Late-onset asthma (LOA) is beginning to account for an increasing proportion of asthma patients, which is often underdiagnosed in the elderly. Studies on the possible relations between aging-related genes and LOA contribute to the diagnosis and treatment of LOA. Forkhead Box O3 (FOXO3) and TP53 are two classic aging-related genes. DNA methylation varies greatly with age which may play an important role in the pathogenesis of LOA. We supposed that the differentially methylated sites of FOXO3 and TP53 associated with clinical phenotypes of LOA may be useful biomarkers for the early screening of LOA.p-value < 0.05) and clinical lung function in LOA patients was evaluated. Next, candidate DMSs combining with age were evaluated to predict LOA by receiver operating characteristic (ROC) analysis and principal components analysis (PCA). Finally, HDM-stressed asthma model was constructed, and DNA methylation inhibitor 5-Aza-2′-deoxycytidine (5-AZA) were used to determine the regulation of DNA methylation on the expression of FOXO3 and TP53.The mRNA expression and DNA methylation of FOXO3 and TP53 in peripheral blood of 43 LOA patients  and 60 healthy controls (HCs) were determined. The association of methylated sites with age was assessed by Cox regression to control the potential confounders. Then, the correlation between differentially methylated sites  and chr17:7591672 (TP53), were associated with the severity of LOA. The combination of the two DMSs and age could predict LOA with high accuracy . In HDM-stressed asthma model, DNA demethylation increased the expression of FOXO3 and P53.The mRNA expression of FOXO3 and TP53 varies significantly in peripheral blood of LOA patients, which may be due to the regulation of DNA methylation. FOXO3 and TP53 methylation is a suitable blood biomarker to predict LOA, which may be useful targets for the risk diagnosis and clinical management of LOA. Asthma symptoms can occur at any time in life though many patients may develop asthma during childhood. The endotype of asthma has a diversity of distinguished clinical phenotypes which is critical for the diagnosis and treatment of different types of asthma. LOA is defined as a kind of asthma with onset of symptoms in adult life in a patient with no pre-existing, persistent respiratory symptoms , 2. WithFOXO3 and TP53 are two classic genes that regulate aging and longevity , 10. FOXLungs exchange gas with the outside world, which are highly susceptible to external environmental conditions . TherefoCriteria for the diagnosis of asthma were recommended by the 2019 Global Strategy for Asthma Management and Prevention . BrieflyLOA is one of the main endotypes of asthma that have been identified . It refe1, FEV1% predicted, FVC, the ratio of FEV1 to the FVC, PEF, FEF75, FEF50 and FEF25. The study was approved by No. 20180308 of the Xiangya Hospital Ethics Review Committee and participants provided written consents to participate in this study.60 healthy controls (HCs) and 43 previously confirmed LOA patients (> 20 years of age) were selected from the Respiratory Department and the Medical Examination Center of Xiangya Hospital in China from October 2018 to January 2019. After Written informed consent was obtained from each patient, questionnaire information , pulmonary function testing, peripheral blood and induced sputum samples were collected. Lung function test included the spirometric values of FEVPeripheral blood and induced sputum samples were collected from the enrolled 60 HCs and 43 LOA patients, respectively. Peripheral blood was collected into 5 ml EDTA anticoagulation tubes and then transferred to a centrifuge tube. After adding 2 volumes of erythrocyte lysate and lysing for 5 min, peripheral blood cells were pelleted by centrifugation.For sputum induction, each volunteer inhaled a 4.5% saline atomized solution three times for 5 min each time and coughed sputum into a separate cup. Then, four-parts 0.1% dithiothreitol was added to one-part sputum and mixed for 15 min before adding four-parts phosphate-buffered saline. Finally, sputum cells were pelleted by centrifugation and used for RNA extraction after filtering .Total mRNA was purified from peripheral blood and induced sputum cells using Trizol (Invitrogen) and quantified by an ultraviolet spectrophotometer  . 1 µg RNGenomic DNA was extracted from peripheral blood with the TIANamp Genomic DNA kit  . DNA quaThis animal study was carried out with the approval No. 2020sydw0178 of Xiangya Animal Protection and Utilization Committee of Central South University. All methods are implemented in accordance with relevant clauses. The mice were placed under the condition of an air‐filtered temperature control unit, where the light and dark cycled alternately for 12 h, with sufficient food and water supply. The HDM‐induced asthma model was constructed according to previous publications . In the Lungs were inflated, fixed in 4% paraformaldehyde, and embedded in paraffin blocks and cut into 5 μm sections. Immunohistochemistry staining was performed with the following antibodies: p53 , FOXO3 . For microscopy, we employed Zeiss Axio Scope.A1 or Zeiss Discovery.V8 Stereo microscopes  integrated with an Axio-Cam ICc3 camera . Images were obtained by AxioVision Rel. 4.7 software from Zeiss.1, FEV1%, FEV1/FVC. Receiver operating characteristic (ROC) curves were obtained to elucidate the accuracy of differentially expressed CpG sites in predicting LOA. Age and the methylation percentage of candidate CpG sites were used in principal components analysis (PCA) to identify LOA. A two-tailed p-value < 0.05 was considered statistically significant.Statistical analyses were conducted using SPSS version 22.0 . The characteristic data of recruited LOA patients and HCs were presented as Mean ± SD and analyzed with Chi-squared test or Mann–Whitney U test. The mRNA expression and the methylation array of FOXO3 and TP53 were analyzed by One-way ANOVA followed by Dunnett's post hoc test. The method of Benjamin Hochberg was used to control the false discovery rate (FDR). Logistic regression analysis was performed on selected differentially expressed CpG sites, with potential risk factor of age, gender and smoking history . PearsonTo detect the expression of FOXO3 and TP53 in LOA patients, we recruited 60 HCs and 43 LOA patients . The percentage of eosinophils and neutrophils in LOA patients was significantly higher than that of HCs. There was no statistical difference in age between these groups  and first exon regions is closely related to gene transcription , we asseTo assess the effect of DMSs more accurately on LOA, logistic regression with age, gender or smoking history was used to determine other possible potential risk factors. The results demonstrated that the age of participants in the study was related to LOA but not to gender and smoking history (data not shown). Hence, a logistic regression analysis with potential risk factors adjustment was further performed. Finally, 7 DMSs in FOXO3 and 1 DMS in TP53 were statistically significant, and the absolute value of methylation difference between the 8 sites ranged from 0.31% to 11.70%  were related to age and lung function indicators  and chr17:7591672 (TP53) were closely related to the severity of LOA. On this basis, we further explored the possible regulation of gene expression by the two DMSs and the feasibility of the two DMSs as LOA biomarkers. Hypermethylation modifications usually result in decreased target gene expression , 64. ConAlthough these specific CpG sites may provide blood biomarker to predict and screen LOA, there are still some limitations in this study. The first one is the relatively limited sample size, a new LOA cohort or Long‐standing asthma cohort (Age of onset < 12 years) should be collected in our further work. Besides, the regulation of the specific CpG site methylation on the expression of FOXO3 and TP53 still needs further verification. Moreover it have been verified that FOXO3 and TP53 are expressed in eosinophils and neutrophils –69. FurtIn summary, this study verified the altered mRNA expression of FOXO3 and TP53 in the peripheral blood of LOA patients, which may be due to the regulation of DNA methylation. Besides, the DNA methylation levels of the chr6:108882977 in FOXO3 and chr17:7591672 in TP53 in peripheral blood were associated with the clinically phenotypic which may be used as effective blood biomarkers of LOA. These results provide some useful insights into the molecular mechanisms of aging-related genes in LOA which provide some potential useful targets for early prediction and screening of LOA.Additional file 1: Table S1. Details of CpG regions in the CpG islands of FOXO3 and TP53.Additional file 2: Table S2. Methylation level of each CpG site in the CpG islands of FOXO3 and TP53.Additional file 3: Table S3. Correlation between mRNA expression of FOXO3 and TP53 and clinical parameters in LOA patients.Additional file 4: Table S4. Correlation between DNA methylation levels and clinical parameters in LOA patients.Additional file 5: Figure S1. The sensitivity and specificity of 6 DMSs in predicting LOA.


BK virus allograft nephropathy is a serious complication after kidney transplantation, and the effect of pre-emptive intervention for high-level BK viruria has been verified, but protocols after kidney transplantation for early identification of high-level viruria are lacking.This was a single-center study. The clinical data of the kidney transplant recipients and their donors in our center from January 1, 2015 to December 31, 2018, were collected. The patients were divided into the high-level BK viruria group (Group A) and a non-high-level BK viruria group (Group B) according to the qPCR results of BK virus DNA loads in urine samples. Significant variables were screened out by univariate analysis, and then the results were incorporated into a multivariate logistic regression model to analyze the independent risk factors for high-level BK viruria.P < 0.001), history of ATG/ATG-F application (P = 0.033), acute rejection (AR) (P = 0.004) and delayed graft function (DGF) (P < 0.001) were related to high-level BK viruria. After the inclusion of the multivariate logistic regression model, the results showed deceased brain and cardiac donors , AR  and DGF .A total of 262 recipients were included in the study. The incidence of high-level BK viruria was 13.4% (n = 35), and the median time of detection was 181 (range 91–1119) days. Univariate analysis showed that donor type (Donation by deceased brain and cardiac patients, history of AR and DGF were independent risk factors for high-level BK viruria after kidney transplantation. BK virus allograft nephropathy (BKVAN) is a severe disease caused by BK virus (BKV) infection or reactivation, which often impairs kidney function irreversibly and is more common in kidney transplant recipients . The usuThis study summarized the incidence of high-level BK viruria from 2015 to 2018 and analyzed its risk factors to identify patients at an early stage and provide treatment to prevent the occurrence of BKVAN.In this study, we retrospectively collected the data of kidney transplant recipients from January 1, 2015, to December 31, 2018, at Jiangxi Provincial People’s Hospital Affiliated with Nanchang University. This study was approved by the Ethics Committee of Jiangxi Provincial People’s Hospital . The patients were divided into two groups according to the different monitoring results of BKV DNA loads in urine after transplantation: high-level BK viruria (Group A) (BKV DNA loads ≥ E + 07 copies/ml) and non-high-level BK viruria (Group B) (BKV DNA loads < E + 07 copies/ml or no BKV DNA in urine samples). BKV evaluation was performed on the day before transplantation to ensure that all patients were BKV negative before transplantation. After transplantation, BKV monitoring was carried out for all patients according to our monitoring protocol. We included patients in group A when the detected BKV DNA load in urine samples was ≥ E + 07 copies/mL and there was no BK viremia. The patients included in group B had a urine BKV DNA load < E + 07 copies/ml or lacked a BKV DNA load. Similarly, we needed to exclude all patients with BK viremia. The exclusion criteria were as follows: 1. the patient was diagnosed with BK viremia; 2. no regular follow-up data were kept after the operation; 3. the patient died or lost the transplanted kidney during the study period.We followed the monitoring protocols recommended by the guidelines published by the American Society of Transplantation . RegularThe clinical data of all the recipients and their donors were collected, including donor factors , recipient factors , and immune factors .P < 0.1 were selected to enter the multivariate analysis. The results of univariate analysis (P < 0.1) were included in the multivariate logistic regression model analysis, and the forward LR method was used to screen the variables and eliminate the unintentional variables. P < 0.05 was considered to indicate statistical significance.The statistical analyses were performed with IBM SPSS software, version 25.0 . The results of continuous variables were expressed as the mean ± standard deviation (SD) or median  and numerical (percentages) values were used for categorical variables. The chi-square test and the independent sample T-test were used for the single-factor comparison between the two groups. In the single factor analysis, the interaction between the factors was not excluded. To avoid missing meaningful variables, the results with a A total of 262 patients were included in this study, including 35 in group A and 227 in group B. All recipients were negative for panel reactive antibody and complement-dependent cytotoxicity before transplant. The demographic and clinical characteristics of all subjects are shown in Table P value < 0.1 were input into the logistic regression model for multifactor analysis to identify the independent risk factors. The treatment history of ATG/ATG-F, AR, DGF and donor type were included in the regression model. The results showed the treatment history of ATG/ATG-F , AR , DGF  and DBCD   were diagnosed with high-level BK viruria at the median time of 181 days in our center. In 2020, several scholars summarized the incidence of BK virus infection in Asia and found that the incidence of BK viruria after kidney transplantation was between 5.9 and 86.9% . AlthougIt is important to identify high-level BK viruria patients. Reischig et al.  have demAs we know, no relevant studies have been reported internationally in this field, so we selected certain variables that may influence the occurrence of high-level BK viruria for analysis. The final results showed that DBCD, AR and DGF were independent risk factors for high-level BK viruria. AR and DGF were the expected results, and AR and DGF were also independent risk factors for BK viremia after renal transplantation . HoweverThis study reports the risk factors for high-level BK viruria after renal transplantation, filling in the gaps in this field preliminarily, but there are also some limitations. In this single-center retrospective study, the insufficient sample size was still a limitation of its quality. Due to the limitation of objective factors, we could not compare all relevant factors. For example, there are limited types of immunosuppressive drugs that we use after transplant, induction therapy is not routine, donor-sourced BK virus surveillance has not been carried out, and so on.In the first 6 months after kidney transplantation, enhanced monitoring of the frequency of BKV infection is necessary, and DBCD kidneys and a history of AR/DGF are independent risk factors for high-level BK viruria. Therefore, we can use DBCD kidneys as a basis to identify patients early on and treat them. In future studies, prospective multicenter studies will always be an important direction in the field of BKV infection.


National guidelines for identifying physiological deterioration and sepsis in hospitals depend on thresholds for blood pressure that do not account for age or sex. In populations outside hospital, differences in blood pressure are known to occur with both variables. Whether these differences remain in the hospitalised population is unknown. This database analysis study aims to generate representative centiles to quantify variations in blood pressure by age and sex in hospitalised patients.Retrospective cross-sectional observational database analysis.Four near-sea-level hospitals between April 2015 and April 201775 342 adult patients who were admitted to the hospitals and had at least one set of documented vital sign observations within 24 hours before discharge were eligible for inclusion. Patients were excluded if they died in hospital, had no vital signs 24 hours prior to discharge, were readmitted within 7 days of discharge, had missing age or sex or had no blood pressure recorded.Systolic blood pressure (SBP) for hospitalised patients increases with age for both sexes. Median SBP increases from 122 (CI: 121.1 to 122.1) mm Hg to 132 (CI: 130.9 to 132.2) mm Hg in men, and 114 (CI: 113.1 to 114.4) mm Hg to 135 (CI: 134.5 to 136.2) mm Hg in women, between the ages of 20 and 90 years. Diastolic blood pressure peaked around 50 years for men 76 (CI: 75.5 to 75.9) mm Hg and women 69 (CI: 69.0 to 69.4) mm Hg. The blood pressure criterion for sepsis, systolic <100 mm Hg, was met by 2.3% of younger (20–30 years) men and 3.5% of older men (81–90 years). In comparison, the criterion was met by 9.7% of younger women and 2.6% of older women.We have quantified variations in blood pressure by age and sex in hospitalised patients that have implications for recognition of deterioration. Nearly 10% of younger women met the blood pressure criterion for sepsis at hospital discharge. Changes in blood pressure by age and sex are currently unknown for the hospitalised population.A large cross-sectional database of 75 342 patients were used to derive blood pressure centiles.Results have implications on how sepsis and other in-hospital deterioration are detected in routine care.Though patterns match those seen in out-of-hospital longitudinal studies, cross-sectional analysis can be affected by survival bias and birth cohort effects.Routine measurement of blood pressure is a key component of patient surveillance and diagnosis in hospitals worldwide. Currently, in-hospital assessment of blood pressure is undertaken by comparison to generic population normal ranges.The ability to use an individual’s physiology to monitor them for signs of deterioration may be improved by taking into account factors that affect these normal ranges.In populations outside hospital, differences in blood pressure are known to occur with both age and sex.To quantify these differences, our objective was to define representative centiles of the stable hospitalised population for SBP, diastolic blood pressure (DBP) and pulse pressure (PP) by age and sex via an analysis of a large near-sea-level database of routinely collected vital signs. Description of this group allows inference about unstable patients via one-class classification (novelty detection), which has previously been used when a clinical outcome of interest is relatively uncommon.We conducted a cross-sectional analysis from a database collated at Oxford University Hospitals (OUH) NHS foundation trust group of hospitals. The OUH consists of four hospitals: an urban teaching hospital, a general district hospital and two specialist hospitals. Our data set included patients admitted to the OUH between April 2015 and April 2017.All adult patients who were admitted to OUH and had at least one set of documented vital sign observations within 24 hours prior to discharge were eligible for inclusion. Patients were excluded from the analysis if they (1) died in hospital, (2) had no recorded vital signs 24 hours prior to discharge, (3) were readmitted within 7 days of discharge, (4) had missing recordings for age or sex or (5) had no blood pressure recorded.We collected the final recorded set of blood pressure observations from a patient’s first attendance to the OUH hospital group during the study period. This ensured that the centiles were not biased towards repeat attenders or patients with longer hospital stays. Blood pressure was measured using automated devices ratified for clinical use as part of routine clinical care and electronically documented using the SEND e-Obs software.Admissions were typed as either elective, emergency or other, according to the admission method code. Codes are defined in full within the NHS data dictionary.12The characteristics of the study population were described using medians and quartiles for the continuous variables, and frequencies otherwise. We calculated median and representative centiles  for blood pressure at all ages between 20 and 90, for men and women. SBP and DBP are presented separately. One further measure, the PP was derived as the arithmetic difference between SBP and DBP and was also analysed using the same methods.In subgroup analyses, we produced separate representative centiles by age and sex for emergency and elective admissions, and for patients without a diagnostic code for hypertension.Centiles were estimated using the Generalised Additive Models for Location, Scale and Shape framework (GAMLSS).All analyses were undertaken using R and the GAMLSS package.16We used all the available data and therefore no formal sample size calculation was undertaken. To ensure that the sample was sufficient, the precision of the centiles was estimated via a bootstrapping procedure, whereby the dataset was sampled with replacement to create a new dataset and the analysis was carried out.10.1136/bmjopen-2019-033618.supp1Supplementary dataPatients or the public were not involved in the design, or conduct, or reporting, or dissemination plans of our researchA total of 83 004 patients were admitted to the hospital trust during the period of study and received at least one observation on the SEND e-Obs system. Of these, 75 342 patients were included in the study. Blood pressure was missing in 885 (1.17%) records. Other reasons for exclusion are shown in Centiles by age for SBP, DBP and PP are shown in For men, there was a modest reduction in median PP between the ages of 20 and 40 years from 55 mm Hg (CI: 54.6 to 55.9) to 50 mm Hg (CI: 49.2 to 50.0), whereas for women PP remained constant at 47 mm Hg . Median A total of 19 669 patients had an ICD-10 diagnostic code for hypertension. Of these, 24.0% (4711) had an SBP of <120 mm Hg and 2.3% (453) had a low SBP of <100 mm Hg at the time of discharge. Per-decade percentages were not calculated as small numbers of patients means that CIs are wider than any differences between decades.We have generated blood pressure centiles for age and sex from a large multi-hospital patient database.Discharge blood pressures  showed clinically significant differences across age ranges and by sex. SBP progressively increased with age for both sexes, but progression was greater in females. DBP increased and then decreased across the life course for both sexes. The fluctuation in DBP was greater for men than for women. These overall trends were visible in both the whole population, and for the cohort that did not have a diagnostic code for hypertension.In populations outside hospital, these patterns are known to exist.22–24While the overall patterns for blood pressure are known to exist for the general population outside hospital, we believe that this is the first time that centiles have been derived from an in-hospital setting.Vital signs were recorded as part of standard clinical practice, so the conditions for data recording were not controlled. This may have directly impacted the measured values. For instance, the state of wakefulness of the patient, which is known to be associated with blood pressure and pulse rate, was unknown.We used the last recorded blood pressure in the 24 hours prior to discharge. While this lessens many biases in comparison to other methods and may represent the blood pressure recording when the patient is most stable, there may be other patterns at different points during a hospital admission.Finally, this study uses a cross-sectional cohort, so the derived centiles may be affected by survival bias and birth cohort effects.28The differences in normal vital sign ranges due to age and sex could have substantial implications for hospital practice. For example, A total of 19 669 patients diagnosed with hypertension had normal or low SBP immediately prior to discharge. This cohort may be reasonably assumed to be prescribed with anti-hypertensives for the purpose of managing blood pressure. While we do not have information on blood pressure medication following discharge, this group may be considered an estimate of the upper-bound of those at risk of medication withdrawal. Inappropriate blood pressure medication withdrawal has been associated with higher risk of further complications.30Another important application for age and sex stratification is Early Warning Scores (EWS). In these systems, integer scores are assigned to each vital sign according to deviation from normality. The aggregate score is then used to guide appropriate clinical care. One such EWS, the National Early Warning Score (NEWS), has been validated in a large in-hospital population and is widely used in the UK and the Ireland.31Based on the results presented, an age-stratified score could strongly affect the quality of care a patient receives. For instance, from Until now, age and sex have not been included within any adult EWS, despite evidence indicating its limitations in predicting deterioration of elderly patients.32The data set was collected from all four hospitals, but we note that there are high proportions of white Caucasian patients. Previous studies have shown correlation between ethnicity and differences in blood pressure trajectory.Our work shows for the first time that meeting current blood pressure criteria for sepsis or early warning system alerts is substantially more likely in younger women than in all other groups in the 24 hours prior to discharge. Exploration of this finding is needed to determine whether adjustment for age and sex can improve discrimination and prevent overdiagnosis.Substantial variations in the final blood pressure recorded in the 24 hours prior to hospital discharge occur with age and sex. These result in large differences in the proportions of patients meeting the blood pressure criterion for sepsis and early warning score alerts. These factors should be examined to understand whether these factors could be used to improve discrimination between stability and deterioration.


Hypertension is a global public health issue and is closely related to chronic kidney disorder (CKD). In people with CKD, strict monitoring of blood pressure is an important part of therapy.The aim of this research was to validate the iHealth Track blood pressure monitoring device for patients with CKD according to the European Society of Hypertension International Protocol 2010 (ESH-IP2).In total, 33 patients who received hemodialysis in Plasencia participated in the study. There were 9 successive measurements made, which conformed to the ESH-IP2. We calculated the differences between the standard reference device (Omron M3 Intellisense) and the test device  for blood pressure and heart rate values. For 99 total comparisons of paired measurements, we classified differences into various categories .In 90 of 99 systolic blood pressure and 89 of 99 diastolic blood pressure comparisons between the devices, measurement differences were within 5 mmHg. In 81 of 99 heart rate comparisons between the devices, measurement differences were within 3 beats per minute. The mean differences between the test and reference standard measurements were 3.27 (SD 2.99) mmHg for systolic blood pressure, 3.59 (SD 4.55) mmHg for diastolic blood pressure, and 2.18 (SD 2.75) beats per minute for heart rate.We also observed that for both systolic and diastolic blood pressure, 31 of 33 participants had at least two of three comparisons between the devices with measurement differences less than 5 mmHg. For heart rate, 28 of 33 patients had at least two of three comparisons between the devices with measurement differences less than 3 beats per minute.To our knowledge, this is the first study to show that iHealth Track meets the requirements of the ESH-IP2 in patients with CKD. Therefore, the iHealth Track is suitable for use in renal patients. Chronic kidney disorder (CKD) is a syndrome defined by persistent alterations in renal function or structure that cause complications in a patient's health. Some of the structural anomalies may be tumors, cysts, malformations, or atrophies. In addition, renal dysfunction can be manifested through alterations in the output or grade of urine, increased risks of intellectual disabilities in children, edema, and hypertension ,2.In fact, the diseases most related with CKD are hypertension and diabetes, especially in high- and middle-income countries ,4. HyperThe interaction between CKD and hypertension is complex and increases the probability of cerebrovascular and cardiovascular problems -11. In sMonitoring of BP should be done with devices that are easy to use and accurate ,15-18. TThe study protocol was approved by the Institutional Local Research and Ethical Committee . In conducting this study, we complied with the ethical principles of the Declaration of Helsinki , includiThe standard device we used for reference was the Omron M3 Intellisense , which has been validated according to the International Protocol for the general population  as well The test device was the iHealth Track automatic appliance with serial number KN-550BT , which registers brachial BP with the oscillometric protocol. It detects BP between the range of 0 mmHg to 300 mmHg (measuring precision ±3 mm Hg) and HRs within the range of 40 to 180 beats per min (measurement precision ±5%). The device’s arm cuff is 22 to 42 cm around.The device’s liquid crystal display screen shows the measured systolic (S) BP, diastolic (D) BP, and HR values. The device unit has enough memory for 99 recordings. Additionally, this device unit can be used with Apple Bluetooth 4.0 devices and certain Android Bluetooth 4.0 cellular phones through an application named Health MyVitals. This means that BP and HR data can be stored on wireless devices connected to iHealth Track and then displayed graphically.We recruited patients with CKD who attended the Fresenius Medical Care dialysis clinic in Plasencia, Spain. A total of 33 patients who met the selection criteria participated. The inclusion criteria were adults 25 years of age or older that received hemodialysis. We sought at least 10 male participants and 10 female participants. The exclusion criteria, which were created according to the ESH-IP2 ,22, wereThe professional validation team consisted of 2 nurses with senior experience (more than 6 years) in BP measurement. The measurement area was correctly conditioned to a suitable temperature, and factors that could affect the records, such as noise or distractions, were removed ,22. All 2. The circumference of the patient’s arm was measured to ensure that the cuff size was adequate.After dialysis, each patient first reported information regarding their sex, age, height, and dry weight. In addition, we calculated participants’ BMI using Quetelet´s index in kg/mNext, patients sat in the measurement room and BP measurements were started after a 10- to 15-minute rest period. Each patient was seated in a standard-size plastic chair with a backrest and armrests.In total, 9 consecutive measurements were made on each participant with the Omron M3 Intellisense (5) and the iHealth Track (4) as follows ,22:(BPA): input BP, by the standard device unit(BPB): device BP detection by the test device unit(BP1): standard device unit(BP2): test device unit(BP3): standard device unit(BP4): test device unit(BP5): standard device unit(BP6): test device unit(BP7): standard device unitDuring the measurements, participants remained calm, quiet, seated, and still. Participants kept their backs straight and feet on the floor in a parallel position . They rested their arms on a flat surface with their palms facing upwards and their elbows slightly flexed so that their fists were at the height of their hearts.BP records were made at heart level on the right arm in 31 participants and on the left arm in 2 participants (because of an arteriovenous fistula on the right arm). The standard cuff size 22-32 cm) for the Omron M3 Intellisense was used for all men (20). For women, the standard cuff (22-32 cm) was used for 11 participants, and the large cuff (32-42 cm) was used for 2 participants. Since the iHealth Track has only one cuff size (22-42 cm), all measurements were taken with it. The interval between one measurement and the next was 30 to 60 seconds [-32 cm foAll measurements were made for each participant during their hemodialysis appointment, after the dialysis was complete. The relative values were then used to calculate the mean difference between the reference device readings and the test device readings. All participants were receiving hemodialysis in the Fresenius Medical Care dialysis clinic in Plasencia.We analyzed the data with the software SPSS Statistics, version 19.0 . We reported the findings as mean (SD).According to the ESH-IP2, the accuracy of a device is based on a comparison between the test device  and the standard reference device (Omron M3 Intellisense) measurements. For each participant, we first compared measurements BP2, BP4, and BP6 with measurements BP1, BP3, and BP5, respectively, and then compared measurements BP3, BP5, and BP7 with each other. The most favorable comparisons were used.In our comparisons, we classified differences for both SBP and DBP, separately, by whether paired values were within 5, 10, or 15 mmHg , and wheMoreover, we produced Bland-Altman plots ,28 to diOf the 34 participants we recruited, 33 completed the study successfully (one was excluded for device failure). The 33 participants included 20 men and 13 women. The iHealth Track BP device validation results were taken in accordance with the ESH-IP2. The mean differences between the reference standard and test devices were 3.27 (SD 2.99) mmHg for SBP and 3.59 (SD 5.28) mmHg for DBP. In 90 out of 99 SBP and 89 out of 99 DBP comparisons between the devices, measurement differences were within 5 mmHg, exceeding the ESH-IP thresholds (>72 comparisons for SBP and >64 comparisons for DBP). Additionally, in 95 out of 99 SBP and 94 out of 99 DBP comparisons between the devices, measurement differences were within 10 mmHg, also exceeding the ESH-IP thresholds (>86 comparisons for SBP and >80 comparisons for DBP). Moreover, in 98 out of 99 SBP and 94 out of 99 DBP comparisons between the devices, measurement differences were within 15 mmHg, which again surpasses the ESH-IP thresholds (>95 comparisons for SBP and >92 comparisons for DBP). Therefore, the iHealth Track passed part 1 of the validation protocol for BP monitoring.Regarding part 2 of the ESH-IP2, 31 out of 33 participants had at least two of the three comparisons between devices with measurement differences within 5 mmHg for both SBP and DBP, exceeding the ESH-IP threshold (>23 participants). One participant had all three comparisons for both SBP and DBP with measurement differences greater than 5 mmHg, which is less than the ESH-IP maximum of 3 participants. Given these results, the iHealth Track also passed part 2 of the validation protocol for BP. Thus, because the iHealth Track passed parts 1 and 2 of the BP validation protocol, it passed part 3 of the protocol, overall validation.The validation findings for the iHealth Track HR monitoring device were taken according to the ESH-IP2. The mean difference between the reference standard and test devices was 2.18 BPM (SD 2.75). In comparisons between devices, 81 out of 99 pairs of measurements were within 3 BPM, 91 out of 99 were within 5 BPM, and 96 out of 99 were within 8 BPM. These results indicate that the iHealth Track passed part 1 of the validation protocol for HR.Regarding part 2 of the ESH-IP2, 28 out of 33 patients had at least two of three comparisons between devices with measurement differences within 3 BPM, which exceeds the ESH-IP threshold (>23 participants). Only 2 participants had all three HR comparisons with measurement differences greater than 3 BPM, which is less than the ESH-IP maximum of 3 participants. Therefore, the iHealth Track passed part 2 of the HR validation protocol and, consequently, part 3  as well.The Bland-Altman graphs -3 give fThis study is the first to validate the accuracy of the iHealth Track device for BP and HR recordings in patients with CKD. The results indicate that the iHealth Track, as used in renal patients, passed the ESH-IP2 validation requirements. We previously validated the iHealth Track device for the general population following ESH-IP2 .This study showed two limitations. Although the iHealth Track has been validated in the general population and now in patients with CKD, we cannot necessarily extrapolate our results to other specific populations. In addition, patients with CKD have stiffer arteries than other people ,11,14. WThe ESH-IP2 for home BP monitoring highlights the need for specific validation in patients with end-stage renal disease ,30 as stHowever, there are few devices that have been validated in patients with renal disease ,31-34. AThe purpose of the ESH-IP2 was to simplify the previous protocols of the British Hypertension Society (BHS)  and the Our study is the first to show that the iHealth Track device meets the requirements of ESH-IP2 in patients with CKD. Future versions of the ESH-IP should include explicit criteria for validation in specific populations. Validation of this device would be valuable in other specific populations such as pregnant women, older adults, and patients with arrhythmias.


The clinical records of dogs receiving intra or postoperative labetalol infusion were retrospectively reviewed. Invasive systolic (SAP), mean (MAP) and diastolic (DAP) arterial pressure and heart rate (HR) before labetalol infusion (T0) and 15, 30, 45 and 60 min  after infusion were retrieved. The dose rate of labetalol infusion and use of concurrently administered drugs that could have potentially affected ABP and/or HR were also recorded. ANOVA for repeated measures and Dunnett’s multiple comparison test were used to determine the effect of labetalol on ABP and HR. Differences were considered significant when p < 0.05), while the effect was not significant at T1 for MAP (p = 0.0519). Median maximum MAP decrease was 31 (20–90) mmHg. Heart rate did not increase significantly during treatment (p = 0.2454). Acepromazine given before or during labetalol treatment did not reduce significantly ABP (p = 0.735).A total of 20 dogs met the inclusion criteria, and hypertension was documented after craniotomy (12/20), adrenalectomy (4/20) and other procedures (4/20). Five dogs received labetalol intraoperatively, 14 postoperatively, and 1 during the surgical procedure and recovery. Median infusion duration and rate were 463 (60-2120) minutes and 1.1 (0.2–3.4) mg/kg/h respectively. Median loading dose was 0.2 (0.2–0.4) mg/kg. Labetalol produced a significant decrease in SAP and DAP at all time points compared to T0 (Labetalol produced a reliable and titratable decrease in ABP with non significant increase in HR. This is a significant advantage when treating acute hypertension, as β- blockade alone leaves α-receptors catecholamine stimulation unopposed, with consequent compensatory increase of systemic vascular resistance. Similarly, α- blockade alone generally results in compensatory tachycardia. Labetalol decreases arterial blood pressure (ABP) and systemic vascular resistance, without significantly affecting cardiac output [Labetalol hydrochloride AH 5158) is a combined alpha- and beta-adrenergic blocking drug 58 is a c. It compc output , 4. The  β1 and β adrenergc output .1 receptors respectively, while the activity of the other two stereoisomers is negligible [Labetalol is a mixture of four stereoisomers present in equal proportion . They block adrenergic receptors to varying degree, and the overall effect mostly depends on the combined action of RR and SR stereoisomers, which are potent antagonists of β- and αgligible , 8.The effect of labetalol and other anti-hypertensive agents has been compared, demonstrating comparable efficacy and side effects profile to β-adrenergic blockers . LabetalIn human medicine labetalol is effective in reducing blood pressure in the short and long-term period, and the onset of effect during a continuous infusion is rapid and controlled , 11.Systemic hypertension is a well-recognized complication during and after intracranial surgery in both humans and dogs, with an incidence of up to 57% reported in human medicine  and 52% Adrenalectomy is also a surgical procedure carrying a significant risk of severe intra and postoperative hypertension in humans and dogs , 18. AdrTo date, all published literature describing use of labetalol in dogs is limited to experimental models. Studies in anaesthetized dogs demonstrated the hypotensive effect of labetalol at the dose of 0.1 to 15 mg/kg intravenously , 20, andThe aim of this manuscript is to report our experience using labetalol to control intra and postoperative hypertension in dogs in a clinical setting.Dogs receiving labetalol during hospitalisation between 2012 and 2017 were identified, searching the practice management system, and case records reviewed retrospectively, if the owner had not declined use of clinical data for research and educational purpose on the general consent form. Animals were included in this retrospective study if a hypertensive episode was documented either during surgery or in the immediate post-operative period, and was treated with labetalol . Dogs were excluded if medical records were incomplete or unclear, if blood pressure was not measured invasively, or if length of infusion was shorter than 60 min.Data including signalment, procedure performed, HR, invasive ABP, body temperature, concomitant administration of drugs potentially affecting systemic or intracranial pressure  and postoperative analgesia were recorded on a spreadsheet. Precalibrated pressure transducers zeroed at the level of the heart (tip of the shoulder) were used to measure invasive blood pressure. They were sterilised between uses and regularly checked against a manometer.The following information were collected for the period of labetalol administration, using a multiparameter monitor : systolic (SAP), MAP and diastolic (DAP) arterial pressure, HR and temperature at the time CRI of labetalol was started (defined as T0), the initial infusion rate and, if any, the loading dose of labetalol; SAP, MAP, DAP, HR and labetalol administration rate 15, 30, 45 and 60 min after infusion started . Duration of labetalol infusion, the maximum and minimum infusion rate, the total amount administered, and the steps made to reduce it were also evaluated.Due to the retrospective nature of the study, the decision of using labetalol to decrease ABP was completely at the anaesthetist’s discretion, both in terms of threshold for intervention and target value. The aim was to maintain MAP below 140 mmHg, as this was considered the value above which autoregulation is likely impaired. Intermittent positive pressure ventilation was used during anaesthesia to maintain normocarbia in all dogs. At the end of the procedure, once spontaneous breathing resumed, animals were transferred to the intensive care unit (ICU) and their vital parameters, including invasive ABP, were monitored continuously and treated as indicated by clinical condition. Postoperative analgesia was assessed every 2 h in all cases by a trained veterinary nurse or veterinary surgeon using the Short-Form of the Glasgow Composite Pain Scale, as per standard operating procedures in our practice . MethadoMonitoring of anaesthesia during surgery was performed by either an anaesthesia specialist or a resident, while data during the recovery phase were collected by a trained nurse, as part of the standard monitoring in our ICU. In all cases, use of labetalol was under the direct supervision of a senior anaesthetist.p < 0.05.All data were analysed using a commercially available statistical software program . Mixed effect ANOVA and a post hoc Dunnett’s multiple comparison test were used to analyse measured cardiovascular variables over time, before and during labetalol administration at different time points. The impact of acepromazine administration on blood pressure was assessed using Mann Whitney test. Results are expressed as mean (standard deviation) or median (range), according to the nature of the data and normality, tested using D’Agostino and Pearson Omnibus test. Differences were considered significant when A total of 33 records of cases that received labetalol as part of the intra and/or post-operative management, were retrieved and analysed. Three cases were excluded due to incomplete records, one because of poor record keeping and nine because labetalol was infused for less than one hour. Twenty dogs were included in the study Fig. . Breeds Pre-anaesthetic stabilization of the animal included administration of corticosteroids or other anti-inflammatory drugs, analgesics, antiemetics, antimicrobials, fluid therapy, anti-epileptic drugs and hyperosmolar agents, such as mannitol. One dog with pheochromocytoma received phenoxybenzamine treatment before surgery.In case of intracranial surgery, according to the course of procedure and the intraoperative findings, mannitol (0.5 ± 0.01 mg/kg) was also administered (12/12 cases); 5 of them received phenobarbital (3.3 ± 1.7 mg/kg), and in 2 dogs dexamethasone (0.13 ± 0.01 mg/kg) was added to the other two drugs. In two cases transient intraoperative hypotension was treated with colloids . Intraoperative analgesia was achieved administering an infusion of dexmedetomidine (0.71 ± 0.27 mcg/kg/h), lidocaine (1.5 ± 0.7 mg/kg/h), an intravenous opioid , or an epidural block in some cases of abdominal surgery .Average labetalol infusion rate was 1.1 (0.2–3.4) mg/kg/h and in 9 cases (45%) a loading dose of 0.2 (0.2–0.4) mg/kg IV was administered. Arterial blood pressure was continuously monitored during labetalol administration, and the dose of labetalol CRI was reduced by sequential steps in case of response, while it was increased if a greater decrease of blood pressure was deemed necessary.Five (25%) animals received labetalol only intraoperatively, 14 (70%) postoperatively, and 1 (5%) received the infusion during the surgical procedure and recovery. Median infusion duration was 463 (60-2120) minutes.The animals in which labetalol was infused intraoperatively were anaesthetized with isoflurane , sevoflurane , or propofol .p = 0.2454)  labetalol infusion rate was halved once, in 4 (20%) twice, in 3 (15%) three times, and 4 and 5 steps reductions were made in one case each; in the remaining cases, infusion rate was kept constant. Maximum and minimum median infusion rates were 2.0 (0.5–3.6) and 0.8 (0.1-2.0) mg/kg/h respectively.p = 0.735).Ten (50%) dogs received acepromazine IV before and/or during the infusion (mean dose 9.2 ± 3.7 mcg/kg), and in 5 cases the drug was administered prior to labetalol in the attempt to control ABP. Maximum MAP decrease was 27.5 (10–90) and 35 (14–55) mmHg in labetalol treated dogs who received acepromazine and those who did not, respectively. The difference was not significant  .In human medicine hypertension is associated with worse surgical outcome, increased risk of myocardial ischemia, cerebrovascular accidents, bleeding, increased duration of hospitalization, and pulmonary oedema in presence of cardiac disease , 25. ForThe ideal agent for treatment of hypertensive emergencies should be rapid acting, predictable, easily titrated, safe and inexpensive. Intravenous infusions of antihypertensive medications present the advantage of a rapid effect, with controlled and steady lowering of blood pressure, with rapid cessation upon discontinuation of infusion . When trp = 0.2454) increased during labetalol therapy, as expected from its reversible antagonism activity at both α- and β-adrenoceptor [Heart rate was only slightly and not significantly , then the parameters gradually decreased over time; in other two an initial decrease was noticed, then the pressure increased (T2) before decreasing again. A paradoxical hypertensive effect of labetalol has been reported in patients with pheochromocytoma. It has been suggested that the weak α-blockade may trigger a paradoxical increase of vascular resistance following β-blockade , 48. For1 receptor alone is probably not potent enough to control the effect of catecholamine release in these cases, or a higher dose may be needed to achieve the desired effect.Acepromazine was administered in 10 50%) patients at variable dose between 5 and 16 mcg/kg; in 5 dogs it was administered as first line treatment to control hypertension, in 4 dogs during labetalol infusion, and 1 dog received it in premedication. Interestingly this drug did not appear to have a significant effect on ABP control strategy. Acepromazine decreases blood pressure blocking vascular α-adrenergic receptors, with consequent vasodilation and decrease in systemic vascular resistance, making it unpopular for the management of unstable patients or those at increased anaesthetic risk % patient. DespiteThe main limitation of this study is its retrospective nature. Intra and postoperative pharmacological managements were not standardized, although type and dose of anaesthetic and vasoactive agents were similar between cases, and this may have influenced the response of the patients to labetalol. Labetalol dose was not standardized; therefore, it is not possible to characterise a dose-effect relationship, yet we have provided a dosing range that may be helpful when treating hypertension. The blood pressure value at which labetalol was started was also decided by the attending anaesthetist and is possible that response to treatment may also depend on severity of hypertension; higher values may take longer to improve, and the 60 min timeframe considered can be insufficient for a satisfactory drug effect. Patients in this study received a number of medications potentially affecting ABP, such as acepromazine, dexmedetomidine, steroids and mannitol, and it is possible that different results may have been obtained with the use of labetalol alone. On the other hand, the fact that labetalol effect was consistent and predictable across such a variety of conditions and concurrent medications, both in awake and anaesthetized dogs is remarkable and highly indicative of the titrability and efficacy of this drug in dogs. A prospective study is however warranted to further confirm our preliminary results.Labetalol was effective in decreasing ABP in dogs experiencing acute intra or postoperative hypertension. Furthermore, prospective studies are needed to better elucidate the potential role of labetalol in the treatment of hypertension.


Renal involvement is common in systemic sclerosis  and includes chronic kidney disease (CKD). We have performed analysis of urinary proteins to gain insight into local molecular pathology of CKD in SSc and identify candidate markers for use in clinical trials.To evaluate urinary proteins that might specifically reflect SSc-related CKD, patients were recruited with confirmed SSc and stratified for the presence or absence of CKD. Controls included patients with CKD and no SSc, in addition to healthy volunteers. Candidate markers were measured in serum and urine by multiplex immunoassay testing for IL6, IL18, TNF-α, monocyte chemoattractant protein 1 (MCP1), monocyte chemoattractant protein 3 (MCP3), VEGF and the soluble adhesion molecules vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1). n = 40), SSc with CKD (n = 39), non-SSc CKD (n = 11) and healthy volunteers (n = 12). Urinary levels of IL6, MCP1, TNF-α, MCP3, IL18 and ICAM-1 were elevated in SSc patients compared with healthy controls. The most significant differences were for MCP1 and ICAM-1 (both P < 0.0001), and these analytes also showed the most significant differences between groups overall (P = 0.003 for MCP1 and P < 0.0001 for ICAM-1). These markers showed a trend  or a significant difference  between SSc–CKD and SSc with normal renal function.One hundred and two subjects were examined, including patients with SSc with no evidence of CKD (Urinary levels of candidate molecular markers appear to reflect SSc–CKD more than serum markers. MCP1 and ICAM-1 are promising molecular markers for SSc–CKD and might be potential biomarkers of SSc renal involvement. This might be explored in future prospective analyses. Elevated inflammatory proteins in serum of SSc patients reflect disease biology at multiple sites.In SSc, urinary proteins can provide insight into local disease mechanisms in chronic kidney disease.This study identifies two candidate renal biomarkers, urinary ICAM-1 and MCP1, that might be specific for SSc.Systemic sclerosis  is a multi-system disease with high mortality owing to the involvement of vital organs, including the heart, lungs, gut and the renal tract . Renal cAlthough recovery from the acute kidney injury of SRC is often excellent, SRC remains an important mechanism leading to chronic kidney disease (CKD) in SSc. In addition, CKD occurs in a large proportion of SSc patients because of systemic vasculopathy, fibrosis and other mechanisms, such as overlap CTD, including SLE and vasculitis. Other pathologies, such as interstitial nephritis or drug toxicity, can also contribute to CKD in SSc. Previous studies of sequential unselected patients suggested that CKD is present in ≤50% of SSc cases [Creatinine-derived measures of glomerular filtration rate (GFR) deteriorate only once there is significant parenchymal abnormality in the kidney; therefore, there is a general need for more sensitive markers of disease activity in CKD, including the context of SSc. Such tests might also help to discriminate SSc from non-SSc-related processes in the kidney in order that management could be more appropriate. In addition, new markers could help in distinguishing clinically important or progressive CKD from more stable cases. Easily measured biomarkers could be applied to outcome assessment in clinical trials or practice and might provide early indicators of therapeutic response that could predict future clinical benefit.There have been many recent studies exploring potential biomarkers in SSc, and these have focused on examination of skin biopsies or peripheral blood, including cell-based approaches such as gene expression, in addition to methods examining serum, plasma or microparticles , 7. CompIn addition to blood, urine offers a highly relevant and accessible substrate to explore candidate biomarkers of SSc-related CKD (SSc–CKD). Urine potentially has significant advantages over serum or plasma as a fluid for renal biomarker investigation. It is produced in direct contact with the epithelial surface of the organ of interest, meaning that relevant proteins expressed in renal injury might be shed directly into the urine. For this reason, urine has been described as a ‘fluid biopsy’ of the kidney and renal tract . FurtherIn the present study, we have analysed candidate urinary proteins in SSc–CKD and compared them with serum levels of these analytes. To determine which urinary proteins might be relevant specifically to SSc–CKD by reflecting local molecular pathology, we have compared levels in urine, after correction for urinary creatinine, for SSc, SSc–CKD and non-SSc–CKD patients. In this way, we expected to differentiate potential markers of SSc pathobiology in the kidney that might be useful in clinical trials and also provide insight into the likely mechanisms of CKD that occur in SSc and might impact on clinical outcomes, such as susceptibility to, or recovery after acute kidney injury occurring in the context of scleroderma renal crisis (SRC).Candidate markers of renal involvement to measure in urine and serum of patients with SSc and controls were defined from previous literature and current concepts of the molecular pathology of SSc and other related multi-system autoimmune rheumatic diseases. The aim was to have a selection of proteins that would help to identify inflammatory, fibrotic and vasculopathic processes in the renal parenchyma that would add to conventional assessment of renal abnormalities, such as serum creatinine and GFR. The following markers were selected for measurement.IL6 is a likely pathogenic mediator of inflammation and connective tissue dysfunction in SSc. Its expression in urine has been correlated with renal disease in several contexts, including the autoimmune CTD SLE . IL18 has been demonstrated to be a mediator of ischaemic damage to the renal tubule in mice , and uriTNF-α is a putative mediator of endothelial damage in SSc . Serum aVEGF is overexpressed in tissue biopsies and sera from patients with SSc . It is eMonocyte chemoattractant proteins 1 (MCP1 or CCL2) and 3 (MCP3 or CCL7) have been described as pathogenic fibroblast activators in SSc , and higSoluble ICAM-1 has been associated with disease severity in SSc in serum and is considered to be a marker of activated endothelium, epithelial cells and fibroblasts . It is eSoluble VCAM-1 has been associated with fibroblast activation and epithelial-to-mesenchymal transition and is a marker of immune cell activation and endothelial cell activation. It is elevated in SRC in other series and has been shown to be increased markedly in the serum in some cases of SRC , 27. UriThis study was approved by the Royal Free Research and Development team and by the Newcastle and North Tyneside Research Ethics Committee. All individuals provided informed consent for their participation, according to the guidance set out by the research ethics committee.2 or <90 ml/min with persistent urinary blood or protein on dipstick were categorized as having CKD, consistent with CKD stages 2–5 in the 2002 guidelines by the Kidney Disease Outcomes Quality Initiative (KDOQI) [The study cohort of 79 adult patients attending the scleroderma clinic at the Royal Free Hospital was recruited prospectively. All had definite SSc classified by 2013 ACR/EULAR classification criteria . Kidney  (KDOQI) . By inclIn addition to demographic data , the medical history was recorded. For the SSc groups, this included disease-specific organ complications, skin subgroup (limited or diffuse) and the disease-specific ANA. These data are summarized in g at 4°C, for 10 min, within 1 h of collection. After centrifuge and serum separation, the serum and urine were divided into aliquots and frozen at −80°C. From each individual recruited to the study, concurrent urine and blood samples were collected. Clotted blood and fresh mid-stream urine were centrifuged at 600 Additional blood and urine samples were sent to the Royal Free Hospital laboratories for routine clinical biochemistry analysis, including serum creatinine, eGFR and urinary albumin:creatinine ratio.Multiplex immunoassay for all eight analytes was performed according to the manufacturer’s protocol . Standards supplied with the assay kits, urine and serum samples were analysed in duplicate wells. Urine biomarker concentrations (in picograms per millilitre) were expressed as a ratio to the urine creatinine concentration (in micromoles per litre) to compensate for diurnal variations in the water concentration of spot urine samples.U-test. Correlation between eGFR and biomarker concentrations was assessed using Pearson’s coefficient. All statistical tests were performed using GraphPad Prism v.8.2.1 for Windows .Four patient groups  were compared for each candidate biomarker. The difference between these groups was assessed using the Kruskal–Wallis test. The SSc–CKD group was also compared individually with each of the other three groups using the Mann–Whitney Overall results for the eight proteins analysed in this study are detailed in Seven of the candidate proteins were detectable in the serum of patients with SSc–CKD and control groups. MCP3 was detectable in the serum of only one subject in the SSc–CKD group and none of the subjects in the three control groups and is therefore of limited value for further analysis. All eight candidate proteins were detectable in urine by our method in subjects in all four groups.Two of the candidate urinary proteins (MCP1 and ICAM-1) showed a higher degree of discrimination between groups, and further analysis was performed to assess whether differences in GFR among our patients would account for the difference in urine concentrations.The detectable concentration of low molecular weight proteins in the urine is inextricably related to the blood concentration of the same proteins and the volume of blood filtered by the kidneys (i.e. the GFR). To investigate the relationship between kidney function and concentrations of MCP1 and ICAM-1 in urine, urinary concentrations were plotted against MDRD eGFR and serum creatinine (from which the eGFR is derived). These data are included in R = −0.42, P = 0.0001) and positive correlation with serum creatinine . The correlation for urinary MCP1:creatinine with eGFR followed the same trend but was weaker . Examination of the dot plots for each marker suggested that the relationship between eGFR and ICAM-1:creatinine was more consistent and robust than that for urinary MCP1. This suggested for MCP1 that raised urinary levels might reflect local pathogenic processes rather than simply lower GFR, consistent with widespread expression of MCP1 in SSc renal biopsy specimens [For the ICAM-1:creatinine ratio, there was a significant negative correlation with eGFR , where MCP1 showed relevant changes after treatment . LikewisIn summary, we have taken a new approach to identify potential urinary protein markers of SSc–CKD. Two promising candidates have been identified, and these should be validated in future cross-sectional and longitudinal studies.Funding: This work was funded by the Medical Research Council (award reference MR/K015230/1) and performed as part of an academic collaboration with AstraZeneca. Additional funding was provided by Scleroderma and Raynaud’s UK.Disclosure statement: R.U. is currently working in Early Clinical Development, Early CVRM , R&D BioPharmaceuticals, AstraZeneca, Cambridge, UK and Gothenburg, Sweden. The other authors have declared no conflicts of interest.The authors agree to make all data, available materials and associated protocols available to readers if requested for academic purposes.


There was a positive correlation between changes in placental thickness and the amount of amniodrainage during intervention . Hematocrit decreased from 33.4 ± 3.8 (%) to 28.4 ± 3.5 (%), i.e., an increase in relative blood volume by 18 ± 10.2% (p < 0.001). Albumin decreased from 37.9 ± 0.9 g/L to 30.7 ± 2.2 g/L, i.e., an increase in relative plasma volume by 24 ± 8.1% (p < 0.001). Amniodrainage leads to uterine decompression, increased placental thickness and subsequent maternal hemodilution. We propose the term “amniodrainage-induced circulatory dysfunction” for these specific maternal hemodynamic changes in the treatment of twin-to-twin transfusion syndrome.Twin-to-twin transfusion syndrome (TTTS) in monochorionic-diamniotic twin pregnancies usually requires fetoscopic laser ablation (FLA) followed by amniodrainage (AD). Perioperative maternal hemodynamic changes and hemodilution have been observed. Little is known about the underlying pathophysiology. We aimed to evaluate the impact of high volume amniodrainage on intrauterine pressure, placental thickness and maternal blood characteristics. A total of 18 cases of TTTS were included in this prospective pilot study. All patients were treated with FLA and subsequent AD. Intrauterine pressure and placental thickness were assessed before, during and after amniodrainage. Maternal hemoglobin, hematocrit and serum albumin were measured at admission and 24 h after the intervention. Amniodrainage led to a decrease in mean intrauterine pressure (from 30.1 ± 8.1 mmHg to 17.6 ± 3.6 mmHg ( Twin-to-twin transfusion syndrome (TTTS) complicates approximately 10% of monochorionic-diamniotic (MCDA) twin pregnancies . FetoscoIn contrast to the high number of scientific reports on fetal outcome and treatment options, data on maternal hemodynamic changes or adverse side effects following FLA procedures are scarce. Clinical signs suggestive of maternal hemodilution were described in cases of mid-trimester TTTS managed by FLA and subsequent amniodrainage, providing evidence that such interventions have a significant impact on the maternal compartment ,6,7. A vIn a previous retrospective study, we observed that high volume amniodrainage was significantly correlated with maternal hemodilution and hemodynamic alterations . Thus, wA total of 18 women at a minimum age of 18 years with MCDA twin pregnancies complicated by TTTS were recruited for this prospective pilot study at the Department of Obstetrics and Gynecology at the Medical University of Graz, Austria, between November 2017 and September 2019. Fetoscopic laser ablation of placental anastomoses was performed between 16 and 26 weeks of gestation following international recommendations . In one At our institution, intrauterine treatment by FLA is generally offered to all cases with Quintero stage I or higher, as defined by polyhydramnios in the recipient‘s sac  measuring more than 8 cm before 20 weeks of gestation and more than 10 cm beyond 20 weeks of gestation) in combination with oligohydramnios in the donor‘s sac  .FLA was performed in a standardized fashion as previously described . All FLAIntrauterine pressure was assessed by a TruWave digital pressure sensor  in 10 consecutive cases. The pressure sensor was directly linked to the aforementioned cannula via a Luer lock connection and a flexible sterile tube without requiring an additional incision for measurements. The sensor was zeroed to the level of the mid-axillary line of the pregnant woman placed in supine position. Intrauterine pressure was recorded under “hands-off” and “stop-flow” conditions before and during amniodrainage. Measurements were conducted in steps of 200 mL of amniodrainage. Pressures obtained during uterine contractions were discarded. Data were digitally processed and stored on a Datex-Ohmeda unit  and recorded in written form as well.We used a standardized approach for the evaluation of the placental thickness. The same expert sonographer conducted all measurements. Placental thickness was defined as the maximum distance from the myometrium–placenta interface to the placenta–amniotic fluid interface. Measurements (n = 18) were performed on an adequately sized and focused ultrasound image in an anterior–posterior view on a Voluson E10 or E6  under the same conditions as the intrauterine pressure measurements. Maximum thickness was obtained in a linear fashion and taken from the very same site of the organ at all timepoints. To ensure measurements from identical placental sites, we linked the position of the ultrasound probe to the site of umbilical cord insertion or to major maternal abdominal vessels in case of posterior wall location of the placenta ,15.Maternal age, gestational age at intervention, maternal body mass index (BMI) and Quintero stage at time of intervention were documented. The total volume of amniodrainage was recorded and maternal hemoglobin (Hb), hematocrit (Hct) and serum albumin were measured at admission (pre) and 24 h after the intervention (post) by standard laboratory techniques. Relative hemodilution for blood (DB) and plasma compartments (DP) were quantified as the ratios of “post to pre” concentrations. The relative expansion of blood (RBV) and plasma volumes (RPV) by hemodilution was measured as the ratio of “pre to post” concentrations as described elsewhere .Data were tested for normal distribution by the Kolmogorov–Smirnov test and the Shapiro–Wilk test. Non-parametric data were tested with the Wilcoxon test. In case of normal distribution, a paired t-test was performed. A probability of < 0.05 was considered statistically significant. Analyses were carried out using SPSS Statistics 25 .The current study was approved by the Ethics Committee of the Medical University of Graz, Austria, registered at Institutional Review Board Registry as IRB00002556 (30-035 ex 17/18). Written informed consent was obtained from all patients before intervention. All patient data were pseudo-anonymized. Only authorized persons had access to the original data sets.The study included 18 women with TTTS treated by FLA and subsequent amniodrainage. Median maternal age at intervention was 28 years (21–39) and maternal BMI was 25.9 (17.4–38.9). Gestational age at intervention was 21.1 (19.3–28.1) weeks . There wp < 0.001) (p < 0.001). Mean placental thickness increased from 16.8 ± 6.4 mm before amniodrainage to 31.8 ± 8.6 mm after amniodrainage (p < 0.001) (p < 0.001)  , corresp< 0.001) , corresp< 0.001) .p = 0.001).We found a significant correlation between the volume of amniodrainage and the effects on placental thickness. From pre- to post-drainage, there was a positive correlation between changes in placental thickness and the amount of amniodrainage during intervention   . Mean heIn this prospective pilot study, we observed a significant increase in placental thickness while intrauterine pressure decreased significantly during high volume amniodrainage. Postoperative maternal hemodilution was found in every study and was linked to these utero-placental changes.Severe polyhydramnios results in increased intrauterine pressure in cases with TTTS , while ap = 0.63). This is an indirect confirmation that the hematocrit dropped as a result of hemodilution and not because of blood loss.In a previous study, we observed that high volume amniodrainage was significantly correlated with maternal hemodilution and hemodynamic alterations . We coulA decrease in intra-uterine and intra-abdominal pressures leads to an increase in the transmural pressure in the vascular system of the uterus and the intra-abdominal vasculature, because the volume within arteries and veins is a function of both the pressure difference across their wall, the transmural pressure, and their distensibility, the vascular compliance . Since vThe following factors are likely to contribute to the observed circulatory findings in the maternal compartment:Firstly, uterine decompression and the subsequent gain of placental thickness increases maternal plasma volume on the basis of modified placental blood rheology and a reduction in effective microfiltration pressure due to the loss of vascular resistance ,32. The Secondly, amniodrainage reduces the physical size of the pregnant uterus. Thus, mechanical decompression of abdominal arterioles and venules may cause splanchnic vasodilatation and increased filling of the vascular bed, comparable to hemodynamic changes observed with large volume paracentesis of tense ascites in hepato-renal syndrome and portal hypertension, also known as paracentesis-induced circulatory dysfunction (PICD) .Thirdly, reduction in uterine size due to amniodrainage may decrease the intraabdominal pressure and therefore increase the venous backflow, especially from the lower extremities of the patient that are placed in a horizontal position during the intervention and during the 24 h observation period after intervention. This might mobilize fluid from the peripheral edema of the lower extremities, i.e., extracellular fluid volume, into the intravascular space, comparable to volume shifts observed during peritoneal dialysis .Although we provided some possible mechanisms of the present phenomenon, we did not formally examine if these really do occur in our patients. Our explanations should provide a common basis for further investigations and an invitation for scientific discussion and co-operation.The positive correlation between the volume of amniodrainage and the increase in placental thickness supports the hypothesis that uterine decompression might provoke placentomegaly resembling placental hydrops. Placental hydrops is frequently reported to occur in the course of Ballantyne´s Syndrome, usually referred to as “mirror syndrome”. In our study, we demonstrated that uterine decompression is linked with the rapid increase in placental thickness and subsequent hemodilution in the very same patient group. This supports the hypothesis that high-volume amniodrainage might cause a subclinical and mild variant of iatrogenic Ballantyne´s Syndrome. To clarify the terminology, we suggest the term amniodrainage-induced circulatory dysfunction (AICD) for this attenuated variant of iatrogenic Ballantyne´s Syndrome in line with findings made during paracentesis for cirrhotic ascites .The study population and patient characteristics were comparable to previous publications on this topic ,18,19,20Amniodrainage is linked with uterine decompression, increased placental thickness and subsequent maternal hemodilution. We propose the term “amniodrainage-induced circulatory dysfunction” for specific maternal hemodynamic changes observed after iatrogenic uterine decompression in the treatment of TTTS.


The relationship between the neutrophil-to-lymphocyte ratio (NLR) and hemorrhagic transformation (HT) in acute ischemic stroke (AIS) treated with intravenous thrombolysis (IVT) remains unclear. This study assessed whether high NLR is associated with HT in this population.  Data were prospectively collected for continuous patients with AIS treated with IVT and retrospectively analyzed. Clinical variables included age, sex, vascular risk factors, National Institutes of Health Stroke Scale (NIHSS) score, onset-to-treatment time, and initial hematologic and neuroimaging findings. HT was confirmed by imaging performed within 3 days after IVT. Symptomatic HT (sHT) was defined as NIHSS score increased by 4 points compared with that on admission according to previously published criteria. The NLR value was based on the blood examination before IVT, and high NLR was defined as ≥75th percentile. P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR ( The study included 285 patients (201 (70.5%) males, the mean age was 62.3 years (range 29–89)). Seventy-two (25.3%) patients presented with HT, including three (1.1%) with sHT. The median NLR was 2.700 . Seventy-one (24.9%) patients had a high NLR (≥4.255) on admission. Univariate analysis indicated that patients with HT had higher NIHSS scores ( High NLR could be a useful marker for predicting HT in AIS patients after IVT. Intravenous thrombolysis (IVT) with recombinant tissue plasminogen activator (r-tPA) is an effective treatment for acute ischemic stroke (AIS) when administered within the hyperacute period , 2. HemoIn recent years, researchers have attempted to identify convenient serum biomarkers to help predict AIS outcomes. Several studies reported that a high neutrophil-to-lymphocyte ratio (NLR) was predictive of HT in AIS patients , 16. HowAIS patients treated with IVT after admission to Dongguan People's Hospital between 1 January 2016 and 31 May 2019 were continuously recruited. The inclusion criteria were as follows: (1) age >18 years, (2) AIS confirmed by magnetic resonance imaging (MRI), and (3) onset of stroke symptoms within 4.5 hours and treated with r-tPA. The exclusion criteria were as follows: (1) hemorrhagic lesions detected on initial computed tomography (CT), (2) temporary or permanent contraindications for MRI scan, (3) no acute lesion on diffusion-weighted imaging (DWI), and (4) additional endovascular therapy after IVT. This study was approved by the hospital ethics committee . The consent of each subject was obtained in accordance with the Declaration of Helsinki.NIHSS score, onset-to-treatment time (OTT), and blood pressure on admission were collected, as well as demographic data including age, sex, and history of hypertension, diabetes mellitus, smoking, AF, antiplatelet therapy, oral anticoagulant therapy, and previous stroke. Initial counts for white blood cells, neutrophils, lymphocytes, and platelets before IVT were also collected, and NLR was calculated accordingly.As MRI is more sensitive than CT for detecting HT in AIS , we usedThe parameters of each sequence were shown as follows: axial SE T1: time of repetition (TR) = 1500 ms, time of echo (TE) = 11 ms, field of view (FOV) = 220 mm, slice thickness/gap = 4 mm/1.2 mm, and time of acquisition = 1 min 26 s; Turbo spin echo (TSE) T2: TR = 4720 ms, TE = 96 ms, FOV = 220 mm, slice thickness/gap = 4 mm/1.2 mm, and time of acquisition = 1 min 50 s; DWI: TR = 4640 ms, TE = 67 ms, FOV = 230 mm, slice thickness/gap = 4 mm/1.2 mm, spin echo planar imaging (EPI) factor = 91, and acquisition time = 1 min 44 s; SWI: TR = 27 ms, TE = 20 ms, FOV = 220 mm, slice thickness/gap = 3 mm/0.6 mm, and time of acquisition = 2 min 28 s.sHT was defined as NIHSS score increase by ≥4 points compared with that on admission .When HT was positive on MRI, the images were categorized into hemorrhagic infarct (HI) and parenchymatous hemorrhage (PH) according to the ECASS II criteria as follows: HI1, small petechiae along the margins of the infarct; HI2, confluent petechiae within the infarcted area but no space effect; PH1, blood clots in ≤30% of the infarcted area with some slight space-occupying effect; and PH2, blood clots in >30% of the infarcted area with substantial space-occupying effect.Small vessel disease (SVD) burden was rated on brain MRI by the presence of lacunes, white matter hyperintensities, cerebral microbleeds, and perivascular spaces. The presence of each SVD feature was summed as an “SVD score” (range 0–4) .HT was defined as the secondary hemorrhage within or away from the infarction area, which appeared as hypointensive lesions on SWI , 19 and Two neurologists (D.H.Q. and J.F.Q.) experienced in neuroimaging and trained by a neuroradiologist (M.Q.L.) evaluated the imaging findings for the presence of HT independently, blinded to the patients' clinical information. After observing the images individually, the two observers reviewed all the images to achieve final interobserver consensus.t-tests, Mann–Whitney U tests, Pearson χ2 tests, or Fisher's exact tests, as appropriate. Variables with P < 0.05 in the univariate analysis were included in further binary multivariate logistic regressions. Statistical significance was defined as P < 0.05 (two-sided).Statistical analyses were conducted using SPSS for Windows . Continuous variables with a normal distribution are reported as mean ± SD, and nonnormally distributed variables as median and interquartile range (IQR). All subjects were divided into two groups based on the presence of HT. Variables were compared using n = 2), permanent or temporary contraindication for MRI (n = 3), and no acute lesion found on DWI (n = 16). A total of 285 patients were ultimately included.During the study period, 306 consecutive patients received IVT with r-tPA within 4.5 hours of stroke onset. In the present study, 21 patients were excluded for the following reasons: additional endovascular therapy after IVT  patients were male. The median interval between stroke onset and MRI scanning was 42  hours. Antiplatelet agents (aspirin 100 mg/day or clopidogrel 75 mg/day) were prescribed 24 hours after IVT when PH  was excluded, and no anticoagulants were prescribed during the acute phase. Among the 72 (25.3%) patients with HT confirmed by MRI, 44 (15.4%) presented with HI1, 21 (7.4%) with HI2, 4 (1.4%) with PH1, and 3 (1.1%) with PH2. Three (1.1%) patients had sHT. No remote HTs were found in our study. The mean OTT was 200.4 ± 55.9 minutes, and the median NIHSS score on admission was 7 . The median NLR was 2.700 . High NLR was defined as an NLR value ≥4.255 (75th percentile). Seventy-one (24.9%) patients had a high NLR before IVT. The demographic and clinical characteristics of this study are shown in P < 0.001), systolic blood pressure (SBP), platelet counts, lymphocyte counts, and NLR (P < 0.05). They also had a greater prevalence of high NLR than those without HT (37.5% vs. 20.7% and P=0.004). Patients with HT were also more likely to have a history of hypertension and AF. The univariable analysis results are shown in Compared with those without HT, patients with HT had significantly higher NIHSS scores (r = −0.499), we used two separate logistic regression models. Besides, hypertension and SBP on admission were also highly correlated (r = 0.412); therefore, SBP on admission was not included in the regression models to avoid the risk of multicollinearity. In model 1 (with high NLR), NIHSS score on admission (odds ratio (OR) = 1.110, 95% confidence interval (CI) = 1.015–1.044, and P=0.001), AF , and high NLR  were significant predictors of HT. In model 2 (with lymphocyte counts), NIHSS score on admission , AF , and lymphocyte counts  were significantly related with HT. Platelet counts and hypertension were not significantly associated with HT in either model. The multivariate logistic regression results for HT risk factors are shown in Variables that were significantly different between the two groups in the univariable analysis were entered into subsequent logistic regression model. Since lymphocyte counts and high NLR were highly correlated (r = 0.676), our findings did not contradict previous studies. NLR is easily evaluated with a routine blood test, making it an economic and effective marker, even in regional hospitals.In our study, high NLR ≥4.255) was significantly associated with HT in AIS patients treated with IVT, which was in accordance with two previous studies , 16. The5 was sigThere were several advantages to our study. First, to the best of our knowledge, it was one of the few that focused on the association between high NLR and HT in IVT-treated AIS patients. Second, all the participants had relatively complete data of neuroimaging including SWI. However, our results should be considered in the context of several limitations. First, repeated MR scanning was not performed, which might have led to underestimation of HT in the subacute phase. Second, we lacked dynamic NLR data, which would be considered a more effective predictor. Third, the numbers of cases with PH1, PH2, or sHT were too small to perform further analyses of these severe HT subtypes.High NLR was a useful predictor of HT in AIS patients after IVT. Further prospective studies with larger sample sizes, repeated MR scans, and dynamic NLR are warranted.


To the Editor,Aspergillus being one of the most common.In asthma, sensitization to fungal, perennial or seasonal allergens increases the risk of uncontrolled symptoms, exacerbations and poor disease outcomes.Severe eosinophilic asthma is characterized by frequent exacerbations and elevated eosinophil counts. Currently, there is limited information on the prevalence of fungal allergen sensitization in patients with severe eosinophilic asthma, and its impact on clinical responses to treatments such as the anti‐interleukin (IL)‐5 monoclonal antibody mepolizumab. In clinical trials, mepolizumab reduced exacerbation frequency and oral corticosteroid use, improved lung function, and health‐related quality of life (HRQoL) and symptoms vs placebo in patients with severe eosinophilic asthma.Aspergillus fumigatus and/or Penicillium chrysogenum (selected because these thermotolerant filamentous fungi are known to colonize the airways and are associated with lung damage in severe asthma),MENSA was a randomized, double‐blind, Phase III trial  in patients with severe eosinophilic asthma.1), St George's Respiratory Questionnaire (SGRQ) score, Asthma Control Questionnaire (ACQ‐5) score and change from baseline in blood eosinophil count. Changes from baseline in eosinophil granule proteins were assessed in patients receiving mepolizumab 100 mg SC or placebo. Descriptive statistical analyses were performed and are described in Appendix End‐points assessed included the prevalence of fungal and/or perennial/seasonal allergen sensitization and response to mepolizumab at Week 32. Mepolizumab response was determined according to the annual rate of clinically significant exacerbations  patients were sensitized to A fumigatus and 58/576 (10%) to P chrysogenum. Overall, 51/576 (9%) patients were sensitized to fungal allergens only, 167/576 (29%) to perennial/seasonal allergens only and 131/576 (23%) to both fungal and perennial/seasonal allergens; 198/576 (34%) patients were not sensitized to allergens. Exacerbation history, baseline SGRQ and ACQ‐5 scores, and blood eosinophil counts were similar across all allergen sensitivity groups  were sensitized to allergens  and those not sensitized to fungal allergens (52%)  Figure . Althoug) Figure .1 from baseline in all patients except those with fungal sensitization only, although this may be due to the small sample size (n = 51) for this group  were funded by GlaxoSmithKline (GSK).App S1Click here for additional data file.


Elderly patients have a relatively high cardiovascular risk due to increased arterial stiffness, elevated blood pressure and decreased amounts of elastin in the arteries. The composition of the media layer in the arterial wall, comprising elastin, collagen, smooth muscle cells, proteoglycans, fibronectin and fibrillin-1, influences its mechanical properties. Mutations in the fibrillin-1 gene leads to increased aortic stiffness, elevated pulse pressure and aortic root dilatation. This study investigates whether there is a sex difference among hypertensive elderly patients regarding blood pressure, arterial stiffness and fibrillin-1 genotypes.A total of 315 hypertensive subjects (systolic blood pressure > 140 mmHg) were included in this study (155 men and 160 women aged 71–88 years). Aortic pulse wave velocity and augmentation index were determined using SphygmoCor, and brachial blood pressure was measured using an oscillometric technique. Fibrillin-1 was genotyped by polymerase chain reaction and with a capillary electrophoresis system.p = 0.008), central mean arterial pressure , central systolic blood pressure  and central pulse pressure  than males. Females with the Fibrillin-1 2/3 genotype showed a significantly higher augmentation index  and systolic blood pressure  than females with the 2/2 and 2/4 genotypes.Females showed a significantly higher peripheral mean arterial pressure (females; 107.20 mmHg, males 101.6 mmHg, The findings of this study may indicate that hypertensive elderly females, especially elderly females with Fibrillin-1 2/3, have increased systolic blood pressure and arterial stiffness. It is still unclear whether arterial wall stiffness is a cause or a consequence of high blood pressure but the two conditions are close associated to each other . IncreasSimilar to arterial stiffness, systolic blood pressure (SBP) increases with age . ElderlyMutations in the fibrillin-1 (FBN1) gene is associated with increased aortic stiffness, elevated PP and aortic root dilatation , 14. SevThe study population was recruited from an ongoing longitudinal study of elderly community-living people from a rural municipality of southeast Sweden. Inhabitants aged 65–82 years were invited to join the original study between 1998 and 2001. A total of 1130 individuals were invited, and 876 of these individuals agreed to participate in the study .− 1 fasting plasma glucose. Ischemic heart disease was defined as a previous diagnosis of ischemic heart disease confirmed by exercise ECG or coronary angiography or electrocardiographically (ECG) verified myocardial infarction. Subjects who reported that they smoked were classified as smokers. Height and weight were measured and used for calculation of body mass index (BMI). All subjects in the study gave their written informed consent [The subjects were instructed not to use tobacco or consume coffee or tea for at least 4 h prior to their examinations. In addition, the participants had a physical examination and were asked about cardiovascular risk factors, cardiovascular diseases and medications. Subjects with previously diagnosed hypertension or with an SBP ≥ 140 mmHg were defined as hypertensive. Diabetes mellitus was defined as a previous diagnosis of diabetes mellitus or with ≥7 mmol l consent , 21. TheAll measurements were performed under standardized conditions. Brachial blood pressure was measured after at least 15 min of rest when the subject was in the supine position using an oscillometric technique . Systolic, diastolic and mean arterial pressures were measured .The ankle brachial pressure index (ABPI) was calculated using the SBP measured from the dorsalis pedis and the posterior tibial arteries recorded by a hand-held 8 MHz Doppler probe and a 12-cm standard cuff divided by the brachial systolic pressures measured in the right and left arm , 21.Radial artery pulse waves were obtained noninvasively using the SphygmoCor system . The SphygmoCor system was equipped with a Millar pressure tonometer. For PWV, the central pressure waveform was derived using a generalized transfer function, calculated from an 11-s recording of the radial artery pressure. For calibration of the pressure wave, brachial systolic and diastolic blood pressure were used. Blood pressure was measured before the pulse wave recordings. Registrations of pulse waves were repeated at least three times, and average data from the three most successful registrations were used for analysis. Only good-quality recordings were used for pulse wave analysis (PWA). The augmentation index (AIx) and pressure augmentation (AG) were calculated from the central pressure waveform. AIx is the pressure augmentation expressed as the percentage of the central pulse pressure (cPP): AIx = (AG/PP) × 100 , 21.Blood samples were taken after overnight fasting and collected in prechilled plastic Vacutainer tubes (Terumo EDTA K-3). Plasma was prepared by centrifugation at 3000 g for 10 min at 4 °C. Blood and plasma were stored at − 70 °C pending analysis. Plasma levels of fasting glucose, low-density lipoprotein (LDL) and high-density lipoprotein (HDL) were measured , 21.n in intron 28 of the FBN1 gene on chromosome 15 using the forward primer 5′ 6FAM – CAG AGT ACA TAG AGT GTT TTA GGG AGA − 3′ and the reverse primer 5′- GTT TCT TCC TGG CTA CCA TCC AAC TCC c-3′. PCR was performed in a volume of 12 μl with predenaturation at 95 °C for 9 min; 35 cycles of denaturation for 30 s, annealing at 65 °C for 30 s (35 cycles), and extension at 72 °C for 30 s (35 cycles); and a final extension at 72 °C. A 1-μl portion of the PCR product was diluted with 9 μl of highly deionized formamide (GENESCAN™ 500ROX™ Size Standard) for electrokinetic injection on the capillary electrophoresis system. The DNA fragments were labeled with the ROX™ fluorophore. The labeling results in a single peak under denaturing conditions were obtained, and the alleles were then identified by the number of base pairs (bp) comparable to each peak  [FBN1 genotyping was performed in 2009, and the investigators performing these assays were blinded to the results of the vascular testing. DNA was prepared for polymerase chain reaction (PCR) by typing the variable tandem nucleotide repeat (VNTR) (TAAAA) 162 bp) .p-values < 0.05 were considered statistically significant. The statistical analyses were performed with IBM SPSS 25.0.All data are presented as the means±SDs. All continuous variables were checked for normality distribution, a log-transformation was used in continuous variables with a non-normality distribution. Student’s t-test and Mann–Whitney U-test were used to calculate differences between sexes. One-way ANOVA with post hoc testing was used to compare the differences between the genotypes. Variance analysis was used to adjust the dependent variables , central mean arterial pressure (cMAP), central SBP (cSBP) and cPP for age, body surface area (BSA) and HR). p = 0.005) and higher creatinine (p <  0.001) than female subjects, whereas females had higher HDL (p <  0.001) and LDL (p = 0.048) than males. Males were more affected by myocardial infarction (p = 0.021) than females. However, regarding heredity for cardiovascular disease, females showed increased family history of CV (p = 0.026) compared to males. Furthermore, there were no significant differences between males and females regarding treatments with statins, beta receptor blockers and ACE inhibitors. However, females were more commonly treated with diuretics than males (p = 0.045).A total of 315 hypertensive subjects  were analyzed, and demographic data according to sex are presented in Table p = 0.008) and cMAP (p = 0.008) than males. These sex differences were also observed for cSBP (p <  0.001) and cPP (p = 0.035). After adjustments for age, HR and BSA, these differences were strengthened . Differences regarding AIx (p <  0.001), AG (p <  0.001) and transit time (TR) (p <  0.001) arterial stiffness were observed between sexes.Data on pressure and arterial stiffness divided by sex are presented in Table p = 0.003), and this difference was maintained after adjustments for age, HR and BSA (p = 0.006). Regarding diastolic blood pressure, no significant difference was found between sexes.Using the oscillometric technique, the SBP was higher among female subjects than among male subjects , and the three most common genotypes  accounted for 79.4% of the population . Subsequent analysis was restricted to individuals with the three most common FBN1 genotypes 2/2, 2/3 and 2/4 and were divided by sex. Data on pressure and arterial stiffness according to FBN1 genotype and sex are presented in Tables p = 0.036) than the 2/3 and 2/4 genotypes. After adjustments for age, the statistically significant difference compared to the 2/3 and 2/4 genotypes disappeared.There was a significant difference in HR between the three main genotypes in the male group. The 2/2 genotype showed a significantly higher HR (p = 0.029) and SBP (p = 0.025) than the 2/2 and 2/4 genotypes. The difference in AIx between the genotypes was still significant after adjustment for age (p = 0.031). Regarding SBP, the 2/3 genotype showed a slight increase in SBP after adjustment for age; however, no significant difference between the 2/3 genotype and the 2/2 or 2/4 genotype was observed. Furthermore, the 2/3 genotype in females had a significantly lower HR (p = 0.048) than the 2/2 and 2/4 genotypes.In the female group, the 2/3 genotype showed a significantly higher AIx . Small but not statistical significantly differences in AIx between the FBN1 genotypes were also seen in male.Increased arterial stiffness results in elevated systolic blood pressure, which is a prognostic indicator of future cardiovascular events , 24. Howp = 0.048) among elderly females than among elderly males, similar to our findings (Table III) [Increases in SBP, cSBP, central and peripheral MAP, and cPP were observed in the hypertensive elderly females of our study compared to males, and these findings are in accordance with those of previous reports –31. We uElderly hypertensive females seem to have higher SBP than males; thus, it is important to discuss the differences between sexes regarding the treatment of hypertension. Ong et al. (2008) studied sex differences in blood pressure rate control among Americans with diagnosed hypertension and showed that blood pressure measurement control rates were not significantly lower among females than among males . The guiFBN1 is a glycoprotein that forms FBN1-rich microfibrils and functions as a skeleton for the deposition of tropoelastin, forming elastic fibers in the arterial wall. FBN1 might affect the maintenance of the integrity of the arterial wall of central arteries, and FBN1 is considered to be a candidate gene of arterial stiffness , 35. TheThe females in the present study with the FBN1 2/3 genotype had a higher AIx and SBP than those with the other common genotypes, namely, 2/2 and 2/4. These differences were not observed in males. One causative explanation of the observed sex difference could be the fact that hypertensive males with high levels of and/or multiple risk factors for cardiovascular diseases already have died in cardiovascular events due to the early onset of cardiovascular morbidity in males , 16; howThe interpretation of the findings from the present study has limitations. First, the different number of subjects in each FBN1 genotype group Tables  and 4 coIn conclusion, the findings of our study indicate that hypertensive females in an elderly population have increased SBP and arterial stiffness compared to males. These findings also indicate that elderly females with FBN1 2/3 have increased arterial stiffness compared to hypertensive elderly females with FBN1 2/2 or FBN1 2/4. Therefore, these findings may be used to design potential therapeutic targets to reduce hypertension in females with the FBN1 2/3 genotype. Measurement of PWV may be a useful tool in clinical practice, especially in female, to select subjects at high risk of developing cardiovascular diseases . However