Background: The impact of the coronavirus disease 2019 (COVID-19) pandemic on people with multiple sclerosis (MS) is a major current concern, in particular the risk of death. Here we describe the impact of the first wave of COVID-19 infections (Mar 2020-July 2020) on the Scottish MS Register (SMSR) population, a cohort of 4702 individuals with MS, all newly diagnosed in the past decade.Methods: We established a clinician alert system, linking the SMSR with the Electronic Communication of Surveillance in Scotland (ECOSS). This allows identification of patients within this cohort who had a positive SARS-CoV-2 PCR test. The SMSR was also linked to death records from National Records Scotland.Results: Of 4702 people with MS, 246 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PCR tests were performed, of which 17 were positive. The proportion of positive tests were similar to the general Scotland population . Between 1st March – 31st July 2020 12 individuals on the SMSR died, 5 of which were linked to COVID-19 . This number of COVID-19-related deaths was higher than expected . All COVID-19-related deaths in the SMSR occurred in individuals with advanced disability , and no deaths occurred in patients receiving disease modifying therapy (DMT) therapies.Conclusion: In this nationally comprehensive cohort of MS patients diagnosed in Scotland within the past 10 years, we observed similar rates of PCR-confirmed SARS-CoV-2 infection compared to the general Scottish population, but a small number of excess COVID-19 related deaths. These deaths occurred in individuals with advanced disability who were not receiving DMTs. An important aspect of the public health response to COVID-19 has been the accurate identification of individuals at risk of severe COVID-19 outcomes, particularly those at high risk of death.Coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), emerged in China in late 2019 and was declared a pandemic by the WHO in mid-March 2020Multiple sclerosis (MS) affects over two million people worldwide and the impact and mortality of COVID-19 on people with MS is a source of major current concern. People with multiple sclerosis are potentially at risk of severe COVID-19 because they are receiving immunosuppressants, may develop significant disability and commonly have comorbidities. Equally, many individuals with MS are of working age and severe COVID-19 avoidance measures may cause social, mental and financial harm.4.During the first wave of the COVID-19 pandemic guidelines were drawn up by national and international neurological and MS societies to advise people with MS of their risk of severe COVID-19 based on theoretical risks posed by immunotherapy, and evidence of respiratory and bulbar failure8. These studies suggest that age, Expanded Disease Severity Score (EDSS) and comorbidities are risk factors for severe COVID-198. These studies also suggest that some immunotherapies such as ocrelizumab may confer modest risk for severe COVID-19 outcomes, but most immunotherapies do not7. Such studies are important for identifying characteristics of affected individuals but rely on spontaneous reporting. There is a need to understand the impact of the pandemic in a representative cohort where cases are ascertained in an unbiased manner, with a focus on severe outcomes and death.As the first wave of the pandemic has emerged, case series and registries of patient-reported and physician-reported COVID-19 in people with MS have enabled the identification of risk factors for severe COVID-19 outcomes9. MS is a lifelong disease, with a clinical course typically lasting over 30 years. Therefore, the Scottish MS register is a nationally comprehensive, incident cohort of people with relatively early MS, in the first decade of disease9. The SMSR can be linked to other healthcare databases across Scotland including infections and deaths.The Scottish MS Register (SMSR) is an NHS Scotland audit tool which has collected data on over 4500 newly diagnosed individuals with MS in Scotland since 2010In light of the recent COVID-19 pandemic, we linked data from the Scottish Multiple Sclerosis Register with the Electronic Communication of Surveillance in Scotland (ECOSS), a Scotland-wide surveillance tool for monitoring infections that are of clinical or public health importance. This flagging system allows neurologists across Scotland to be informed when people with MS under their care develop COVID-19, based on a positive nasopharyngeal PCR test. In addition, linkage to Scottish death records from National Records Scotland was also performed during the period of the pandemic first wave.st March – 31st July 2020.In this manuscript we report the findings from this surveillance system during the first wave of the COVID-19 pandemic in Scotland over the period 110. The Scottish MS Register is an established Scottish national NHS audit, and as such does not require research ethics approval. The aim of the register, which was established in 2010, is to improve the NHS care of people with MS in Scotland. The aims, objectives, permissions and data governance of the SMSR are available from theRegister site.The research and governance framework of the SMSR has been previously describedPatient Information Sheet.Patient information about how the data is collected and used is provided in thest 2020. Local neurologists were informed by the SMSR when a patient under their care had developed PCR confirmed SARS-CoV-2 infection, or had died. This information was used in the routine clinical care of the patient and neurologists fed back deidentified data to the SMSR. Deaths in the absence of a positive COVID-19 PCR results were determined to be COVID-19 related in the clinical judgement of the local clinical neurologist. Collated data was reviewed periodically during the pandemic.Following internal governance review, the ECOSS and SMSR databases were electronically linked in real-time, permitting patients appearing on both databases to be identified weekly using the Scottish national patient unique identifier system  starting March 1https://www.msr.scot.nhs.uk/Reports/Dashboard-2020.html and was cross-referenced with national death records to exclude individuals who died prior to 1st March 2020.The population structure of the SMSR was taken frompublished PHS datasets. These age and sex-specific rates were used to estimate expected number of positive tests and deaths within the SMSR. COVID-19 test data were taken from the Public Health Scotland Weekly COVID-19 report dated August 2 and included the period 1st March – 31st July 202011. This contains information on all positive and negative nasopharyngeal COVID-19 tests carried through NHS Scotland laboratories and includes results from hospitals, GP practices, drive-through centres, mobile units, and home testing kits. Data on COVID-19 related deaths from 1st March to 31st July 2020 were obtained from the National Records of Scotland, defined as deaths occurring in any location where COVID-19 was recorded on the death certificate12.National background rates of COVID-19, including mortality data, were identified fromR version 3.6.3 using packageepiR version 1.0-15. Confidence intervals were approximated using the method of Rothman and Greenland assuming observed events to be Poisson variates and expected events invariate. Under the same assumptions, p values (H0: ratio of observed to expected = 1), were calculated by chi-squared test. Exact confidence intervals and hypothesis testing, where expected counts were low (≤5), were calculated using the mid-P exact method. For reporting statistical significance, the threshold (α) was set at 0.05.Statistical analyses were performed in2(d.f. = 1) = 0.014,p=0.90).Of the 4702 people diagnosed with MS since 2010 on the SMSR, 246 (5.2%) underwent SARS-CoV-2 PCR testing during the first wave. Over the same period, 6.7% of the Scottish population underwent testing. Of the 246 SARS-CoV-2 PCR tests carried out in the SMSR population, 17 (6.9%) were positive . The nump=0.01 (mid-P exact)). One individual died after positive PCR-confirmation, and 4 died of clinically suspected COVID-19 but were not tested. The timings of these deaths relative to the wave of COVID-19 attributable deaths in Scotland are shown inGiven that the proportion of PCR-confirmed cases were similar between the SMSR and the general population, we next asked whether there were differences in COVID-19 related mortality. During the first pandemic wave 12 deaths of individuals on the SMSR were recorded, 5 of which were identified to be COVID-19 related. This observed number of COVID-19-related deaths in the SMSR cohort is higher than expected, based on COVID-19 related death rate in the general Scottish population over the same time period.  and none were receiving disease modifying therapies.Using the Scottish MS Register COVID-19 surveillance system, we found that the proportion of SMSR with PCR-confirmed SARS-CoV2 infection was similar to that of the general Scottish population. However, we observed 5 deaths linked to COVID-19, when ~1 death was expected, based on the Scottish reference population. These deaths all occurred in patients with advanced disability and no patients receiving immunotherapy died.13.The SMSR was established in 2010 to capture all incident MS cases in Scotland. The register therefore has the advantage of being nationally comprehensive and captures data from individuals who are in the first decade of the disease, who will have lower disability scores, and higher immunotherapy treatment rates than the prevalent populationOur data provide particular insight into the impact of COVID-19 on mortality within the SMSR and suggest that this burden is falling upon patients with advanced disability. However, our analysis has a number of important limitations. Firstly, during the majority of the time period covered by this analysis, SARS-CoV2 PCR swab testing was restricted to a hospital setting or healthcare workers. Throughout the first wave of COVID-19 infection the availability of SARS-CoV2 PCR nasopharyngeal swab tests changed, as did the threshold for testing: initially only hospitalised patients were eligible for tests, followed by healthcare professionals, and finally widespread community testing was introduced. Therefore, many mild cases occurred in the community without PCR confirmation and would not be identified using this surveillance system. Secondly, while the ECOSS database is comprehensive, we cannot be sure that all cases are captured through record linkage, such as those having privately organised tests or not being tested at all. Thirdly, the data linkage described occurs within Scotland only and will not capture COVID-19 diagnoses or deaths outside Scotland. Finally, since the SMSR is primarily an audit tool, detailed clinical data such as severity scores and immunotherapy details are not currently collected across the SMSR.Caution is also needed in interpretation of our observed versus expected analyses for COVID-19-related deaths because of slight differences in ascertainment between SMSR and National Records Scotland. In the general Scottish population, a COVID-19-related death is defined by National Records of Scotland as a death which records COVID-19 on the death certificate. In the SMSR population, a death was recorded as COVID-19-related by the neurologist.6. Our data do not permit quantification of the risk of death with individual immunotherapies since the SMSR holds only very limited data on the current DMT usage of the population, although over 60% of newly diagnosed patients in the SMSR are offered a DMT at the point of diagnosis9. While it is reassuring that no patients on DMTs in the SMSR cohort died of COVID-19 it is important to bear in mind that UK Government and Scottish Government advice was for all patients with MS to perform stringent social distancing during much of the period captured here. These results are consistent with an emerging body of evidence derived from other cohort and registry studies which suggest that the burden of severe COVID-19 is falling on individuals with advanced disability rather than those receiving immunotherapiesThese results may be relevant to informing the optimisation of COVID-19 avoidance measures for people with MS in the event of future pandemic waves. In Scotland during the first wave of the pandemic, all patients with MS were advised to perform particularly stringent social distancing. In addition, people with MS who (i) had recently received treatment with alemtuzumab or cladribine or (ii) experienced bulbar or respiratory dysfunction, were advised to adopt even more stringent “shielding” measures. Our analysis is not intended to evaluate the complex risk-benefits outcomes of such interventions. However, it is clear that, within the SMSR cohort, those with advanced neurological disability are a particularly vulnerable population.The number of confirmed SARS-CoV-2 infections in the SMSR were observed at a similar frequency to the general population. However, we observed a small number of excess deaths due to COVID-19 in people with MS compared to the general population. These deaths occurred in individuals with advanced disability who were not receiving immunotherapy. These results may help identify people with MS who are vulnerable to severe/fatal COVID-19.https://publichealthscotland.scot/). The specific dataset used is the “Public Health Scotland Weekly Covid-19 Report August 9” ;https://beta.isdscotland.org/find-publications-and-data/population-health/Covid-19/Covid-19-statistical-report/. [Accessed: 16-Aug-2020]).Scotland-wide COVID-19 PCR test data are available online from Public Health Scotland (https://www.nrscotland.gov.uk/). The specific dataset used is the “Deaths involving coronavirus (COVID-19) in Scotland Week 31”;https://www.nrscotland.gov.uk/files/statistics/covid19/covid-deaths-report-week-31.pdf. [Accessed: 16-Aug-2020].Scotland-wide COVID-19 related mortality data are available online from the National Records of Scotland . All personal details on the register are stored in accordance with Information Services Division (ISD) Guidelines. The most recent Scottish MS Register report and publicly available data are available from:nss.isdscottishmsregister@nhs.net. No identifiable information will be passed to any individual or organisation outwith the National Health Service.The information is collected by the hospital and is collated by the Information Services Division (ISD) at NHS Scotland. ISD has well established systems to protect the privacy of data held on patients and staff. The General Data Protection Regulation (GDPR), the Data Protection Act 2018 and Public Benefit and Privacy Panel for Health & Social Care (PBPP) Guidelines. Application for access to data from the SMSR can be made to This very clearly written report describes rates of Covid infection, and Covid-related mortality, in individuals with multiple sclerosis (MS) on the Scottish MS Register (SMSR) during the first wave of Covid infections (March 2020-July 2020).  The study concludes that individuals on the SMSR had similar rates of PCR-confirmed SARS-CoV2 infections as the general population. But that there was a small number of excess Covid-related deaths (n=5), all of which occurred in individuals with advanced disease and high levels of disability (EDSS >7.0), who were not receiving DMTs. These findings agree well with other reports.  The main strength of this study is the unique SMSR resource, and how this was linked by the research team to an electronic surveillance system for SARS-CoV2 testing results, as well as deaths reported to the National records registry. Limited PCR testing during the first wave (meaning that mild community cases would not have been identified) and the fact that 4 out of the 5 Covid-related deaths were diagnosed clinically. It is also unclear to me why Covid-relatedness for the SMSR cohort was determined by a neurologist rather than by the death certificate . This may have introduced bias. Limited clinical details held by the SMSR (which is primarily an audit tool) - for example regarding use of DMTs, co-morbidities and level of disability. Given this, some caution must be applied when extrapolating these data to other clinical groups. It would also have been interesting for the authors to speculate on how these data may have been influenced by the initial shielding guidance given to individuals with MS. For example, I am unsure if the authors have access to information regarding what proportion of the SMSR cohort would have been advised to shield. But if significant, the risks calculated in this manuscript may be an underestimate. It would also be interesting to know what proportion of patients with advanced disease were living in a care-home setting.  As pointed out by the authors, the study has a number of limitations (most of which are beyond the control of the research team), including: In conclusion, this is a very interesting paper that makes excellent use of a unique clinical resource (SMSR). It adds further weight to the argument that for further waves/future pandemics greater emphasis should be placed on protecting individuals with advanced MS and higher levels of disability.Is the work clearly and accurately presented and does it cite the current literature?YesIf applicable, is the statistical analysis and its interpretation appropriate?YesAre all the source data underlying the results available to ensure full reproducibility?YesIs the study design appropriate and is the work technically sound?YesAre the conclusions drawn adequately supported by the results?YesAre sufficient details of methods and analysis provided to allow replication by others?YesReviewer Expertise:Immunology, T-regulatory cell biology, Tissue Immunity, Multiple Sclerosis, Autoimmunity, Clinical Trials.I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. A national epidemiological survey which aims to estimate the risk of COVID susceptibility and death in Multiple Sclerosis (MS) employing a national registry of early MS as its population frame. Scotland enjoys, almost internationally, unique health care system data linkage capacities. The joined up nature of their health care system and almost complete population coverage enables rapid comprehensive studies such as this. Its also heartening to see how these facilities can be used to immediately inform patient care as evidenced by the flagging system documented in this manuscript. The analysis is a useful contribution which provides data which aligns with earlier, less comprehensive reports in this space. With the rapid evolution of discovery in this space, it would be helpful for these finding to be contextualized with the most up-to-date extant literature. More details regrading the clinical characteristics of the MS registry would be helpful to understand the prevalence of known COVID risk factors. Further, details of what proportion of patients fall into the UK government's extreme social isolation category would help assess whether the calculated risk is an underestimate.  Finally, given the known geographical risk factors for both MS and COVID (which conceivably could conflict), a future effort to match controls according to post code (a feasible undertaking) would enhance confidence around the risk estimates.Is the work clearly and accurately presented and does it cite the current literature?PartlyIf applicable, is the statistical analysis and its interpretation appropriate?YesAre all the source data underlying the results available to ensure full reproducibility?NoIs the study design appropriate and is the work technically sound?YesAre the conclusions drawn adequately supported by the results?YesAre sufficient details of methods and analysis provided to allow replication by others?YesReviewer Expertise:EpidemiologyI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. In this paper, Fernandes and co-workers report outcomes with Covid-19 in a Scottish MS population. A great deal of attention has been given to the possible impact of disease-modulatory treatments (DMT) for MS on COVID-19 outcomes. However, most existing data comes from spontaneous reports, constituting a limitation for extrapolating results on a population level. In this study, the Scottish MS registry (SMSR) was linked to an electronic surveillance system for Sars-CoV2 testing results, as well as deaths reported to the National Records registry. Top line results comprise a similar rate of positive Sars-CoV2 testing results as the general population, but increased mortality compared to age and sex matched controls. All five deaths reported occurred in individuals with a high disability level who were not currently treated with DMT. The authors correctly mention certain limitations with the study, such as that a relatively small proportion of individuals were tested for Sars-CoV2 with PCR, the reporting system for test results not covering testing in private clinics, the fact that four out of five deaths connected to Covid-19 were clinically diagnosed and that the SMSR does not include details on DMT and severity scores. In particular, without knowledge about disability levels and DMT status, caution must be exerted before extrapolating results to other populations that may be structured in a different way. In addition, the SMSR includes individuals being diagnosed with MS in the last decade. It is unclear to this reviewer if the coverage has been validated by some means. Also, the age structure of the reported population is quite high given the disease duration, likely reflecting a delay between disease onset and diagnosis. Only age and sex are reported for the five identified deaths, and other factors, such as co-morbidity and other concomitant treatments apart from DMTs, may also have been of relevance. Nevertheless, population-based studies, such as this, are still rare and the presentation of results, statistical methods and resulting conclusions are relevant given the available data.Is the work clearly and accurately presented and does it cite the current literature?YesIf applicable, is the statistical analysis and its interpretation appropriate?YesAre all the source data underlying the results available to ensure full reproducibility?YesIs the study design appropriate and is the work technically sound?YesAre the conclusions drawn adequately supported by the results?YesAre sufficient details of methods and analysis provided to allow replication by others?YesReviewer Expertise:multiple sclerosis, clinical and experimental aspects, including treatmentsI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.


Following the initial identification of the 2019 coronavirus disease (covid-19), the subsequent months saw substantial increases in published biomedical research. Concerns have been raised in both scientific and lay press around the quality of some of this research. We assessed clinical research from major clinical journals, comparing methodological and reporting quality of covid-19 papers published in the first wave (here defined as December 2019 to May 2020 inclusive) of the viral pandemic with non-covid papers published at the same time.The BMJ, Journal of the American Medical Association (JAMA), The Lancet, and New England Journal of Medicine, from first publication of a covid-19 research paper (February 2020) to May 2020 inclusive. Paired reviewers were randomly allocated to extract data on methodological quality (risk of bias) and reporting quality (adherence to reporting guidance) from each paper using validated assessment tools. A random 10% of papers were assessed by a third, independent rater. Overall methodological quality for each paper was rated high, low or unclear. Reporting quality was described as percentage of total items reported.We reviewed research publications (print and online) from From 168 research papers, 165 were eligible, including 54 (33%) papers with a covid-19 focus. For methodological quality, 18 (33%) covid-19 papers and 83 (73%) non-covid papers were rated as low risk of bias, OR 6.32 (95%CI 2.85 to 14.00). The difference in quality was maintained after adjusting for publication date, results, funding, study design, journal and raters (OR 6.09 (95%CI 2.09 to 17.72)). For reporting quality, adherence to reporting guidelines was poorer for covid-19 papers, mean percentage of total items reported 72% (95%CI:66 to 77) for covid-19 papers and 84% (95%CI:81 to 87) for non-covid.Across various measures, we have demonstrated that covid-19 research from the first wave of the pandemic was potentially of lower quality than contemporaneous non-covid research. While some differences may be an inevitable consequence of conducting research during a viral pandemic, poor reporting should not be accepted.The online version contains supplementary material available at 10.1186/s12916-021-01920-x. The severe acute respiratory syndrome coronavirus 2 (SARS COV-2), and the resulting clinical coronavirus disease 2019 (covid-19) have disrupted all aspects of healthcare [The main platform for sharing results of scientific research remains the peer reviewed, biomedical journal. Biomedical publishers reported a substantial increase in submissions in early 2020, with most of the content related to covid-19. Journals responded to the increasing volume of covid-19 research with rapid publication of these papers . HoweverThere are many aspects of a scientific paper that contribute towards the overall ‘quality’ . Two of We aimed to assess methodological quality (risk of bias) and reporting quality (compliance with reporting guidelines) for papers published in the highest impact biomedical journals during the first wave (here defined as December 2019 to May 2020 inclusive) of the covid-19 pandemic. We compared results for covid-19 and contemporaneous non-covid research.Although not a systematic review in the classical sense, where appropriate we adhered to the relevant sections of the Preferred Reporting Items for Systematic reviews and Meta-analyses (PRISMA) guidance . No ethiThere was no funding source for this study. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication.The BMJ , The Journal of the American Medical Association , The Lancet (Elsevier) and The New England Journal of Medicine . We favoured these four titles as they publish weekly print editions, have international readerships and are generally considered to have the highest publication standards. These were also some of the first journals to publish clinical research on covid-19 during the first wave of the pandemic.We included the following journals, chosen as representing the highest impact clinical research titles in the category of ‘Medicine, General and Internal’ ): Each print journal was hand searched by a single reviewer, beginning from January 2020 to identify the first clinical covid-19 research papers. These publications were the inception point for further hand searching to collate all research papers from that journal. As an internal validity check, the journals’ online search facility and/or any specific covid-19 resource hubs were checked to ensure no relevant content had been missed. Paper selection was from the first week that the relevant journal published a covid-19 related clinical research paper and concluded on Sunday 17th of May inclusive. On this date, journal websites were searched for all covid-19 papers available online prior to print. These dates were chosen to represent the first wave of the covid-19 pandemic.We assessed all articles labelled as original clinical research, including Brief Reports. We did not include non-research content such as editorials or commentary. Within the original clinical research remit, we excluded papers that were not suitable for assessment with our chosen quality tools, for example pre-clinical or translational science. Decisions on exclusion were made by raters as part of the initial assessment of potentially eligible papers.We classified included papers based on the primary study method and this classification informed the choice of the quality assessment tools used. We pre-defined six categories chosen to encompass the most commonly used clinical research designs: diagnostic test accuracy; observational studies ; prognosis; qualitative; randomised controlled trials (RCT); and systematic review.We created two groups to facilitate comparisons, ‘covid-19’ research and ‘non-covid’. The covid-19 label was applied where the exposure, intervention, test or outcomes related to covid-19/SARS COV-2.We assessed the two quality measures (risk of bias and reporting) separately, using validated tools suited to our pre-specified study designs –21. ChoiFor methodological quality assessment, we assessed risk of bias at the level of pre-specified domains and at the level of the complete paper with a final overall grading of ‘low’, ‘high’ or ‘unclear’ risk of bias. Decisions on overall risk of bias were made by the rater pairs, informed by the domain level assessments. Overall risk was not defined by a threshold of number of individual domains that scored low or high risk; rather, overall risk of bias was considered on a paper by paper basis.For reporting quality, we assessed adherence with reporting guidance at an individual item level with reporting scored as ‘yes’ (where reporting was deemed adequate), ‘no’ or ‘not applicable’. This allowed calculation of proportional adherence  as the number of adequately reported items against the number of relevant items for the paper.random.org online resource [The team were all researchers with experience and training in meta-research. From a pool of 12 reviewers, we created six reviewer pairs, consisting of one experienced and one less experienced researcher. Each pair had recourse to the senior author (TQ) for advice or where there was disagreement on assessment. Assessor pairs were allocated a test set of six papers (covid-19 and non-covid) for review and calibration within pairs and within the group. Pairs were then randomly allocated eligible papers in blocks. Reviewer pairs extracted descriptive and outcomes data independently and compared results. In the case of disagreement, papers were discussed with a third reviewer (the experienced reviewer from another reviewer pair). As a validity check, a random selection of 10% of the included papers was selected for further review by an independent reviewer . We collected data on domain level disagreement requiring discussion for each assessor pair to calculate percentage disagreement within the group. All random allocation used the resource .Reviewers used standardised data extraction forms, piloted on two studies  , 26. We Our primary outcome was methodological quality, based on assessment of overall risk of bias for each included paper. Our co-primary outcome was reporting quality, based on adherence to reporting guidelines and quantified as proportion of relevant reporting items completed for each included paper.We locked the database on 11th of June when the last review was submitted, and quality control checks were complete. A statistician independent of the main review group conducted the analyses. Covid-19 status was numerically coded so that this variable was not obvious to the statistician.We tabulated descriptive statistics for covid-19 and non-covid research, comparing features of the included papers using non-parametric and proportional statistics as required. We collated data on inter-rater agreement from reviewer pairs to calculate summary reliability for the team.We created graphical illustrations of methodological and reporting quality at paper level and in aggregate . We creaWe compared proportions of ‘low risk of bias’ in covid-19 and non-covid research across all included papers. We fitted a mixed effects logistic regression describing the odds of study level low risk of bias, where the rater pairs were fitted as a random effect to control for the heterogeneity introduced by scorers. The multivariable analysis was adjusted for time of publication, funder (academic or industry), study results (positive or negative), study design and journal. Stata version 15 (StataCorp) was used for the primary quantitative analyses.n = 6 (11%) for covid-19 v n = 60 (53%), difference 42% (95%CI 28 to 53)) and more likely to use case series or other observational designs (n = 46 (85%) for covid-19 papers v n = 37 (32%), difference 52% (95%CI 37 to 63))  and shorter follow-up  ), to have an accompanying editorial ) and to have a retraction or major post-publication correction ). Covid-19 papers had smaller sample size  were rated as a low risk of bias. Of the non-covid studies, 83  were rated as low risk of bias, compared to 18  in the covid-19 group .p < 0.001). Bias differed between study designs. Compared to observational designs, RCTS were almost three times more likely to have low risk of bias . The time of publication, the journal of publication, the study design, study findings and source of funding were all not associated with study level risk of bias.Covid-19 study status was associated with study-level risk of bias. Low risk of bias was 6 times more common in the non-covid group than the covid-19 group  with none of the other covariates exhibiting evidence of association with risk of bias.In multivariable analysis, covid-19 papers remained associated with almost six times higher risk of bias  than covid-19 papers  . Low risk of bias in observational studies was also more common in non-covid studies  than in covid-19 studies  .All domains for observational studies and all but one domain for RCTs suggested lower quality scores in the covid-19 papers, although with small sample sizes not all reached significance at the conventional level Figs. a,b.Proportional adherence to reporting guidelines differed between covid-19 and non-covid research. Mean percentage adherence to reporting guidance for non-covid was higher  than for covid-19 studies , with a mean difference of 13% (95%CI 0 to 27).In analyses restricted to RCTs (CONSORT) and observational studies (STROBE), differences between covid-19 and non-covid research were apparent but did not reach statistical significance. For RCTs, mean adherence to reporting guidelines for non-covid studies was 90% and for covid-19 papers this was 87% . For observational studies, mean adherence for non-covid studies was 75% (95%CI 69 to 81) and for covid-19 studies was 69% (95%CI 63 to 75) .All reporting domains for observational studies suggested lower quality scores in the covid-19 papers, differences were less obvious for RCT reporting and small sample sizes preclude meaningful comparative testing at this level  and poorer reporting of the published covid-19 research. Given the time pressures to produce data and the novelty of the virus, some pragmatism can perhaps be allowed around research design, for example it seems intuitive that the first papers describing a new pathogen are more likely to be descriptive. However, there is no reason to sacrifice comprehensive, transparent reporting.In a time-sensitive publishing space, some may argue that publication standards may have been relaxed for publications on covid-19 during the first wave, with the anticipation that later papers would be more robust. We would argue that there is a scientific and ethical imperative to maintain standards of conduct and reporting. We explored the effect of time, and within our first wave sampling frame, we found that publication date did not have a major effect on quality. Many of the covid-19 papers were published as ‘Brief Report’ format. Limits on word count could compromise quality, but we did not find that format of submission explained variation in scores. Planning, conducting and reporting a trial take time and it seems intuitive that early covid-19 research may favour observational designs. However, our finding of lower quality in covid-19 was not solely driven by an excess of early observational studies. Observational studies were seen throughout the period of assessment, and within those observational papers, quality was poorer for covid-19 papers. Regardless our measures of quality and reporting were bespoke to the trial design and the imbalance of trials to observational data for covid-19 research will not fully explain the findings.To ensure a platform for the important emerging research, standards for covid-19 submissions may have been relaxed compared to non covid-19 submissions. It could be argued that we should be more tolerant of potential risk of bias when assessing covid-19 studies. We acknowledge that covid-19 research was at an early stage when we performed our review and methods may have been modified accordingly. For example, given the uncertain nature of covid-19 and potential treatments, full blinding may have been considered too risky by investigators and for observational studies it would be difficult to know the most important confounder variables to correct for in covid-19 analyses.We followed best practice guidance for meta-research and used validated and objective scoring tools. Within the time constraints of this rapid study, we created training and calibration processes to ensure consistency in assessment. It would not have been possible to mask our raters to the covid-19 status of the papers under review. Consciously or subconsciously, raters may have preferentially marked down covid-19 research. To mitigate this risk, we embedded multiple checks of internal validity throughout the review process.Our choice of quality assessment tool was based on explicit criteria and agreed by the group but we recognise that there are a variety of tools available with no consensus on the preferred approach . While oA selection bias is possible. We did not attempt a comprehensive analysis of all clinical research published in the first wave of the pandemic, as this would involve thousands of papers. Rather we selected exemplar, high profile, weekly clinical journals who had published substantial numbers of covid-19 research papers. Although the early covid-19 research was often shared with these journals, we recognise that our included papers represent only a fraction of the totality of covid-19 research published in the first wave.Our study followed a pre-specified and publicly available protocol and was designed to assess a selection of covid-19 papers published during the first wave of the pandemic. Other approaches to an assessment of methodological and reporting quality are plausible. For example, we anticipated an imbalance of trials to observational research methods in covid-19 and non-covid research and did not attempt a matched case-control design where the only difference was the covid-19 subject matter. It could be argued that the ideal method to assess the quality of covid-19 research would be to compare the covid-19 papers with contemporaneous research describing an established but similarly infectious and dangerous viral agent. While theoretically possible, these designs would have been almost impossible to deliver in practice during the first wave.Quality of science is more than just valid methods and transparent reporting. Indeed, in the covid-19 context, other important factors like inclusiveness, data sharing and clinical urgency may take precedence over rigid rules on method and reporting. Even within our chosen remit of methodological quality, there are many facets. Some of our tools have a strict focus on internal validity (bias) while others also consider external validity . Accepting all this, we do not claim to have described a definitive measure of overall study quality, but we have described and quantified fundamental aspects of conduct and reporting that should be maintained in all scientific publications. We believe this is especially true when the topic matter is so important to health and society.The volume of published clinical research is constantly increasing, even more so, if we consider the pre-print servers. It would not have been possible to assess the entirety of the biomedical literature published during the first wave of the pandemic. We limited to those journals with the greatest clinical impact and considered to have the highest standards. If there are concerns around quality in these four flagship journals, it seems likely these issues will also permeate other journals. There is research to support this view, studies of research methods, for example prognostic modelling  and pharThe story of hydroxychloroquine as a potential covid-19 treatment is a pertinent example of the need for rapid communication of science and the attendant risk of sharing potentially biased research. Based on predominantly observational studies, the drug achieved substantial visibility and entered clinical practice . FurtherQuality concerns are not an exclusive covid-19 phenomenon, and it is worth noting that our non-covid papers also had biases. Issues with reporting, which were prevalent in both covid-19 and non-covid studies, seem less forgivable. Although differences in reporting quality were not of the same magnitude as differences in methodological quality, if anything, where research is produced at speed to answer an urgent clinical question, then the need for comprehensive and fully transparent reporting is even greater. All the journals assessed in this analysis adhere to CONSORT recommendations for RCTs and completion of a CONSORT checklist is mandatory. Experience tells us that mandating a checklist is not a panacea. Since the introduction of CONSORT, there have been improvements in trial reporting, but poor reporting remains prevalent in contemporary research , 41.We offer only a snapshot of the covid-19 evidence base from the first wave of the pandemic. Our analyses of the effects of time are limited by the relatively short period within which we selected papers. As we move into second wave of the pandemic and beyond, it would be interesting to repeat our analysis looking for longer term temporal trends in quality. Even within the period of this review, new concerns around research publications are emerging. Using our approach to look at pre-prints, inclusiveness or data sharing arrangements would all be informative. We assessed various factors that may be associated with research quality, but recognise that many other plausible factors exist, some specific to covid-19 and some common to all clinical research .Covid-19 research published in major journals during the first wave of the pandemic had methodological and reporting issues that ultimately compromise the utility of the research and may cause harm. The clinical response to covid-19 has seen many examples of sacrifice and tragedy. In clinical science we must not sacrifice research quality in the race to publish data and it would be further tragedy if current researchers and publishers do not learn from the first wave of covid-19 research.Additional file 1: Supplementary text S1. Description of Study team. Supplementary text S2. Categories of research design assessed in the project. Supplementary text S3. Data dictionary for ‘included studies’ master sheet. Figure S1-S4 Traffic light visual summary of risk of bias (ROB). S1a) Traffic light summary of ROB for RCTs at individual study level. S1b) Traffic light summary of ROB for RCTs as aggregate scores. S2a) Traffic light summary of ROB for observational studies at individual study level. S2b) Traffic light summary of ROB for observational studies at aggregate level. S3a) Traffic light summary of ROB for test accuracy studies at individual study level. S3b) Traffic light summary of ROB for test accuracy studies at aggregate level. S4a) Traffic light summary of ROB for systematic reviews at individual study level. S4a) Traffic light summary of ROB for systematic reviews at aggregate level. Figure S5. Reporting guideline adherence (CONSORT) for RCTs. Figure S6. Reporting guideline adherence (STROBE) for observational studies. Figure S7 Reporting guideline adherence (STARD) for test accuracy studies. Figure S8 Reporting guideline adherence (PRISMA) for systematic review. Figure S9 Modified star plot describing overall and individual item level reporting adherence for STROBE  reporting. Figure S10 Modified star plot describing overall and individual item level reporting adherence for CONSORT (RCT) reporting.


This case series analyzes whether supplements categorized as containing novel ingredients in the National Institutes of Health Dietary Supplement Label Database are accurately labeled. In an effort to provide a resource for researchers, clinicians, and the public, the National Institutes of Health (NIH) maintains an online searchable database of supplement labels.2 To our knowledge, whether supplements categorized as containing novel ingredients in the NIH database are accurately labeled has not been previously studied.Dietary supplements are estimated to be responsible for tens of thousands of emergency department visits each year in the US.3 on the Dietary Supplement Ingredient Advisory List to alert the public that it does not appear to be a lawful supplement ingredient.4 We analyzed the actual contents of supplements categorized as containing 5-alpha-hydroxy-laxogenin in the NIH database to determine whether they are accurately labeled.In October 2019, the US Food and Drug Administration (FDA) placed 5-alpha-hydroxy-laxogenin, a synthetic analog of a plant steroid that is marketed as a natural alternative to anabolic steroids,2 were purchased online in November 2018 and analyzed from January to June 2019 for their chemical contents  according to previously described methods . The statistical prerequisite established was the determination of goodness of fit  to conclude that linear calibration lines were fit for estimation purposes within the chosen concentration interval.In this case series, supplements categorized as containing 5-alpha-hydroxy-laxogenin in the NIH Dietary Supplement Label DatabaseA total of 6 supplements were categorized in the NIH database as containing 5-alpha-hydroxy-laxogenin; 4 (67%) were available for purchase . In total, 16 ingredients, excluding expedients, minerals, and plant species, were declared on the 4 product labels.Compared with their actual chemical contents, no product was accurately labeled . Of the Only 1 product (25%) contained 5-alpha-hydroxy-laxogenin, an ingredient with purported anabolic effects in rats that, to our knowledge, has not been studied in humans. The supplements contained phenibut, an unapproved drug with abuse potential that is used in Russia to treat a variety of neurologic conditions; arimistane, a designer steroid that may function as an aromatase inhibitor; 7-keto dehydroepiandrosterone, a minor metabolite of dehydroepiandrosterone; ecdysterone, a steroid-like compound found in many insects and plants that may have anabolic effects in humans; and diosgenin, a steroid-like compound found in trace amounts in several plants and synthetically produced for use in the industrial production of some commercial steroids . None of2 However, this statement does not accurately capture the breadth of gaps in the database, including supplements that contain ingredients not listed on the label, do not contain ingredients declared on the label, provide inaccurate quantities of ingredients on the label, and contain prohibited ingredients.Supplements categorized in the NIH supplement database as containing 5-alpha-hydroxy-laxogenin were inaccurately labeled and contained a variety of potentially pharmaceutically active compounds with unpredictable health effects. The NIH database contains the disclaimer that “label information has not been verified or checked for conformity with existing US Food and Drug Administration (FDA) regulations.”6Our study was limited because we analyzed only supplements categorized as containing 1 new ingredient at 1 point. Ingredients may vary over time within the same product.Our analysis suggests that consumers and clinicians cannot obtain accurate information on novel supplement ingredients with the NIH supplement database. Future efforts to design and update supplement databases should take steps to assure accurate results.


Successful embolization of a basilar bifurcation aneurysm associated with a persistent primitive hypoglossal artery (PPHA) using Y-stent-assisted coiling. Carotid–vertebrobasilar anastomoses, known as presegmental arteries in the embryonic period, supply blood from the internal carotid artery (ICA) to the primitive vertebrobasilar system . PersistWe report a patient with an unruptured wide-neck basilar bifurcation aneurysm accompanied by right PPHA. Although vertebral arteries were present, both were hypoplastic. We advanced a microcatheter via the PPHA, with the help of Y-stent-assisted coiling and successfully treated this aneurysm. Follow-up 6 months after the endovascular treatment using brain digital subtraction angiography (DSA) showed no recanalization of the aneurysm. To our knowledge, the use of Y-stent-assisted coiling to treat PPHA accompanied by basilar bifurcation aneurysm has not been reported previously. Here, we describe the details of the patient's clinical course with a brief review of the relevant literature.2 and she weighed 78 kg; she had no family history of intracranial aneurysm. Magnetic resonance imaging (MRI) detected a basilar bifurcation aneurysm  was 32.5 kg/maneurysm . During aneurysm . DSA demaneurysm  measurinFor such complex wide-neck bifurcation aneurysms, the Woven EndoBridge  system is a good choice –14. BecaIn this patient, the left vertebral and posterior communicating arteries (PcomAs) were hypoplastic ; therefoDual-antiplatelet therapy  was administered 3 days prior to the operation. Under general anesthesia, a 6-Fr sheath was introduced into the right femoral artery; then, we guided a 6-Fr Cook shuttle  to the C1 segment of the right ICA. Next, we introduced a 6-Fr Navien  intracranial support catheter to the V2 segment of the right vertebral artery (VA). A Headway 17 microcatheter  was navigated over a Traxcess 14 guidewire (MicroVention Inc.) into the P1 segment of the right posterior cerebellar artery (PCA). Next, with the help of another Traxcess 14 guidewire, we navigated an Echelon 10 microcatheter  into the lumen of the aneurysm. To preclude coil detachment, we “wove a basket” with a coil ; the firPPHA is a primitive communication between the ICA and the BA. Normally, the embryonic anastomotic vessels disappear after the formation of the posterior communicating artery (PcomA). When a segmental anastomotic branch does not disappear and persists to adulthood, it is called a persistent carotid–vertebrobasilar anastomosis. In 1889, the first report of PPHA was presented by Batujeff . Begg fiThe anatomic and angiographic definition of PPHA is according to the following diagnostic criteria: (1) the PPHA originates from the cervical segment of the ICA at the CI–C3 level; (2) the PPHA along with the twelfth cranial nerve enters the skull via traversing the hypoglossal canal, joining with the BA at the level of the occipital foramen; (3) the BA is filled only beyond the point where the artery joins it; and 4) the absence of a PcomA, and the ipsilateral VA is hypoplastic .PPHA is usually asymptomatic, and most cases are encountered incidentally during angiography. Nonetheless, the most common clinical characteristic of PPHA is aneurysm . AlthougOur literature review identified several reports of the treatment of PPHA accompanied by a basilar bifurcation aneurysm  3, 5–7,,7, 18–21Surgical clipping and endovascular treatment remain the predominant modality in the management of intracranial aneurysms , 27. ManTo our knowledge, ours is the first report of successful embolization of a basilar bifurcation aneurysm associated with a persistent PPHA treated with Y-stent-assisted coiling.In this case, both clinical outcomes and short-term follow-up findings were encouraging. Y-stent-assisted coil embolization for an unruptured basilar bifurcation aneurysm via PPHA is feasible. However, long-term observation will be required for this single case.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by Beijing Tiantan ethics committee. The patients/participants provided their written informed consent to participate in this study. Written informed consent was obtained from the individual(s) for the publication of any potentially identifiable images or data included in this article.LZ and XC performed the manuscript writing. LJ acquired the data. LD and JW analyzed and interpreted the data. LZ edited the figure of the article. ML and PL conceived and designed the research. All authors contributed to the article and approved the submitted version.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.


The therapeutic capacity of mesenchymal stem cells  depends on their ability to respond to the need of the damaged tissue by secreting beneficial paracrine factors. MSCs can be genetically engineered to express certain beneficial factors. The aim of this systematic review is to compile and analyze published scientific literatures that report the use of engineered MSCs for the treatment of various diseases/conditions, to discuss the mechanisms of action, and to assess the efficacy of engineered MSC treatment.We retrieved all published studies in PubMed/MEDLINE and Cochrane Library on July 27, 2019, without time restriction using the following keywords: “engineered MSC” and “therapy” or “manipulated MSC” and “therapy.” In addition, relevant articles that were found during full text search were added. We identified 85 articles that were reviewed in this paper.Of the 85 articles reviewed, 51 studies reported the use of engineered MSCs to treat tumor/cancer/malignancy/metastasis, whereas the other 34 studies tested engineered MSCs in treating non-tumor conditions. Most of the studies reported the use of MSCs in animal models, with only one study reporting a trial in human subjects. Thirty nine studies showed that the expression of beneficial paracrine factors would significantly enhance the therapeutic effects of the MSCs, whereas thirty three studies showed moderate effects, and one study in humans reported no effect. The mechanisms of action for MSC-based cancer treatment include the expression of “suicide genes,” induction of tumor cell apoptosis, and delivery of cytokines to induce an immune response against cancer cells. In the context of the treatment of non-cancerous diseases, the mechanism described in the reviewed papers included the expression of angiogenic, osteogenic, and growth factors.The therapeutic capacity of MSCs can be enhanced by inducing the expression of certain paracrine factors by genetic modification. Genetically engineered MSCs have been used successfully in various animal models of diseases. However, the results should be interpreted cautiously because animal models might not perfectly represent real human diseases. Therefore, further studies are needed to explore the translational potential of genetically engineered MSCs. Mesenchymal stem cells (MSCs) or mesenchymal stromal cells are adult multipotent stem cells that can be differentiated to other cell types such as bones, cartilage, skeletal muscles, and connective tissues. MSCs can be isolated from various sources, including bone marrow, adipose tissue, and the mucoid tissue within the umbilical cord (Wharton’s jelly) . A largeThis systematic review complies with the Preferred Reporting Items for Systematic Review (PRISMA) guidelines. The protocol was submitted to the PROSPERO database of systematic review protocols (registration number: CRD 42020163707). We have checked the Cochrane Library to ensure that there was no systematic review on a similar topic.We performed a PubMed/MEDLINE and Cochrane database search for relevant published studies on July 27, 2019, without time restriction using the following search terms: “engineered MSC” AND “therapy” or “manipulated MSC” AND “therapy.” In addition, relevant articles found during full text search were reviewed.All original articles on the therapeutic use of engineered MSC or manipulated MSC were included. Studies on engineered MSC or manipulated MSC therapy that did not provide data on the treated conditions or disease models, type of MSC manipulation, detail information on expressed protein, administration route, and outcome of the treatment were excluded from this review. Article selection was conducted by examination of the titles, abstracts, and full texts.The data collected include the following: the treated disease or disease models, the species used, the number of animals/participants, the source of the MSC, MSC passage, the type of gene insertion and expressed protein, the outcomes of the treatment, adverse events, and mechanism of action.Data analysis: the studies were grouped and tabulated according to the treated conditions/disease models, species, number of animals, MSC source and passage, vector used, types of gene insertion/expressed proteins, outcome, adverse event, and mechanism of action.Data were analyzed descriptively to determine the safety and efficacy of a certain engineered MSC on a certain disease and the possible mechanism.Using the keywords “engineered MSC” AND “therapy,” we identified 26 articles from the PubMed database, of which 17 articles were included for review. Six articles from the Cochrane Library database were identified using the same keywords (of which one was selected for review). Using the keywords “manipulated MSC” AND “therapy,” we identified 50 articles from the PubMed database, of which six papers were selected for review. The same keywords were used in the Cochrane database, and we have identified five articles, but none of them met the inclusion criteria for review. During full text search, we found 61 relevant articles that met the inclusion criteria, and thus, the total number of articles included in the review was 85 .The conditions/disease models treated by engineered MSC were grouped into two main groups: (i) tumor, cancer, malignancy, and metastasis  and (ii)Engineered MSCs were mostly used to treat tumor/cancer/malignancy/metastasis  by inserting suicide gene carriers that will activate a non-toxic pro-drug to become a cytotoxic substance that can kill the tumor cells, (ii) by using MSCs as vehicles to deliver cytokines for cancer immunotherapy; and (iii) by using MSCs as agents to induce tumor cell apoptosis .One of the main challenges of the current first-line cancer therapies, especially chemotherapy, is systemic cytotoxicity, and limited delivery to the tumor sites. To tackle these problems, genetically modified MSCs carrying suicide genes have been employed to convert non-toxic pro-drugs into active agents for selective elimination of cancer cells. Various genes encoding these “suicide proteins” were used: cytosine deaminase (CD) or cytosine deaminase-uracil phosphoribosyltransferase (CD-UPRT), which converts the pro-drug 5-fluorocytosine (5-FC) into an active agent 5-fluorouracil (5-FU) ; herpes in vitro and in vivo models (A study showed that human AT-MSCs expressing cytosine deaminase (CD) could trigger dose-dependent apoptosis of human melanoma A375 cells in both o models . Subcutao models . MoreoveOther pieces of evidence showing the promising therapeutic potential of MSCs in suppressing melanoma tumors have been provided by a study using a more aggressive variant of melanoma cells, that is, EGFP-A375/Rel3, which exhibited altered cell adhesion and tumorigenic and metastatic properties . The comAs mentioned previously, one major issue on the use of MSCs in combination with chemotherapeutic substances is the toxicity of the chemotherapeutic drugs on the MSCs. MSCs have been shown to have some degree of resistance against alkylating agents such as cyclophosphamide, melphalan, and busulfan . It has In addition to the application in delivering “suicide genes,” engineered MSCs have been used as tools to deliver anti-cancer cytokines to the tumor local environment. Thus, treatment using engineered MSCs has been regarded as one important approach in the field of cancer immunotherapy In a study on 786-0 renal cancer cell xenografts, MSCs expressing interleukin-12 could migrate to the tumor site and inhibit tumor growth through the activation of the natural killer cells and secretion of interferon γ at 14 days after MSC administration . In a moin vitro  subcutaneous tumor in NOD/SCID mice and induce 65% tumor regression without causing liver toxicity compared to 42.7% of tumor regression in groups receiving MSC-TRAIL only .So far, most studies using pre-clinical cancer models have shown consistent results, i.e., the engineered MSCs could inhibit tumor growth and enhance the survival rate of the tumor-bearing animals. However, a recent clinical report of a patient treated with engineered MSC-CD-UPRT and 5-FC for metastatic head and neck cancer has shown that there was no sign of tumor regression at 6 days after intravenous injection of the therapeutic stem cells. Moreover, tumor metastasis further progressed after 40 days of the treatment . TherefoFormation of new blood vessels (angiogenesis) has been regarded as one of the key factors in the progression of cancers. Treatment with anti-angiogenetic agents appears to become one promising approach in cancer therapy. MSCs have been considered as effective vehicles to deliver such factors. Several lines of evidence have shown the use of genetic modification to generate MSCs that are capable of sending anti-angiogenetic factors to the tumor sites. For example, MSCs overexpressing endostatin have been reported to be effective in controlling the growth of ovarian cancer. It was shown that the beneficial effect was likely due to the inhibition of neovascularization, enhanced apoptosis, and inhibition of cell proliferation . EquallyAnother strategy in the development of anti-cancer MSCs is by using these cells to deliver oncolytic adenovirus. One example of this strategy is the use of MSCs to carry and deliver the Delta-24-RGD adenovirus, which has a potent anti-glioma property . Experimin vivo can induce the growth of the tumor cells. This has been observed in models of ovarian  to increase the median survival time of the animals receiving organ transplantation. Foxp3 caused an increase in Treg level, which has a strong immunomodulatory effect. The increase was T cell contact dependent and was associated with the upregulation of programmed death ligand 1 (PD-L1) expression in MSC . AnotherEngineered MSCs have been used in a study using a mouse model of experimental autoimmune encephalomyelitis (EAE). Wang and co-workers generated engineered MSCs that secrete nerve growth factor (NGF) and used them to treat nerve injury in a rabbit model. They showed that treatment with these cells (MSC-NGF) increased the formation of nerve fibers and the density of myelinated nerve fibers. It also decreased the deposition of myelin debris compared to control animals. The study demonstrated that the MSC-NGF cells enhanced the nerve regenerative capacity .Another study used stromal cell-derived factor-1 (SDF-1) secreting MSCs (MSC-SDF-1) to treat full thickness wound in a rat model. Wound closure and blood vessel formation were significantly improved in the MSC-SDF-1-treated group. Animals treated with non-engineered MSCs have also shown improvement compared to the control group, although not as significant as in the MSC-SDF-1-treated group. SDF-1 increased vascular endothelial growth factor (VEGF) expression, which induced angiogenesis. Furthermore, there was an increase in dermal fibroblast migration in treated animals .The study by Using a mouse model with acute kidney injury, a study used lipocalin-2-secreting MSCs (MSC-Lcn2). The results showed that the number of cast and tubular necrosis was significantly decreased compared to mice treated with non-engineered MSCs and with the control groups. At day 21 following treatment, the serum creatinine level in the MSC-Lcn2-treated group decreased significantly to a level comparable to those of normal mice. The blood urea nitrogen level was also decreased, although the level was still higher than normal. Lipocalin-2 has been shown to upregulate the expression of various growth factors, such as hepatocyte growth factor (HGF), insulin-like growth factor, fibroblast growth factor, and VEGF. All of them have the capability to trigger kidney regeneration. This was indicated by the increase in the expression of kidney regenerative markers  and the decreased expression of kidney injury markers such as KIM-1 and cystatin C .Our systematic literature search has indicated four studies on the use of engineered MSCs to treat myocardial infarction. One study was conducted using a mouse model, and the others used a rat model. The first study used VEGF-secreting MSCs and showed that treatment with these cells improved heart function compared to the control group. It was reported that the improvement was likely due to the increased in capillary density because VEGF is known as a potent angiogenic factor .Two studies used engineered MSCs that produce and secrete CXCR4 (MSC-CXCR4) . In a stThe fourth study in the field of myocardial infarction indicated by our literature search involved the use of engineered MSCs overexpressing Bcl2 (MSC-Bcl2). Bcl2 is known as a potent anti-apoptotic agent. In this study, Li and co-authors demonstrated that treatment with MSC-Bcl2 resulted in the improvement of heart function following myocardial infarction compared to the control group. Bcl2 was shown to induce cell engraftment and VEGF expression level, which, in turn, increased angiogenesis and eventually improved heart function .We found 13 studies on the use of genetically engineered MSCs for the treatment of various types of bone defects . In mostA study examined a combination treatment of MSC-BMP2 with the addition of various tissue scaffolds, such as ultrapure alginate, alginate, arginine–glycine–aspartic acid (RGD)–alginate, and collagen-1 . The autSeveral other studies examined the differential effects of the transient and constitutive expression of the BMP2 and VEGF combination in the MSCs. The data indicated that the constitutive expression of the osteogenic factor showed better therapeutic effects , 2012a.BMP2-engineered MSCs seemed to produce better therapeutic effects compared to TGF-β3-engineered MSCs in a rabbit model of calvarial bone defect. A study reported that implantation of BMP2-engineered MSCs promoted bone formation and significantly improved bone volume and bone density. The better therapeutic effect of these cells was likely due to the ability of BMP2 to promote osteogenesis and chondrogenesis as well as to stimulate the formation of the dura mater, which is important in calvarial bone healing. The study also compared the use of two types of tissue scaffolds, i.e., poly(L-lactide-co-glycolide) (PLGA) and gelatin. It was shown that the gelatin sponge produced better chondroinductive ability than PLGA and subsequently improved bone healing .A recent review from Freitas et al. summarized the diverse approaches in using genetically modified MSCs for the treatment of critical-size, non-union, and delayed bone defects. Typically, the MSCs were engineered with genes responsible for osteogenesis , angiogenesis , anti-apoptosis , or non-coding RNA (microRNAs). The MSCs can be implanted in the lesions, or alternatively, the secretomes can be given either locally or systemically. The critical questions in utilizing engineered MSCs are how to ensure the safety of the genetic modification and how to standardize the end products of the therapy .A study showed that MSCs overexpressing angiotensin-converting enzyme 2 (ACE 2) produced a better therapeutic effect in a bleomycin-induced lung fibrosis model in mice . Mice trAnother condition that has been successfully treated by genetically engineered MSC is radiation-induced toxicity. A study using a mouse model receiving total body irradiation showed that genetically modified MSCs overexpressing extracellular superoxide dismutase (ECSOD) could alleviate the toxic effects of the radiation, resulting in an increase in the 35-day survival rate to 52%, which was significantly better compared to control (10%). The mechanism of action was likely due to the improved scavenging process of the superoxide anion by ECSOD, which might prevent hematologic toxicity .Similarly, a study using a radiation-induced lung injury model in mice demonstrated that treatment with MSCs expressing soluble transforming growth factor-ß type II receptor (sTGF-ß-R) resulted in significantly higher survival (80%) compared to mice treated with native MSCs , and none of the control animals (no MSC treatment) survived. It is believed that the fibrinolytic effect of MSC in combination with the anti-inflammatory effect of sTGF-ß-R is responsible for preventing inflammatory cell infiltration, pro-inflammatory cytokine secretion, and collagen overproduction in mice treated with MSC-sTGF-ß-R .In keeping with those results, another study using a mouse model of radiation-induced lung injury analyzed the use of MSCs overexpressing CXCR4. The genetically engineered MSCs displayed better performance compared to native MSCs as CXCR4 expression facilitated MSC migration and homing to the injury sites. CXCR4 is a receptor for stem cell stromal cell-derived factor-1 (SDF-1), and the activation of CXCR4 by SDF-1 modulates the signaling pathway important for stem cell migration and homing .Two other studies related to radiation-induced toxicity involved the use of HGF. Wang et al. showed that MSCs expressing HGF showed a significantly better therapeutic capacity in alleviating radiation-induced intestinal injury compared to native MSCs. It is thought that the regenerative and anti-apoptotic effects of HGF might be the responsible factors in enhancing intestinal epithelial cell regeneration . MSCs ovHuntington’s disease is caused by mutation in the huntingtin (HTT) gene resulting in the production of mutant huntingtin (mutHTT) protein. mutHTT may induce the death of the medium spiny neurons in the brain striatum. The mechanism of neuronal cell loss involves the reduction of neurotrophic factors, such as brain-derived neurotrophic factor (BDNF). A study using a mouse model of Huntington’s disease demonstrated that treatment with MSCs overexpressing BDNF and/or NGF produced better therapeutic effects compared with treatment with naive MSCs .A study on rat and mouse models of arthritis showed that genetically modified MSCs expressing human soluble tumor necrosis factor receptor type 2 (hsTNFR) could partially prevent arthritis symptoms and produced better therapeutic effects than naive MSCs. However, MSC-based treatment was still less effective than treatment with dexamethasone. In addition to the intrinsic anti-inflammatory properties of the MSCs, the expression of hsTNFR might inhibit the pro-inflammatory effects of TNF-α. hsTNFR could bind and decoy TNFα and thus block the TNF-α-mediated inflammatory effect .More recently, MSCs expressing HGF were reported for the treatment of rheumatoid arthritis in mice. The study showed that MSC-HGF could alleviate arthritis symptoms, but at the later phase of the disease, the effect was not better than treatment with naive MSCs. HGF at the early phase of the disease displayed an immunosuppression effect, but at a later phase, HGF induced the activation of fibroblast-like synoviocytes. These cells had the capacity to produce Il-6, which induced inflammation, cell proliferation, and decreased apoptosis .Engineered MSCs have been used successfully to treat various diseases and conditions in animal models. However, it is important to note that animal models might not be similar to human diseases/conditions. Therefore, the results should be interpreted cautiously as a translational study in human so far did not show any positive result. Nevertheless, MSC-based therapy is still a promising new therapeutic modality for various conditions, and it is clear that enhancement of its therapeutic effects can be achieved by inducting the expression of beneficial paracrine factors. Further studies are still needed to accelerate the pre-clinical findings into clinical application.JP conceived the idea, performed the literature search and data entry on tumor/malignancy treatment, analyzed and compiled the data, and wrote the manuscript. TB performed a review on cancer treatment. WM performed a review on engineered MSCs to prevent rejection in organ transplantation, treatment of autoimmune encephalomyelitis tissue injuries, ischemia, and myocardial infarction. RA performed a review on a metastatic cancer model. RN performed the data entry and a review on engineered MSCs for bone regeneration. ID conceived the main idea and performed a review on clinical trials. DO conceived the idea, compiled the data, and wrote and edited the manuscript. All authors contributed to the article and approved the submitted version.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.


The mortality of COVID-19 patients with cerebrovascular events can be as high as 38%.2 As of this writing, there has been no reported case of a COVID-19 patient with concurrent acute ischemic and hemorrhagic strokes.Cerebrovascular disease has been described as a potential sequela of the novel coronavirus disease of 2019 (COVID-19) with one study reporting its occurrence in 106 patients. In this rapid review, 83% of the patients had ischemic strokes, while 17% were hemorrhagic.We report a case of a 64-year-old male, hypertensive and smoker, who came to our emergency department for sudden onset right-sided weakness, numbness, and dysarthria. There was no history of trauma or respiratory symptoms prior to admission. He had no prior use of antithrombotics. He works as a utility worker in the designated COVID-19 area of the hospital. On examination, he had a blood pressure of 150/90 with a normal heart rate and regular rhythm right superior quadrantanopia, right hypesthesia, and right hemiparesis. His National Institute of Health Stroke Scale (NIHSS) score was 7. A 12-lead electrocardiogram showed sinus rhythm. A plain cranial CT scan revealed an acute infarct on the left thalamus and left temporo-occipital lobe, as well as a small subarachnoid hemorrhage on the left parietal sulcus (Figure 1 The virus can bind to the ACE-2 receptors of the cerebrovascular endothelium and causes endothelial damage and induces cytokine storm, which can then lead to vasculitis and subsequent stroke.3 Endothelial dysfunction and the consequent hyperinflammatory reaction may also increase the risk of plaque rupture and thrombosis.4 A hypercoagulable state is also another mechanism by which the virus can cause stroke, as evidenced by the increase in D-dimer seen in our patient.6 These mechanisms may explain why our patient had ischemic stroke. The disease is also associated with a downregulation of the ACE-2 receptors, elevated angiotensin II levels, and aforementioned endothelial damage, which can cause an increase in blood pressure and subsequently increase the risk for hemorrhagic stroke.4 The presence of other well-known risk factors for stroke, such as hypertension and smoking in our patient may have placed him at a greater risk for acquiring COVID-19 and developing cerebrovascular disease.Recent studies reported that COVID-19 increases the risk of cerebrovascular diseases. The exact mechanism by which the virus can involve the nervous system is still unclear. However, the proposed pathophysiology suggests that it may be a multifactorial process. Possible routes of SARS-CoV-2 in the cerebrovascular system include the olfactory epithelium and the respiratory tract with subsequent viremia.7 However, the co-occurrence of acute ischemic stroke and non-aneurysmal SAH has not been documented previously. The location of the patient’s subarachnoid hemorrhage is far from the infarcted tissue, making the diagnosis of a hemorrhagic infarct or a delayed cerebral ischemia from vasospasm unlikely. This suggests the possibility that the proposed mechanisms in COVID-19-related stroke can happen simultaneously, predisposing the patients to both types of stroke. Whether this finding is associated with COVID-19 or just coincidental is yet to be elucidated in future researches.Non-aneurysmal SAH has previously been reported in a COVID-19 patient.


A 43-year-old man was introduced to us for the treatment of an infected wound after below-knee (B-K) amputation . Four moHow can this stump be treated?What is the preferred level of amputation?What are the risks of preserving the proximal lower-leg region?What measures are available for preventing the development of pressure injuries?The opened wound was washed and treated with silver sulfadiazine cream. On the 17th postoperative day, we initiated negative pressure wound therapy (NPWT) at −125 mm Hg, under a diagnosis of disappearance of infection. After 8 days of NPWT, the wound showed good granulation. Successive treatment using basic fibroblast growth factor (bFGF) and bucladesine sodium ointment was performed to promote capillary formation and accelerate wound healing. We observed epithelialization of the ulcer on 35th postoperative day. Considering the bone stump was covered only by scar, we initially used an Iceross Dermo cushion liner ; however, the stimulation caused by the loss of soft tissue made continuous wearing difficult. Thus, the liner was changed to a 6Y92 Copolymer Liner , which he was able to wear continuously. Next, a patella tendon-supporting load-relieving brace was made; however, the stump moved in the socket, resulting in skin damage. In contrast, an all-surface loading prosthesis fit well . At 18 mSurgeons need to choose whether to perform A-K amputation, to achieve early wound resolution, or to preserve the B-K limb, which results in a better function. In B-K amputation, covering a tibial stump with well-vascularized soleus and gastrocnemius muscles is important for preventing the development of pressure ulcers and preserving the knee joint function.We were anxious about 2 risks when preserving the proximal lower-leg region. The first risk was that the preserved knee would lose its function and cause increased instability during walking. A-K amputation is associated with high energy consumption and difficult rehabilitation, even in younger patients.,The second risk was the potential for the development of pressure ulcers on the tibial stump because of lost muscle coverage. The management of stumps has been improved with recent silicon liners. The sleeve-like structure lined with silicone inside can reduce friction between skin and socket while maintaining an excellent suspension action.5Even if all of the lower-leg muscles are lost, we can preserve a functional lower leg. It is necessary to understand the advantages and disadvantages of active preservation of lower-limb length.


Borrelia burgdorferi, is the most common tick-borne infection in many regions of Eurasia. Antibody detection is the most frequently used laboratory test, favoring a two-step serodiagnostic algorithm; immunoenzymatic detection of antibodies to C6 has been shown to perform similarly to a standard two-step workflow. The aim of this study was the performance evaluation of the C6 Lyme ELISA kit compared to a standard two-step algorithm in three laboratories located in the northeastern region of Italy which cater to areas with different LD epidemiology. A total of 804 samples were tested, of which 695 gave concordant results between C6 testing and routine workflow . Wherever available, clinical presentation and additional laboratory tests were analyzed to solve discrepancies. The C6 based method showed a good concordance with the standard two-step algorithm (Cohen’s κ = 0.619), however, the distribution of discrepancies seems to point towards a slightly lower specificity of C6 testing, which is supported by literature and could impact on patient management. The C6 ELISA, therefore, is not an ideal stand-alone test; however, if integrated into a two-step algorithm, it might play a part in achieving a sensitive, specific laboratory diagnosis of LD.Lyme disease (LD), caused by infection with  Borrelia burgdorferi sensu lato spirochetes, which are transmitted by the bite of ixodid ticks where Ixodes ricinus is the main vector found in Europe [Lyme disease (LD) is the most common tick-borne infection in many regions of Eurasia . It is cn Europe .B. burgdorferi DNA in samples such as skin biopsy or blood specimens would be an ideal choice in terms of specificity, the sensitivity of molecular testing is not sufficient for a negative result to rule out infection, especially in the cases of suspected neuroborreliosis [Lyme borreliosis can be diagnosed through a combination of clinical observations and laboratory testing. The presence of erythema migrans (EM), an expanding circular skin rash, is considered sufficient to diagnose early Lyme disease in known endemic areas. However, later manifestations of LD are less specific, so highly reliable laboratory tests are necessary to support the diagnosis of suspected late LD . Althougreliosis ,4.Infection with other spirochetes or Epstein-Barr virus (EBV) and conditions of altered immunological reactivity, such as autoimmune diseases, may give false-positive results due to cross-reactions, an issue that was especially pronounced in first-generation LD diagnostic tests. The use of recombinant antigens or synthetic peptides has improved both the sensitivity and specificity of LD serological testing, but the issue is not yet solved .To this purpose, laboratory testing in the USA  and in pB. burgdorferi, which has been proposed as a diagnostic antigen in enzyme-linked immunosorbent assay (ELISA) tests, both in its full-length form and the invariable region 6 (IR6) portion. Over the years, EIA detection of antibodies to C6, a synthetic peptide that reproduces the sequence of IR6, has been shown to perform similarly to standard 2-step workflow [The variable surface protein VlsE is an immunogenic molecule of workflow ,10,11,12Borrelia burgdorferi, compared to a standard two-step algorithm in three Italian laboratories which cater to areas with different LD epidemiology: the Romagna area, where LD reporting is scarce, but the presence of infected ticks has been ascertained [The aim of this study was the performance evaluation of the C6 Lyme ELISA kit  for the presumptive detection of IgG and IgM antibodies against ertained ; the Giuertained ; the Altertained . A seconThe study was conducted on serum specimens deriving from the routine diagnostic procedures and collected from January to October 2019. The laboratories involved were the Great Romagna Area Hub Laboratory, Unit of Microbiology for the Romagna region, the Azienda Sanitaria Alto-Adige-Bolzano Microbiology Laboratory for the Alto Adige region, and the Azienda Sanitaria Universitaria Integrata di Trieste (ASUIT) Laboratory for the Giuliano-Isontina region.B. burgdorferi IgM and IgG antibodies, of which at least 50 were from patients presenting potential interfering factors that could potentially give false-positive results due to cross-reactions . Testing for interfering factors was conducted through using VCA IgM serology.The samples were anonymized using the current procedure  checked by the local Romagna Ethical Board, keeping exclusively relevant data for the purpose of this study . Each of the three laboratories involved in the study collected at least 200 samples routinely tested for anti-All three laboratories followed the same 2-step routing algorithm, used as a reference: a first-tier immunoassay with Liason XL CLIA  and a confirmation of positivity using recomLine Borrelia IgG/IgM . Serum samples were then tested with the C6 Lyme ELISA kit .Borrelia antibodies to the 2-tier algorithm, whenever available. This was done to better evaluate discordant results between the standard workflow and C6 testing, as the Giuliano-Isontina region is characterized by the highest rate of LD prevalence in Italy.While the specimens collected at the Great Romagna Area Hub Laboratory and the Azienda Sanitaria Alto Adige-Bolzano Microbiology Laboratory were selected consecutively amongst all samples with clinical suspicion of LD, the ASUIT Laboratory selected samples positive for anti-Clinical presentation was requested along with laboratory testing in order to solve potential discrepancies between the two workflows.To determine the concordance between the two workflows, Cohen’s κ coefficient was calculated in addition to the raw agreement rate, as the two-step diagnostic algorithm cannot be considered the gold standard. Given the differing LD epidemiology and collection criteria, κ was calculated both separately for each center and collectively. Data were processed using the STATA v14 software .A total of 804 samples were tested, of which 296 at the Great Romagna Area Hub Laboratory (Romagna area), 294 at the Azienda Sanitaria Alto-Adige-Bolzano Microbiology Laboratory (Alto Adige area), and 214 at the Azienda Sanitaria Universitaria Integrata di Trieste Laboratory (Giuliano-Isontina area). Out of the total 804 samples, 695 (86.4%) gave concordant results between the two methods, of which 564 were negative and 131 positive. Wherever available, clinical presentation and additional exams were analyzed to solve discrepancies and discussed below.The Great Romagna Area Hub Laboratory showed a total of 29 discrepancies. Of those, seven specimens testing negative for C6 were positive with the reference workflow, all presenting conditions which could potentially give rise to aspecific reactions . Twenty- two samples were positive with C6 testing and negative with the standard diagnostic algorithm; five of those were positive for interfering factors, while no additional clinical information was available for the remaining 17.T. pallidum infection); 11 of those samples could be demonstrated as actual positives pairing these results with clinical presentation, while clinical data was unavailable or not indicative of LD in the remaining cases.The Azienda Sanitaria Alto Adige-Bolzano Microbiology Laboratory tested 294 samples, 56 of which gave discordant results. Thirty-six specimens were positive for C6 but negative with the standard workflow; of these, seven of which were positive for interfering factors  compared to C6 positive-routine diagnosis negative (n = 9). All discrepancies were found in samples presenting no interfering factor. In each discordant group, three specimens could be identified as true positives when taking clinical presentation into account; in all other cases, data was unavailable or unhelpful to clarify the final serum status.The Azienda Sanitaria Universitaria Integrata di Trieste (ASUIT) Laboratory analyzed a total of 214 samples, selecting positives specimens for the standard diagnosis when possible. Unlike the two other laboratories, TS showed a higher number of C6 negative-routine diagnosis positive discordants . However, while clinical data could not solve all discrepancies, a greater proportion of true positives was found in the C6 negative/2-tier positive group. This would seem to point in the direction of a higher specificity of the currently recommended workflow, which is supported by the literature [The highest number of discordant results was found in the C6 positive/2-tier negative group , [Borrelia spirochetes should encompass the spectrum of Borrelia organisms capable of causing disease, including Bbss, B. burgdorferi sensu lato (Bbsl) and the Relapsing Fever Borrelia (RFB). Ideally, testing should detect both LD and RFB and should differentiate between the two.On the other hand, false negatives resulting from the two-tier algorithm are common, resulting from factors such as patient seronegativity, and genetic diversity among disease-causing strains. ,22,23 ELstrains.  and abougdorferi . To this (Bbss), ,27 sacriRecently, modified two-tiered testing (MTTT) algorithms that utilize two sequential first-tier tests and eliminate immunoblotting have been evaluated. Specifically, all MTTTs resulted in a significantly higher proportion of correct classifications than standard two-tier testing ,29.It must always be kept in mind that a laboratory diagnosis of LD is imperfect and no test available at the moment is reliably sensitive and specific.