of potential divergence in reported outcomes. RWE has broader inclusion criteria, accounting for factors like non-standard dosing, and is not limited by scope of disease, thereby improving ecological validity . HoweverRWE can bring further clarity on questions that remain unanswered in RCTs. A recent study utilised anonymised surveys of patients with fibromyalgia who consumed cannabis flower . In addiRWE can improve the efficiency of clinical trials by generating hypotheses, refining eligibility criteria, and exploring drug development tools. Registries can be used to form an infrastructure to conduct a clinical trial, lowering costs whilst maintaining high evidence quality . In suppRWE, however, does have limits to its utility. There is variation in the quality and provenance of the data stored in electronic medical records . FurtherAnother limitation is the lack of randomisation, controlled variables and internal validity. This can make it more difficult to derive causative mechanisms behind clinical outcomes. However, this is also one of the strengths of these studies, allowing for generalisability to true clinical practice . TreatmeCBMPs are a complex range of pharmaceuticals which pose challenges to traditional pathways of drug development and translation. Development of CBMPs requires novel approaches of evidence collection to address these challenges. RWE can be used in conjunction or as an extension to RCTs to both broaden and streamline the process of evidence generation. Currently, there is an abundance of potential data, however, it is important the right tools and analysis are utilised to unlock potential insights from these.


Late inflammatory reactions (LIRs) are the most challenging complications after filler use. The immune system plays a prominent role in its etiology, albeit to an unknown extent. Bacterial contamination in situ has been hypothesized to be causative for LIRs. How this relates to the immunological processes involved is unknown. This article aims to provide an overview of immunological and bacterial factors involved in development of LIRs.We undertook a systematic literature review focused on immunological factors and microbiota in relation to LIRs after filler use. This systematic review was performed in accordance with the PRISMA guidelines. PubMed, EMBASE and the Cochrane databases were searched from inception up to August 2019. Included studies were assessed for the following variables: subject characteristics, number of patients, primary indication for filler injection, implant type/amount and injection site, type of complication, follow-up or injection duration, study methods, type of antibiotics or medical therapies and outcomes related to microbiota and immunological factors.Staphylococcus epidermidis within biopsies were often associated with LIRs.Data on immunological factors and bacterial contamination were retrieved from 21 included studies. Notably, the presence of histocytes, giant cells and This review provides a clear overview of the immunological factors associated with LIRs and provides a hypothetical immunological model for development of the disease. Furthermore, an overview of bacterial contamination and associations with LIRs has been provided. Follow-up research may result in clinical recommendations to prevent LIRs.www.springer.com/00266..This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors-The online version contains supplementary material available at 10.1007/s00266-021-02306-3. Fillers, also referred to as dermal fillers, soft tissue fillers or dermal implants have been used for decades, mainly for cosmetic reasons, but also to improve aesthetic outcome after trauma, cancer or malformations . They arAlthough manufacturers and several studies claim that adverse events after dermal fillers are very uncommon, unwanted adverse events do occur with all fillers products Fig. a, b 6–1–16–12. AA foreign body reaction is a natural tissue response after soft tissue filler injection . The immBacterial contamination of the filler material has frequently been proposed as a likely cause of LIRs and some indications for this theory exist , 8, 28. A systematic review was performed of the literature pertaining to immunological factors and bacterial contamination/infection of LIRs after soft tissue filler use. This systematic review was carried out in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Statement (PRISMA) guidelines . An exteWe searched for original studies on LIRs after soft tissue filler use which reported on microbiota and/or immunological factors. All patient studies as well as relevant animal studies were included. Of these studies, all types of filler products were included, as well as all sites of injection. In vitro studies and studies which investigated the normal immune response of filler products were excluded. Case reports less than 5 patients and isolated abstracts were excluded as well as non-original studies such as reviews, editorials, communications, correspondence, discussions and letters.The search was executed in cooperation with a medical information specialist. The article selection was performed by two reviewers in two steps: in the first step eligibility of the articles was screened based on the title. Subsequently, the abstracts of the selected articles were evaluated, and, in case of doubt, the full article was reviewed. Articles which met the inclusion criteria were included in the systematic review.Data were extracted using standardized tables developed for this purpose. Data included the following variables: authors, year of publication, study design, number of patients or samples, type/amount and location sites of injected filler, included group of subjects, primary indication for injection, type of complication, follow-up or injection duration, microbial/immunological methodologies, type of antibiotics/medical therapies, main outcomes and the conclusion. Included studies were assigned a level of evidence according to the Oxford Centre for Evidence-Based Medicine .Due to heterogeneity of the studies, statistical meta-analysis of the data is impossible. Instead, we performed qualitative and descriptive analyses of the outcomes.The literature search produced 2340 articles after removal of duplicates. Of these, 487 remained after title screening, and 197 after evaluation of the abstracts. Full text review yielded 21 studies which met the inclusion criteria  have investigated the presence of bacteria on samples from LIRs after filler use , 31–33. We found 14 studies with regard to a variety of immunological factors in relation to LIRs , 32–35. The exact aetiopathology of LIRs caused by filler is still poorly understood. Several authors have suggested that delayed-onset complications can occur in response to local or systemic infections,  injuries, systemic medication or vaccinations, or invasive treatments  in the vicinity of the filler deposits , 36–44.Staphylococcus spp. (mainly Staphylococcus epidermidis) and Propionibacterium acnes, which were detected in up to 98% of positive cultures, and which lead to the hypothesis that the presence of these bacteria is involved in development of LIRs [Numerous human studies have shown the presence of bacteria in material derived from patients with LIRs Table 7 in ESM) [ in ESM , 31, 32. of LIRs , 31, 32. of LIRs .In a human study by Christensen et al., they showed that bacterial presence varies among different filler compounds, with an increased contamination within the permanent filler material . This fiDespite this evidence for the presence of bacteria around filler material and support for a primary role in the development of LIRs, some authors postulate that these findings are not enough to conclude LIRs have an infectious aetiopathology . They stMultiple studies indicate that extrinsic factors such as (micro-) trauma, infections and drugs or vaccinations seem to be capable of activating certain mechanisms leading to delayed-onset complications such as (low-grade) inflammatory nodules and abscess formation Fig. . AlthougThe generation of a granulomatous foreign body reaction (GFBR) is a ‘normal,’ physiological response from the host to any foreign body. All filler agents used for soft tissue augmentation are thought to elicit some degree of granulomatous inflammatory reaction following injection , 51. To Current insights differentiate several steps in host reaction on biomaterial implantation. Within minutes after implantation, host plasma components such as albumin, fibrinogen, fibronectin, lipids, sugars, ions and platelets are absorbed on the surface of the implant , 22, 53.The release of immune-regulatory signals by neutrophils attracts monocytes, macrophages, immature DCs and lymphocytes, promotes monocytes differentiation into ‘M1’-type macrophages secreting pro-inflammatory cytokines (Interleukin (IL-) 1β, IL-6, necrosis factor alpha (TNF-α)) and downregulates the presence and activity of neutrophils themselves. The next step in the healing process is a shift toward ‘M2’-type macrophages secreting anti-inflammatory cytokines  and recruiting fibroblasts for effective tissue regeneration. The shift toward an anti-inflammatory ‘M2’-type healing response can be altered by specific characteristics of the implanted material. For example, particles larger than 5 μm require the presence of aggregated macrophages, or giant cells, to be phagocytosed –58. PartNumerous studies have been performed on the different structural properties of filler, such as chemical composition, electrical charge, surface irregularities or particle size and the presence of contaminants, which are of known influence on host immune responses , 50 Fig, Table 3The relation with time can also be of importance. Most studies show a decreased inflammatory reaction after several months; however, this also depends on the type of filler used . WhereasImmune-mediated hypersensitivity to fillers in patients with LIRs has been investigated by one study. Micheels et al. showed that skin tests in patients with adverse events were positive for one or the other or both of the injectable hyaluronic acid preparations . Also, sh1 inflammatory cytokines [Alijotas-Reig recently postulated that fillers may act as adjuvants, rather than as antigens . Adjuvanytokines . They alytokines , 64. Cerytokines , 63–66. ytokines , 66. Sevytokines , 68  , 33, 45.Also an antimicrobial approach for the treatment of LIRs has been widely addressed (Table 8b in ESM). The hypothesis stating bacteria in situ are involved in the development of adverse events is further investigated by Marusza et al. who treated patients with an antibiotic scheme of whom all experienced resolution of symptoms and with no recurrence of their complication . HoweverAlthough a lot has been written on the best treatment approach for LIRs, to date no evidence-based effective therapy for LIRs exists. To achieve this, first the aetiopathological uncertainties most be solved, keeping in mind that maybe not all LIRs have the same aetiopathology or cause. Also, although bacterial presence might not necessarily prove an infectious process, bacterial contamination of the filler material might act as stimulus for other (innate) immune response. This is why many practitioners use and will keep using immune modulating antibiotics as first line therapy in case of HA-induced LIRs. This way treats both an infectious and an inflammatory reaction.The best approach, however, remains prevention. For this reason, possible causes or triggers for an infectious or inflammatory process should be avoided. Filler treatments should therefore be performed in under aseptic conditions by well-trained physicians with knowledge of skin preparation, facial anatomy and potential complications. Like described by De Boulle and Heydenrych, special considerations are necessary for patients with preexisting autoimmune or -inflammatory diseases .It must also be empathized that  permanent fillers have been proven to have a significantly higher risk for adverse events such as bacterial contamination, inflammatory reactions, abscesses and migration up to decades after injection. Their use should therefore be discouraged, especially for aesthetic indications.As previously mentioned, dermal fillers can be used for cosmetic but also for medical reasons. For both indications, there might be a loss of elasticity and collagen in the dermis, volume loss of the subcutaneous fat and bone structures. As a result of this, injection of fillers needs a three-dimensional approach. The injection of fillers is therefore divided into three injection levels: deep dermal, subdermal and supraperiosteal . AlthougBased on this review, we propose the following treatment algorithm for LIRs Fig. . As statS. epidermidis is the bacterium most often found in LIRs although its role is still debated . Several substances, such as antibiotics and corticosteroids, seem to be effective in the treatment of LIRs, although the mechanism is not fully understood, and more research is needed.This review has analyzed the aetiopathological hypotheses of LIRs that can currently be found in the literature. A major role is seen for a  foreign body reaction led by activated histocytes and giant cells, eventually resulting in a chronic inflammation. Supplementary file1 (DOCX 32 KB)Below is the link to the electronic supplementary material.


Clarity regarding accuracy and effectiveness for interventional procedures around the foot and ankle is lacking. Consequently, a board of 53 members of the Ultrasound and Interventional Subcommittees of the European Society of Musculoskeletal Radiology (ESSR) reviewed the published literature to evaluate the evidence on image-guided musculoskeletal interventional procedures around this anatomical region.We report the results of a Delphi-based consensus of 53 experts from the European Society of Musculoskeletal Radiology who reviewed the published literature for evidence on image-guided interventional procedures offered around foot and ankle in order to derive their clinical indications. Experts drafted a list of statements and graded them according to the Oxford Centre for evidence-based medicine levels of evidence. Consensus was considered strong when > 95% of experts agreed with the statement or broad when > 80% but < 95% agreed. The results of the Delphi-based consensus were used to write the paper that was shared with all panel members for final approval.A list of 16 evidence-based statements on clinical indications for image-guided musculoskeletal interventional procedures in the foot and ankle were drafted after a literature review. The highest level of evidence was reported for four statements, all receiving 100% agreement.According to this consensus, image-guided interventions should not be considered a first-level approach for treating Achilles tendinopathy, while ultrasonography guidance is strongly recommended to improve the efficacy of interventional procedures for plantar fasciitis and Morton’s neuroma, particularly using platelet-rich plasma and corticosteroids, respectively.The expert panel of the ESSR listed 16 evidence-based statements on clinical indications of image-guided musculoskeletal interventional procedures in the foot and ankle.• • Strong consensus was obtained for all statements.• The highest level of evidence was reached by four statements concerning the effectiveness of US-guided injections of corticosteroid for Morton’s neuroma and PRP for plantar fasciitis.The online version contains supplementary material available at 10.1007/s00330-021-08125-z. Eur Radiol). We report the evidence-based statements for clinical indications of image-guided musculoskeletal interventional procedures around the foot and ankle listed by an expert board of the ESSR after a Delphi method performed a review of current literature.Musculoskeletal interventional procedures are routinely performed around the foot and ankle, including injections and variable interventions on tendons, bursae, plantar fascia, nerves, and joints –3. Some Institutional Review Board approval was not needed as no patients were directly involved. This paper arises from the review of published evidence on image-guided interventional musculoskeletal procedures around the foot and ankle. Similar to previous ESSR consensus papers –16, a liImage-guided injections are safe and feasible to treat Achilles degenerative tendinopathy (AT), but there is insufficient evidence from randomized controlled trials (RCT) to support them over conservative therapies.Level of evidence: 1n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, Both the Cochrane Review of 2015  and Maff2.Image-guided anesthetic-corticosteroid injections into the anterior and posterior tibial tendon sheaths are safe and may provide effective diagnosis and treatment in patients with anterior and posterior tibial tendon tenosynovitis.High-volume injections seem to be effective at treating AT, but confounding factors such as corticosteroid, aprotinin, dry needling, or eccentric exercise prevent assessment of this method alone , 35.2.ILevel of evidence: 3n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 3.US-guided corticosteroid injections are more effective than palpation-guided injections to treat plantar fasciitis (PF) providing significant short-term pain relief, particularly when combined with strength training and stretching.Several studies reported complete or near-complete prolonged symptom relief after anesthetic-corticosteroid injections for tenosynovitis unresponsive to conservative management –40. A stLevel of evidence: 2n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 4.US-guided PRP injections for PF are safe and provide significant pain relief in chronic PF, with better clinical outcome at mid- and long-term follow-up if compared with corticosteroid injections.A meta-analysis included 5 RCTs with 149 patients and concluded that US-guided injections were superior with regard to VAS, tenderness threshold, response rate, plantar fascia thickness, and hypoechogenicity. There was no difference in heel pad thickness and heel tenderness index . A compaLevel of evidence: 1n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 5.The effectiveness of US-guided injections with ozone, hyaluronic acid, or botulinum toxin type A has not still sufficiently proven to be recommended for PF.Four meta-analysis studies having compared US-guided PRP and corticosteroid support the use of PRP instead of corticosteroids, highlighting the favorable and long-lasting clinical outcomes in patients with chronic PF , 49–51.Level of evidence: 3n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 6.US and fluoroscopy guidance improves the accuracy of joint injections in the foot and ankle although these procedures can be safely performed with palpation alone.Limited number of outcome-based investigation studies reported the effectiveness of those methods to treat painful PF without inducing fat pad atrophy –54. MostLevel of evidence: 4n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 7.Image-guided corticosteroid injections for midfoot joint osteoarthritis might provide short-term improvement, but further studies are warranted to support their clinical use.Intra-articular injections of the foot and ankle using palpation and US were conducted on a cadaver model and were both 100% accurate . The risLevel of evidence: 4n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 8.PRP and prolotherapy are safe methods to treat osteochondral lesions of the talus with promising results, but evidence on efficacy is limited.Two uncontrolled studies of intra-articular corticosteroid injections for midfoot osteoarthritis led to short-term improvement with poor long-term outcome , 60. SymLevel of evidence 4n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, n = 27 prolotherapy, n = 22 PRP) [9.Intraarticular foot and ankle anesthetic injections performed under imaging guidance offers precise information about pain source.PRP and prolotherapy are shown to be safe methods to treat talus osteochondral lesions and might be considered treatment options for younger patients and for patients with early stage and small lesions, as reported in a single retrospective cohort study including 49 patients ( 22 PRP) . Proloth 22 PRP) . HoweverLevel of evidence: 4n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 10.US-guided tarsal tunnel decompression is a feasible procedure in cadavers, but clinical value is unknown.Fluoroscopically guided foot and ankle joint anesthetic injections aid diagnosis and have a potential for targeted therapy when combined with corticosteroids. Large and small joints in the foot and ankle are readily accessed , 63. AneLevel of evidence: 4n = 52; disagree, n = 1; abstain, n = 0. Agreement = 98%Agree, 11.US-guided injections of bursae around the foot are technically feasible, but clinical value is unknown.US-guided tarsal tunnel decompression associated with flexor retinaculum release was feasible in a cadaveric study without any vascular or neurological damage . HoweverLevel of evidence: 5n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 12.US-guided deep retrocalcaneal bursa injections are feasible.Foot bursae aspiration and injection could be performed in a wide series of rheumatologic and degenerative conditions , but eviLevel of evidence: 4n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 13.US-guided corticosteroid injections are the most effective image-guided interventional procedure to improve pain in patients with Morton’s neuroma, especially in the first 3 months.Deep retrocalcaneal bursa injections are feasible and effective both under US  and fluoLevel of evidence: 1n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 14.US guidance improves the effectiveness of different interventional procedures for Morton’s neuroma if compared to palpation guidance, particularly for corticosteroid injection.A systematic review identified eight different types of interventions for Morton’s neuroma. The most frequent procedure was US-guided corticosteroid (7 studies) and sclerosing injections (7 studies). A meta-analysis of two RCTs conducted in the same study found that US-guided corticosteroid injections decreased pain more than controls. Other RCTs reported the efficacy of mobilization or extracorporeal shockwave therapy versus control. Treatment success was evaluated for extracorporeal shockwave therapy versus control and for corticosteroids versus footwear padding. Sclerosing and Botox injections, radiofrequency ablation, and cryoneurolysis had been investigated only by case studies with limited methodological quality . ResultsLevel of evidence: 1n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 15.US-guided ethanol injection of Morton’s neuroma is relatively safe and well-tolerated, but further investigations are required to clearly demonstrate its clinical value prior to supporting this procedure.When compared to palpation guidance, US was proven to increase the effectiveness of interventional procedures in patients with Morton’s neuroma. A systematic review  reportedLevel of evidence: 2n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, 16.Image-guided thermal ablation of Morton’s neuroma is safe with promising initial results and might reduce the need for surgery in the short term.A systematic review  found thLevel of evidence: 4n = 53; disagree, n = 0; abstain, n = 0. Agreement = 100%Agree, Case series reported the use of radiofrequency ablation as a safe and efficient potential treatment for Morton’s neuroma , 91. A cThis panel listed 16 evidence-based statements on clinical indications of image-guided musculoskeletal interventional procedures in the foot and ankle, with strong consensus reached for all statements. The highest level of evidence was reached by four statements concerning the effectiveness of US-guided injections of corticosteroid for Morton’s neuroma and PRP for PF, and also concerning the limited role of injection therapies in AT.Despite the safety and efficacy of image-guided injections for AT, the superiority of these procedures over conservative therapies has not been proven, leading us to consider them a second-level approach for treating patients with AT (statement #1).Regarding PF, prospective RCTs demonstrated the added value of US guidance to improve the clinical outcome of interventional procedures over blinded injections (statement #3). The highest level of evidence paper showed that US-guided PRP injections have better and long-lasting effects than US-guided corticosteroid injections in patients with PF (statement #4). In patients with Morton’s neuroma, the highest level of evidence papers showed that US guidance improves the efficacy of several interventional procedures (statement #14), particularly corticosteroid injections used to improve pain in the short-/mid-term (statement #13). Although ethanol injections, radiofrequency, and cryoablation seem to be safe alternatives for treating Morton’s neuroma, the clinical value of these interventions still needs further clarification (statements #15 and #16).Anecdotal reports and small case series have shown the feasibility, safety, and effectiveness of image-guided injections of ankle/foot joints (statements #6–9) and bursae (statements #11–12), with too limited evidence to support these in routine clinical practice.In conclusion, 16 statements about image-guided musculoskeletal interventional procedures around the foot and ankle have been produced by an expert panel of the ESSR. Image-guided interventions should not be considered a first-level approach for AT. US is strongly recommended as a guidance for interventions for PF and Morton’s neuroma to improve the effectiveness of the procedures, particularly using PRP and corticosteroids, respectively.ESM 1(DOCX 20 kb)


Scientific evidence related to environmental exposures continues to mount. Tools such as evidence mapping support decision making, but can be resource- and time-intensive. We explored “rapid evidence mapping” to efficiently map scientific evidence using rigorous and transparent methodologies. We undertook a proof-of-concept case study on the topic of low-calorie sweeteners. Our intent was to conduct a traditional evidence map based on the same evidence base from a prior rapid evidence map case study to compare approaches, findings, and conclusions. We searched the literature, screened full text of studies, manually tagged and categorized articles, and created visualizations to map the evidence.We conducted full-text screening of studies from the prior rapid evidence map and identified 255 relevant studies. Our findings corroborated those of the rapid evidence map, identifying most studies as short-term conducted in healthy individuals studying outcomes of appetite, energy sensing and body weight. We identified gaps in research areas related to outcomes of appetite and dietary intake, particularly in study populations with diabetes. Our findings illustrate the promise of rapid evidence mapping as a rigorous approach that can summarize scientific evidence, identify knowledge gaps, and identify areas for a future systematic review in a time-efficient manner.The online version contains supplementary material available at 10.1186/s13104-022-05926-3. A recurring challenge in the field of environmental health is for stakeholders to efficiently summarize the ever-expanding scientific evidence and come to defendable scientific consensus to support decision-making –3. ToolsWe previously developed “rapid Evidence Mapping” (rEM), a knowledge synthesis approach where the evidence mapping review process is simplified in a resource-efficient manner . We applHere, our main goal is to compare the rEM to traditional evidence mapping, by creating a traditional evidence map on the same evidence base used to generate the rEM and comparing the results as well as the time and resource commitments required for each.To create the traditional evidence map, we started with the results from an rEM of human exposures to LCS published by Lam et al.  and applLam et al.  searchedhttps://www.sciome.com/swift-review/). Software functionalities were used to automatically classify outcome, baseline health, and comparison categories [Lam et al.  importedtegories  from refWe started with all references included from Lam et al.  followinStudy design characteristics Study population characteristics Study interventions/exposures and comparisons Number of people analyzed, form of administrationOutcome information Aim or hypothesis and funding source of the studyThe following data were manually extracted from the full text of included studies:Data was extracted by a single reviewer , with an independent second reviewer reviewing and verifying the data. A pilot test of 10 studies was conducted to calibrate screeners and resolve any confusion interpreting directions. Extracted data were limited to information available in the published manuscripts; additional details were not sought from study authors.The same frequency tables and bubble plot visualizations generated by Lam et al.  were ultLam et al.’s  originalLam et al.  reportedAmong our 255 included studies, 60% reported energy sensing-related outcomes, 26% reported appetite-related outcomes, 31% reported glycemic-related outcomes, 27% reported dietary intake, and 9% reported body weight/composition-related outcomes (Table The majority of our included studies 61%) reported sample size ranging between 11 and 50 human participants. This aligned with findings from Lam et al. [1% reportWe generated an evidence map consisting of frequency tables and bubble plots displaying the categorical features of included studies. Our results illustrate a consistent pattern with greater number of studies investigating comparisons of LCS versus sugars across all outcome categories. This difference was consistent, with the greatest difference in the body weight/composition outcome with 83% of studies comparing LCS vs. Sugars. The glycemic outcome demonstrated the lowest difference, with 70% of studies comparing LCS vs. Sugars. Lam et al.  reportedWe also categorized studies by study and outcome group  were excluded at this step. This was not surprising—reviewing the full text provides additional details for reviewers to determine relevance. This illustrates one limitation of the rEM approach that an evidence map generated based on solely titles and abstracts of records will almost certainly include irrelevant references.Our overall conclusions from the traditional evidence map corroborated those reported by Lam et al. . AlthougWe estimated that the traditional evidence map required an additional 65 person-hours over the rEM approach. Although this additional required time may seem minimal, taken in the context of the relative size of the evidence base  indicates that for a larger evidence base the potential time savings from implementing an rEM over a traditional evidence map could be significant.Alternative approaches to evidence mapping such as rEM are a promising approach to identify, collect, and evaluate scientific evidence that reduces the required time, effort, and resources. From our direct comparison to a traditional evidence map, we identified strong concordance with the ultimate conclusions when utilizing rEM. Since the main goal for evidence mapping is to quickly identify research gaps as well as opportunities for systematic review, an rEM may be able to do so in a more efficient manner that requires less time and resources.Our traditional evidence map has several limitations, most of which could be readily addressed with increased time and resources. First, we only utilized references retrieved from searching one database (PubMed). By not including additional databases or the grey literature, we risk missing relevant studies. A broader literature search could have potentially retrieved a higher number of relevant studies and altered our final evidence map and conclusions.We also utilized single-screening for records beyond the first 500 records . This reduced the time and resources required for screening, but introduces the possibility that a single screener could have erroneously excluded a relevant study. This limitation could be addressed by employing duplicate screening of references; however, this would have doubled the required time and resources.Lastly, evidence mapping does not include the evaluation of study quality or risk of bias . Thus, it is unknown whether the included studies are of high or poor quality. For instance, in categories where there was a high volume of included studies, it is possible that these studies are of variable quality. This also hinders limits our ability to state conclusions regarding the health effects of low calorie sweeteners; as expected, with an evidence map the end result is a visual depiction of where evidence exists and where it is lacking, not what conclusions one might ultimately draw regarding the body of scientific evidence. This could potentially be addressed by evaluating each included study using one of several available validated quality assessment tool; however, this would again require additional time and resources to complete this step.Additional file 1. A comparison between a traditional evidence map  and a rapid evidence map .Additional file 2. List of inclusion and exclusion criteria for article screening.Additional file 3. Outcome group categories defined by Lam et al. (2019) for included health outcomes.Additional file 4. Table categorizing studies by study duration and outcome group comparing Lam et al. (2022) and Lam et al. (2019).Additional file 5. Lam et al. (2019) search terms implemented in PubMed on January 24, 2018.


Poly(lactic acid) (PLA) is a biobased polyester with ever-growing applications in the fields of packaging and medicine. Despite its popularity, it suffers from inherent brittleness, a very slow degradation rate and a high production cost. To tune the properties of PLA, block copolymers with poly(propylene adipate) (PPAd) prepolymer were prepared by polymerizing L-lactide and PPAd oligomers via reactive extrusion (REX) in a torque rheometer. The effect of reaction temperature and composition on the molecular weight, chemical structure, and physicochemical properties of the copolymers was studied. The introduction of PPAd successfully increased the elongation and the biodegradation rate of PLA. REX is an efficient and economical alternative method for the fast and continuous synthesis of PLA-based copolymers with tunable properties. The most popular approach to minimize the problem, which is recycling, is only performed in less than 10% of the plastic waste ); the majority of it still ends up in landfills [The negative effects of conventional petrochemical-based plastics on the environment, namely the accumulation of waste , marine andfills . In addiandfills . HoweverPlastics are considered one of the world’s most important environmental problems, which led to EU regulations that are designed to help transition to a sustainable plastics economy . To reduSome of the approaches to overcome the brittleness and slow degradation rate of PLA are copolymerization, blending with other polymers, and addition of plasticizers or fillers ,17,18,19co-terephthalate) (PBAT) finds applications in packaging and as a mulch film [g = −57 °C, Tm = 29 °C, Tcc = −8 °C) [Adipic acid (AdA) is considered the most important dicarboxylic acid by the International Energy Agency, because of its use in the synthesis of polyurethanes, polyamides, and its applications as a plasticizer, as well as in food additives and in the cosmetics industry. The production of biobased adipic acid is under development, and the currently known processes are both chemo-enzymatic (with glucose as precursor) and fermentation ,29,30. Alch film . Poly(pr= −8 °C) . It also= −8 °C) ,34.To incorporate fillers, additives or comonomers into PLA, reactive extrusion (REX) is commonly employed on an industrial scale . Besidesco-PPAd copolymers for the first time via REX in a torque rheometer as a preliminary step for the scale-up of the process to continuous REX in a twin-screw extruder. The mass of PPAd in the formulation varied from 5 to 25 wt%. Neat PLA from L-lactide (with 5% D-isomer) was also synthesized as a control. Two different reactions temperatures, namely 180 °C and 190 °C were tested. The resulting polymers were characterized by means of their physicochemical properties, including molecular weight determination, evaluation of thermal transition temperatures, tensile properties, hydrophilicity, and hydrolysis rate measurements. In this work, poly(propylene adipate) (PPAd) pre-polymer was synthesized with the two-stage melt polycondensation of AdA and 1,3-propanediol  and used in the synthesis of PLA-4, purity: >97.0%), was purchased from Sigma Aldrich Chemical Co . L-Lactide (LA) Purlact B3  was purchased from Corbion N.V.  and Tin(II) 2-ethylhexanoate (Sn(Oct)2, purity > 92.5%) from Merck KGaA, .Adipic acid (AdA) (purity >99.5%) and 1,3-propanediol  (purity > 99.6%) were purchased from Fluka . Titanium butoxide (Ti(OBu)4. The reaction mixture was heated at 180 °C, under N2 atmosphere and continuous stirring. The reaction lasted until the theoretical amount of H2O produced during the reaction was collected (3.5 h). In the second stage (polycondensation), vacuum (5.0 Pa) was slowly applied to minimize the creation of foam and to avoid PPAd oligomer sublimation, and the polycondensation procedure was carried out at 230 °C for 15 min. The classic two-stage melt polycondensation method was used for the synthesis of PPAd aliphatic polyester ,43,44. I® Plasti-Corder® Lab-Station torque rheometer. The ROP of lactide was catalysed by Sn(Oct)2 in a lactide/Sn(Oct)2 molar ratio 1000/1. Triphenyl phosphine (TPP) was used as co-catalyst, in equimolar amount to Sn(Oct)2 [co-PPAd copolymers. Blank samples without an initiator (ROP initiated by traces of H2O) were also prepared. The polymerization was carried out at temperatures of 180 °C and 190 °C in the mixing chamber, with a 100 rpm screw speed. A small stream of dry N2 was circulated to minimize the presence of air in the mixing chamber. The premix of lactide/PPAd/catalyst solution was fed to the rheometer at t = 0. Motor torque (α viscosity) and temperature were monitored vs. time during the polymerization. The samples prepared are listed in PLA and its copolymers with PPAd were prepared by REX in a BrabenderSn(Oct)2 . The catThe molecular weight of the materials was determined using gel permeation chromatography (GPC) with a Waters 600 high-performance liquid chromatographic pump, Waters Ultrastyragel columns HR-1, HR-2, HR-4E, HR-4 and HR-5, and a Shimadzu RID-10A refractive index detector. For the calibration, 9 polystyrene (PS) standards of molecular weight between 2.5 and 900 kg/mol were employed. The prepared solutions had a concentration of 15 mg /mL, the injection volume was 100 μL and the total elution time was 50 min. The oven temperature was 40 °C. Nuclear magnetic resonance (NMR) spectra were recorded in deuterated chloroform for the structural study of monomer and polymer, respectively. An Agilent 500 spectrometer was utilized , at room temperature. Spectra were calibrated using the residual solvent peaks. The morphology of cryofractured cross sections of the samples was studied with a JEOL  JSM 7610F field emission scanning electron microscope (SEM) operating at 5 kV. c) was calculated with equation X:Hm, Differential scanning calorimetry (DSC) analysis was performed using a PerkinElmer Pyris Diamond DSC differential scanning calorimeter  calibrated with pure indium and zinc standards. The system included a PerkinElmer Intracooler 2  cooling accessory. Samples of 5 ± 0.1 mg sealed in aluminium pans were used to test the thermal behavior of the polymers. Crystallinity degree  with Cu Ka radiation (λ = 0.154 nm).X-ray Diffraction (XRD) measurements of the polymers and copolymers were performed over the 2−1 with 32 scans and resolution 4 cm−1.Infrared (IR) spectra were recorded with a Cary 670 , Agilent Technologies, equipped with a diamond attenuated total reflectance accessory, ATR, model GladiATR, Pike Technologies. The spectra were recorded in the range 4000–400 cmTensile tests were performed using an Instron 3344 dynamometer , in accordance with ASTM D638 using a crosshead speed of 5 mm/min. Dumb-bell-shaped tensile test specimens  were prepared by compression moulding in a thermopress at 180 °C, cooled rapidly and cut in a Wallace cutting press. At least five measurements were conducted for each sample, and the results were averaged to obtain the mean values of Young’s modulus, tensile strength at yield and breakpoint, and elongation at break. A student’s T-test was used to determine the statistical significance. p-value ≤ 0.05 was considered statistically significant.Water contact angle was measured with an Ossila contact angle goniometer L2004A1 at room temperature (25 °C). Contact angle was measured by gently placing a water droplet (5 µL) on the surface of films of the samples prepared by compression moulding. At least three measurements were performed, and the mean value is reported. A one-way ANOVA test with post hoc Bonferroni correction was performed to determine the effect of the presence of PPAd on the water contact angle of PLA. The software used was IBM SPSS Statistics . A Pseudomonas cepacia and 0.1 mg/mL lipase from Rhizopus oryzae. The flasks were then incubated at 36.6 ± 1 °C in an oven for several days while the media were replaced every 3 days. After a specific period of incubation (every 5 days), the samples were removed from the flasks, washed with distilled water, dried under vacuum, and weighed until they remained at a constant weight. The degree of enzymatic hydrolysis was estimated from the mass loss of the samples. Blank incubations were also carried out with the samples in the same buffer without enzyme addition. The degree of biodegradation was estimated by the sample weight loss, according to the following equation:For the enzymatic degradation testing, thin films prepared by compression moulding (thickness 0.8–1 mm) were cut in squares with dimensions ~1.5 cm × 1.5 cm and were placed in test tubes containing 10 mL phosphate buffer solution (pH 7.4) with 0.01 mg/mL lipase from Each sample was hydrolyzed in triplicate and the mean value was calculated.2O traces present in raw lactide. The rheograms obtained at 180 °C and 190 °C are shown in The polymerization of PLA was carried out initiated by HAt 190 °C, the ROP of lactide was faster in comparison to 180 °C, reaching the maximum torque at 8.5 min and 14 min, respectively. In addition, the torque  was also higher at higher temperature (10.1 Nm at 190 °C vs. 5.9 Nm at 180 °C). However, depolymerization started right after the torque peak was reached, when the mixing chamber temperature was fixed at 190 °C, because the actual melt temperature increased above 200 °C. co-PPAd copolymers were prepared at 180 °C and 190 °C using 5 wt% of PPAd as macroinitiator for the ROP of L-lactide.Following the same procedure, PLA-co-PPAd copolymers was fixed at 180 °C. The torque–temperature curves of all the polymers prepared at 180 °C are shown in For PLA-PPAd 95/5, the polymerization was faster at 190 °C, reaching stable torque after 11 min, whereas at 180 °C torque stability was reached after 14 min b. HoweveAs PPAd acts as initiator of the ROP of lactide, the higher content of initiator results in faster initiation of the polymerization reaction; however, the maximum Torque (viscosity) is lower due to a larger number of initiation points.co-PPAd is still good for polymer transformation, for example, for injection moulding applications.It must be highlighted that at 5 wt% of PPAd, the time required for torque stabilization is similar to blank PLA (12–14 min), and viscosity and melt strength of the PLA-co-PPAd copolymers with low viscosity and melt strength. Those copolymers would have poor processability by either extrusion or injection moulding technologies; however, they can find applications as base resins in formulations of hot melt adhesives where high viscosity or molecular weight is not required.For higher PPAd contents, the required residence time to achieve torque stabilization was >20 min and resulted in PLA-1H NMR spectra of the PLA-co-PPAd copolymers  and PPAd  are visible in the polymers a. Additi spectra b, among 1H NMR spectra (2OC(O) methylene group of a 1,3-PD unit, which is adjacent to a PLA ester, confirming successful copolymerization. Due to their low intensity and some overlapping, the peak could not be integrated to calculate the randomness of the copolymers. Nevertheless, judging from its low intensity, we can safely conclude that the copolymers exhibit a blocky structure with long PLA sequences. The composition of the copolymers was calculated by comparing the integrations of peaks B and 4 of the  spectra a. It was spectra . Finally−1 assigned to O–H bending, at 2994–2944 cm−1 of C–H stretching, at 1747 cm−1 caused by C=O stretching, at 1452 cm−1 of –CH3 asymmetric bending, at 1182 cm−1 and 1040 cm−1 because of the stretching of the –C–O–C– groups [2 bending at 1467 cm−1 and the C–O/C–C stretch at 1255 cm−1. Additionally, the peak of the carbonyl group of PPAd appears at 1720 cm−1. The spectra of the copolymers do not show any significant differences from the spectrum of PLA. After closer examination of the carbonyl’s region The ATR spectra of PPAd, PLA, and their copolymers are shown in – groups , and theco-PPAd copolymers had The molecular weight and polydispersity index (PDI) of all the polymers prepared were measured by GPC. The resulting chromatographs are shown in oduction . Similarterature ,36,40. Wco-initiator for the ROP, resulting in the observed smaller molecular weight values. While PLA synthesized at 190 °C has higher molecular weight than when synthesized at 180 °C, the first copolymer that was tested (PLA-PPAd 95/5) had a better molecular weight when the ROP took place at 180 °C. In agreement with the motor torque monitoring during reactive extrusion, the molecular weight of the samples decreases after the addition of PPAd, and with increasing the PPAd content. The free hydroxyl groups of the PPAd oligomers act as a 2 with the addition of an initiator. Sn(Oct)2 catalyzes the ROP via a coordination-insertion mechanism extruder ,38. AddiWhen adding the PPAd oligomers in the reaction mixture, their hydroxyl and carboxyl end groups are expected to participate in the reaction. By increasing the concentration of hydroxyls, the number of initiation sites is increased, resulting in more inactive PLA chains and fewer active tin alkoxide species, ultimately reducing the molecular weight . Therefoco-PPAd copolymers with PPAd content up to 15 wt%. The surface of PLA-PPAd 75/25 . All heating steps were performed with a rate of 20 °C/min. The DSC traces of PLA and its copolymers with PPAd are presented in m) was observed at 41 °C, and its Tg after quenching was −57 °C. These values are consistent with previous reports [cc = 6.8 °C and melt crystallization at Tc = −10.6 °C. PPAd exhibited a multiple melting behavior, since a secondary melting peak is recorded in its reheating scans at about 30 °C, right after the end of cold crystallization. This phenomenon has been observed before for PPAd [PPAd a was rec reports ,51,52,53for PPAd  and is ag of the melt-quenched PLA was observed at 55.6 °C; it crystallizes during heating with a peak at Tcc = 109.7 °C and the formed crystals partially melted at 162.2 °C, and finally at 169.9 °C, where the main melting endotherm is positioned. Multiple melting of PLA is ascribed to crystal reorganization during heating, and it is affected by the crystallization temperature [The DSC traces of PLA 180 b show thperature . The as-perature . cc, Tm and Tg were obtained from the heating of the melt-quenched samples with a rate of 20 °C/min, and the Tc from the cooling run with rate 10 °C/min. Tcc, Tm and Tg decreased by increasing the PPAd content. PPAd both reduces the molecular weight and acts as a plasticizer to PLA, which results in the decrease of the Tg and Tm. In contrast with neat PLA, which crystallized insignificantly during cooling with the applied rate, all copolymers showed detectable melt crystallization peaks at around 85–90 °C, which may be a result of the lower molecular weight of the copolymers, as well as the mobility of the aliphatic macromolecular chains of PPAd. The effect of PPAd content on the characteristic thermal transitions of PLA, deducted from the DSC curves, is presented in DSC curves of the materials prepared at 180 °C were also recorded after cold crystallization at 85 °C for 1 h . Cold crX-ray diffraction patterns of the materials as-received from the extruder and of PPAd are presented in The PLA homopolymer was very brittle and could not be tested, because the specimens would break during handling. The two samples PLA-PPAd 85/15 and PLA-PPAd 75/25 were the only ones that could be molded into dumbbell shapes specimens for testing; therefore at least 15 wt% of PPAd oligomer must be added in the synthesis of PLA copolymers to yield adequately plasticized materials. The small amounts of unreacted lactide see  can alsoThe stress–strain curves are shown in The factors that affect the biodegradability of polymers are their chemical structure, hydrophilicity, molecular weight, melting point and crystallinity ,63. PLA To explore the effect of hydrophilicity on the hydrolysis rates of polymers, their water contact angle was measured. The effect of PPAd content on the water contact angle of PLA is shown in The biodegradation of polymers can be monitored with several different methods, including measuring their weight loss during incubation in the presence of enzymes, and microscopic observation of their surfaces. To diminish the effect of crystallinity and dimensions, the specimens used were amorphous films of the same size and similar thickness. In co-PPAd copolymers lose more weight as they are more prone to be attacked by the enzymes in comparison with neat PLA. Similar results were obtained in previous work on PLA-PPAd blends [The effect of the presence of the enzymes of the final mass loss of the polyesters is presented in d blends .The morphology of the sample’s surfaces after 30 days of enzymatic hydrolysis was studies with SEM . The surco-PPAd block copolymers were successfully prepared by REX in a torque rheometer, showing potential to be scaled-up by REX in a twin-screw extruder, in an economic and fast way. The optimum reaction temperature was 180 °C, and the reactions were completed in very short times (up to 20 min) in comparison with the typical batch-type processes. The molecular weight of the materials was comparable to similar copolymers synthesized with typical ROP and melt polycondensation reactions that require several hours to be completed. The PLA moieties are those that were responsible for crystallization. The addition of 15 wt% PPAd in the feed successfully increased the elongation of PLA to ~100%, making the addition of a plasticizer unnecessary. PPAd acted as a macroinitiator that reduced the molecular weight of PLA; however, the produced copolymers can find applications as films or base resins in formulations of hot melt adhesives where high viscosity is not required. Finally, PPAd significantly increased the hydrolysis rates of PLA copolymers, which can be controlled by adjusting the PPAd content. PLA-


Modified vaccinia Ankara (MVA) is a promising vaccine vector due to its highly attenuated phenotype and good immunogenicity. However, obtaining a new recombinant MVA remains a tedious and laborious procedure involving many rounds of plaque purification. Recombinant MVA generation can be greatly improved and facilitated by different selection techniques. Here, we describe a comparison between techniques based on K1L, F13L and D4R genes. For many years, the modified virus Ankara (MVA) has been widely used as a viral vector for the development of numerous vaccines that are presently at different stages of clinical and preclinical trials. Vaccine candidates against tuberculosis, malaria, flu, HIV and MERS expressing transgenic antigens have been developed .Some features of the virus have contributed to that popularity of the vector. MVA was obtained in Germany as a result of multiple passages of vaccinia virus (VACV) Ankara in chicken embryo fibroblasts, which led to the loss of some part of the genome and the ability to replicate in mammalian cells, including human cells (except for the cell line BHK-21) ..12].To evaluate the effectiveness and convenience of this selection method, we designed a shuttle vector pShuttle tk RFP K1L that contained a marker gene of the red fluorescent protein (RFP), K1L gene, and homology arms to the thymidine kinase MVA locus a. To obtWe did not obtain a pure clone of rMVA-K1L in the three separate experiments starting from recombination. As shown in This selection is based on obtaining the strain MVA ΔF13L that does not contain the native gene F13L  and furtThe first step in F13L-based selection is obtaining the virus MVA ΔF13L with a deletion of this gene in the genome. To obtain MVA ΔF13L, we designed a shuttle vector pShuttle F13Ldel, which contained the RFP gene flanked by homology arms to locus F13L. During recombination with a shuttle vector, the gene F13L in the MVA genome was replaced with a marker gene RFP. Pure MVA ΔF13L virus was isolated by several rounds of plaque purification. PCR analysis of the obtained clones is presented in To obtain recombinant MVA with restored growth properties, we designed the shuttle vector pShuttle F13LrevGOI that contained arms of homology to the locus F13L and a native gene F13L. Recombination between pShuttle F13LrevGOI and MVA ΔF13L genome mediates the replacement of the RFP gene with the F13L gene, restoring the F13L gene into its original locus. The lack of RFP fluorescence allows the differentiation of a recombinant virus from parent MVA ΔF13L. Moreover, after obtaining a pure recombinant rMVA F13Lrev, additional deletion of the RFP sequence is not required.To perform recombination, pShuttle F13LrevGOI was transfected into BHK-21 cells preliminarily infected with MVA ΔF13L. Further, we purified the recombinant MVA using two different protocols proposed by  Figure . The firAs shown in We also tested the possibility of accelerating the rMVA purification using the faster purification protocol. We tried to reduce the number of passages for a viral mixture in the first protocol from four to two. After that, separate clones were isolated by the limiting dilution method. There were small fluorescent plaques of the parent virus in numerous wells, although we managed to select 5 wells with target single nonfluorescent large plaques from 16 wells. The obtained clones were analyzed by PCR. As a result, two clones of pure recombinant were selected c. Thus, Method of selection based on the vaccinia virus D4R gene was first described by . D4R encTo obtain a BHK-21-D4R cell line that expresses D4R, we designed a lentivirus vector LV D4R containing the D4R gene controlled by its promoter and EGFP controlled by the PGK promoter. The vector was transduced into BHK-21 cells, and EGFP-positive clones were selected and grown into cell lines.To obtain the MVA virus with D4R deletion, we designed a shuttle vector pShuttle D4Rdel, which contained the RFP gene flanked by homology arms to locus D4R. For recombination, a shuttle vector was transduced into BHK-21-D4R cells preliminarily infected with MVA. Pure virus MVA ΔD4R was obtained by plaque purification. The results of PCR analysis of the obtained clones are presented in To obtain MVA recombinants with restored replication, we designed the shuttle vector pShuttle D4RrevGOI containing flanking sequences of D4R along with the complete D4R gene. Recombination between the pShuttle D4RrevGOI and MVA ΔD4R genome mediates the replacement of the RFP gene with the D4R gene, restoring the D4R gene into its original locus. Therefore, after obtaining pure recombinant MVA, the additional removal of a fluorescent marker is not required.To perform recombination, pShuttle D4RrevGOI was transfected into BHK-21-D4R cells preliminarily infected with MVA ΔD4R. After recombination, the viral mixture underwent two blind passages in BHK-21 cells. The authors of this selection method noted that after two passages the viral mixture contained only a recombinant. We analyzed the PCR results of the viral mixture after the 2nd passage and revealed that there were residual traces of the parent MVA ΔD4R virus a. Thus, To obtain an MVA-based vaccine, it is necessary to integrate the sequence encoding antigens into the viral genome. It is carried out by homologous recombination between the initial virus and shuttle vector with further isolation of the target recombinant MVA by plaque purification. This process is rather laborious and inefficient. Previously, we tried to optimize a conventional method of rMVA selection by adding a real-time PCR test of single plaques , which aSeveral methods of recombinant virus purification based on the enrichment of rMVA during replication have been developed to simplify the process. However, until now these methods have not been compared directly. Thus, the aim of the present study was to compare the efficiency and usability of some of them.+) allows the rescue of the wild-type virus (K1L−) in the same cell.The first tested method of rMVA isolation was based on the host range gene, K1L, as the selective growth marker in rabbit kidney RK-13 cells. We showed that such a method of rMVA selection did not perform well. The publications mentioned various periods required for isolation of pure recombinant virus (from 2 to 5 selective passages in RK-13) ,19. In oFurther, we tested two similar systems of selection based on the deletion of F13L or D4R from the MVA genome and further reinsertion of these genes, which are necessary for successful replication. The main difference between MVA ΔF13L and MVA ΔD4R was in the rate of replication. MVA ΔF13L replication is dramatically reduced in the BHK-21 cell culture compared to wild-type MVA, while MVA without essential D4R gene could not replicate in these cells at all. Since MVA ΔD4R was not capable of replicating in cells, for its propagating it was required to additionally generate a cell line expressing the D4R to complement the absent gene function.We successfully obtained pure recombinant viruses without admixtures of the parental viruses using both systems. Although D4R-based selection was more effective. Nine out of ten (90%) viral clones, isolated by the limiting dilution method, did not contain the admixtures of the initial MVA ΔD4R after two blind passages. On the other hand, for F13L selection approximately 12% and 50% of such samples were obtained after two and four blind passages correspondingly.Both systems were simple and convenient in practice. First, there is no need for the application of antibiotics, selective agents, agar medium, etc. Second, a recombinant virus can be obtained after two to four blind passages without laborious plaque isolation and analysis of plaques at each stage. Third, since passaging is not time-consuming, several recombinant viruses can be obtained simultaneously. Fourth, the advantage of this method compared to the conventional method is that it does not require additional marker genes to select a recombinant sequence and thus does not need further removal of these sequences.The disadvantage of both systems is that the insertion of the target gene is possible only in the initial locus, where the genes F13L and D4R are located. Furthermore, this type of selection can be used for the integration of only one insert because the reinsertion of F13L/D4R along with the target gene restores the native growth properties of the virus.When it is necessary to obtain rMVA with the additional insert in a different locus, the methods of rMVA selection should be combined. For example, F13L/D4R-based selection can be used for the first integration. For the second integration, it is possible to use an accelerated method of selection of single plaques using real-time PCR . The appAdditionally, the approach based on the application of MVA with the deletion of both genes (MVA ΔF13L ΔD4R) is promising because it can reduce the time for isolation of double recombinants. However, the prospects of this approach have yet to be elucidated.


Randomized controlled trials (RCTs) are considered to represent the gold standard of scientific studies and paved the way for evidence-based medicine (EBM). Besides the initial aim to improve the quality of patient care, EBM is used in the meanwhile for political and economic decision-making and legal issues as well. A review of the literature was performed, followed by a search using links and references of the detected articles. Additionally, homepages for German institutions of public health were screened. Substantial limitations of RCTs and EBM health care could be identified. Based on the selected literature, 80% of the medical treatments have low evidence. RCTs are expensive and are mainly performed by the industry nowadays. A publication bias for positive results exists. Some RCTs are of low external validity. Many studies have a low fragility index. Nonetheless, negative RCTs could be of benefit for the patients. The results of RCTs, gained in a distinct patient population, are partially generalized. RCTs should be analyzed critically before adopting the results to daily clinical routine. It is not really justified to use RCTs and EBM for political and economic decision-making and legal issues as seen today. Clinical Judgment and by Cochrane in 1972, who published the book Effectiveness and Efficiency: Random reflections on Health Services, and paved the way to the concept of EBM [The improvement of existing and the development of new therapies are of genuine interest to scientists and clinicians in the different medical disciplines. Knowledge gain is reached by experimental and especially clinical trials of different retrospective and prospective study designs. Retrospective and prospective non-randomized studies are prone to bias, inaccuracy, and missing data and are therefore labeled to be inferior to randomized controlled trials (RCT). The exact design of RCTs was already developed in the 1940s but was largely considered being unethical because for many diseases; at that time, a therapeutic alternative was lacking. In these years, mostly case reports and case series and a few case-control studies were published [t of EBM , 7. Sinct of EBM .Hence, RCTs are considered to provide the best available medical knowledge (evidence class I), with the aim to mainly enhance the accuracy of medical decision making and to improve medical therapies . DespiteA search of the available literature on RCTs and their limitations was performed using the National Library of Medicine (PubMed), Google, Google Scholar, and Wikipedia. The main search tool was the National Library of Medicine (PubMed). The following MeSH terms were taken: “randomized controlled trial/economics, epidemiology, ethics, legislation, and jurisprudence,” “evidence-based medicine/economics, ethics, history,” and “fragility index.” Only articles in English were included without time restriction. Abstracts were read if the title suggested that the article critically discusses the role of RCTs. The complete paper was read, if the abstract was considered being relevant by the two authors. Additionally, the homepages of the three major German institutions for quality in health care  were searched for their positioning concerning RCTs and evidence-based medicine (EBM). Because of the very high number of potentially retrievable publications and the blurring difference between more scientific manuscripts and more political statements, expressing the personal opinion of the author, no attempt was made to achieve completeness of the search. Accordingly, the references of the retrieved articles were not used for completing the search.Despite the strong focus on RCTs and EBM that could be witnessed in the last 20 years, approximately 50% of all medical therapies are still performed without class I evidence nowadays. Three factors explain this high percentage. (1) Most frequently, the superiority of a therapy/a medical measure is obvious and already proven by studies of lower evidence. Without equipoise, the results of a RCT are predictable, alleviating the reasonableness and possibly the ethical tenability of a study . A neuroThe execution of a RCT is complex and, as a consequence, expensive. The expenses for a phase III prospective, randomized drug trial had been 30 million US dollar at the turn of the millennium ; today, p value of 0.05 is used to determine a statistical significance. The fragility index is the minimum number of events that convert a statistically significant into a statistically insignificant result [Before the initiation of a RCT, the number of patients that is required to proof or refute the study hypothesis in a statistically meaningful way has to be calculated. This size of the study population on the one hand and the need to control the costs and to provide results that are still considered to be up-to-date by the time of publication often require a multicenter patient recruitment. Unfortunately, criteria for the selection of participating centers are often not clearly defined. This is especially important for RCTs, in which operative procedures or non-operative management strategies are compared with each other . The indt result . Many RCt result . The medt result .Primary study endpoints of many RCTs are distinct and easy to evaluate, such as overall survival or progression-free survival in several oncologic trials. Patient-related outcome parameters (PROMs), if assessed at all, are mostly secondary study endpoints . This faRCTs are designed to answer a distinct medical question in a defined study population. Despite, both negative and positive study results are transferred to patient populations that were not subjects of the trial. An example for such a generalization could be witnessed after the International Subarachnoid Aneurysm Trial (ISAT) . In the The authors acknowledge the important role of RCTs and EBM for improving diagnostics and treatment in medicine, but also believe that certain skepticism should be retained, considering the results of the literature research. Because of the high costs, many RCTs are performed by the industry, which introduces a bias in favor of reporting positive results as witnessed recently for TDR. On the other hand, negative results of industry-sponsored RCTs, which are likewise important from a scientific standpoint, are underreported with a subsequently presumed effect on meta-analyses (which are required for class Ia evidence) towards better results . Less inhttps://www.iqwig.de/en/methods/basic-principles.3314.html). That evaluation guides the decision for or against covering the treatment costs by medical insurance companies, which might result in the loss of the patients’ perspective  [http://www.awmf.org/leitlinien/awmf-regelwerk/einfuehrung.html), negating the fact that the relevance of RCT results in the non-academic setting, the “real world” is often unclear [The intention of the protagonists of RCTs and EBM is considered to be the improvement of medical decision-making, but nowadays RCTs/EBM have gained a substantial political, economic, and legal dimension. In Germany, for example, the IQWiG evaluates the efficacy of new therapies based on RCTs/EBM (e versa) . RCTs/EB unclear . Despite unclear . Given tWhile RCTs are designed to answer a distinct medical question in a defined study population, we sometimes witness a generalization of the results after the completion of the trial. Typical examples are ISAT and the randomized trial of unruptured brain AVMs (ARUBA) that resulted in an unjustified change of patient management fueled by the interests of neurologists, interventionalists, and neurosurgeons plus the industry (in ISAT). We have to be aware that that generalization of RCTs is scientifically not justified.In many instances, RCTs represent the best available scientific evidence. However, RCTs have to be analyzed in detail, and a healthy level of skepticism should be retained, because economic aspects, especially industry funding and methodological flaws, can largely influence the results. The increasing tendency of using RCTs for justification of political, medico-legal, and economic decisions as well as generalizing the results should be seen with caution.