Central systolic and pulse pressures are stronger predictors of cardiovascular risk and hypertensive organ damage than brachial blood pressure. It is suggested that isolated systolic hypertension typically seen in adolescents is associated with normal central blood pressure and does not lead to organ damage and this phenomenon is called spurious hypertension.We assessed the prevalence of spurious hypertension and analyzed utility of pulse wave analysis as determinant of hypertensive organ damage in 294 children  diagnosed as primary hypertension. White coat hypertension, ambulatory prehypertension, ambulatory hypertension, and severe ambulatory hypertension were diagnosed in 127, 29, 41, and 97 patients, respectively.p = 0.0001). Children with severe ambulatory hypertension had higher central systolic and pulse pressure, pulse wave velocity, and greater prevalence of left ventricular hypertrophy than white coat and prehypertensive children (p < 0.05). Left ventricular mass index and carotid intima-media thickness correlated with central systolic and pulse pressure . Receiver operating curve area was similar for augmentation pressure (0.5836), 24-h ambulatory systolic blood pressure (0.5841), central systolic blood pressure (0.6090), and central pulse pressure (0.5611) as predictors of left ventricular hypertrophy.Normal central blood pressure was found in 100% in patients with white coat hypertension, 93% in pre-hypertensives, 51.2% in those with ambulatory hypertension, and 27.8% with severe ambulatory hypertension (These findings suggest that pulse wave analysis is complementary to ambulatory blood pressure monitoring in assessment of risk of organ damage in hypertensive adolescents. By definition, arterial hypertension is based on brachial artery blood pressure measurements. Because of the white coat effect, elevated blood pressure found by means of office or home measurements should be confirmed by 24-h ambulatory blood pressure monitoring (ABPM) , 2. StudTwo-hundred and ninety-four children  referred with the diagnosis of arterial hypertension based on office blood pressure readings exceeding 95th percentile for age, sex, and height on at least three independent occasions and confirmed by auscultatory blood pressure measurements were included in the study. None of the patients received antihypertensive drugs. All patients underwent full diagnostic evaluation according to the recently published pediatric guidelines of the European Society of Hypertension [All ABPM measurements were assessed oscillometrically by means of the SpaceLabs Monitor 90207 using the most appropriate cuff fitted to the non-dominant arm. Readings were taken every 20 min during daytime and every 30 min at night. Recordings lasting ≥ 20 h with ≥ 80% of readings were considered valid and were included in the analysis. Patients completed a diary for the identification of activity (wake) and sleep periods.We used a recently published classification system based on ABPM to classify patients as having white coat hypertension (WCH), ambulatory prehypertension (AmbpreHT), ambulatory hypertension (AmbHT), and severe ambulatory hypertension (AmbsevereHT) . In shorPulse wave velocity and PWA were measured non-invasively with oscillometric method using Vicorder®  system device. This system has been validated against applanation tonometry systems (Sphygmocor®) and invasive measurements of central blood pressure. It was found to be a reliable and simple alternative to tonometry , 17. MorThe Vicorder system provides a simple and quick non-invasive oscillometric method of obtaining PWV for an arterial segment. Measurement was performed in the supine position after 5 min of rest by means of the Vicorder device according to the actual guidelines , 20. A 1The waveform of brachial artery pulse obtained oscillometrically was analyzed and then by means of the transfer function the aortic waveform was calculated. The analysis of a waveform of the aortic wave enables calculation of some parameters describing the characteristics of the arterial system including aortic central blood pressure, AugPress, AugInd, cPP, cardiac output, and total peripheral resistance –21. BothAll echocardiography examinations were performed by 1 examiner who knew the clinical diagnosis, but was not aware of the severity of hypertension and the results of PWA. Echocardiography measurements were performed according to the American Society of Echocardiography guidelines , 23. To Carotid intima-media thickness was evaluated by ultrasound according to the methodology described previously. The mean WCSA was calculated from the equation: WCSA = π (dD/2 + IMT) 2 − π (dD/2) 2, where dD is the mean diastolic diameter , 26.Plasma glucose level, lipid profile, and serum uric acid were assessed at diagnosis. Blood samples were taken after 12 h of fasting. After separation of serum without anticoagulant at vacuum tubes, routine clinical chemistry parameters were measured with use of Cobas 6000® . We used unit c501 as biochemistry analyzer for spectrophotometric, immunoturbidimetric, and ion-selective determination of biochemical analytes: uric acid, ions, C-reactive protein, glucose, lipids, and creatinine.The anthropometrical indices, cIMT, WCSA, LVMi, and PWV values, were expressed as absolute values and standard deviation score (SDS) from the mean of the normal values according to the referential normative values published recently –27. The p values < 0.05 were considered statistically significant, and values between 0.05 and 0.1 were considered as demonstrating trend toward significance.Variables with normal distribution were presented as mean and SD values, whereas variables with non-parametric distribution were presented as median and range values between the 5th and 95th percentiles. The correlation analysis was performed by means of the Spearman test for non-parametric distribution. Variables with significant correlation including changes in anthropometrical parameters and changes in BP and metabolic parameters were then included in the step-wise multiple regression analysis. Sensitivity and specificity of cSBP, cPP, AugPress, AugInd, and ABPM as predictors of TOD were determined by means of the receiver operator curve (ROC) analysis.Out of 308 children referred because of the diagnosis of arterial hypertension based on at least three independent office blood pressure measurements and in whom secondary causes of arterial hypertension were excluded, 294 patients in whom all measurements have been completed were included in the analysis. In 127 (43.2%) subjects, WCH was diagnosed and 29 had AmbpreHT (9.8%). In 138 (47%) patients, the diagnosis of arterial hypertension was confirmed by ABPM, and ultimately, PH was diagnosed. In 41 (29.7%) of them AmbHT and in 97 (70.3%) SevereambHT was diagnosed . Overall, 48 of 138 (35%) children in whom ABPM confirmed arterial hypertension had normal cSBP. Out of 92 patients with ISH, 56 (61%) had elevated cSBP, 4 out of 6 (61%) of those with IDH, and 30 out of 40 (75%) patients with SDH.All patients with WCH and 27 of 29 (93.1%) patients with AmbpreHT had normal cSBP in comparison with 21 out of 41 (51.1%) patients with AmbHT and 27 out of 97 (27.8%) patients with SevereambHT  . On the average, patients with LVH had greater BMI-SDS , waist-SDS , 24 h SBP , AugPress , AugInd  . There was a trend to higher cSBP in those with LVH .In 291 out of 294 patients, results of echocardiographic examination were reliable and LVMi was assessed. Prevalence of LVH increased from 21.6% among WCH patients to 32% in patients with SevereambHT (n = 202) and elevated cSBP (n = 92) revealed significantly greater values of 24 h SBP and DBP, cIMT, LVMi, and PWV-SDS in children with the elevated cSBP , cIMT , WCSA , PWV , and PWV-SDS . cPP correlated with cIMT , WCSA , and LVMi . AugPress correlated with LVMi  and cIMT . AugInd was associated with LVMi  and cIMT-SDS .There were mild to moderate correlations between indices of cSBP and markers of TOD. cSBP correlated with LVMi  than 24 h SBP 0.5840). However, markers of obesity (BMI-SDS) and visceral obesity (WC-SDS) had greater area under curve  (Fig. 840. HoweThe main finding of our study is that one third of children diagnosed according to the current recommendations as PH had normal cSBP and low risk of hypertensive target organ damage. Second, although we found that there is steady increase in cSBP and cPP and of indices of backward wave such as AugPress and AugInd across blood pressure strata from ambulatory normotension to severe ambulatory hypertension, normal cSBP was found even among patients diagnosed with severe ambulatory hypertension. Third, assessment of PWA, including cSBP, cPP, AugPress, and AugInd, had at least the same or higher power as ABPM in predicting presence of hypertensive TOD in children with PH. However, obesity and visceral fatness had even greater predictive value.7. However, it was suggested that the assessment of cSBP and cPP may have greater importance in adolescents and young adults than in the middle age and older people [There are only few pediatric studies analyzing hemodynamic phenotype and PWA in children with PH. Most of the studies analyzing significance of PWA in assessment of cardiovascular risk in hypertensive patients were performed in adults. In the recently published meta-analysis of studies looking for association between cSBP and hypertensive TOD, it was found that the mean age of a participant was above 40 years. The general conclusion of studies carried out in adults is that cSBP and cPP had better predictive value than brachial SBP and brachial PP in the assessment of hypertensive TOD with the exception of albuminuriar people , 14. It r people . In our r people . In contr people . When blCentral systolic blood pressure increased steadily from normal in WCH subjects to elevate in SevereambHT patients. The differences were significant but still 28% of patients with SevereambHT had normal cSBP. Nevertheless, this finding supports rightness of the current pediatric classification of blood pressure status based on ABPM. A diagnosis of hypertension confirmed by ABPM differentiated patients with WCH and sustained hypertension. The same was found also by Lurbe et al. . HoweverAlthough cSBP has been linked with cardiovascular events in adults, there are no data on such association from pediatric studies. It is because hypertensive children present the first stages of cardiovascular disease and may present only with subclinical markers of hypertensive TOD such as hypertensive arteriopathy expressed as elevated cIMT and/or WCSA and LVH , 31. cSBAccording to the current recommendations, pharmacological therapy is suggested in children with PH who had TOD and/or stage 2 hypertension and/or when non-pharmacological therapy is not effective , 2. In vThe main weak point of our study is its cross-sectional nature. Thus, it is not possible to assess the risk of development of sustained hypertension and/or TOD in children with spurious hypertension. However, this issue was not studied in children in whom diagnosis of PH was confirmed by ABPM. The strong point of our study is large group of children in whom arterial hypertension was diagnosed and in whom full set of investigations toward TOD were carried out. To the best of our knowledge, it is the largest study to date involving children with arterial hypertension and with full set of data on TOD in whom PWA was assessed.Although PWA is recommended as the diagnostic approach to hypertensive adults, there are still only few data from pediatric studies. Thus, it is important to evaluate evolution of cSBP, cPP, AugPress, and AugInd during antihypertensive treatment and their relation to a regression of TOD . Second,ABPM alone is not a sufficient tool in a diagnosis of young patients with PH. New methods of cardiovascular risk assessment, such as cSBP, cPP, and AugPress, may help in discriminating patients with sustained hypertension and hypertensive TOD from patients with WCH, ambulatory prehypertension and mild hypertension without TOD. It may be an argument for a wider or even routine use of PWA in the diagnosis and treatment of hypertensive children and adolescents.


MAX) reflects inspiratory weakness in late-onset Pompe disease (LOPD). However, static pressure tests may not reveal specific respiratory muscle adaptations to disruptions in breathing. We hypothesized that dynamic respiratory muscle functional tests reflect distinct ventilatory compensations in LOPD. We evaluated LOPD (n = 7) and healthy controls  during pulmonary function tests, inspiratory endurance testing, dynamic kinematic MRI of the thorax, and ventilatory adjustments to single-breath inspiratory loads . We observed significantly lower static and dynamic respiratory function in LOPD. PIMAX, spirometry, endurance time, and maximal diaphragm descent were significantly correlated. During single-breath inspiratory loads, inspiratory time and airflow acceleration increased to preserve volume, and in LOPD, the response magnitudes correlated to maximal chest wall kinematics. The results indicate that changes in diaphragmatic motor function and strength among LOPD subjects could be detected through dynamic respiratory testing. We concluded that neuromuscular function significantly influenced breathing endurance, timing and loading compensations.Maximal inspiratory pressure (PI A shortage or absence of GAA leads to glycogen accumulation and ultimately impairment of the contractile units present in skeletal, smooth, and cardiac muscle cells2. In the late-onset Pompe phenotype (LOPD), respiratory muscle dysfunction progresses over years to decades and leads to sleep disordered breathing, ineffective airway clearance, and progressive ventilatory insufficiency4. Preferential phrenic paresis occurs early in Pompe5 and is closely followed by abdominal muscle dysfunction7. As in most neuromuscular diseases, respiratory insufficiency is the primary contributor to morbidity and mortality9.Pompe disease is a hereditary lysosomal storage disorder prompted by a mutation of the gene that encodes acid α-glucosidase (GAA), an enzyme essential for the degradation of lysosomal glycogenMAX) is the clinical estimate of inspiratory muscle strength. The PIMAX maneuver is noninvasive and does not require bulky or expensive equipment, which makes it useful for patient care and multi-site clinical trials. However, PIMAX is influenced by learning as well as the starting lung volume, making it prone to measurement error10. Moreover, the static PIMAX maneuver does not reflect the dynamic properties of diaphragm and respiratory accessory muscle contractions or take into account the effects of airway and pulmonary mechanics. In ventilator-dependent patients with infantile Pompe, we found that neither initial PIMAX nor changes in PIMAX corresponded with a reduced requirement for mechanical ventilation or airway clearance assistance12.Despite the high prevalence of respiratory problems in patients with LOPD, there are limited methods to comprehensively and objectively evaluate changes in respiratory muscle function. Maximal inspiratory pressure , depend upon the nature and magnitude of the imposed load. During ILC and respiratory muscle fatigue tests, pressure threshold loads deliver a flow-independent pressure load7. Dynamic mode (cinematic) thoracic MRI has also been employed to quantify rate of change of diaphragm descent and disease severity in pulmonary disease, scoliosis, and Duchenne muscular dystrophy20. Dynamic scans are advantageous because they do not require static breath-holds at fixed volumes to calculate respiratory muscle excursions, and multiple repeated breaths can be acquired and averaged over a short duration. Moreover, dynamic imaging can discern changes in the configuration of the chest related to diaphragm descent versus expansion of the chest wall.A second method to evaluate the dynamic function of the respiratory pump is with magnetic resonance imaging (MRI). Static MRI images of the thorax at total lung capacity and residual volume indicate that lung volumes and diaphragm descent are reduced in LOPD, and these appear to correlate with clinical pulmonary function tests such as forced vital capacityMAX and with MRI measurements of diaphragm descent. Further, we hypothesized that dynamic respiratory muscle function in LOPD would differ significantly from unaffected controls.This study compared respiratory function in LOPD to that of unaffected, age- and gender-matched controls, using ILC and dynamic MRI testing. The primary purpose of this project was to determine the relationships between pulmonary function tests, ILC responses to mechanical loads, and dynamic-mode MRI as assessments of dynamic function of the respiratory muscles in adult Pompe disease. We hypothesized that ILC volume responses and endurance will correlate both with PI1 , PIMAX , and PEMAX  were significantly below the control subjects. LOPD subjects averaged 5.7 (±4.4) years from diagnosis at testing, 5 were treated with enzyme replacement therapy (ERT), and 4 required nighttime ventilatory assistance . Body mass index, ETCO2, and minute ventilation did not differ between groups.The demographic characteristics of the sample are listed on Table Table Significant single-breath ILC responses are depicted in Fig. There were significant breath-load interaction effects for VTI  and VTE  during single-breath ILC. Neither group preserved breath volume during the 40% and 60% load magnitudes, when compared to lower loads, unloaded control breaths, or recovery breaths (p < 0.005). Despite a lower PIF during loaded breaths, both Pompe and control subjects minimized the loss of volume by tending to lengthen TI . Additionally, both groups significantly shortened TE during loaded breaths , as compared to the unloaded control and recovery breaths.Figure MAX and FVC correlated significantly , along with maximal diaphragm descent and Tlim . Inspiratory volume ILC significantly correlated to flow , as well as inspiratory and expiratory timing . In LOPD, volume and inspiratory flow ILC correlated with maximal chest wall expansion  but not with diaphragm descent .For the entire sample, PIThe key findings of these research tests of dynamic inspiratory muscle function indicated kinematic evidence of diaphragm paresis, accompanied by altered inspiratory endurance and strength in patients with LOPD. The ILC tests indicate an impaired ability for LOPD patients to generate inspiratory flow with an unanticipated inspiratory load. However, a compensatory prolongation of TI helped to preserve volume during the loaded breath. We also observed flow and volume ILC were associated with MRI tests of maximal chest wall expansion, but not with diaphragm kinematics, of patients. While likely more useful as research tools than in daily clinical practice, these tests appear to capture distinct aspects of dynamic respiratory muscle function beyond standard pulmonary function tests, which could help clarify how patients maintain minute ventilation despite gradual LOPD disease progression.We used dynamic MRI to evaluate kinematics of breathing at rest and during inspiratory capacity. The C-C kinematic reserve represented the difference between resting diaphragm excursion and maximal diaphragm excursion, and it indicated the presence of diaphragmatic paresis in LOPD. On the other hand, A-P resting and reserve excursion did not differ significantly between patients and controls, indicating relative preservation of chest wall expansion in patients.7, but many prior efforts focused on static breath-holds. Our findings of early diaphragmatic paresis in LOPD were also in agreement with existing respiratory studies using opto-electronic plethysmography (OEP)21 and static MRI6. Additional advantages to using dynamic MR modes include the ability to average the kinematics over multiple breaths in a short period of time and the removal of possible confounding effects of elastic recoil on breath-holding22. MRI may be suitable for multi-site research studies that cannot accommodate the substantial equipment costs and data analyses required with OEP. For example, children with infantile-onset Pompe disease tolerated dynamic respiratory MRI in a clinical trial of AAV1 gene therapy11, while on ventilatory support and for brief periods of CPAP or independent breathing.Existing studies in adult onset Pompe also contrasted MRI to conventional lung function tests24. However, there are limitations to MRI, particularly in the context of clinical care. In the current study, one LOPD subject with inspiratory muscle weakness and absent C-C excursion required the use of BiPAP to image tidal breathing. Therefore, while ultrasonography can yield lower tissue resolution and an insufficient window in some patients, it may be preferred in clinical settings or to monitor moderate to severe diaphragmatic involvement26. Additionally, sniff nasal inspiratory pressure (SNIP) reflects pressure associated with a maximal effort sniff. While the current study focused on MIP due to its common inclusion as a clinical study endpoint, the SNIP test is well tolerated in neuromuscular diseases27 with potential predictive value for respiratory morbidity28.Thoracic MRI may also offer scientists insights regarding the presence of fat or fibrotic incursions in the respiratory muscles29. Since the respiratory pattern and mouth pressure were standardized, the endurance test has been considered less susceptible to learning effects, in healthy subjects and patients with neuromuscular disease31. As loads increase in magnitude or are sustained to fatigue, inspiratory flow and inspiratory time progressively diminish32. Subjects reached task failure (Tlim) when they could not generate sufficient pressure to continue to open the inspiratory valve and receive airflow32. Longitudinal changes in Tlim may indicate an external factor  altered the endurance Tlim, and further study is needed on whether tests of threshold loading can longitudinally track changes respiratory muscle function with either progressive disease or in response to therapeutic interventions such as inspiratory muscle training. Indeed, recent reports suggest inspiratory muscle training can lead to strength gains, in some individuals with LOPD33, We previously observed that difficult to wean ICU patients who failed to wean after inspiratory muscle training had lower flow/volume ILC that failed to improve with training34. We speculate that Tlim and/or ILC tests could help scientists further identify inspiratory flow and volume signatures associated with an incomplete strengthening or functional benefit from inspiratory muscle training31.Tlim was restricted in LOPD and correlated strongly with conventional lung function tests as well as maximum diaphragm excursion during MRI. In contrast to the MRI tests, all LOPD subjects were able to complete Tlim. Advantages of endurance tests are their ability to discern changes in pressure generation that do not involve a static hold and become less effort-dependent with approaching task failure35. We observed these modifications during single-breath ILC, with some distinctions in the LOPD group. Consistent with other neuromuscular diseases36, both TI and TE were slightly shorter in LOPD subjects than CON. As the magnitude of a single breath load increased, both PIF and volume dropped significantly in LOPD subjects. With inspiratory challenges, the diaphragm responds by increasing the firing frequency of active motor units and by recruiting previously inactive diaphragm motor units37. Additionally, accessory muscle motor units that are largely inactive at rest are recruited39. In healthy individuals, recruitment of chest wall muscles is thought to offset diaphragm fatigue during sustained inspiratory loading40. In LOPD, we noted a strong association between increased chest wall kinematic motion and the PIF and VTI generated during single-breath ILCs. We speculate that chest wall respiratory recruitment may influence single-breath ILC responses in LOPD, because early, severe diaphragm paresis minimizes the ability of the diaphragm to compensate to the transient, unexpected load.Compensatory responses to a single-breath threshold load include reductions in inspiratory flow and volume, with prolongation of TI and reduction of TE41. Both neurological and muscular contributions to respiratory dysfunction in LOPD have been reported in the literature. Diaphragm dysfunction is a distinguishing characteristic of LOPD identified even among mildly affected LOPD patients42, but an enhanced inspiratory neural drive may be able to preserve independent breathing in early diaphragm dysfunction43. Neuromuscular activity, as estimated by dP/dt, was similar for Pompe and control subjects during ILC44. ILC loading was administered as a proportion of MIP, rather than a fixed value, indicating that relative proportional loads elicited a similar proportional neuromuscular activation. While the small sample did not permit a more rigorous comparison of neuromuscular activation in the patients who used nighttime support48, the use of nighttime ventilation in LOPD improves both ventilation and oxygenation during sleep, without an appreciable change in respiratory muscle strength49. In the current study, the similar dP/dt and prolongation of TI with threshold loading in both LOPD and control subjects indicates that both groups used a similar motor control strategy to overcome inspiratory loads, and we speculate this could indicate a loss of respiratory motor units in Pompe, as opposed to an altered functioning of the motor neurons48.Fifty-seven percent of the LOPD subjects required nighttime non-invasive respiratory support, which itself indicates a loss of respiratory neuromuscular reserve50. It could be argued that both the Tlim test could be considered as effort-dependent as lung function and maximal respiratory pressure tests51. A distinguishing factor is the binary nature of the threshold valve, which blocks inspiratory flow for inspiratory pressures below the threshold. Additionally, the threshold valve mechanics and limited compensatory strategies for clinical populations reduce effort and learning effects on ILC and Tlim. Finally, the finite time and energy of LOPD patients required us to limit the range of respiratory tests. Specifically, invasive tests of evoked diaphragmatic pressure would provide additional mechanistic insights on non-voluntary activation of the diaphragm, while SNIP and diaphragm ultrasound tests offer clinical insights into dynamic diaphragm function.We acknowledge some limitations to this study. FVC and respiratory pressures were not tested in supine. Subject fatigue was the primary consideration for this decision, and IC in the MRI was computed instead. While the requirement to maintain a consistent respiratory rate and duty cycle during the endurance test has been criticized by some, it reduces variability in the breathing pattern, namely an early prolongation of exhalation, which could artificially prolong TlimIn summary, our findings provide evidence of early dynamic diaphragmatic dysfunction in LOPD, compared with controls. This provides critical evidence that diaphragm dysfunction in Pompe alters not only static force generation, but also impairs the ability to preserve minute ventilation during transient or sustained inspiratory loads. In turn, a lowered dynamic inspiratory muscle reserve may ultimately affect a patient’s ability to resist physiological stressors such as bronchoconstriction or a narrowed airway during sleep, placing them at higher risk for failure. Future studies should distinguish how dynamic respiratory muscle function changes with disease progression.MAX and the volume ILC, at least 6 subjects per group would be needed for α = 0.05, (1-β) = 0.9, (ρ = 0.68). The study design and procedures were approved by the University of Florida Institutional Review Board and conducted in accordance with principles established in the 1964 Declaration of Helsinki and its subsequent amendments. All participants provided their informed consent prior to inclusion in the study. The study was registered in February 2015 (NCT02354664).Fourteen subjects volunteered to participate. Seven subjects had a diagnosis of LOPD confirmed by genetic testing or enzyme activity assays , and the remaining seven were age- and gender-matched unaffected control subjects (age 20–61 years). Based upon a power analysis of preliminary data on the correlation between PIExclusionary characteristics for patient participation included: pre-existing obstructive lung disease or asthma, a forced vital capacity (FVC) < 30% of age/gender predicted values, requirement for mechanical ventilation while awake and upright, inability to travel to the study site, pregnancy, or any other chronic medical condition that would interfere with study participation.Each subject participated in a single 6-hour study visit, divided into two days. The following tests were included: (1) Clinical pulmonary function and maximal respiratory pressure tests, (2) Dynamic MRI of the thorax, (3) Resting breathing pattern, (4) ILC to unexpected, single-breath loads, (5) ILC to sustained loads, and (6) inspiratory endurance.52 and compared to references for age, gender and ethnicity53. Tests were repeated until three technically-acceptable efforts were acquired within 5% variability. In addition, subjects performed PIMAX maneuvers from residual volume and maximal expiratory pressure (PEMAX) from total lung capacity, per American Thoracic Society testing guidelines52. Each trial was repeated after a>30 second break, until <10% variability was achieved between three trials .Upright forced vital capacity (FVC) was completed per American Thoracic Society guidelines55. Triplanar localizer images were acquired during a maximal inspiratory breath-hold, to establish the optimum field of view for the dynamic scans. A T2-weighted, balanced, gradient echo sequence with short repetition time and retrospective gating was employed to obtain fast imagery with excellent tissue resolution56, . Separate dynamic scans captured resting tidal breathing and inspiratory capacity maneuvers, in the coronal and right sagittal planes. Each tidal breathing acquisition lasted one minute and captured 12–15 tidal breaths.Patients were positioned supine in a 3.0T research MR scanner (Philips Achieva) using a 32-channel cardiac coil. We customized a dynamic MR sequence, based on techniques validated in pulmonary diseaseDuring each MRI scan, respiratory parameters were collected simultaneously, using a flowmeter  connected to the patient via facemask and placed in series with a respiratory monitor  and laptop computer. Respiratory data acquisition occurred at 100 Hz frequency. Extended tubing was required to maintain the data acquisition equipment behind the 5 Gauss line, which added 24 mL of dead space to the respiratory circuit.Tidal breathing pattern was assessed with subjects seated wearing a sealed facemask and in a relaxed state listening to music. Approximately 10 minutes of steady-state, resting data was sampled at 100 Hz with a respiratory monitor  attached to a laptop. The resting respiratory rate was used as the breath rate for the endurance test.2O). Inspiratory loads of either 10, 20, 40, or 60% of PIMAX (or a no-load control) were applied during exhalation, for a single inspiratory effort. Subjects were separated from the testing apparatus by a screen. Six presentations of each load were applied in random order, and a minimum of 4 and maximum of 12 unloaded breaths separated each ILC breath.Single-breath ILC was tested using a custom apparatus that contained 4 pressure-threshold devices  connected to the patient circuit, which consisted of a pneumotachograph  with phalange mouthpiece and nose clip. Some of the pressure-threshold devices contained a modified spring to provide higher threshold loads . A pressure-threshold inspiratory load equivalent to 40% of PIMAX was placed on the inspiratory port of the mouthpiece. The respiratory rate was set to each subject’s self-selected resting breath rate with a 50% duty cycle, and a two-tone metronome timed inhalation and exhalation. Oxygen saturation, end-tidal CO2 (ETCO2), heart rate, tidal volume, and respiratory rate were continuously monitored, with blood pressure readings every 2 minutes. Subjects received encouragement to maintain the target rate and mouth pressure. The test ended when one of the following conditions was met: (1) the inspiratory port did not open for 3 continuous breaths, (2) significant changes in heart rate or blood pressure outside of conventional exercise testing limits, (3) the subject reported intolerance or inability to continue, or (4) the subject reached 10 minutes of continuous loaded breathing.Endurance was evaluated by identifying the time limit (Tlim) that a participant could maintain breathing with a submaximal inspiratory threshold load, as described previously in Duchenne muscular dystrophy57. The first ten uninterrupted breaths in each breathing sequence were evaluated. The cranio-caudal length (C-C) and anterior-posterior width (A-P) of the thorax were measured in each breath at end-inhalation and end-exhalation and represented the positions of the diaphragm and chest wall, respectively. Kinematic excursion was calculated as the difference between inspiration and expiration distance. Figure MRI images were uploaded to a secure database and then analyzed offline with an open-source DICOM viewer  by a trained assistant blinded to subject group. We evaluated the right sagittal plane, since height of the diaphragm differs by ventral-dorsal position See Fig. 57. The f44. In addition, we averaged the breaths before (unloaded control) and after (recovery) the loaded breath.At each load magnitude, the breaths with the highest and lowest volume were excluded. The remaining 4 presentations were averaged for peak inspiratory and expiratory flow , inhaled and exhaled volume , inspiratory and expiratory times , and maximal date of inspiratory pressure development (dP/dt). The dP/dt was the positive peak of the pressure-time derivativeAnalyses were performed with the IBM SPSS Statistics 21.0 statistical package . Group demographics were compared using 2-sample t-tests. For MRI studies, we used 3-way repeated measures ANOVAs to evaluate the kinematic excursion of the respiratory muscles , and 2-way ANOVA to compare the volumetrics . Single-breath ILC was assessed using a 3-way ANOVA . A 2-way ANOVA tested dP/dt of the loaded breaths. With ANOVA testing, we corrected violations of sphericity with Greenhouse-Geiser adjustments. Tukey’s HSD post hoc test was used to assess individual ANOVA differences. Two-sample t-tests were used to compare the Tlim of the Pompe subjects and controls. Pearson’s correlations were used to evaluate the linear relationships between the pulmonary function, imaging, ILC, and endurance variables. The alpha level was 0.05, with the exception of correlations, which used a more stringent threshold of p < 0.005 to correct for multiple correlations.


Although patients were carefully treated in outpatient clinics and adequate pharmacological therapy was prescribed, we noted an increase in total cholesterol (p = 0.001), LDL cholesterol (p = 0.002), HbA1C (p = 0.054), and diastolic blood pressure (p = 0.047) However, multivariate analysis revealed that this increase in cardiovascular risk factors could not explain the observed deterioration in endothelial function. In patients with STEMI, we observed a significant deterioration in endothelial function during 12 months after PPCI that could not be explained by changes in the traditional cardiovascular risk profile.Knowledge about the changes in endothelial function after ST-elevation myocardial infarction (STEMI) is of substantial interest, but serial data are scarce. The aim of the present study was to noninvasively evaluate whether endothelial function, as assessed shortly after primary percutaneous coronary intervention (PPCI) for STEMI, may improve until 12-month follow-up. This prospective observational cohort study was performed in patients in the RESPONSE randomized trial who participated in a substudy and underwent noninvasive assessment of endothelial function at 1 (baseline), 6, and 12-month follow-up after treatment of a STEMI by PPCI. The reactive hyperemia peripheral artery tonometry (RH-PAT) method was used to assess endothelial function (higher RH-PAT index signifies better function). Of the 70 study participants, who were 57.4 ± 9.7 years of age, 55 (78.6%) were male and 9 (13%) had diabetes. The endothelial function deteriorated significantly during follow-up: the RH-PAT index at baseline, 6, and 12-month follow-up was 1.90 ± 0.58, 1.81 ± 0.57, and 1.69 ± 0.49, respectively ( Coronary heart disease remains the leading cause of morbidity and mortality in societies with a Western lifestyle . PatientSeveral techniques are available for a noninvasive assessment of peripheral endothelial function, which has been shown to correlate well with coronary endothelial function , 7. WhilIn the present prospective substudy of the RESPONSE  trial , we obtaThis prospective cohort study was performed in 70 STEMI patients in the RESPONSE trial , who undPatients were eligible for enrollment if they: were between 18 and 80 years of age; were available for and willing to adhere to the follow-up procedures; had no surgery or additional PCI planned within 8 weeks from PPI; and had a life expectancy of at least 2 years in the absence of heart failure NYHA class III or IV. Details of the randomized RESPONSE trial have previously been reported .As inflammation and repair processes of the infarcted myocardium might have disturbed endothelial function measurements during the first weeks after the STEMI and endothelial dysfunction has not been fully recovered under medication, endothelial function was first assessed during 1-month follow-up  –14. All Patients were treated in the ambulance with an intravenous bolus of 5.000 IE of unfractionated heparin, a loading dose of ≥ 300 mg of acetylsalicylic acid  and an oral loading dose of 600 mg of clopidogrel. In 50 patients, a weight-adjusted intracoronary bolus of abciximab was administered after visualizing the culprit coronary artery. PPCI procedures were generally performed via the femoral route through 6F sheaths. Pharmacological therapy, use of aspiration catheters, lesion preparation (vs. direct stenting), and stent postdilatation were performed according to current guidelines and the operator’s judgment and discretion. Following primary PCI, a heart team carefully assessed the coronary angiographies and, if required, patients underwent a staged PCI for additional coronary lesions, which was generally performed within 1–2 weeks from primary PCI.Endothelial function was evaluated with the RH-PAT method. The finger pulse wave amplitude was assessed with the EndoPAT-2000 sensing device and finger plethysmographic probes , both at baseline and during ischemia-induced hyperemia. All measurements were performed in the early morning in a dedicated laboratory after patients had fasted for at least 8 h. Patients also had to refrain from caffeine consumption, smoking, and vasoactive medications. At least 15 min prior to testing, blood pressure was measured and a blood sample was drawn in the control arm. Before any measurement, patients had an acclimatization period of 20 min in a quiet room, lying in a hospital bed at an ambient temperature of 21–23 °C.The RH-PAT method has previously been reported in detail , 16. In 2); arterial hypertension (blood pressure of > 140/90 mmHg or treatment with anti-hypertensive medication); history of smoking (previous or current smoker); history of previous myocardial infarction and/or coronary revascularization by means of PCI or bypass surgery; the presence of diabetes mellitus ; and history of hypercholesterolemia or treatment with lipid-lowering drugs. Procedure details were also documented. At follow-up, RH-PAT and systolic blood pressure, diastolic blood pressure, and BMI were measured; current smoking habit and medication use were documented. Blood samples were analyzed for the measurements of total cholesterol, HDL cholesterol, LDL cholesterol, triglycerides, fasting glucose, and HbA1c. Patients were instructed to fast for a period of 8 h prior to drawing of the blood sample. At the three times of follow-up, we assessed the proportion of patients on adequate cardiovascular risk factor control, according to the 2003 European guidelines of cardiovascular disease prevention [The following information was documented at baseline: RH-PAT; age; sex; body mass index .Data are presented as frequencies (%) or mean ± standard deviation (SD). Between-group comparisons of RH-PAT were conducted using longitudinal mixed models. Time was treated as a categorical variable . Compound symmetry covariance structure was used for time. Only those variables that significantly modified over time were included in the subsequent multivariate analyses. As repeated covariates, we included Hb1Ac, diastolic blood pressure, total cholesterol, and LDL cholesterol. A 2 at inclusion; 40.0% were smokers while 12.9% had diabetes, 28.6% hypertension, and 24.3% hypercholesterolemia. Eight patients (11.4%) had a history of previous PCI. The index PPCI procedure was performed through the femoral route in 98.4% of the patients with an ischemia time of 155.9 ± 105.5 min. Of all patients, 73.9% had single-vessel disease and 89.9% had a left ventricular ejection fraction > 50% prior to discharge (Table Patient characteristics at baseline are displayed in Table p = 0.04 in mixed model). Further details of the RH-PAT index measurements are displayed in Table The endothelial function decreased over time, starting at a mean RH-PAT index of 1.90 ± 0.58 at baseline, to 1.81 ± 0.57 at 6-month follow-up, and eventually to 1.69 ± 0.49 at 12-month follow-up (p < 0.01).Table A longitudinal mixed model analysis shows that, of all cardiovascular risk factors assessed, the following worsened significantly over time: HbA1c; diastolic blood pressure; total cholesterol; and LDL cholesterol Table . Howeverin STEMI patients during a period of 12 months after PPCI.Following STEMI treated by PPCI, endothelial function—as measured by RH-PAT—did not improve from baseline at 1–12-month follow-up, despite the prescription of guideline-recommended medical therapy  in the mThrough the years, several methods have been developed to identify subjects with disturbed endothelial function, as they are at an increased risk of cerebrovascular  and cardAnother method of measuring the reactive hyperemic response to ischemia is RH-PAT. Data obtained with RH-PAT were shown to correlate well with both findings of the high-frequency ultrasound method and coronary endothelial function , 10, 20.Other research groups have performed serial endothelial function measurements in different study populations, assessing changes during a period of 3–6 months. Kitta et al.  previousThe phenomenon of persistent impairment of endothelial function was also seen in 245 patients with chronic heart failure, in whom endothelial function was serially assessed during 6 months by flow-mediated arterial vasodilatation . DespiteOur present study (in STEMI patients) as well as most of the previous studies (in other patient populations than STEMI patients) suggest that the continuum of endothelial dysfunction can ultimately reach a stage from which—despite adequate or even optimal medical therapy and risk factor control—restoration is unlikely or even impossible. At that point, the total load of atheroma, genetic predisposition, or unknown other risk factors may become increasingly important for the further course of endothelial function. While we found endothelial function to decrease over time, the majority of patients did not fulfill the RH-PAT method-based criterium of a significant endothelial dysfunction  . In addiUse of the femoral access route in 98% of our patients may be considered an advantage . NeverthIn patients with STEMI, we observed during 12 months after PPCI a significant deterioration in endothelial function, as measured by the RH-PAT method, which could not be explained by changes in traditional cardiovascular risk profiles. These findings suggest that the continuum of endothelial dysfunction can ultimately reach a stage from which restoration is unlikely.


Little is known about factors that affect the composition of contracted blood clots in specific diseases. We investigated the content of polyhedral erythrocytes (polyhedrocytes) formed in blood clots and its determinants in type 2 diabetes (T2D) patients.1c, 6.4% (interquartile range 5.9–7.8)], we measured in vitro the composition of blood clots, including a clot area covered by polyhedrocytes using scanning electron microscopy and the erythrocyte compression index (ECI), defined as a ratio of the mean polyhedrocyte area to the mean native erythrocyte area. Moreover, plasma fibrin clot permeability (Ks), clot lysis time (CLT), thrombin generation, oxidative stress , and platelet activation markers were determined. The impact of glucose concentration on polyhedrocytes formation was assessed in vitro.In 97 patients with long-standing T2D [median HbAs or CLT. Patients who formed more polyhedrocytes (≥ 10th percentile)  had higher glucose , fibrinogen , lower red blood cell distribution width , reduced plasma clot density  and impaired fibrinolysis  when compared to patients with lesser amount of polyhedrocytes (< 10th percentile). ECI and the content of polyhedrocytes were strongly associated with total PC . In vitro an increase of glucose concentration by 10 mmol/L was associated with 94% higher polyhedrocytes content (p = 0.033) when compared to the baseline (7.1 mM). After adjustment for age, sex and fibrinogen, multiple regression analysis showed that RDW was the only independent predictor of polyhedrocytes content in T2D .Polyhedrocytes content in contracted clots was positively correlated with glucose , glycated hemoglobin , total cholesterol , TBARS , P-selectin  and platelet factor-4, PF4 , but not with thrombin generation, platelet count, KPoor glycemic control, together with enhanced platelet activation and oxidative stress, increase the content of polyhedrocytes in blood clots generated in T2D patients.The online version of this article (10.1186/s12933-018-0789-6) contains supplementary material, which is available to authorized users. Type 2 diabetes (T2D) is associated with a hypercoagulable state involving increased platelet activation, thrombin generation and unfavorable fibrin clot properties –4. T2D pTo our knowledge, there have been no reports on determinants of polyhedrocyte formation in clots of patients with T2D. RBCs are extremely sensitive to oxidative stress or cytokine upregulation. This usually accompanies systemic inflammation in most diseases, including T2D . We hypoWe enrolled 97 consecutive T2D patients with high cardiovascular risk hospitalized from August 2015 to April 2016 in John Paul II Hospital in Krakow, Poland and 30 apparently healthy controls. All patients fulfilled the American Diabetes Association criteria for diagnosis of T2D and were treated for at least 12 months. The exclusion criteria were arterial or venous thromboembolic events within previous 6 months, current anticoagulant therapy, known cancer, signs of acute infection, chronic inflammatory disorders (e.g. rheumatoid arthritis), liver injury , estimated glomerular filtration rate (eGFR) < 30 mL/min, and pregnancy. Current smoking was defined as smoking at least one cigarette daily. Arterial hypertension was defined as a systolic and/or diastolic blood pressure of ≥ 140 mmHg or ≥ 90 mmHg, respectively or a history of arterial hypertension with or without taking antihypertensive agents. Coronary artery disease (CAD) was established based on a documented history of myocardial infarction (MI) or a positive result of ECG stress test or gated single photon emission computed tomography with Tc-99 m-MIBI (SPECT) or a coronary angiography. Previous MI was established based on medical records.The Jagiellonian University Ethical Committee approved the study and all the participants provided their written informed consent.g and 20 °C for 10 min to obtain platelet poor plasma (PPP), snap-frozen within 60 min, and stored in small aliquots at − 80 °C until analysis. The activated partial thromboplastin time (aPTT), creatinine, eGFR, serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), glucose and thyroid stimulating hormone (TSH) were assayed by routine laboratory techniques. Glycated hemoglobin A1c (HbA1c) was measured using immunoturbidimetry . Complete blood count including white blood cells (WBC), RBC, hemoglobin, hematocrit, red blood cell distribution width (RDW), platelet count and platelet distribution width were assayed. Fibrinogen was assessed using the Clauss method. The hsCRP was determined using immunoturbidimetry . Immunoenzymatic assay was used to determine plasminogen activator inhibitor-1 (PAI-1) antigen  in citrated plasma. Plasma α2-antiplasmin (α2AP) and plasminogen were measured by chromogenic assays . Markers of platelet activation, P-selectin (CD62P) and platelet factor-4 (PF4), were determined in citrated plasma by ELISAs . The interassay and intraassay coefficients of variation for all the ELISAs were < 8%.Fasting venous blood was drawn between 7 and 10 A.M. and was kept at a room temperature. Blood samples were collected into citrated tubes (9:1 of 0.106 M sodium citrate), centrifuged at 25002  and human thrombin  at final concentrations of 0.01 M and 1 U/mL, respectively) to 48 μL of whole blood from the antecubital vein that was prewarmed for 5 min at 37 °C. The samples were incubated at room temperature for 24 h.Clotting was initiated by addition of 2 µL of activation mixture (CaClScanning electron microscope (SEM) analysis was performed as previously described . Blood c2) of native RBCs and polyhedrocytes. Results were presented as means ± standard deviations (SD) of about 100 consecutive RBC areas obtained from each SEM image.For the semiquantitative analysis, each picture was divided into 400 squares and the dominant composition  of each was assayed Fig. . The visErythrocyte compression index (ECI) was determined as previously described . Briefly2, 60 mg/mL bovine albumin, and 2.5 mmol/L Z-Gly-Gly-Arg-amidometylcoumarin). The reference ranges of thrombin generation potential were as follows: Lag time, 1.57 ± 0.33 min; ETP, 1694 ± 373 nM × min; Peak thrombin generation, 360 ± 108 nM; ttPeak, 3.96 ± 0.71 min [Plasma thrombogenic potential was assessed using calibrated automated thrombography (CAT)  in a 96-well plate fluorometer  at 37 °C according to the manufacturer’s instructions. Eighty microliters of platelet poor plasma were diluted with 20 µL of a commercially available tissue factor (TF)-based activator  containing 5 pM recombinant TF, 4 micromolar phosphatidylserine/phosphatidylcholine/phosphatidylethanolamine vesicles, and 20 µL of FluCa solution , which indicates the pore size, was calculated from the equation:2), and Δp, a differential pressure (in dyne/cm2).Fibrin clot permeability was determined using a pressure-driven system . Briefly−9cm2; CLT, 100 ± 9.7 min [To assess efficiency of clot lysis, citrated plasma was mixed with 15 mM calcium chloride, 6 pM human tissue factor , 12 μM phospholipid vesicles and 60 ng/mL recombinant tPA . The mixture was transferred to a microtiter plate and its turbidity was measured at 405 nm at 37 °C. Clot lysis time (CLT) was defined as the time from the midpoint of the clear-to-maximum-turbid transition, which represents clot formation, to the midpoint of the maximum-turbid-to-clear transition (representing the lysis of the clot). The reference ranges of fibrin clot permeability and lysis were as follows: Ks, 7.01 ± 2.06 × 10 9.7 min , 31.−1 cm−1. The results, expressed as nM/mL of PC, were then normalized for protein concentration. The reference range of total PC was 1.82 ± 0.2 nmol/mg [Oxidative modification of plasma proteins was assessed based on carbonyl content using 2–4 dinitrophenylhydrazine (DNPH), as reported by Becatti et al. . DNPH re nmol/mg .Thiobarbituric acid reactive substances levels were measured in citrated PPP using a TBARS assay kit  in accordance with the manufacturer’s instructions as previously reported . BrieflyThe ORAC method , based on the inhibition of the peroxyl-radical-induced oxidation initiated by thermal decomposition of azo-compounds, like 2,2icazobis(2-amidinopropane) dihydrochloride (AAPH), was performed as reported by Becatti et al. . BrieflyThe oxidation parameters, platelet markers, and fibrinolytic proteins were measured in a subset of patients (n = 23).To assess the direct influence of glucose concentration on polyhedrocytes formation, we added 10 mM glucose solution in 0.9% sodium chloride to blood obtained from randomly selected 10 healthy  and 7 randomly selected T2D subjects . The samples were incubated at 37 °C for 1 h. Then, the whole blood clots were prepared in duplicate and analyzed as described above by two independent investigators blinded to the clinical data.2 test of Fisher’s Exact Test. Pearson’s or/and Spearman’s correlations were calculated when applicable for measuring the association between two continuous variables. The change in analyzed variables due to the increase in polyhedrocytes by 1% was assessed using linear regression. Several logistic regression models with production of polyhedrocytes as a dependent variable (production occurred or not occurred). The best model was then obtained stepwise backwards minimalizing Bayesian Information Criterion (BIC) with variables gender, age and level of fibrinogen locked in the model for adjustment. Association between the variables was expressed as odds ratio (OR) with corresponding 95% confidence intervals (CI). A p-value < 0.05 was considered statistically significant.Continuous variables were presented as mean and standard deviation or median with the first and the third quartile depending on data distribution assessed using Shapiro–Wilk test and compared using Student’s t test or Wilcoxon test. Nominal variables were described using counts and percentages and compared using χ1c, 6.4%), long-standing  T2D  were considered as a low-polyhedrocytes group (LP), in contrast to the rest of the patients  who were considered as a high-polyhedrocytes group (HP). Median percentage of polyhedrocytes in blood clots in the HP group was 20%  and it was 51% lower than in healthy controls . There were no differences between the HP and LP groups in demographic and clinical characteristics including the medications used , but not with other lipid parameters. Compared with patients with glucose < 6 mmol/L, those with higher glycemia were more prone to form greater amounts of polyhedrocytes .Platelet markers, fibrinolytic proteins and oxidation parameters are presented in Additional file In the HP patients the median of the new parameter describing compression of erythrocytes, ECI was 59.4 (53.7–69.6)%. There were no associations between ECI and demographic or clinical parameters in those patients. ECI was strongly associated with total PC  and with fibrinogen level .1c   showed that their major components were native erythrocytes 32.6 ± 16.6%), transitional erythrocytes (28 ± 12.6%), and fibrin (24.5 ± 9.4%). Polyhedrocytes accounted for nearly 8% and platelets for 1% of clot composition. Interestingly, the percentage of polyhedrocytes was positively associated with HbA6.6%, traThe in vitro increase of glucose concentration from 4.9 to 14.9 mmol/L in blood obtained from healthy subjects was associated with higher content of polyhedrocytes . Simple linear regression models showed that 1% increase in percentage of polyhedrocytes was significantly associated with the increase in TBARS by 0.4%), total PC (by 0.5%), P-selectin (by 0.5%), and PF4 (by 0.3%)  . In contRBCs are especially sensitive cells, therefore they are important health indicators . AssessmThe clot’s ability to contract is a feature of each patient and could be modulated by many transient factors . The majTutwiler et al. reported that higher hematocrit along with an increased RBC rigidity were associated with limited clot contractility . In our 1c and accumulation of intracellular sorbitol or to the stiffness of red blood cell membranes probably due to oxidative modification of the proteins and to an imbalance of the cholesterol/phospholipids ratio [We have shown positive association of polyhedrocytes formation with worse glycemia control. Moreover, in the vitro experiment we demonstrated, that higher glucose concentration is associated with higher polyhedrocyte content in blood clots. Erythrocytes of patients with T2D exhibit impaired deformability related to increased HbAds ratio . Moreoveds ratio  and wereds ratio . Therefos or prolonged CLT. We have previously demonstrated that prothrombotic fibrin clot phenotype and impaired fibrinolysis might be associated with prolonged T2D duration [Patients who formed more polyhedrocytes in our study showed also prothrombotic fibrin clot properties, such as decreased Kduration . Moreoveduration . Carrollduration . The cur1c was measured using immunoturbidimetry, but not by high-performance liquid chromatography. Third, we did not measure the RBCs membranes rigidity or assess function of platelets such as platelet aggregation. It is unclear whether polyhedrocytes formation affects the risk of cardiovascular events or thrombosis in T2D as suggested by other studies with regard to MI [First, a limited number of patients have been included in the study, therefore subgroup comparisons should be analyzed cautiously. Second, HbArd to MI  or strokrd to MI . FinallyIncreased glucose levels, enhanced oxidative stress and platelet activation were associated with increased formation of polyhedrocytes in contracted blood clots. The significance of in vivo polyhedrocytes in T2D and in other diseases remains to be established and further studies are necessary to demonstrate the role of polyhedrocytes in thromboembolic events.Additional file 1: Table S1. Platelet markers, fibrinolytic proteins and oxidation parameters in a subset of the study group T2D patients (n=23).


Heparinized solution (HS) use for the maintenance of arterial cannulas has been associated with coagulation disorders and has not been conclusively shown to confer additional benefits over normal saline (NS) alone. We tested the hypothesis that in adult patients admitted to the cardiac ICU (T0) after cardiac surgery, HS arterial cannulas might be independently associated with increased heparin-induced coagulation disorders and might not be superior to NS arterial cannulas in reducing arterial cannula occlusion.In this cohort study, 170 patients who received NS arterial cannulas during the period from T0 to ICU discharge were included in this study from June 1, 2017, to May 1, 2018 (NS group). There were 2930 patients who used HS (2.5 U/ml) arterial cannulas from January 1, 2015, to December 31, 2016 (heparin group). To address indicated biases, we derived a propensity score that predicted the functions of NS and HS in the patency of arterial cannulas.P = 0.4) and arterial cannula occlusion (P = 0.5) showed no differences between the NS and heparin groups. However, the heparin group had a significantly increased activated clotting time (P < 0.05), activated partial thromboplastin time (P = 0.01) and allogeneic red blood cell utilization . Compared with the NS group, the heparin group had more drainage from chest tubes from T0 to T48  and had more allogeneic red blood cells transfused .There were 296 patients (148 in the NS group and 148 in the heparin group) with similar risk profiles in the propensity-score matched cohorts. In the propensity-matched patients, the duration of arterial cannulas (Based on the results of our study, the addition of heparin to normal saline for flushing arterial pressure monitoring cannulae did not reduce the incidence of catheter thrombosis and result in a very small but statically significant in increase in activated clotting time and activated partial thromboplastin time. Cardiac surgery is increasingly performed for high-risk patients referred for operations . Many atRoutine arterial cannula maintenance includes regular flushing with heparin solution  to prevent cannula occlusion . Other cSome studies, including a guideline , severalThis study was approved by the ethical committee of Nanjing Drum Tower Hospital on February 28, 2017. From January 1, 2015, to May 1, 2018, 3100 patients aged > 18 years old and requiring NS and HS arterial cannulas were enrolled following surgery for the repair of heart disease in our Hospital. Patients with aortic diseases, acute mesenteric ischaemia, pregnancy, cancers, chronic obstructive pulmonary disease (COPD) and extracorporeal membrane oxygenation (ECMO) initiation before surgery, known allergies to heparin, platelet treatment, a history of HIT, coagulation disorder before using heparin and thrombocytopenia with platelets less than 100,000 mm3 were also excluded. After the study was approved by the ethical committee of our hospital, we reviewed hospital medical records, nursing records, the laboratory database, and the cardiac surgical database.In our hospital, the replacement of heparinized arterial cannulas was routinely implemented before 2017. Therefore, based on the retrospective review of our institution’s database, 2930 patients who received heparinized arterial cannulas met the criteria from January 1, 2015, to December 31, 2016 (heparin group). To investigate whether NS arterial cannulas could improve outcomes, we selected 170 patients who underwent “simple” cardiac surgery and received postoperatively indwelling NS arterial cannulas since June 1, 2017 (NS group). These “simple” cardiac surgery cases included isolated coronary artery bypass grafts (CABGs), single-valve surgeries (SVS), and isolated CABG+ SVS.An arterial measurement transducer was acquired Edwards Lifesciences . The types of arterial cannula were 20 G, 22G and 24 G . The arterial cannulas were made by Becton Dickinson Medical Devices Co. Ltd. . The transducers were calibrated, zeroed and levelled at the beginning of each working shift and also when the measurements demonstrated any problems. The radial artery was used for catheterization in all of the patients. To maintain homogeneity between the groups, radial arterial cannulas were replaced in these patients they had femoral or brachial arterial cannulas. The data collected by radial arterial cannulas were used for this study. Inflating the pressure infuser bag to a constant 300 mmHg has been routinely used to prevent retrograde blood flow. The catheterization technique was the same between the NS and heparin groups. Furthermore, arterial cannulas could be an appropriate response to the flush test. In the heparin group, heparin  was added to 500 ml of normal saline . Finally, the heparin concentration was 2.5 U/ml in the heparin group. In the NS group, 500 ml of normal saline was used. In our hospital, blood samples were obtained from arterial cannulas for blood gas analysis. Therefore, flushing was routinely implemented after collecting blood samples. Additionally, flushing of the cannula, mobilization or both were performed if cannulas were potentially occluded.After six months of ICU training, bedside nurses undertook the assessments of the arterial lines. To assess the usefulness of arterial cannulas, three parameters were used. (i) Patency: The cannula was sufficiently patent when the blood was back-drawn using a syringe. Otherwise, flushing or movement of the cannula was used and duly noted. (ii) Reliability of arterial pressure (AP): AP using the intra-arterial pressure measurement, compared with brachial artery cuff pressure measurement, is regarded as appropriate when the values have differences of less than ±10 mmHg between the mean AP obtained via the cannula vs the cuff . (iii) AHoste EA et al. reported that the incidence of “heparin-induced coagulation disorders” was 23% . The folt-test, or the Mann-Whitney U nonparametric method was used for non-normally continuous variables. Continuous variables were determined to be normal in distribution by the Shapiro-Wilk test. Categorical data were equated using the chi-square test or Fisher’s exact test. The two groups were analysed using repeated measures analysis of variance (ANOVA). Differences between two groups were determined by repeat measures ANOVA with subsequent Bonferroni correction, with P < 0.05/n considered significant.IBM SPSS statistical software was used  for analysis. Continuous variables were portrayed as the mean ± SD or, if appropriate, as the median with interquartile ranges (IQR). Discrete variables were depicted as frequencies . Normally continuous variables were evaluated using Student’s P ≤ 0.1 in univariate analysis, as presented in Table P value < 0.05 was regarded as significant.Some bias could exist in our study. The conclusions would be not scientific and reasonable if these biases were not eliminated. Adjustment for indication bias was further assessed using a propensity score. With the help of this method, a comparison between patients receiving HS arterial cannulas and those who received NS arterial cannulas with similar risk profiles was made possible  and arterial cannula occlusion  showed no differences between the NS and heparin groups. Furthermore, flushing as well as cannulae size were not different between the NS and heparin groups. The detailed variables associated with the main variable in propensity-matched patients are presented in Table P < 0.05) and higher APTT  during the period from T0 to T48, compared with the NS group. After Bonferroni correction, heparin group had higher APTT during the period from T24 to T48 (P < 0.05/4), and had higher ACT at T48 (P < 0.05/5) compared with NS group. The heparin group had more allogeneic RBC utilization  and more drainage to chest tubes . In addition, more patients used RBC  than in the NS group.In propensity-matched patients, the duration of arterial cannulas  as a benchmark, heparin (200–400 U/kg) was used to achieve anticoagulation. At the end of CPB, protamine was initially administered to reverse heparin at a 1:1 ratio, so the activated clotting time was returned to preoperative values. Additional doses of protamine might be required if the ACT is not appropriate. In theory, we should remove previously applied heparin before applying new heparin to prevent the accumulation of medication that exceeds the prescribed dose. However, HS was routinely used to prevent arterial cannula occlusion. It can lead to unnecessary heparin exposure. Heparin, even at a low dose, can cause thrombocytopenia and bleeding , 23. UpoDel  et al. dSome drugs can affect coagulation function, and they were used for prophylaxis of deep venous thrombosis. In our centre, low molecular weight heparin (LMWH) was used to prevent deep venous thrombosis, which should be monitored for daily by ultrasonography to assure that the LMWH is successfully administered early. Moreover, early rehabilitation intervention was usually implemented as soon as possible. Further, we would encourage patients to walk if they had been weaned from mechanical ventilation. LMWH was only prophylactically used to prevent deep venous thrombosis in long-term bedridden patients. Table A systematic review  reportedOur study design involves one centre’s experiences with the inherent disadvantages of an observational study, which is highly prone to bias. This observational study could be influenced by potential biases. We used propensity score matching to avoid these biases. However, factors that affect assignment to treatment and outcomes but that cannot be observed cannot be accounted for in the matching procedure. Any hidden bias due to latent variables might remain after matching, which could lead to some statistical faults. Furthermore, with this analysis, we remove a large amount of patients from the analysis, but may have elevated statistical errors. The multiple statistical analyses was also undertaken in our study. The outstanding criticism is the inflated risk of Type I error because of multiple statistical analyses. It may have adverse effects on our study. Additionally, the reliability of the pressure measurement was tested against the brachial artery pressure measure by cuff and sphygmomanometer, which is questionable in the setting of cardiac surgery because patients often experience vasodilation as a result of systemic inflammatory response syndrome, and there could be a large difference in arterial blood pressure measured at the radial artery compared with more centrally, such as the brachial artery. The cuff arterial pressure measured on the arm is dependent on the ratio of the cuff size to arm size. These limitations of this assessment are important and should be acknowledges.Based on the results of our study, the addition of heparin to normal saline for flushing arterial pressure monitoring cannulae did not reduce the incidence of catheter thrombosis and result in a very small but statically significant in increase in ACT and APPT.


We performed a post-hoc analysis of the OLiVIA-study investigating whether current and ex-smoking asthmatics with small airways dysfunction (SAD) show a better response in airway hyperresponsiveness (AHR) to small particle adenosine after treatment with extrafine compared to non-extrafine particle inhaled corticosteroids (ICS), and to investigate which clinical parameters predict a favorable response to both treatments. We show that smoking and ex-smoking asthmatics with and without SAD have a similar treatment response with either extrafine or non-extrafine particle ICS. We also found that lower blood neutrophils are associated with a smaller ICS-response in smokers and ex-smokers with asthma, independent from the level of blood eosinophils.The online version of this article (10.1186/s12931-018-0961-2) contains supplementary material, which is available to authorized users. Asthma patients who smoke experience more severe symptoms as well as airflow limitation, and benefit less from treatment with inhaled corticosteroids (ICS) compared with non-smoking asthmatics . Additio1 (forced expiratory volume at 1 s) from baseline to post-treatment (ΔPD20). Small particle adenosine is a provocative agent that acts indirectly via the release of mediators from inflammatory cells. It was chosen rather than methacholine as it is a more sensitive measurement to detect improvement in AHR after treatment with ICS in patients with asthma  < lower limit of normal (LLN) from spirometry; ratio of residual volume to total lung capacity [RV/TLC] > upper limit of normal (ULN) from body plethysmography; difference between resistance at 5 Hz and 20 Hz [R5-R20] > 0.1 kPa sL− 1 from IOS [acin] and conductive airways [Scond] > ULN from MBNW [Since there is no clearly defined golden standard for SAD, we applied various parameters next to the PDrom MBNW . These m20 adenosine after treatment with extrafine compared to non-extrafine particle ICS in patients with SAD and patients without SAD. PD20 values were log2-transformed prior to analyses. To investigate the difference in ΔPD20 between patients with SAD and without SAD within one treatment group, we performed a student t-test or Mann-Whitney test. Next, we explored which baseline variables  were associated with ΔPD20 adenosine after extrafine and non-extrafine particle ICS treatment using univariate linear regression analysis. We finally performed multivariate linear regression analyses for each treatment type to identify independent clinical predictors for ΔPD20 including age, sex, smoking status, numbers of neutrophils and eosinophils, and completed with those variables showing a trend towards association with ΔPD20 (p < 0.1) in the univariate analysis. We allowed a maximum of 7 variables to be included in the multivariate model. The variables in this final model were selected based on the following criteria: (1) the variable changed regression coefficients (B) > 10%, (2) the model had highest total explained variance (R2), and (3) the model preferably included the highest number of subjects.We performed a two-sided paired t-test or Wilcoxon test to assess the difference in ΔPD1 83 (14.5) % predicted, and PD20 2.69 (3.41) mg. Baseline characteristics of the study population are presented in Additional file 20 adenosine to extrafine (QVAR) and non-extrafine particle ICS treatments (Clenil and Flixotide) , 42% being male, with mean (standard deviation (SD)) age 45 (12.6) years, FEV20 adenosine, independently from the level of blood eosinophils, in patients treated with QVAR, Clenil and Flixotide . Higher blood eosinophils were also associated with a larger increase in PD20 adenosine in patients treated with Flixotide (p < 0.05). Finally, younger age tended to be associated with a larger increase in PD20 adenosine after using Clenil (p = 0.05).Results of the univariate linear regression analyses are presented in Additional file 1 after 2-week ICS-therapy [We did not confirm our hypothesis that current and ex-smoking asthmatics with SAD had a better treatment response to QVAR compared to Clenil and Flixotide. In line with previous reports on better clinical effects of ICS in asthmatics with eosinophilia –11, we s20 adenosine between patients with and without SAD. With a standard deviation of one doubling dose, β = 0.8 and α = 0.05, we would need 16 patients per group while in our study, 18 patients with and 23 patients without SAD were investigated. Further studies with larger sample sizes would provide a clearer picture of SAD in smokers and ex-smokers with asthma. Another possible limitation is that we treated for two weeks which may have been too short to improve SAD.A strength of the study is that we defined SAD by cut-off levels applying multiple measurement techniques including spirometry, body plethysmography, impulse oscillometry and multiple breath nitrogen washout. A limitation is the relatively small sample size of patients with SAD and non-SAD which limits the power of the study and raises the risk of type I error. However, a post-hoc power analysis indicated that we had a sufficient number of patients to detect a difference of one doubling dose increase in PDIn conclusion, we show that smoking and ex-smoking asthmatics with and without SAD have a similar response to small particle adenosine after treatment with either extrafine or non-extrafine particle ICS. These findings suggest that clinicians may not need to consider SAD in order to decide whether current- and ex-smoking asthmatics would benefit preferentially from treatment with extrafine rather than non-extrafine particle ICS. Of importance, we find that lower blood neutrophils is a favorable predictor of ICS response, independent from the level of blood eosinophils.Additional file 1:Table S1. Baseline characteristics. Table S2. Baseline values of measurements of small airways dysfunction. Table S3. Difference in ΔPD20 adenosine in patients with and without small airways dysfunction after treatment with QVAR, Clenil and Flixotide. Table S4. Univariate linear regression analysis of baseline variables and change in airway hyperresponsiveness (ΔPD20) in patients treated with QVAR, Clenil and Flixotide. Figure S1. Scatterplots of significant correlations between baseline variables and ΔPD20 adenosine. (PDF 731 kb)


Moreover, baseline 24-h urinary metanephrines significantly correlated with wSD of both 24-h systolic and diastolic BP. Our study highlights as PPGL patients, after proper treatment, show a significant decrease in some short-term BPV markers, which might represent a further cardiovascular risk factor.Data on short-term blood pressure variability (BPV), which is a well-established cardiovascular prognostic tool, in pheochromocytoma and paraganglioma (PPGL) patients is still lack and conflicting. We retrospectively evaluated 23 PPGL patients referred to our unit from 2010 to 2019 to analyze 24 h ambulatory blood pressure monitoring (24-h ABPM)-derived markers of short-term BPV, before and after surgical treatment. PPGL diagnosis was assessed according to guidelines and confirmed by histologic examination. The 24-h ABPM-derived markers of short-term BPV included: circadian pressure rhythm; standard deviation (SD) and weighted SD (wSD) of 24-h, daytime, and night-time systolic and diastolic blood pressure (BP); average real variability (ARV) of 24-h, daytime, and night-time systolic and diastolic BP. 7 males and 16 females of 53 ± 18 years old were evaluated. After surgical resection of PPGL we found a significant decrease in 24-h systolic BP ARV (8.8 ± 1.6 vs. 7.6 ± 1.3 mmHg,  NF1, RET, VHL, SDHD, SDHB, etc.) and subjects with hereditary forms typically show multifocal diseases at younger age than those with sporadic neoplasms  for SBP and [(daytime DBP − night-time DBP)/daytime DBP × 100%] for DBP. Patients were classified as dippers if BP falls ≥10% and <20% of daytime average BP or non-dippers . Nocturnal BP falls ≥20% and <0% identified “extreme” dipper and “reverse” dipper subjects.Degree of nocturnal BP fall ((ii)Standard deviation (SD) of 24-h, daytime, and night-time SBP and DBP;(iii)Average of daytime and night-time SD, each weighted for the duration of the day and night periods [24-h “weighted” SD of BP (wSD)], which allows for removing the mathematical interference from night-time BP fall;(iv)N−1N denotes the number of valid BP measurements, and k is the order of measurements.ARV for 24-h SBP and DBP, i.e., the average of the absolute differences between consecutive BP measurements over 24-h, according to a mathematical algorithm: ARV = 1All data were acquired and analyzed according to the current recommendations . DevicesTo prevent perioperative cardiovascular complications, during 2–3 weeks before surgery, patients have been treated with α-adrenergic receptor blockers . Preoperative co-administration of β-adrenergic receptor blockers was performed in patients with uncontrolled BP values and/or tachycardia , only after administration of α-adrenergic receptor blockers [®, SPSS® Statistics, Italy) and GraphPad Prism software . Significance was set at p < 0.05.Data were expressed as mean and standard deviation (SD). Before statistical analysis, variables that showed a non-Gaussian distribution at Kolmogorov–Smirnov test were transformed to achieve a normal distribution and they were analyzed by non-parametric tests. Categorical variables were compared with Fisher and chi-square tests. Continuous and categorical variables were compared at baseline and follow-up by the Mann–Whitney test. ANOVA with Fisher’s Least Significant Difference post-hoc tests was used when needed. Relationships between continuous variables were assessed calculating the Pearson’s correlation coefficient. Linear regression models and multivariate analysis were performed to determine the combined effect of several variables on BPV markers. Statistical analysis was performed using SPSS software  successful surgical treatment in patients affected by PPGL is associated with decreasing of two of the most accurate indexes of short-term blood pressure variability, (ii) there is a significant relationship between the weighted standard deviation of 24-h systolic and diastolic BP and 24-h urinary metanephrines. Therefore, it is supposable that the excessive and pulsatile production of plasma catecholamines and their metabolites in PPGLs has a role in the increased BPV markers, which is involved in the increasing of cardiovascular risk. The clinical relevance of our findings relies on the fact that these patients might take particular benefit from surgical removal of the catecholamine-secreting tumor. Moreover, short-term blood pressure variability may be used as a screening tool in the work-up of PPGLs, especially in those cases with high suspicion for which the diagnosis is challenging. Further prospective studies with a larger number of patients free from cardiovascular events and risk factors are necessary to confirm our findings.


Higher CoV of SVdiff and CIdiff were significantly correlated with longer CV event-free survival, and the area under the receiver operating characteristic (ROC) curve showed fair overall discriminative power . The responses of hemodynamic control mechanisms can be independent predictive indexes for lower hospitalized CV events, which implies that these patients who have better autonomic control systems may have better CV outcomes.Dialysis-induced hemodynamic instability has been associated with increased risk of cardiovascular (CV) mortality. However, the control mechanisms beneath the dynamic BP changes and cardiac function during hemodialysis and subsequent CV events are not known. We hypothesize that the impaired hemodynamic control can be prognostic indicators for subsequent CV events in end stage renal diseaes (ESRD) patients. To explore the association of hemodynamic parameters and CV events in hemodialysis patients, we enrolled ESRD patients who received chronic hemodialysis without documented atherosclerotic cardiovascular disease and hemodynamic parameters were continuously obtained from the impedance cardiography during hemodialysis. A total of 35 patients were enrolled. 16 patients developed hospitalized CV events. The statistical properties [coefficient of variance ] of hourly beat-to-beat dynamics of hemodynamic parameters were calculated. The CoV of stroke volume (SV) and cardiac index (CI) between the 1 More than half of the CV events are the result of fatal arrhythmia and congestive heart failure, and some are the result of myocardial infarction3. In addition to the already identified CV risks including hypertension, hyperlipidemia, diabetes4 and electrolytes imbalance5, the intradialytic hypertension/hypotension or autonomic instability were thought to worsen their CV outcome in dialysis patients. Dialysis-induced hemodynamic instability was one of the most common complications, and those patients with unstable hemodynamics during hemodialysis were associated with worse outcomes6. A large retrospective cohort showed that the modest decline of BP between initiation and the end of hemodialysis was accompanied by the most favorable outcomes7. The relationship between the pre- and post-hemodialysis BP changes and all-cause mortality in the end-stage renal disease (ESRD) patients was described as “U- or J-shaped associations with lowest risk around −20 mm-Hg between post- and pre-dialysis BP in two observational studies ”9. Furthermore, the greater fluctuation of systolic BP (SBP) measured at 30-min intervals during dialysis was shown to be associated with higher risk of all-cause mortality and CV mortality in these patients10.The risk of cardiovascular (CV) mortality in dialysis patients is approximately 9 times higher than that of the general population12. The relatively stable BP in a constantly changing environment is the physiologic response of continuously fine-tuning the hemodynamic variables including cardiac output [(stroke volume (SV) * heart rate (HR)] and systemic vascular resistance (SVR) by the underlying control mechanisms. Increased beat-to-beat BP variability is not only a sign of impaired control systems but also a risk factor for CV events in hypertensive patients14. In addition, evidence has shown that the dynamics of beat-to-beat SV or HR can serve as earlier precursors to fluid responsiveness for several critical conditions17 before the actual change of BP. The temporal changes of hemodynamic variables in patients undergoing dialysis can be regarded as how the control systems respond19 while being exposed to continuous fluid shifts and osmolarity changes. However, few studies focused on continuously monitoring the hemodynamic variables other than BP during hemodialysis, and the relationship between the alteration of cardiovascular systems during hemodialysis and CV events is yet to be reported. We hypothesize that the impaired hemodynamic control can be prognostic indicators for subsequent CV events in ESRD patients and the dynamics of the intradialytic hemodynamic parameters derived from impedance cardiography were quantified to explore the association of hemodynamic parameters and CV events in hemodialysis patients.The BP homeostasis is one of the most sophisticated control mechanisms that incorporates several systems interacting with each other continuouslyA total of 35 patients were enrolled, and the flow chart of this study is shown in Fig. .For patients with or without CV outcome, the hourly mean SV gradually dropped over the course of dialysis , while the hourly mean HR, CI , and SVRI  were not significantly changed. In addition, the direction changes of hourly CoV of SV  and CI  in the early course of hemodialysis were significantly different between the two groups , 66 = 4.diff  and CIdiff  were independent predictors of better prognosis in these patients (Table 3). In addition, history of diabetes mellitus was a prognostic indicator. The optimal cut point of CoV of SVdiff and CIdiff were −0.009 and −0.006 (40th and 60th percentile of the corresponding variables), which for the most part predicted the differences in survival curves. Figure diff and CIdiff with the dichotomized threshold and higher CoV of SVdiff and CIdiff correlated with longer event-free survival . In ROC analysis, the AUCs of the CoV of SVdiff (0.783) and CIdiff (0.796) showed fair overall discriminative power in predicting patents with poor CV outcome. Added-on diabetes with CoV of SVdiff and CIdiff increased little discriminative power with an AUC of 0.822 .In multivariate analysis, higher CoV of SV14, and patients with poor cardiac function20. However, the clinical meaningful events of hemodynamic instability were presented by intradialytic or pre/post-dialytic BP changes, and we often ignored the underlying dynamics of the control mechanisms. In this study, we applied the dynamics of several hemodynamic parameters on a beat-by-beat basis, including HR, SV, CI and SVRI recorded from an impedance cardiograph, to explore the underlying mechanisms. The main findings are that the enhanced regulation of SV and CI in the early course of hemodialysis is associated with better CV outcomes in ESRD patients.Decrease in blood volume during the course of hemodialysis inevitably elicited compensatory mechanisms responding to a hemodynamic challenge. This hemodynamic instability was often a manifestation of the deteriorated control system particularly in the elderly21 but also with cardiac structural changes such as left ventricular hypertrophy in chronic hemodialysis patients22. This implies that the declining homeostatic control mechanisms may be associated with poor prognosis. The impairment of homeostatic control mechanisms can contribute to larger differences in pre-to-post dialytic BP9 and the increased intra-dialytic BP measured during hemodialysis10. However, the mechanisms of increased BP variability are not well known. The compensated mechanisms in response to rapid fluid shifting during hemodialysis included increased CI or increased SVRI19. In this study, the patients presented gradual reduction of hourly SV during dialysis while maintaining relatively stable HR, CI, and SVRI. This may indicate that the average values of the hemodynamic parameters can be insensitive to the variability or dynamics generated from the physiological systems as suggested by several studies on HR and BP variability24.Longitudinal studies found that greater BP variability at pre-dialysis was not only associated with increased risk of all-cause mortality25. CoV is a normalization function that estimates the ratio of SD to its mean. Since a higher CoV represents a greater percentage of variability irrespective of the different mean values, it can avoid the potential issue related to the larger mean values with greater SD in physiological signals. That is, increased BP variability can result from elevated BP, but lower CoV reflects declining control mechanisms such as decreased baroreflex sensitivity26 and autonomic system dysfunction27. In the early course of hemodialysis, the patients with increased hourly CoV of SV and CI between the 1st and 2nd hour were associated with better CV outcomes, while hourly SD of the hemodynamic parameters showed no associations. In addition, the patients with unstable BP had lower CoV of SVRI and CI during hemodialysis compared to those with stable BP28. Properly enhancing the regulatory responses during hemodialysis can be a sign of better underlying control systems. A study that enrolled 109 patients showed worsening of left ventricular diastolic function early during hemodialysis session29. The mean early diastolic velocity (e’) change at 60 minutes of dialysis was not correlated with volume indexes. It may underlie the non-volume related mechanisms involved during early hours of hemodialysis. This may explain why the CoV change in SV and CI were only found in the early course of hemodialysis.In hypertensive patients, the variability of diastolic BP continuously recorded by arterial line quantified at 30-minute intervals was increased significantly in hypertensive patients compared to normotensive subjects, but the CoV of continuous systolic and diastolic BP over the long term (24 hours) or short term (30 minutes) significantly decreased in hypertensive patientset al.30 defined the stroke volume variation (SVV) recorded by impedance cardiography between the maximal and minimal SV over 10 consecutive beats to assess the SV dynamics during dialysis. The study showed that higher SVV is an independent predictor of unstable BP . Numerous studies in the non-dialysis population have shown that higher respiratory-related SVV during surgery16 or hypovolemia31 can be an indicator of inadequate blood volume. The discrepancies between hourly CoV of SV and SVV may arise from the SVV quantifying the mechanisms related to certain time scales such as respiration-driven SV fluctuations and averaging them over the course of dialysis. The hourly CoV of SV could contain additional mechanisms and their hourly changes together. Most patients developed intra-dialytic hypotension due to blood volume decrease without timely refills. A study showed that patients with post-dialysis hypotension were related to an altered response in peripheral resistance but not a change in cardiac performance32. This may explain why higher SVV can predict unstable BP. However, Titapiccolo et al.24 reported that renal failure patients with peripheral vascular disease had reduced cardiac baroreflex and reduced sympathetic activity. Several studies also proposed BP variability might be due to less control of peripheral vascular resistance by reduced cardiac baroreflex during volume depletion26. Moreover, insulin-dependent diabetes itself is an independent risk factor for CV events, which could be partly attributed to the autonomic neuropathy that consequently cause higher risk of arrhythmia, QT prolongation33, and impaired BP control mechanisms34. This warrants further studies on exploring the temporal changes of the hemodynamic parameters during dialysis to further clarify the linkage of specific underlying mechanisms to the dynamical patterns. Nevertheless, we demonstrate that the dynamics of the control mechanisms during the early course of dialysis is crucial. Poor control of the hemodynamic variables is a risk factor for CV events.More recently, Yoshihara F 35. Further validation cohorts with a larger sample size or increased hemodialysis session observations are needed to overcome this and ensure the findings.Several limitations are present in this study. First, our results only demonstrated the association. The causal effect of BP, hemodynamic parameters on the CV outcomes may only be established by randomized controlled trials. Second, this non-invasive device was not correlated with other fluid measurement devices, such as a bioimpedance spectroscopy device, or cardiac sonography as a validation. The volume status is pre-load, which is also an important parameter influencing the SV. Fluid management depends on the physical examination of edema status and absence of intra-dialytic hypotension. This volume status and ultrafiltration rate may affect the heart function as well. Third, our study was a single center design, and the sample size is small. Monitoring only one hemodialysis session may be confounded by the variability in volume and condition, although we chose the mid-week session to avoid this. In addition, a vast number of covariates were corrected in the model, though it may induce a type I error. We also did not record the changes of dialysate temperature, though whether it affect the MACE or all-cause mortality may need more evidencesIn conclusion, change in SV and CI in the early period of dialysis may have predictive values for CV outcomes in hemodialysis patients, which implies that patients with better autonomic control systems may have better CV outcomes.The observation, prospective cohort was performed from May 2015 to May 2018. Patients were recruited from the dialysis center of Taipei Medical University Hospital. The inclusion criteria were patients with ESRD receiving regular hemodialysis three times weekly for more than 3 months. The exclusion criteria were patients who were suffering acute illness or in hospital with documentation of atherosclerosis cardiovascular disease, aged older than 80 or younger than 20. This study was approved by the Institutional Review Board of Taipei Medical University (N201404050) and written informed consent was obtained from each participant before enrollment in this study. Furthermore, all methods were performed in accordance with the approved guidelines.® machine , Osypka Medical, Inc., CA, USA). The definition of HR is the rate at which the heart beats; SV is the volume of blood pumped from the ventricle of the heart in one beat; CI is the measure of cardiac output per square meter of body surface area36. This non-invasive method was described for hemodynamic monitoring37. Analysis was performed on an hourly basis, and several statistical properties of those hemodynamic parameters such as the mean value, standard deviation (SD), and coefficient of variance (CoV) were carried out to explore the dynamical changes of these parameters during dialysis. The measurement of “CoV” was calculated as the ratio of SD to the mean, which represents the degree of variability in proportion to its mean, and a higher CoV indicates better control of the homeostatic system25. The trend of these hemodynamic parameters were described and the difference of coefficient variance between hourly-based changes of SV and CI were studied (SVdiff and CIdiff).Patients’ clinical parameters, co-medications, and biochemistry results were collected. Risks for cardiovascular disease were recorded, including smoking, body mass index, hypertension, diabetes mellitus and hyperlipidemia. In addition, the hemodialysis parameters including blood flow, dialysate calcium concentration, dialysis duration and ultrafiltration rate were recorded. After enrollment, we chose the mid-week dialysis session as the observation period. During hemodialysis, BP was measured with an electric sphygmomanometer in the supine position at 10-min intervals starting from 10 minutes before initiation of hemodialysis, and it was ended 10 minutes after the end of hemodialysis. Systemic vascular resistance indexes (SVRI) were calculated as the ratio of mean arterial pressure to cardiac index  to normalize the differences in patients’ size. Continuous impedance cardiography was recorded and the beat-to-beat hemodynamic parameters including HR, SV, CI were derived from the ICONWe prospectively followed these patients until the occurrence of CV events or the end of study in May, 2017. The prespecified CV events were defined according to the primary diagnosis of the discharge note. The CV events included the composition of major adverse cardiovascular events (MACE)  or hospitalization for a cardiovascular-related illness, including heart failure, symptom-driven revascularizations, and acute limb ischemia.2 or Fisher’s exact test. Hourly based parameters were compared by a 2 × 3 two-way ANOVA with repeated measures to examine the time course of hemodialysis and the outcome on the changes in hemodynamic parameters. The multivariate Cox model was performed, adjusted for age, gender, BMI, underlying diseases , smoking, and electrolyte  to evaluate the effects of the hemodynamic parameters on outcome. Furthermore, the optimal cut point of each selected variable was determined by the maximal hazards ratio of the dichotomized threshold calculated from the values between the 25th to 75th percentile with a 5 percentile step. The Kaplan-Meier survival analysis and log-rank analysis were performed to test whether the event-free probabilities over time of the stratified groups were significantly different. The receiver operating characteristic (ROC) curve of the selected variables were constructed, and generalized linear model with binary regression were applied to combine multiple variables. The areas under the ROC curve (AUC) were calculated to evaluate the overall predictive power. All statistical analyses were performed by using R software, version 3.5.0. and IBM SPSS Statistics for Windows Version 22 . A p value of <0.05 was considered statistically significant.Baseline demographic characteristics are represented as the mean ± SD for continuous variables, and as proportions for categorical variables. The between-group comparisons of categorical variables were calculated by χThis study was approved by the Institutional Review Board of Taipei Medical University .Written informed consent was obtained from all patients.