Elevated cortisol levels have been reported in Alzheimer’s disease (AD) and may accelerate the development of brain pathology and cognitive decline. Dehydroepiandrosterone sulfate (DHEAS) has anti-glucocorticoid effects and it may be involved in the AD pathophysiology.To investigate associations of cerebrospinal fluid (CSF) cortisol and DHEAS levels with (1) cognitive performance at baseline; (2) CSF biomarkers of amyloid pathology (as assessed by CSF Aβ levels), neuronal injury (as assessed by CSF tau), and tau hyperphosphorylation (as assessed by CSF p-tau); (3) regional brain volumes; and (4) clinical disease progression.n = 145) with mild cognitive impairment or dementia or with normal cognition were included. Clinical scores, AD biomarkers, brain MRI volumetry along with CSF cortisol and DHEAS were obtained at baseline. Cognitive and functional performance was re-assessed at 18 and 36 months from baseline. We also assessed the following covariates: apolipoprotein E (APOE) genotype, BMI, and education. We used linear regression and mixed models to address associations of interest.Individuals between 49 and 88 years (Higher CSF cortisol was associated with poorer global cognitive performance and higher disease severity at baseline. Cortisol and cortisol/DHEAS ratio were positively associated with tau and p-tau CSF levels, and negatively associated with the amygdala and insula volumes at baseline. Higher CSF cortisol predicted more pronounced cognitive decline and clinical disease progression over 36 months. Higher CSF DHEAS predicted more pronounced disease progression over 36 months.Increased cortisol in the CNS is associated with tau pathology and neurodegeneration, and with decreased insula and amygdala volume. Both CSF cortisol and DHEAS levels predict faster clinical disease progression. These results have implications for the identification of patients at risk of rapid decline as well as for the development of interventions targeting both neurodegeneration and clinical manifestations of AD. Different steroids including cortisol, dehydroepiandrosterone (DHEA) and its sulfate ester (DHEAS) have been shown to exert crucial actions on the brain, translating into effects on brain aging, neurocognition, and the pathogenesis and course of neurocognitive disorders, in particular Alzheimer’s disease (AD) .Cortisol has been associated with impeding normal amyloid β (Aβ) and tau processing in the brain, exacerbating Aβ and tau toxicity, promoting oxidative stress and neurodegeneration, inducing synaptic dysfunction, and reducing dendritic plasticity in the hippocampus and prefrontal cortex in animal models . Higher The role of DHEAS on the brain is complex as it has both direct and indirect effects following conversion into testosterone and estrogens . DHEAS iGiven that DHEAS and cortisol often display antagonistic actions in the brain, cortisol/DHEAS ratio has been used to address interrelated effects. Plasma cortisol/DHEAS ratio was reported to be elevated in patients with dementia , and speDespite the growing body of evidence that steroids, in particular cortisol and DHEAS, might be incriminated in the etiopathogenesis of AD and might be considered potential therapeutic targets, there have been very few studies that assessed both cortisol and DHEAS levels in human subjects with MCI or AD type dementia . While aIn the present paper, we aimed to investigate whether CSF Cortisol and DHEAS were associated with (1) cognitive and functional performance at baseline; (2) with amyloid pathology (as assessed by CSF Aβ levels), neuronal injury (as assessed by CSF tau), and tau hyperphosphorylation (as assessed by CSF phosphorylated tau or p-tau); (3) regional brain volumetry; and (4) with clinical disease progression at 18 then at 36 months from baseline.We recruited 145 community-dwelling individuals aged between 49 and 88 years in an observational study on biomarkers of cognitive decline and AD conducted at the Department of Psychiatry and the Department of Clinical Neurosciences, Lausanne University Hospital, Switzerland.n = 93) were recruited among outpatients with cognitive impairment (MCI or mild dementia) referred to the Memory Clinics, Department of Psychiatry and the Department Clinical Neurosciences, Lausanne University Hospital for investigation of cognitive complaints. The diagnosis of MCI or of dementia was based on neuropsychological and clinical evaluations according to published criteria (n = 52) were recruited through journal announcements and word of mouth. They underwent the same clinical and neuropsychological examination as the cognitively impaired participants. They had no history or clinical signs of cognitive decline and the absence of cognitive impairment was confirmed by a multi-domain cognitive and functional neuropsychological assessment (as detailed below) that allowed to confirm that their CDR score was 0. Clinical examination and the Hospital Anxiety and Depression Scale , a detailed neuropsychological evaluation was completed and information on the participants’ daily life activities was collected by a trained psychologist blinded to the investigated biological markers, as previously described .The neuropsychological assessment included:•Mini Mental State Examination (MMSE) for global cognitive performance.•episodic memory.Grober and Buschke Double Memory Test  for epis•verbal fluency.DO40 picture-naming test , the pho•executive functions.Stroop Test for •visuospatial construction.Figures from the Consortium to Establish a Registry for Alzheimer’s Disease (CERAD) neuropsychological test battery  for visuThe functional assessment included the Katz Index of Independence in ADLs  for basiThe questionnaires and tools above were used to determine the overall cognitive and functional status by the CDR scale. CDR is widely used for the clinical staging of cognitive impairment . The CDRIn addition to the CDR-SB scores derived from the detailed neuropsychological evaluation and used as an outcome variable to assess cognitive and functional decline, MMSE scores were also used as an outcome variable. MMSE is a measure of global cognition, not allowing for comprehensive evaluation of subdomains of cognition. It is also of limited value in cognitively normal people because of floor effect. However, MMSE score change in time is widely used in clinical practice as well as in research to assess global disease progression .Ten to twelve mL of CSF were collected by lumbar puncture between 8 and 9 AM after overnight fasting. The CSF samples were spun down at 4°C, immediately aliquoted, and snap frozen at −80°C until assay . Blood s1–42 (Aβ1–42), Aβ1–40, tau and p-tau were measured using commercially available enzyme-linked immunosorbent assay kits . In addition, the levels of Aβ1–42 and Aβ1–40 were measured with immunoassays from IBL International  according to the manufacturer’s protocols.Cerebrospinal fluid Amyloid-β1–42 ratio with a cut-off of 0.0779 to define the presence/absence of an AD CSF profile, previously determined as the center cutoff indicating the concomitant presence of amyloid and tau pathology . Since DHEAS is a robust measure of DHEA production, we focused on DHEAS rather than DHEA in this study . The intAn MRI of the brain was obtained at baseline for each of the participants.Gray matter (GM) and white matter (WM) regions of interest (ROIs) were defined and segmented using the MorphoBox prototype as described in Here we used regional volumetric data normalized by total intracranial volume  to determine relative volumes of ROIs.Here we used regional volumetric data normalized by total intracranial volume  to determine relative volumes of ROIs.apolipoprotein E (APOE) genotype on the addressed relationships, leukocyte genomic DNA was isolated from EDTA blood with the Qiagen blood isolation kit  and the APOE genotype was determined as previously described  was also considered as a covariate because cortisol levels have been shown to depend on the BMI and BMI change has been associated with AD pathology .For categorical variables, we calculated absolute and relative frequencies. For continuous variables, we calculated means and standard deviations. Median values were also determined whenever the distribution was not gaussian (as per the Shapiro–Wilk test).For each of the analyses below, we chose the following covariates:–For associations only involving biological measures , we used age and sex as covariates.–For associations also involving cognitive performance, we used age, sex, years of education, BMI, and APOEε4 status as covariates. Apart from the level of education which obviously influences the results of most cognitive tests, we added APOE and BMI as covariates because they were shown to influence cognitive performance both in AD and in normal aging .t-test to compare CSF cortisol, CSF DHEAS, and CSF cortisol/DHEAS ratio between participants with baseline cognitive impairment (CDR score > 0) and those without (CDR = 0).We used the independent samples We also constructed separate multiple linear regression models with CSF cortisol and DHEAS as independent variables, and with the baseline MMSE score and CDR-SB, each as a dependent variable in separate analysis, controlling for age, sex, years of education, BMI, and APOEε4 status.1–42 ratio > 0.0779) and those without, before then after stratification by the presence of baseline cognitive impairment. We used the independent samples t-test whenever valid (sample size ≥ 30 in each group), and Mann–Whitney U-test otherwise.We compared CSF cortisol, CSF DHEAS, and CSF cortisol/DHEAS ratio between participants with an AD CSF profile , controlling for age and sex.Since most of the variables did not follow a normal distribution, we used Spearman’s non-parametric correlations to examine the associations between CSF cortisol, DHEAS and cortisol/DHEAS ratio and the CSF AD biomarkers  as within-subject variables in a first model then MMSE scores  in a second model–with sex and APOEε4 as between-subject factors–and with age, years of education, and CSF cortisol and CSF DHEAS as covariates in a first model, then with age, years of education, and CSF cortisol/DHEAS ratio as covariates in a second model.2).Global analyses were followed by between-subject and within-subject analyses to highlight significant interactions. Greenhouse–Geisser correction was applied whenever the sphericity assumption was not fulfilled. Effect sizes were determined using partial eta squares  did not differ from participants without cognitive impairment (n = 52) regarding sex, education level, and BMI. Participants with cognitive impairment were significantly older, and more likely to be APOEε4 carriers than participants without cognitive impairment , and with poorer baseline cognitive and functional performance as measured by the CDR-SB , after controlling for age, sex, years of education, BMI, and APOEε4 status.Higher CSF cortisol was associated with poorer baseline cognitive performance as measured by the MMSE score (Neither CSF DHEAS nor CSF cortisol/DHEAS ratio were significantly associated with baseline MMSE or CDR-SB.p = 0.011), while CSF DHEAS and CSF cortisol/DHEAS ratio did not differ significantly between groups. When comparing groups with/without cognitive impairment, and those with/without CSF AD profile, CSF cortisol was significantly higher in patients with cognitive impairment and CSF AD profile than in patients without cognitive impairment and without CSF AD profile . No other differences between subgroups were found   .No significant associations were found between CSF DHEAS or cortisol/DHEAS ratio and any of the brain volumes.When we analyzed the associations between baseline CSF cortisol and DHEAS and baseline brain volumes in each group (with vs. without CI at baseline) separately, no significant associations were observed.F = 4.5, p = 0.019, η2 = 0.045) and CSF DHEAS  = 3.7, p = 0.037, η2 = 0.037).Mixed design repeated measures ANOVAs found significant interactions between CDR-SB changes over time and CSF cortisol  = 3.7, p = 0.040, η2 = 0.039) but not with CSF DHEAS  = 1.6, p = 0.216, η2 = 0.017).Similarly, we found significant interactions between MMSE score changes over time and CSF cortisol  = 4.2, p = 0.021, η2 = 0.104).When we stratified the analysis by the presence of baseline CI, we found that the interactions between MMSE changes over time and CSF cortisol were not significant in any of the groups. However, the interactions between MMSE changes over time and CSF DHEAS were significant only in the group without CI (Higher CSF cortisol levels were associated with more pronounced cognitive decline (as evidenced by MMSE score change) and progression in disease severity (as evidenced by CDR-SB score change), while higher CSF DHEAS was only associated with more important progression in disease severity over 36 months.Cerebrospinal fluid cortisol/DHEAS ratio was not found to be longitudinally associated with the change in MMSE or CDR-SB.In this prospective study, we found that higher CSF cortisol, but not CSF DHEAS, was associated with poorer baseline cognitive and functional performance. At baseline, CSF cortisol was higher in participants with an AD CSF biomarker profile, in particular in those who also had cognitive impairment. Baseline CSF cortisol and cortisol/DHEAS ratio were associated with CSF tau and p-tau levels, as well as with the baseline volumes of the amygdala and the insula. DHEAS was not associated with any of the AD CSF biomarkers. Furthermore, higher baseline CSF cortisol predicted more pronounced cognitive decline and disease progression over 36 months. Higher baseline CSF DHEAS predicted more pronounced disease progression (as measured by CDR-SB) but not faster MMSE score decline over 36 months.Most previous studies investigating the links between cortisol and cognitive impairment or dementia of AD type used peripheral measures of cortisol, and many reported similar cross-sectional associations between increased cortisol levels  and poorer cognitive and functional performance and clinically diagnosed AD . A furthIn sum, higher cortisol appears to be cross-sectionally associated with lower cognitive performance, but this association is significant only in subjects with cerebral AD pathology. It is possible that the detrimental effects of cortisol on cognition are only clinically relevant in individuals with developing brain AD pathology. It is also possible that AD pathogenesis itself entails early HPA axis dysfunction. One of the factors leading to this dysfunction might be the early neurodegeneration of structures involved in cognitive performance.Previous studies found DHEAS levels to be reduced in the CSF and in the brain of AD patients . StudiesSince DHEAS has been shown to counter certain neurotoxic effects of cortisol, cortisol/DHEAS ratio was suggested as a potential marker of glucocorticoid vs. anti-glucocorticoid effects in the brain, and was shown to better distinguish patients with dementia from controls than cortisol or DHEAS alone . In the 1–42  , as wellWe found that higher baseline CSF cortisol independently predicted more marked decline in global and progression of disease severity over 36 months. This confirms the findings of our previous study in an independent cohort where baseline CSF cortisol levels in subjects with AD at the MCI stage predicted faster cognitive decline over time . SimilarMost studies which investigated the relationship between cortisol levels and cognitive decline over time examined plasma or salivary cortisol. Two previous studies found that higher plasma cortisol levels predicted faster cognitive decline in cognitively healthy older individuals  as well We found that elevated baseline CSF DHEAS predicted more pronounced progression of disease (as measured by CDR-SB) over 36 months. This finding may seem unexpected given the previously described neuroprotective effects of DHEAS. Indeed, a nested case-control study found lower plasma DHEAS in participants who developed AD 3 years later than in those who did not . One posThe association between CSF cortisol levels and more pronounced progression of AD can have therapeutic implications. Indeed, one may hypothesize that lowering cortisol levels or inhibiting cortisol effects on the brain may help prevent cognitive decline. In this context, several non-pharmacological as well as pharmacological interventions were studied. Even though the initial findings from animal studies were promising for glucocorticoid receptor antagonists or modulators as well as for compounds targeting key enzymes for cortisol synthesis, we are yet to see any convincing evidence from clinical trials .We measured central rather than peripheral steroid (cortisol and DHEAS) levels, which better reflects the central nervous system action of these hormones. The free  forms of cortisol and DHEAS represent only small fractions in the plasma but are the most prevalent fractions in the CSF . CorrelaThis study is, to the best of our knowledge, the first prospective study to examine the relationship between CSF DHEAS and cognitive decline in humans. Moreover, we addressed here for the first time relationships of both CSF cortisol and DHEAS with the “core” AD pathology using well-established CSF biomarkers of amyloid pathology, neuronal injury, and tau pathology. In addition, we assessed relationships with clinical manifestations and disease progression considering measures of both global cognition and disease severity . This isNevertheless, a few limitations need to be acknowledged. CSF cortisol and DHEAS levels were measured only at baseline. Hence, we were unable to examine any potential associations between changes in cognition and changes in cortisol and DHEAS levels. In addition, we did not measure saliva or serum/plasma levels of steroids. While these measures can be less accurate than their CSF counterparts, their use in clinical practice might be much easier. Furthermore, brain volumetry was only obtained at baseline, which did not allow us to examine the changes in the structure volumes over time as the cognitive decline occurs. The associations observed at baseline were cross-sectional, which does not allow to identify their direction, let alone to assume causality. As cognitive decline often occurs over a long time span, a longer follow-up period might allow to elicit clinical and biological changes that might be too subtle to detect over 3 years. Last, our choice of analyzing all participants  in a single cohort might have led to a heterogenous sample. When we conducted the main analyses for each clinical group separately, our small sample size probably led to type II errors.Our findings indicate that higher CSF cortisol levels are involved in the pathophysiology of AD, and are related in particular to neuronal injury and tau pathology, as indicated by CSF tau and p-tau levels, and decreased regional brain volumes. Higher CSF cortisol and DHEAS are associated with more rapid cognitive decline. Modulation of their CNS levels and activity may represent intervention targets to improve symptoms and slow down clinical disease progression.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Lausanne University Hospital . The patients/participants provided their written informed consent to participate in this study.SO: conceptualization, methodology, data curation, formal analysis, writing–original draft, review and editing, and visualization. CC, PL, JK, and CK: methodology and writing — review and editing. JR and BM: methodology, writing — review and editing and visualization. JP: conceptualization, methodology, data curation, writing — original draft and review and editing, funding acquisition, and supervision. All authors contributed to the article and approved the submitted version.JP received consultation honoraria from Nestle Institute of Health Sciences, Innovation Campus, EPFL, Lausanne, Switzerland, Ono Pharmaceutical, OM Pharma and from Fujirebio Europe, all unrelated to the present work. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Mycobacterium tuberculosis, and is characterized by high morbidity and mortality. Unfortunately, it is difficult to distinguish TBM from bacterial meningitis (BM) based on clinical features alone. The latest diagnostic tests and neuroimaging methods are still not available in many developing countries. This study aimed to develop a simple diagnostic algorithm based on clinical and laboratory test results as an early predictor of TBM in South China.Tuberculous meningitis (TBM) is the most serious form of extrapulmonary tuberculosis caused by A retrospective study was conducted to compare the clinical and laboratory characteristics of 114 patients with TBM and 47 with BM. Multivariate logistic regression analysis was performed on the characteristics of independently predicted TBM to develop a new diagnostic rule.Five characteristics were predictive of a diagnosis of TBM: duration of symptoms before admission; tuberculous symptoms; white blood cell (WBC) count, total cerebrospinal fluid WBC count, and cerebrospinal fluid chloride concentration. The sensitivity and specificity of the new scoring system developed in this study were 81.6 and 93.6%, respectively.The new scoring system proposed in this study can help physicians empirically diagnose TBM and can be used in countries and regions with limited microbial and radiological resources. M. tuberculosis culture and polymerase chain reaction are the “gold standard” for detecting M. tuberculosis in the cerebrospinal fluid. However, the possibility of identifying acid-fast bacilli (AFB) in CSF smears is very low, and the culture cycle of M. tuberculosis in CSF is very long  is the most serious form of extrapulmonary tuberculosis (TB), especially in many countries with a high burden of pulmonary TB . Howeverery long . Althougive rate . Therefoive rate . The aimAll patients admitted to the Neurology ward of Jiangxi Provincial People's Hospital  and Infectious Diseases Hospital of Jiangxi Province with a suspected clinical diagnosis of meningitis were included in this study. These hospitals admit patients referred from the province, which has a population of ~45.2 million. Clinical and laboratory data , and radiographic imaging  results from patients with suspected intracranial infection between 2004 and 2019 were collected and retrospectively analyzed. .All patients enrolled in the study underwent standard history-taking and examination. Clinical signs and symptoms, including fever, headache, nausea and vomiting, disturbance of consciousness, duration of illness, psychiatric symptoms, cranial nerve palsies, and focal neurological impairment were recorded. Blood was drawn from all patients for routine hematological and biochemical examination, which included: erythrocyte sedimentation rate (ESR); C-reactive protein (CRP), hemoglobin, anti-TB antibody; and T cell enzyme-linked immuno-spot assay. All patients were screened for HIV infection, and each underwent routine lumbar puncture. CSF was analyzed for chloride, glucose, total and differential lymphocyte counts, and total protein concentration. CSF samples were centrifuged and stained with Gram, India ink, and Ziehl-Neelsen, and cultured on blood and chocolate agar, and Lowenstein-Jensen media. Patients underwent CXR and, if clinically necessary, cranial CT or MRI was also performed. Data were extracted from patient hospital files or computer records.Patients were categorized into two groups based on baseline investigations. TBM diagnosis was classified as “definite,” “probable,” and “possible,” based on the case definition proposed by Marais et al. , who desThe diagnosis of BM was made according to the following criteria [10]: pathogenic bacteria isolated from the CSF; or clinical meningitis with all of the following: lymphocytes and neutrophils in CSF; low glucose concentration in the CSF (<50% of that in blood); sterile blood and CSF cultures; and full recovery (without anti-TB chemotherapy) 3 months after admission (these criteria were modified to full recovery at the time of discharge).t-test or Mann–Whitney U-test was used to compare continuous variables between the two groups. Frequency (%) for categorical variables and qualitative data were analyzed using the chi-squared or Fischer's exact test. Differences with p < 0.05 were considered to be statistically significant. The odds ratio and corresponding 95% confidence interval were calculated.Data were entered into a spreadsheet  and analyzed using SPSS version 19.0  for Windows . The 25 clinical and laboratory parameters of those who met the diagnostic criteria for BM and TBM were compared . Data weTo identify features that can be used to distinguish TBM from BM, two statistical methods were used. Firstly, variables significantly associated with TBM were included in a logistic regression analysis. Validation of the diagnostic algorithm was then performed using the logistic regression method. To determine cut-off values from the data, the value chosen was that which was closest to the upper left corner of the receiver operating characteristic (ROC) curve. Logistic regression analysis was used to build a diagnostic model. The diagnostic index (DI) of each clinical variable in the model was defined by relying on rounded β-coefficients of the model. The diagnostic index derived was: 6.645 × tuberculous symptoms + 0.196 × DSBA – 0.506 × blood WBC – 0.003 × CSF total WCC – 0.203 × CSF chloride (tuberculous symptoms were coded 1 if present and 0 if absent). Secondly, the use of a classification and regression tree (CART) model. The model was built on the basis of considering all variables individually. The range of each variable was divided into two groups to obtain the optimal separation of TBM patients from BM patients.M. tuberculosis and/or CSF Gram stain or acid-fast stain positivity was obtained in a total of 12 patients. There were no HIV-positive patients. Forty of the 161 adults were tested using the T cell enzyme-linked immuno-spot assay (T-spot. TB): 23 were positive . Among patients with TBM, 45 (45.5%) had a positive anti-TB antibody (TB-Ab), 33 (28.9%) exhibited low hemoglobin, high ESR in 38 of 86 (44%), and high CRP in 25 of 53 (47.2%). Eighty-nine of the 161 adults underwent electroencephalogram testing: 77 were positive . CXR and cranial CT or MRI scans were performed in all patients; chest radiographs with signs suggestive of active TB were found in 52 (46%) patients with TBM. Cranial CT/MRI scans of the patients presenting with TBM revealed hydrocephalus in 36 (31.9%), cerebral infarction in 57 (50.4%), meningeal reinforcement in 74 (66.1%), and tuberculoma in 12 (10.6%).During the 15-year period in question (2004–2019), 380 adults with meningitis were admitted to the participating hospitals, of which 219 were excluded due to insufficient data being available to establish a definitive diagnosis or to apply a DI. Ultimately, 161 patients fulfilled the diagnostic criteria for inclusion in this study: 114 with TBM and 47 with BM. A positive CSF culture for On univariate analysis, factors with a significant difference between the two groups included TB symptoms , focal neurological impairment, duration of symptoms before admission (DSBA), blood white cell count (WBC), blood % neutrophils, serum sodium, CSF total white cell count (WCC), CSF % neutrophils, CSF % lymphocytes, and CSF chloride . Variabl9/L), CSF total WCC (> or < 435 × 106/mL), and CSF chloride (> or < 119.5 mmol/L).A classification and regression tree model was derived using the following variables: tuberculous symptoms, DSBA, blood WBC, CSF total WCC and CSF chloride. Meanwhile, the optimum cut-off value for the total diagnostic parameters to diagnose TBM patients was determined using ROC curve analysis : tubercu9/L) + DI  + DI (CSF chloride of <119.5 mmol/L). The DIs for the five variables are summarized in The total DI (TDI) was calculated by aggregating all variables DIs. TDI = DI (tuberculous symptoms presence) + DI (DSBA exceeding 11 days) + DI , negative likelihood ratio (NLR), positive predictive value (PPV), negative predictive value (NPV), and accuracy were 96%, 86, 7.07, 0.04, 0.90, 0.95, and 92%, respectively .M. tuberculosis detection and diagnostic confirmation of TBM   . In addin-making . Some stn-making . Furthern-making .Worldwide, TBM and BM are serious life-threatening infectious diseases that require early diagnosis and treatment to reduce death and disability. Typical BM is not difficult to diagnose; however, with the widespread use and abuse of antibiotics, a large proportion of patients with BM have been treated with antibiotics in primary hospitals before admission, resulting in atypical CSF examination results and a low positive rate in bacteriology. High levels of CSF protein and low CSF glucose are commonly present in these two diseases. Meanwhile, the early clinical presentation of TBM, lack of specificity and difficult to distinguish from BM, especially partially treated BM. At the same time, clinicians are reluctant to administer patients antibacterial drugs for weeks or anti-TB drugs for months, without evidence supporting etiology, which has been a challenge for clinicians.To facilitate clinicians in better distinguishing TBM from BM in clinical practice, some scoring systems using clinical and laboratory characteristics have been developed –29. In a9/L); CSF total WCC (<435 × 106/mL); and CSF chloride (<119.5 mmol/L)  .The present study, however, had some limitations, the first of which was its retrospective design and that data for many features were unavailable. In addition, 129 patients were excluded due to incomplete data and other reasons. Second, HIV-1 antibody testing was negative for all patients included in this study. Third, the low positivity rate of CSF culture/staining makes the final diagnosis of TBM relatively weak.The clinical features of TBM are non-specific, conventional bacteriology is widely regarded to be insensitive, and the validation of newer diagnostic methods is not complete , 30, 31.The scoring system proposed in this study may help clinicians to empirically diagnose TBM, especially the high incidence of TB, and can be used in settings where microbiological diagnostic support is limited, thus ensuring rapid anti-TB treatment before other confirmation methods are available. Nevertheless, further studies are needed to validate this diagnostic scoring system.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 study was conducted according to the guidelines of the Declaration of Helsinki and approved by the Ethics Committee at Jiangxi Provincial People's Hospital Affiliated to Nanchang University. The patients/participants provided their written informed consent to participate in this study.AW, E-LL, and S-ML designed the study. The manuscript was written by AW, E-LL, and S-ML with the final approval. W-FC and Y-LZ did the data searches and study selection. C-QL and Z-bX did the data synthesis and created the tables and figures. All authors contributed to the article and approved the submitted version.This study was supported by Jiangxi Provincial Health Commission Science and Technology Foundation Grants 20204024 and 20204025.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Dear editor,. ABO incompatibility (ABOi) or positive complement-dependent cytotoxicity (CDC) crossmatch between the candidate recipient and the prospective donor is a major obstacle to living donor kidney transplant, and consequently many recipients remain on the waiting list for years. Based on a simulated model, Bastos et al. recently showed that with the implementation of the Kidney Paired Donation (KPD) program in Brazil, the number of transplants with living donors could increase by 23%, using an optimized algorithm and the number of sensitized recipients transplanted could increase by 70.7% when using the prioritizing model. In KPD, a potential kidney recipient that has a willing but incompatible living donor receives a kidney from the donor of another incompatible pair and vice versa.The supply of kidneys from deceased and living donors in Brazil cannot meet the growing annual demand for transplantsSince 2011, we have started the discussion and preparation for the implementation of a local KPD program at the Hospital das Clínicas - FMUSP. We discussed the topic internally with our Bioethics Commission, Medical Ethics Commission, Organ and Tissue Transplant Commission, and Clinical Board. In Brazil, Law No. 9,434 regulates the removal of organs, tissues, and parts of the human body for transplant purposes, without contemplating the activity of paired donation. Therefore, we proposed and obtained ethical approval (CAAE 83469418400000068) for a research project to assess the effectiveness of transplantation in patients with ABOi living donor or positive CDC/flow cytometry crossmatch by donor exchange. At the end of the medical, psychological, social, and immunological evaluation of the pairs and after the consent terms were signed, approval was obtained from the Ethics Committee that evaluates transplants with unrelated donors to perform the paired donation. Subsequently, we also obtained the approvals of the Public Prosecutor’s Office, the judicial approval and that of the General Coordination of the National Transplant System - Ministry of Health, for the first transplant with KPD in Brazil.On March 10, 2020, the 38-year-old recipient (recipient 1) with chronic glomerulonephritis, undergoing dialysis treatment, on a deceased transplant waiting list for 8 years and with the previous exclusion of 7 living donors, including his wife (donor 1), because of ABOi, received the kidney of the 45-year-old donor of the second pair. The second recipient, 57 years old, with chronic glomerulonephritis, undergoing dialysis treatment, on a deceased transplant waiting list for 1.9 years, and with exclusion of his only donor, wife (donor 2) because of ABOi, received the kidney from the 39-year-old donor of the first pair (donor 1). Pair anonymity was assured until the time of admission, as was reciprocal compatibility between the pairs and simultaneous surgery in 4 operating rooms, which allowed donors to withdraw consent at any time before anesthesia. After 12 months, recipients had adequate kidney function and their donors were doing well.. We expect that this HCFMUSP program provide the impetus for the national development of DRP.Efforts to increase organ supply have a significant source of potential donors in the paired donation programs. Today, all countries that are world leaders in transplantation are practicing and developing this modality in an attempt to reach more and more recipients because of its excellent results


C. striatum pneumonia who require intensive care unit admission is currently limited.The clinical information on patients with severe Corynebacterium striatum pneumonia during a 6-year period at Asan Medical Center in comparison with severe pneumonia associated with MRSA.We investigated the incidence and characteristics of severe C. striatum pneumonia. The majority of the cases (70.4%) were hospital-acquired pneumonia (HAP), and about half of the patients (51.9%) were immunocompromised. The incidence of C. striatum HAP significantly increased from 1.0% (2/200) in 2014-2015 to 5.4% (10/185) in 2018-2019 (P < 0.001), while the incidence of severe methicillin-resistant Staphylococcus aureus (MRSA) HAP significantly decreased from 12.0% to 2.7% during the same period. Of the 75 HAP cases whose bacterial pathogens were identified in 2018–2019, C. striatum was responsible for 13.3% of the cases. The 90-day mortality rates were similarly high in the C. striatum and MRSA groups .Between 2014 and 2019, there were 27 adult cases of severe C. striatum was a major pathogen of recent severe HAP and was associated with a substantially high mortality rate.In conclusion, All Authors: No reported disclosures.


Senile plaques, which are mostly composed of beta-amyloid peptide, are the main signature of Alzheimer’s disease (AD). Two main forms of beta-amyloid in humans are 40 and 42-amino acid, long; the latter is considered more relevant to AD etiology. The concentration of soluble beta-amyloid-42 (Aβ42) in cerebrospinal fluid (CSF-Aβ42) and the density of amyloid depositions have a strong negative correlation. However, AD patients have lower CSF-Aβ42 levels compared to individuals with normal cognition (NC), even after accounting for this correlation. The goal of this study was to infer deviations of Aβ42 metabolism parameters that underlie this difference using data from the Alzheimer’s Disease Neuroimaging Initiative cohort. Aβ42 is released to the interstitial fluid (ISF) by cells and is removed by several processes. First, growth of insoluble fibrils by aggregation decreases the concentration of soluble beta-amyloid in the ISF. Second, Aβ42 is physically transferred from the brain to the CSF and removed with the CSF flow. Finally, there is an intratissue removal of Aβ42 ending in proteolysis, which can occur either in the ISF or inside the cells after the peptide is endocytosed. Unlike aggregation, which preserves the peptide in the brain, transfer to the CSF and intratissue proteolysis together represent amyloid removal. Using a kinetic model of Aβ42 turnover, we found that compared to NC subjects, AD patients had dramatically increased rates of amyloid removal. A group with late-onset mild cognitive impairment (LMCI) also exhibited a higher rate of amyloid removal; however, this was less pronounced than in the AD group. Estimated parameters in the early-onset MCI group did not differ significantly from those in the NC group. We hypothesize that increased amyloid removal is mediated by Aβ42 cellular uptake; this is because CSF flow is not increased in AD patients, while most proteases are intracellular. Aβ cytotoxicity depends on both the amount of beta-amyloid internalized by cells and its intracellular conversion into toxic products. We speculate that AD and LMCI are associated with increased cellular amyloid uptake, which leads to faster disease progression. The early-onset MCI (EMCI) patients do not differ from the NC participants in terms of cellular amyloid uptake. Therefore, EMCI may be mediated by the increased production of toxic amyloid metabolites. Dr. Alois Alzheimer described a certain kind of dementia (later named after him ) as havipostmortem. With the increased sensitivity of analytical methods, measuring the concentration of various Aβ peptides  in ceregy of AD . Then, pgy of AD . It was gy of AD .The concentration of beta-amyloid in CSF and the density of amyloid deposits are interconnected, as high levels of amyloid deposits are associated with significantly decreased Aβ42 levels in the CSF –10. Due Based on an analysis of data from the Alzheimer’s Disease Neuroimaging Initiative (ADNI), Sturchio et al  reportedThis observation can be deduced from and, thus, confirmed by the data of Mattsson et al. . In theiThus, CSF-Aβ42 concentration and beta-amyloid deposits are partially independent correlates of cognitive function . More spTo interpret the interactions between these two major biomarkers of AD, we estimated the parameters of beta-amyloid turnover by fitting a mathematical model to the data from the ADNI dataset. The modeling framework we used is typical for pharmacokinetic studies. Previously, a similar approach was used for studying the turnover of beta-amyloid in individual patients –16. The We considered that Aβ42 released by cells to the interstitial fluid (ISF) can be removed via several processes. First, Aβ42 is transferred to the CSF and is physically removed from the brain with the CSF flow. Second, soluble beta-amyloid aggregates and forms insoluble fibrils. This process is slow in the absence of aggregated peptide but accelerates after amyloid seeds are already formed. The aggregation process decreases the concentration of soluble beta-amyloid in the ISF, even though it does not remove the peptide from the brain. Finally, Aβ42 can be digested inside the brain through proteolytic degradation. The proteolysis can occur either in the ISF or inside the cells after the peptide is endocytosed by the cells. The SILK technique in its original form , 17 can http://adni.loni.usc.edu/). The ADNI was launched in 2003 as a public-private partnership, led by Principal Investigator Michael W. Weiner, MD. The primary goal of ADNI has been to test whether serial magnetic resonance imaging, positron emission tomography (PET), other biological markers, and clinical and neuropsychological assessments can be combined to measure the progression of mild cognitive impairment (MCI) and Alzheimer’s disease (AD). The study protocol for ADNI was approved by the local ethical committees of all participating institutions and all participants signed informed consent forms which included the consent for de-identified data being shared with the general scientific community for research purposes (https://adni.loni.usc.edu/wp-content/uploads/how_to_apply/ADNI_DSP_Policy.pdf). The authors were approved by ADNI for receiving and analyzing de-identified data. The manuscript was approved for publication in written in accordance with the policies of ADNI.We used non-personalized data obtained from the Alzheimer’s Disease Neuroimaging Initiative (ADNI) (www.adni-info.org). Given the use of two different amyloid PET-tracers, SUVR levels were converted to centiloids (CL) using tracer-specific formulas provided by ADNI , is defined by several processes: 1) synthesis by cells, 2) filtration of the protein into the CSF, 3) aggregation into non-soluble plaques, and 4) intratissue removal , which we denote as [oval see . The modSynthesis rate (FLOWCSF) and CSF-Aβ42 ([CSF]): FLOWCSF·[CSF].The rate of removal of the protein through the CSF is a product of the CSF removal rate , and the coefficient of aggregation Ka: Ka·[PET]·[ISF].Existing plaques serve as seeds for the aggregation of soluble Aβ42 in the ISF. The rate of loss of soluble Aβ42 in the ISF due to aggregation is the product of Aβ42 concentration in the ISF, the concentration of plaques /Ka, the equation take the following form:After introducing new parameters, SYN and KF, which will be referred to as the amyloid synthesis rate and the amyloid removal rate, respectively, in the rest of this manuscript. Note that the removal rate KF is a sum of two terms: (FLOWCSF·KT/Ka), which is proportional to the amyloid removal rate through the CSF, and the intratissue amyloid removal rate divided by the aggregation rate, (KR/Ka).The equation has two parameters, KT and aggregation rate Ka are invariant over the population, so these two components of the amyloid removal rate are defined by the rate of removal by CSF and intratissue removal, respectively. Units for all parameters and variables are provided in We assume that the transfer coefficient SYN and KF for each of the four groups , we used the CSF-Aβ42 and the amyloid deposit density of each research subject (based on data provided by the ADNI). We assumed that the conditional probability distribution p([CSF]|[PET]) is log-normal. Therefore, the posterior probability density function (PDF) of the parameters can be represented asσ is an unknown standard deviation, N is the number of participants in the group, and ln(.) is the natural log function. To calculate the marginal PDF for SYN and KF, we integrated the posterior PDF over σ as follows.k = (N−1)/2 and Γ(k) is the gamma function. Therefore, the following holds.To infer the values of the parameters SYN, KF)-plane, as well as their standard errors.This marginal parameter distribution was used to calculate the confidence regions for the individual parameters on the  as a function of their CSF-Aβ42 levels . The firCSF]) and the density of amyloid deposits ([PET]):SYN is a parameter representing the synthesis rate and KF is a sum of the rate of amyloid removal through the CSF and the cellular amyloid uptake rate  for ance see .SYN and KF (see previous section). The three curves in To elucidate these differences, we fitted the model described in the previous section to the data for each group see also  and calcTo illustrate that our model accurately predicts average CSF-Aβ42 based on amyloid load, we separated all amyloid-positive subjects in each group into those with low and high amyloid load . For each research subject, we used either the measured CSF-Aβ42 value or the value calculated based on their [PET] and the model parameters for the corresponding group. The averages of the values calculated using the model  in the NC group, but 75% more (140 CL) in the AD group. As previously noted, our model does not allow for the independent estimation of the two components of KF–the rate of intratissue beta-amyloid removal and the rate of removal of beta-amyloid through CSF. As we speculate in the Discussion, however, the latter component is either not changed in AD patients [Our calculations show that the amyloid removal rate, patients  or decrepatients . TherefoThe estimate of a difference in the intratissue beta-amyloid removal rate between NC and AD subjects depends on the distribution of the amyloid removal rate between intratissue removal and removal through the CSF see . For theUsing this approach, it is possible to compare the intensity of the amyloid removal process as a product of the removal rate and the amyloid concentration in the CSF for individuals in the AD and NC groups from the ADNI cohort . Again, The brain amyloid load and CSF-Aβ42 have a strong negative correlation , 9, 24. Our analysis of the ADNI dataset confirms that while the biomarkers have a strong negative correlation described by a single compartment model, CSF-Aβ42 in fact provides independent clinically valuable information. In cohorts with a similar amyloid load, a higher CSF-Aβ42 clearly correlates with a better clinical outcome. CSF-Aβ42 levels above 1000 pg/ml are associated with a low ratio of AD patients, even in the group with high amyloid loads . Lower CSYN, should be lower, or the rate of removal, KF, should be higher, in order for the CSF-Aβ42 to be lower,. The suggestion of a lower synthesis rate in AD is not only counterintuitive, but is also not supported by experimental data [Sturchio et al.  suggestetal data . With thIn scenarios where intratissue removal is between 50% and 25% of the beta-amyloid removal rate, a 75% increase in the aggregation-independent amyloid removal rate translates into a 2.5 to 4-fold increase in the intratissue amyloid removal rate. Furthermore, amyloid removal through CSF could be decreased in AD patients due to a lower CSF flow . TherefoAccording to the model, the increased amyloid removal rate leads to a lower amyloid concentration in the CSF (see Eq ). The toin vitro and in vivo [The progression of Alzheimer’s disease is associated with neuronal death. The severity of the disease correlates with the density of beta-amyloid deposits. Although the aggregated amyloid is not toxic to cells, the synaptic loss and neuritic dystrophy are highest in anatomical proximity to the senile plaques . On the  in vivo –31; howe in vivo , 33.Recently, we introduced the amyloid degradation toxicity hypothesis, which explains known facts about beta-amyloid toxicity and suggests specific pathways leading to cell death . AccordiThe degradation of beta-amyloid is mediated by proteases . HoweverThe idea that neurotoxicity in AD is mediated by the cellular uptake of beta-amyloid could be key to understanding the non-effectiveness of the plaque dissolving approach in the treatment or prophylaxis of AD. According to our model, dissolving the aggregated beta-amyloid would decrease the removal of soluble beta-amyloid through aggregation and, thus, it would increase the interstitial Aβ concentration. In fact, the treatment with plaque-dissolving monoclonal antibodies does result in a significant increase of CSF-Aβ42 . The incOur results suggest that AD and LMCI groups are characterized by an increased cellular amyloid uptake rate, which presumably results in an increased formation of toxic amyloid species and leads to irreversible neuronal damage. In contrast, beta-amyloid turnover in patients with EMCI does not appear different from that in NC subjects. Specifically, there is no tendency towards higher intratissue amyloid removal in EMCI patients . Is it pTo answer this question, it is necessary to consider how amyloid toxicity develops at the cellular level. The production of peptide fragments is a part of lysosomes’ physiological function, which is to degrade large molecules such as proteins. Unlike short amyloid fragments, full-length Aβ42 is not able to form membrane channels in cell-like structures such as liposomes , 37. TakBased on the above, the rate of channel formation depends on the concentration of toxic fragments. This concentration, in turn, depends not only on the concentration of the full-length peptide but also on the balance between the formation and degradation of these fragments. Hypothetically, most toxic amyloid fragments are produced by endoproteases, while their degradation is performed by exoproteases. Therefore, the degradation is mediated by proteases that are not necessarily the same enzymes as the ones producing toxic amyloid fragments. A change in the balance between exo- and endoproteolytic activities of lysosomal proteases would affect the probability of membrane channel formation.Thus, enzymatic disbalance (an increased rate of toxic fragment production and/or a slower degradation of toxic fragments) can be a mechanism for EMCI that is not associated with an increased cellular uptake rate of beta-amyloid. Alternatively, the concentration of toxic fragments can be increased due to a higher amyloid endocytosis rate, which appears to be the mechanism of AD and LMCI.The single compartment model which we used in this study is oversimplified. First, we assumed that CSF-Aβ42 levels can be used as a surrogate for interstitial concentrations of Aβ42. This assumption is based on the correlation of the concentrations of the peptide in these two bioliquids , while tThe precision of estimates is dependent on the number of observations, so increasing the size of the cohort can demonstrate statistical significance for the differences between the characteristic parameters of the groups. However, using a statistical approach does not allow for the estimation of the parameters of interest in specific patients. Establishing novel biomarkers, which directly characterize the rates of relevant processes such as cellular amyloid uptake and metabolic production of toxic amyloid species, can be a breakthrough for the early and/or differential diagnosis of Alzheimer’s disease.Based on this modeling study, we conclude that lower levels of soluble Aβ42 in CSF, considered in combination with the amyloid load, is an important biomarker suggesting an increased intratissue amyloid removal rate. We hypothesize that increased amyloid removal can be the consequence of increased cellular amyloid uptake in the brain, resulting in an increased production of neurotoxic amyloid degradation products. This suggests that the rate of beta-amyloid uptake can be tested as a novel pathophysiologically relevant biomarker of Alzheimer’s disease as well as a predictor of late-onset AD. A differential diagnosis of AD also requires an assessment of lysosomal proteolytic activity that can lead to the production of toxic amyloid fragments.S1 Data(XLSX)Click here for additional data file.


The cerebrospinal fluid (CSF) space is convoluted. CSF flow oscillates with a net flow from the ventricles towards the cerebral and spinal subarachnoid space. This flow is influenced by heartbeats, breath, head or body movements as well as the activity of the ciliated epithelium of the plexus and ventricular ependyma. The shape of the CSF space and the CSF flow preclude rapid equilibration of cells, proteins and smaller compounds between the different parts of the compartment. In this review including reinterpretation of previously published data we illustrate, how anatomical and (patho)physiological conditions can influence routine CSF analysis. Equilibration of the components of the CSF depends on the size of the molecule or particle, e.g., lactate is distributed in the CSF more homogeneously than proteins or cells. The concentrations of blood-derived compounds usually increase from the ventricles to the lumbar CSF space, whereas the concentrations of brain-derived compounds usually decrease. Under special conditions, in particular when distribution is impaired, the rostro-caudal gradient of blood-derived compounds can be reversed. In the last century, several researchers attempted to define typical CSF findings for the diagnosis of several inflammatory diseases based on routine parameters. Because of the high spatial and temporal variations, findings considered typical of certain CNS diseases often are absent in parts of or even in the entire CSF compartment. In CNS infections, identification of the pathogen by culture, antigen detection or molecular methods is essential for diagnosis. Streptococcus agalactiae (n = 146), CSF routine parameters were even found to be normal in 6% of reported patients  after intrathecal administration are not included here , 44.CCSF/CS) and the ratio of the areas under the concentration–time curves in CSF and serum (AUCCSF/AUCS) after a short-duration infusion depend on the molecular mass, lipophilicity and binding to plasma proteins of the compound studied as well as on the state of the blood–CSF barrier and the CSF flow [CCSF/CS and AUCCSF/AUCS are higher in the lumbar than in the ventricular CSF leading to higher lumbar than ventricular drug concentrations. With the exception of small, moderately lipophilic drugs, elimination from the blood is faster than elimination from the CSF, resulting in a lag in the concentration–time curve in CSF in comparison to the respective curve in serum [After an intravenous injection, the time it takes to reach equilibrium between blood and CSF depends on the molecular mass and lipophilicity of the compound studied and on the size of the host organism: ions and small moderately lipophilic drugs rapidly equilibrate, whereas large hydrophilic compounds need up to several days to fully equilibrate –47. EquiCSF flow , 13, 48.In humans, after an intravenous infusion of radiolabeled albumin, it takes approximately 3 days to establish an equilibrium between blood and CSF, due to the dynamic exchange of albumin between these two compartments . The entInjection into the lumbar CSF does not ensure a homogeneous drug distribution in the whole CSF space: after intralumbar injection, the drug concentrations in ventricular CSF are highly variable. Often, therapeutic concentrations are not achieved –52. AfteUnless there is a complete block of the CSF circulation, e.g., in cerebral aqueduct stenosis or after an obstruction of the cervical, thoracic or lumbar spinal canal, intraventricular drug injection ensures its distribution throughout the entire CSF space. Peak aminoglycoside concentrations in lumbar CSF were 2–10 times lower than the simultaneous intraventricular levels, but still in the therapeutic range . In infaScedosporium apiospermum meningitis, daptomycin concentrations measured in the CSF from the right and left ventricle were discordant by a factor of about 3 after injection of daptomycin into the right ventricle [In neonates with bacterial meningitis, injection of amikacin into one ventricle resulted in approximately equal drug levels in both ventricles 10 h after dosing . Conversentricle .Elimination from the CSF compartment occurs through: (a) CSF bulk flow with peripheral outflow of CSF along all nerves and  by the arachnoid granulations; (b) diffusion across the blood–brain and blood–CSF barrier , 48, 55,1/2β CSF of 3.15 h. However, in view of recent data on the CSF production rate and the volume of the CSF space, this estimate appears to be too high. Assuming an average production of 20 ml of CSF/h and an average CSF volume of 200 ml would lead to an estimated CSF turnover rate  of 0.1/h (10%/h) and of t1/2β CSF of 6.9 h. The average time a “γ-globulin molecule” spent in human CSF was estimated to be 17.3 h which, when assuming first order kinetics, corresponds to a t1/2β of 12.0 h [1/2β CSF of large hydrophilic antibiotics correspond well to this estimate: t1/2β CSF of vancomycin (molecular mass 1486 g/mol) ranges from approximately 2 to 20.5 h [1/2β CSF of colistin (molecular mass 1155 g/mol) after intraventricular administration in humans [1/2β CSF estimated in clinical studies in critically ill patients originates from several variables: (1) inter- and intra-individual differences of the CSF volume, (2) inter- and intra-individual variations of the CSF production and absorption, and (3) the rate of CSF drainage by an external or internal ventricular catheter [1/2 β) of hydrophilic molecules in the CSF after intrathecal administration shows a great interindividual variation and in most cases lies between 6 and 12 h.In humans, an early estimate of the CSF turnover rate was approximately 22%/h  correspof 12.0 h . After io 20.5 h , 67–72. n humans  was simin humans . The higcatheter . GeneralMany ventricular CSF specimens contain blood, either because of the underlying disease or as a consequence of the implantation of the ventricular catheter. When dealing with blood in CSF samples in several clinical settings, it must be taken into account that blood can heavily disturb CSF analysis of routine parameters. Blood in the CSF can originate (a) from an intracranial spontaneous or traumatic hemorrhage; (b) from surgery including ventriculostomy insertion, or a traumatic lumbar  puncture. Blood in the CSF usually is detected by counting erythrocytes or by measuring free hemoglobin . It can 6/μl erythrocytes and 5 × 103/μl leukocytes, i.e., a leukocyte/erythrocyte quotient of 1/1000 in blood: 5000 erythrocytes/μl CSF indicate a blood contamination of 0.1%. For the assessment of inflammation 1 leukocyte/μl is subtracted from the CSF white blood cell count for each 1000 erythrocytes/μl counted. The main advantages of this method of correction are: (1) It is easy to perform and (2) the laboratory analyzing the CSF does not need to know the true leukocyte and erythrocyte densities  in the patient`s blood. A more sophisticated method using the variable leukocyte/erythrocyte quotient in blood is the cell index proposed by B. Pfausler and colleagues [Whether CSF findings should be corrected for blood contamination and if so, by which formula, is a matter of debate. The easiest correction method is based on the assumption of a density of 5 × 10lleagues . Here, tlleagues . Moreovelleagues , 75 , calculated for several classes of immunoglobulins  as:is a useful method to diagnose CNS infections and autoimmune diseases of the CNS , 85. AftIn a prospective study in patients with external ventriculostomy and with sterile CSF, sampling of ventricular CSF was followed by a 2nd sample 10 min later. During these 10 min either a standard clinical re-positioning was performed , or patients were not moved for 2 h before the 1st sampling and during the 10 min interval between 1st and 2nd sampling (control = C-samples). In 162 pairs of CSF samples from 51 patients, both re-sampling and re-positioning influenced CSF leukocyte and erythrocyte densities: the effect of re-positioning was stronger than the effect of re-sampling with a constant head position, the direction of change was random, variability increased with the leukocyte and erythrocyte count in sample 1, and leukocytes, granulocytes and erythrocytes showed greater variability than lactate or glucose . When thIn a microminipig, after placement of a ventricular drain into the cisterna magna, melatonin in CSF showed a ≥ tenfold higher value at night than during the day, whereas no substantial diurnal variation in the CSF lactate dehydrogenase, creatine kinase, glucose, sodium, potassium, chloride, calcium, glutamine or alanine concentrations were noted. Peak CSF melatonin concentrations at night were 2–3 times higher than the respective blood concentrations during the day. CSF leukocytes, protein and lactate were not determined . This stSince it would be unethical withdraw CSF without a clear indication, we are unaware of comparisons of the composition of ventricular, cisternal and lumbar CSF in healthy volunteers after simultaneous ventricular, cisternal and lumbar puncture. In clinical routine CSF analysis, the normal values of the CSF-to-serum albumin ratio and of the total protein concentration in ventricular CSF are considered to be 2.5 times lower than the normal lumbar CSF values , 85. In In an evaluation of routine data from a large neurochemical laboratory using mainly ventricular and lumbar CSF from different patients , the concentrations of proteins originating from neurons or glial cells  decreased from ventricular to lumbar CSF. In contrast, concentrations of blood-derived proteins and primarily leptomeningeal proteins (beta-trace protein and cystatin C) increased from ventricular to lumbar CSF .rS 0.95 and 0.91; p < 0.0001 each, n = 16 pairs of samples). As expected for brain-derived proteins, median tau, phosphorylated tau (ptau) and β-amyloid1-42 concentrations were slightly lower in lumbar than in “cisternal” CSF , whereas this gradient was absent for β-amyloid1-42 [Since cisternal CSF samples are very rare in clinical routine due to the risk of damaging the brain stem by suboccipital puncture, we attempted to collect samples representing an approximation to cisternal CSF in patients with clinical and radiological signs of normal pressure hydrocephalus by serial sampling of 40 ml in 5 ml fractions. Fraction 1 represented lumbar CSF, whereas fraction 8 was assumed to primarily contain CSF from regions close to the cisterna magna and cisterna pontis . Normal loid1-42 . As expeloid1-42 –94.In a large study, analyzing uninflamed CSF from different patients , the median ventricular CSF-to-serum glucose ratio was 0.70 (5th–95th percentile: 0.48–0.93), and the median lumbar ratio was 0.60 (5th–95th percentile: 0.41–0.81). There was thus a small ventriculo-lumbar decrease, which failed to reach statistical significance in a multivariate linear regression analysis [In normal pressure hydrocephalus, leukocyte count in lumbar CSF is normal (≤ 4/μl), and CSF-to-serum albumin ratio and CSF protein content are normal or slightly elevated. One year after placement of a ventriculoperitoneal shunt, in 7 of 8 patients leukocyte density in lumbar CSF was normal (≤ 4/μl), and in one patient it was 5/μl, whereas the lumbar CSF-to-serum albumin ratio and the lumbar CSF protein content had increased substantially  Table 11 . Of 8 simultaneous comparisons between ventricular and lumbar CSF, the difference in 5 pairs of samples was ≤ 1 log CFU/ml, and in 2 of the 3 discordant pairs of CSF samples, bacterial densities in the ventricular were greater than in the lumbar CSF [H. influenzae meningitis, CSF leukocyte concentrations in ventricular CSF ranged from 1 to 3980/μl. Of 20 simultaneous comparisons between ventricular and cisternal CSF, leukocyte concentrations were about equal in 5, but were lower in the ventricular than in the cisternal CSF in 15 pairs of samples . CSF protein, glucose and lactate and CSF-to-serum albumin ratio were not determined [In an infant rhesus monkey model of mbar CSF . In expeIn humans, differences in the composition between ventricular and lumbar CSF in CNS infections were first noted by Merritt and Freemont-Smith in 1937 . In 3 parS = 0.37), between ventricular and lumbar total protein concentration intermediate (rS = 0.42), and highest between ventricular and lumbar lactate concentrations (rS = 0.79) [In 41 pairs of samples from 26 patients with a suspected bacterial CNS infection , who received a lumbar and ventricular CSF analysis within 24 h, the correlation between ventricular and lumbar leukocyte count was lowest (Spearman`s rank correlation coefficient  = 0.79) . On aver = 0.79) . It was  = 0.79) .rS = 0.455), intermediate for CSF total protein and albumin (rS = 0.624 and 0.525), and highest for the CSF lactate concentration (rS = 0.767). The correlation of the lumbar versus ventricular erythrocyte density (rS = 0.224) was even lower than the correlation between lumbar and ventricular total leukocyte density and the percentages of the differential white blood cell counts (rS 0.271–0.644). On average, glucose concentrations were slightly higher in the ventricular than in the lumbar CSF, and lactate and erythrocyte density were approximately equal. The lumbar concentrations of albumin, total protein, IgG, IgA, IgM, and lumbar leukocyte densities were higher than the ventricular concentrations [Because the interval of ≤ 24 h between the analysis of lumbar and ventricular CSF is rather long in view of the rapid temporal changes occurring in CNS infections, a prospective study was performed with an interval between lumbar and ventricular sampling < 30 min in 25 patients  . Again, trations . These dn rS = 0.67. The cr = 0.944), protein (r = 0.679) and glucose (r = 0.805), whereas no statistically significant correlation of the ventricular and lumbar erythrocyte density was found. Again, on an average, the ventricular was higher than the lumbar glucose concentration, and the lumbar protein and leukocyte concentrations were higher than the respective ventricular concentrations. A reversal of these gradients, however, was present in at least one pair of samples [A recent study on 12 patients with an external ventriculostomy and an external lumbar drain  analyzing pairs of samples obtained at an interval < 24 h found a high correlation between cranial and spinal CSF leukocyte density ( samples . In 3 of samples ).In 16 patients with hydrocephalus as a consequence of neurocysticercosis, who received a ventriculoperitoneal shunt and a lumbar puncture at an interval of approximately 30 min, mean white blood cell density was 14 ± 5/μl in the ventricular and 72 ± 28/μl in the lumbar fluid. The protein content in the ventricular CSF was 280 ± 50 mg/l (mean ± SD) compared to 780 ± 120 mg/l in the lumbar CSF. Immunodiagnostic tests for cysticerci antigens (enzyme immunoassay and complement fixation) were positive in 12 of 16 ventricular and in all 16 lumbar samples. Again, glucose was higher in the ventricular than in the lumbar CSF .In contrast to albumin, which is synthesized by the liver, immunoglobulins can be synthesized by B-lymphocytes in the systemic circulation or in the CNS. For the detection of an inflammatory process involving immunoglobulin synthesis in the CNS it is insufficient to merely measure CSF immunoglobulin concentrations. For this reason, nomograms using CSF/plasma ratios for albumin and immunoglobulins in persons without diseases of the nervous system were established as references to discriminate between local immunoglobulin synthesis in the CNS and an increase in CSF protein concentration for other reasons . In contn = 25) [The Reiber–Felgenhauer nomograms are susceptible to errors, when the CSF is contaminated by blood Table . As shown = 25) , the eryn = 25) . The prol-lactate, whereas some bacterial strains causing meningitis can produce d-lactate. Since even in experimental meningitis caused by d-lactate-producing bacteria the d-lactate fraction in CSF was below 5% [l-lactate is usually measured in the clinical routine. As a consequence of anaerobic energy production in the CNS, under normal conditions the CSF levels of lactate are higher than the respective serum concentrations [Lactate is a small hydrophilic molecule (molecular mass 89 g/mol). As outlined, it distributes more evenly in the CSF than larger molecules in health and disease . Its conbelow 5% , l-lactatrations , 106. Sitrations . In expetrations . Lactatetrations . In a metrations .Since under anaerobic conditions the brain and CSF leukocytes produce lactate, lactate is not a specific marker of bacterial or fungal infection, but may be elevated in various conditions with elevated ICP or other causes of reduced cerebral perfusion. In 51 patients with subarachnoid hemorrhage and an external ventriculostomy, CSF lactate ranged from 1.9 to 6.2 mmol/L (median 3.2 mmol/l) . The lacDuring the course of bacterial meningitis, the most important CSF parameter predicting therapeutic efficacy or failure are the CSF-to-serum glucose ratio and CSF lactate concentrations . The CSFCompared to CSF lactate, CSF glucose possesses disadvantages: (1) The CSF lactate concentration is less dependent on the respective serum concentration than the CSF glucose concentration. Therefore, often the CSF-to-serum glucose concentration ratio is used instead of the CSF glucose level. (2) Serum glucose may rapidly change dependent on carbohydrate intake and insulin therapy. This probably impairs the stability of the CSF-to-serum glucose concentration ratio. (3) Unlike lactate, CSF glucose (and the CSF-to-serum glucose concentration ratio) possess a small, but not negligible ventricular–lumbar gradient.Temporal and spatial variations of CSF parameters and an impaired immune response can complicate the etiological diagnosis of CNS infections based on the CSF routine parameters leukocyte count, differential white blood cell count, protein, lactate, glucose and the CSF-to-serum rations of albumin, immunoglobulins and glucose. Patterns typical of certain infections may provide diagnostic hints, but are neither sensitive nor specific enough for effective clinical decision-makings. In clinical routine, the parameter which most readily equilibrates among the different parts of the CSF space, and has a relatively high sensitivity and specificity in the discrimination of bacterial/fungal versus viral meningitis and meningoencephalitis, is CSF lactate at a cut-off of 3.5(− 4) mmol/l. In severe non-infectious CNS diseases including intracranial hemorrhage, however, CSF lactate as a non-specific parameter of a suffering brain can also be elevated ≥ 3.5 mmol/l.Thus, establishing a definite diagnosis of a CNS infection by clinical chemical or cytological routine parameters is still not possible at present. Diagnosis of infections relies on the identification of the causative pathogen by culture or molecular methods (e.g., 16 s rRNA amplification ; multipl


Alzheimer's disease (AD) impacts the daily life of aging people. Oligomerized amyloid β (Aβ)‐associated neuronal senescence is involved in the pathological mechanism of AD. Blockage of neuronal senescence has been considered an important strategy for the treatment of AD. Midazolam is a hypnotic‐sedative drug with pleiotropic properties.1.42‐induced neurotoxicity have not been reported previously. Here, we investigate whether Midazolam possesses a beneficial effect against oligomerized Aβ1.42 in SH‐SY5Y neuronal cells.However, the effects of Midazolam in oligomerized AβCellular senescence was assessed using senescence‐associated β‐galactosidase staining. Telomerase activity was measured using the TeloTAGGG Telomerase PCR ELISA.1.42 significantly increased cellular senescence of SH‐SY5Y cells, but it was significantly alleviated by Midazolam. Additionally, Midazolam restored the oligomerized Aβ1.42‐induced reduction of telomerase activity. Interestingly, we found that oligomerized Aβ1.42 remarkably reduced human telomerase reverse transcriptase (hTERT) gene expression but increased the telomeric repeat‐binding factor 2 (TERF2) expression. However, treatment with Midazolam reversed the effects of oligomerized Aβ1.42 on the hTERT and TERF2 gene expressions. Importantly, the presence of Midazolam attenuated Aβ1.42‐induced p53 and p21 expressions. Mechanistically, Midazolam repressed the level of cyclooxygenase‐2 (COX‐2) and the release of prostaglandin E2. Importantly, overexpression of COX‐2 abolished the impact of Midazolam against oligomerized Aβ1.42 in neuronal senescence.First, the lactate dehydrogenase release assay demonstrates that 10 and 20 µM are the optimal concentrations of Midazolam used for cell cultures. Senescence‐associated β‐galactosidase staining results indicate that exposure to oligomerized AβWe conclude that the usage of Midazolam is a potential treatment strategy for AD.  As described before  with or without Midazolam  for 7 days.SH‐SY5Y neuronal cells were obtained from Procell  and cultured in MEM/F12 medium containing 15% FBS and 1% penicillin–streptomycin under the condition of 5% CO2.2SH‐SY5Y neuronal cells were plated in a 6‐well plate. The confluent cells were stimulated with 1, 2, 10, 20, 100, and 200 μM Midazolam for 24 h. To obtain the lactate dehydrogenase (LDH) level from cell culture, 120 μl cell supernatant was loaded into a 96‐well plate, then 60 μl LDH solution was added to each well. After incubating for half an hour in the dark, the absorbance value of each well was detected at 490 nm. The detected optical density value of each well was subtracted from that of the blank control well, and then the absorbance value of each group was standardized after normalization with the control group.2.32 and incubated for 12–15 h, followed by sealing to prevent evaporation of the dyeing solution in the culture plate. The number of positive cells was checked under the microscope .Cells were rinsed with 2 ml PBS, and then 1 ml fixing solution was added to each well, followed by being fixed for 15 min. After washing with 2 ml PBS, 1 ml senescence‐associated β‐galactosidase  staining solution was introduced into the wells. The culture plate was placed in an incubator with a constant temperature of 37°C without CO2.4g for 90 min. Isolated protein was quantified using the bicinchoninic acid (BCA) method, and the TeloTAGGG Telomerase PCR ELISA Plus Kit  was then utilized to detect the activity of telomerase. Subsequently, the telomeric repeat amplification protocol–PCR reaction system was introduced together with samples and primers. Lastly, RT‐PCR was used to perform the quantification.We measured the telomerase activity followed by the method described before . The following primers were used in this study: human telomerase reverse transcriptase (HTERT), forward, 5′‐GCCGATTGTGAACATGGACTACG‐3′, reverse, 5′‐GCTCGTAGTTGAGCACGCTGAA‐3′; telomeric repeat‐binding factor 2 (TERF2), forward, 5′‐GTGGAAAAGCCACCCAGAGAAC‐3′, reverse, 5′‐TGCAAAGGCTGCCTCAGAATCC‐3′; p53, forward, 5′‐CCTCAGCATCTTATCCGAGTGG‐3′, reverse, 5′‐TGGATGGTGGTACAGTCAGAGC‐3′; p21, forward, 5′‐AGGTGGACCTGGAGACTCTCAG−3′, reverse, 5′‐TCCTCTTGGAGAAGATCAGCCG−3′; GAPDH, forward, 5′‐GTCTCCTCTGACTTCAACAGCG‐3′, reverse, 5′‐ACCACCCTGTTGCTGTAGCCAA‐3′.Briefly, the TRIzol reagent was used to isolate RNAs from SH‐SY5Y neuronal cells, followed by transcribing into cDNA with a HiScript II Q RT SuperMix for qPCR (+gDNA wiper) . Subsequently, the 2 × SYBR Green PCR Master Mix  was used to perform the PCR reaction with specific primers to determine the gene levels. To monitor the gene expression, the relative RNA level was presented using the 22.6Cells in each group were lysed with lysis buffer, and the protein concentration was determined with the BCA method. The same amount of protein (20 μg) was added to the prepared gel, and electrophoresis was performed at 150 V at a constant voltage for 60 min. After electrophoresis, proteins were transferred to the polyvinylidene fluoride (PVDF) membrane, which was quickly put into 1 × TBST for rinsing .The cell supernatant was centrifuged at 4°C for 10 min at 1000 2.8p < .05 was considered a significant difference.All the experiments have been repeated at least three times. Data were presented as mean ± standard deviation and analyzed utilizing the GraphPad Prism software . The statistical analysis was performed with the one‐way analysis of variance method using the Tukey postK hoc test. 31–42‐treated SH‐SY5Y neuronal cells. We found that 10 and 20 μM were the two safe concentrations of Midazolam not threatening cytotoxicity in cell cultures. Moreover, Midazolam significantly alleviated senescence progression, including the increase in telomerase activity and the reduction of p53 and p21. Mechanistically, Midazolam repressed COX‐2 to alleviate Aβ1–42‐induced neuronal senescence.In this study, we investigated the beneficial effects of Midazolam in oligomerized Aβ3.1SH‐SY5Y neuronal cells were stimulated with 1, 2, 10, 20, 100, and 200 μM Midazolam for 24 h, followed by checking for its toxicity on SH‐SY5Y neuronal cells. The release of LDH Figure  was main3.21–42 (5 μM) with or without Midazolam  for 7 days, followed by evaluating cellular senescence. We found that the number of SA‐β‐gal staining positive cells  axis is involved in mediating cellular senescence  with oligomerized Aβ1–42 (5 μM) for 7 days. The efficacy of transfection was verified by real‐time PCR  and the catalytic protein subunit (hTERT) (Lingner et al., 2 axis critically regulates cellular senescence by shaping the composition of the senescence‐associated secretory phenotype and increasing senescence surveillance (Goncalves et al., 2 axis in Aβ1–42‐challenged SH‐SY5Y cells was profoundly repressed by Midazolam. Furthermore, the inhibitory effect of Midazolam on cellular senescence in Aβ1–42‐challenged SH‐SY5Y cells was largely rescued by the overexpression of COX‐2, suggesting that it might mediate the impact of Midazolam on cellular senescence in Aβ1–42‐challenged neurons. However, the effects of COX‐2 on p53 and p21 expression are still unknown. A further study will explore the underlying mechanism.Recently, it is reported that the COX‐2/PGEA receptors, promotes the binding of GABA to receptor A, and facilitates the opening of chloride channels to increase the inward chloride influx. In immature neurons, Midazolam altered the mRNA GABA receptor subunit expressions (Sinner et al., The limitation of the current study has to be addressed. First, the molecular mechanism of Midazolam in neurons remains to be fully understood in the setting of amyloid accumulation. Midazolam activates GABA51–42‐induced neuronal senescence by suppressing the COX‐2/PGE2 axis. These data suggest that Midazolam possesses neuroprotective effects in the setting of Aβ challenge in vitro; however, its efficacy remains to be investigated in an AD model.In summary, our data reveal that the hypnotic‐sedative drug Midazolam had a beneficial effect on alleviating cellular senescence in brain neuronal cells. Our study confirms that Midazolam ameliorated AβThe authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.All the authors agreed to publish this article.https://publons.com/publon/10.1002/brb3.2822The peer review history for this article is available at:


P < 0.001), compared with their position when the CFL was detached. The actual lift amount reached 2.0 ± 0.8 mm for the PLT and 1.9 ± 1.0 mm for the PBT. These results strongly suggest that a tensed CFL can lift the peroneal tendons and may act as a “tensioner” for the effective transmission of muscle contraction. This phenomenon contributes to postural control, especially in regaining balance on uneven terrain, and provides a new perspective for the exercise methods or understanding the ankle joint instability due to sprains.The ligaments are believed to have a role in stabilizing joints and regulating joint motion. Here, we propose a novel function of the calcaneofibular ligament (CFL), which stabilizes the ankle joint. In human bipedal locomotion, the peroneal muscles maintain mediolateral stability and prevent involuntary ankle inversion. To investigate the functional relationship between the peroneal longus tendon (PLT), brevis tendon (PBT), and CFL, we quantitatively analyzed the positional changes of the tendons by using a contactless three-dimensional optical scan system. Eighteen cadaveric specimens were included in the study. Interestingly, with increased tension of the CFL, the tendons significantly moved toward the lateral direction ( However, a precise understanding of the ligament has not been fully achieved. Several cadaveric studies have revealed structural variations in the CFL regarding length, width, running angle, or size of the attachment area4. Among the parameters, the running angle of the CFL ranges from 0° to 90°, as reported by Ruth5, and the angle shows a difference in tensing behavior, as noted in a simulation study by Edama et al.6. In contrast, Golanó et al.7 noted that the running angles of the CFL vary during the ankle movements because the CFL becomes horizontal in plantar flexion and vertical in dorsi flexion. Recently, Hattori et al.8 categorized the CFL structure into the mid-substance and fibular attachment. They described that direction of the mid-substance changed solely with the ankle joint position, and the CFL running angle variation could be interpreted as the different directions of the mid-substance in the various ankle position.The calcaneofibular ligament (CFL), which is a part of the lateral ligament complex, attaches the lateral malleolus to the calcaneus and stabilizes the ankle joint together with the anterior talofibular ligament (ATFL) and posterior talofibular ligament (PTFL). Injuries involving the CFL are usually severe4. An interesting finding is that histological examinations reveal fibrocartilaginous changes on the contact surface of the CFL with the peroneal tendons9. Benjamin and Ralphs10 mentioned that when tendons and ligaments are subject to compression, they are frequently fibrocartilaginous. Therefore, a hardened CFL, which indicates some mechanical interactions between the ligament and tendons, is highly possible. With this point of view, Shinohara et al.9 speculated that a stretched CFL lifted both tendons during ankle inversion. This speculation is quite interesting because the increased tendinous tension due to lifting may contribute to the contractile efficiency of the peroneal muscles. Thus, the CFL may function as a tensioner of the peroneal tendons to control ankle joint stability by enhancing muscle contraction.Regardless of the morphological variety, the CFL always closely crosses under the deep side of the peroneus longus tendon (PLT) and peroneus brevis tendon (PBT) Fig. 4. An intThis cadaveric study aimed to investigate whether the PLT and PBT are both lifted when the CFL is taut. Both tendons were assumed to change on an extremely small scale; therefore, we intended to use a contactless three-dimensional (3D) optical scan system to detect changes in positional coordinates set on the tendons.Before the examination, we assessed 26 ankles from 13 formalin-fixed Japanese cadavers (6 men and 7 women) that had been subjected to a gross anatomy course at the Faculty of Medicine, Saga University, Saga, Japan. The exclusion criteria were as follows: evidence of a previous injury or surgical treatment of the ankle region, severe deformity of the plantar skin, and a severe decrease in the extensibility of the peroneal muscles. Finally, 18 ankles from 12 cadavers (6 men and 6 women) were included in this study . Informed consent for the storage and use of the cadavers for research purposes was provided in advance by the donors and their relatives. All methods were performed in accordance with the relevant guidelines and regulations.11. The ATFL, PTFL, and deltoid ligament were also removed to ensure mobility of the talocrural and subtalar joints.During the gross anatomy course, the skin, subcutaneous tissue, and deep fascia of the lower legs and feet were removed, whereas the peroneus longus, peroneus brevis, and CFL were left intact. After the amputation of the lower legs at the knee joint, the PLT, PBT, and CFL were carefully exposed to measure the CFL dimensions accurately Fig. . In briex–y–z coordinate system defined by three sides of an alignment cube (3 cm on a side) placed behind the calcaneus in such a way as to contact one vertical side to the foot.To examine whether the PLT and PBT were lifted when the CFL was tensed, the positional changes of both tendons were evaluated using a contactless 3D optical scan system , which was set on an automatic turntable to allow rotation during scanning. In this step, the two markings on the foot and one side of the cube were aligned on a straight line of a grid sheet attached to the floor of the apparatus . The centers of the pinhead were specified as the positional coordinates, and distances from the origin’s coordinates were automatically calculated using WorkXPlore 2019 R1  on the 13, whereas the portion that intersected with the PLT and PBT was defined as the width  was calculated by subtracting the value under the condition in which the CFL was detached from the value of the intact CFL. The LA of the peroneal tendons was calculated using the following formula: LA = Each change in the positional coordinates on the P value of 0.05. The Shapiro–Wilk test was used to evaluate normality. The positional coordinates were compared using the paired t-test. Pearson’s correlation coefficient or Spearman’s rank correlation coefficient was calculated to investigate the relationships between the measured data and CFL dimensions. These statistical analyses were conducted using JMP Pro 15.2.0 .The level of statistical significance was set at a 15 to evaluate whether our data had sufficient verification power.Post-hoc power analysis was performed by using G* power version 3.1.9.6y-axis, the positions of the PLT and PBT were significantly higher with the intact CFL than with the detached CFL . These results demonstrated a statistical power value greater than 99% by a post-hoc power analysis. Furthermore, in all 18 samples, Δy had positive values, ranging from 0.2 mm to 2.8 mm for the PLT and 0.1 mm to 3.3 mm for the PBT. The average Δy was 1.5 mm for both tendons, and the 95% confidence intervals for the PLT and PBT were 1.1–1.9 and 1.0–1.9 mm, respectively  (range) of the LA, indicating the actual moving distance of the tendons, reached 2.0 ± 0.8 mm (0.8–3.5 mm) for the PLT and 1.9 ± 0.9 mm (0.5–3.8 mm) for the PBT.A comparison of the mean positional coordinates of the peroneal tendons before and after the detaching operation of the CFL is shown in Table ns Table . For they and the LA of both tendons and calculated the correlation coefficients for each parameter  and LA .To confirm the synchronous movement of the PLT and PBT, we prepared scatter plot diagrams of Δter Fig. . The diaThe mean values ± SD (range) of the measured CFL dimensions were: 15.1 ± 2.6 mm (10.0–20.0 mm) for length, 4.8 ± 0.6 mm (4.0–6.0 mm) for width, and 50.9° ± 9.8° (22.9°–61.6°) for the running angle , width (4.8 ± 0.6 mm vs. 4.68–5.64 mm17), and running angle (50.9° ± 9.8° vs. 43°–47°18). The CFL with two crossing fiber bundles6 was not observed. Thus, the samples were morphologically unbiased in this study.When the mean dimensional values of the CFL were compared with those of past cadaveric studies that employed the same measuring method, the values were similar for length  since up to 40% of patients with lateral ankle sprain develop CAI24. The CFL has an important role in the talocrural and subtalar joints stability25; therefore, CFL injury or CAI due to lateral ankle sprain would result in loss of “joint stabilizer” and “tensioner” functions. While many studies have revealed the specific pathological effects of the joint stabilizer function25, many reports have also demonstrated data which are interpreted that the tensioner function exists. That is, the reaction time of the peroneal muscles is significantly longer in patients with CAI than in those without CAI26. This finding could be explained by the fact that a slack ligament due to injury may delay the reaction time because of the loss of tensioner function. In that case, reacquisition of tensioner function by surgical treatment will also be of interest. Furthermore, no consensus exists regarding the optimal training method for the peroneal muscles27. Recently, Bavdek et al. proposed walking on a medially incline ramp or changing incline in the frontal plane as a strengthening exercise for peroneal muscles22. This method seems very effective because the tensioner function is exerted maximally. Thus, the present results would provide a new perspective on broad issues.In human bipedal locomotion, the main function of the peroneal muscles is mediolateral stability and prevention of involuntary ankle inversion at foot strikeThis study has some limitations. The position that induces maximum CFL tension may vary between individuals; however, all samples were examined under a specified position to achieve reliable coordinate measurements. Another approach should be considered to clarify interindividual differences. Second, in the cadaveric studies, the lack of detailed records regarding sprain or CAI during the lifetime and some variations in the hardness of the ligament and tendons caused by formalin fixation may have affected the accuracy of our findings. Third, the in vivo CFL and tendons were thicker because the surrounding connective tissue, including the tendon sheath, was completely removed during the sample preparation. Therefore, a more apparent lift-up behavior will likely occur in living bodies. However, the fibular retinaculum and skin may inhibit the tensioner effect by reactive compressive force. To verify the hypothesis of the current paper, functional studies using living bodies and non-invasive modalities, such as ultrasonography, will be required.To the best of our knowledge, we believe this study is the first to demonstrate an undiscovered function of the ligament in the lateral ankle region. Our results showed that the tensed CFL lifts the peroneal tendons and strongly suggested that the CFL acts as a “tensioner” for the effective transmission of contractile activity of the peroneal muscles. This phenomenon may contribute to ankle joint stability and postural regulation during gait or exercise when the foot is in the inversion position. These findings provide a new perspective to develop or modify exercises for the peroneal muscles and better understand ankle joint instability caused by sprains.Supplementary Information.