EEOT) and aortic flow (∆AoFEEOT) during the EEOT. PPV and SVV were obtained by the variation of beat-to-beat PP and SV, respectively. Baseline PPV, SVV, ∆SVEEOT, and ∆AoFEEOT increased significantly after BW, with a decrease of aortic flow (P < 0.05). PHE induced a significant decrease of PPV and SVV, but without affecting ∆SVEEOT, and ∆AoFEEOT. We conclude that ∆SV and ∆AoF during EEOT kept the ability to predict fluid responsiveness during PHE infusion in a rabbit hemorrhage model. This result may suggest the advantage of EEOT with respect to SVV and PPV in predicting fluid responsiveness during vasomotor tone increase.End-expiratory occlusion test (EEOT) has been proposed as a preload responsiveness test that overcomes several limitations of pulse pressure (PPV) and stroke volume (SVV) variations. We compared the ability of EEOT versus SVV and PPV to predict fluid responsiveness during the increase of the vasomotor tone in a rabbit model of hemorrhage. Ten rabbits were anesthetized, paralyzed, and mechanically ventilated during basal load (BL), after progressive blood withdrawal (BW), and after volume replacement. Other two sets of data were obtained during vasomotor increase by phenylephrine (PHE) infusion in BL and BW. We estimated the change of stroke volume (∆SV Several indices are available at the bedside to predict fluid responsiveness. Dynamic parameters based on heart-lung interactions have been proposed to predict fluid responsiveness and to promote more rational fluid administration2. Nevertheless, respiratory variations of stroke volume (SVV), pulse pressure (PPV), and vena cava diameter have numerous limitations that reduce their accuracy and applicability in clinical scenarios3. By interrupting cyclic ventilation at end-expiration for a few seconds , abolishing the inspiratory increase in intrathoracic pressure, and by passive leg raising tests, we can produce a self-volume challenge and increase cardiac output (CO) by a rapid and reversible increase of venous return (VR) in those patients preload-responsive5. Both tests have shown an excellent predictive value keeping their accuracy in patients with cardiac arrhythmias, with spontaneous breathing activity, with low compliance of the respiratory system and independently of positive end-expiratory pressure in acute respiratory distress syndrome patients7. However, passive leg raising presents some false negatives in patients with intra-abdominal pressure ≥16 mmHg, is contra-indicated to perform in patients with head trauma and intracranial hypertension and may be influenced by the use of compression stocking9. Further, the EEO technique does not have the technical constraints of passive leg raising, such as the measurements of CO by a fast and direct technique10.The main goal of fluid therapy is to correct hypovolemia avoiding both under and over-resuscitation, as well as the excessive use of inotropic and vasopressor drugs. Moreover, the comorbidities and type of disease determine that not all patients are volume responsive. The preload responsiveness should be assessed to decide both to administer fluid in the early resuscitation phase of shock states and to stop fluid administration in the de-escalation phase of shock management11. Comparing the CO during and before the expiratory hold could be advantageous for the EEO test over SVV and PPV, maintaining the ability to predict fluid responsiveness beyond the vasomotor tone. The study aimed to examine the ability of the EEO test to predict fluid responsiveness under the increased vasomotor tone in a rabbit model of hemorrhage and to compare it with SVV and PPV.In an animal model of hemorrhage, we have shown that the infusion of phenylephrine (PHE) (a pure α1-receptor agonist) blunts the dynamic preload indices increase after bleeding. This effect was mainly due to an increase in vasomotor toneTable /kg) induced a decrease of central venous pressure, inferior vena cava (IVC) flow, stroke volume (SV), and aortic flow, although only in SV and aortic flow reached statistical significance. Meanwhile, systemic arterial pressure (mean and pulse pressure) did not show a significant change.Hemorrhage . During basal load , PHE increased central venous pressure (P < 0.05) with a concomitant decrease of aortic flow and IVC flow. After blood withdrawal , PHE kept low values of the aortic flow, SV, and central venous pressure (P < 0.05) . However, after hemorrhage, PHE infusion decreased SVV and PPV , without affecting ∆SVEEOT and ∆AoFEEOT  decreases first, and it has been related both to an increase in central venous pressure and to the compression of the vena cava due to the increase in pleural pressure. Concomitantly, right ventricular afterload increases secondary to the increase in transpulmonary pressure that opposes right ventricular ejection16. Both mechanisms result in a decrease in right ventricular SV and pulmonary artery blood flow that finally leads to a reduction in left ventricular filling and output, which is expressed during the expiratory period, a few heartbeats later17. Other two mechanisms that are expressed during the inspiration phase include the left ventricular preload increase due to squeezing the blood out of the capillaries toward the left ventricle  and the left ventricular afterload decrease due to the reduction of the systolic cardiac transmural pressure18. The EEO test consists of comparing the SV or CO during and before the expiratory hold. The interruption of the mechanical ventilation at end-expiration stops the cyclic increase of right ventricular impedance and the impediment in VR, which determines an increase in right ventricular SV that is observed at a systemic level  in the following heartbeats. We cannot discard a concomitant left ventricle preload increase secondary to the maintenance of pulmonary capillaries opened inside the deflated alveoli by the decreased transpulmonary pressure during the end-expiratory hold5.We must recognize that the mechanisms involved in the variation of the SV during mechanical ventilation and the EEO test are not the same. Mechanical ventilation induces cyclic changes in blood flow in vena cava, pulmonary artery, and aorta successivelyEEOT and ∆AoFEEOT12.In hypovolemic conditions, the extent of the respiratory variation in SV is significant, mainly due to the expiratory decrease in left ventricular output that follows the higher inspiratory decrease in right ventricular output (often assessed as the “delta-down” of the systolic blood pressure) what determines an increase of both SVV  and the ∆SVEEOT and ∆AoFEEOT high in BW condition. Although BW and PHE decrease the VR, its absolute change was preserved, allowing to keep the predictive value of the EEO test.During the infusion of PHE, the rise of vasomotor tone would eliminate the expiratory component of the SVV or PPV (secondary to the inspiratory decrease of the right ventricular SV) and, it could attenuate the inspiratory increase of left ventricular VR (by hindering left ventricular filling due to the rise of ventricular diastolic pressure). However, it would not prevent the increase of the right ventricular SV secondary to the absence of the rise of transpulmonary pressure corresponding to the inspiratory period, keeping ∆SV18. Hence, one drug with even affecting only one type of adrenoreceptors can induce hemodynamic changes hard to predict. α1-adrenoreceptor agonists (such as PHE) constrict the arteries in systemic circulation, leading to a decrease in blood flow through the pre-capillary vessels and thereby decreasing the VR, and constrict the arteries of the splanchnic vasculature reducing the splanchnic arterial flow (leading to veins elastic recoil by a passive mechanism) and venoconstriction of vena cava and peripheral large and medium-sized veins (active mechanism) that work in concert to shift splanchnic blood volume to the systemic circulation increasing VR20. Approximately two-thirds of the increase in blood pressure during PHE administration resulted from an increase in VR with only one-third resulting from an increase in arterial tone. However, the concomitant increases in intrahepatic venous resistance can impede this blood volume shift, leading to blood sequestration within the splanchnic vascular bed and relative systemic hypovolemia18. We showed that PHE decreased both aortic and IVC flows, increasing mean and pulse aortic pressure  in BL condition. After BW, the decrease of aortic and IVC flows could not be modified by PHE. So, we can assume that the main effects of PHE administration in our model and the dose used are the increase of vasomotor tone ruling out any increase of VR, probably by a prevailing constriction of hepatic veins and vasculature within the liver.The effects of vasoactive drugs on hemodynamics and especially on VR/CO interactions depend on many factors, including the plasma concentrations of the drugs, the relative density of α1, α2, β1, β2 adrenoreceptors and affinity for the receptor subtype21. The difference between MCFP and CVP is the pressure gradient for VR and is related to Rv by the following formula:At steady state, VR (IVC flow) and CO (aortic flow) are equal. According to Guyton, VR is defined by three variables: the mean circulatory filling pressure (MCFP), the right atrial pressure, which is clinically measured as the central venous pressure (CVP), and the resistance of venous return (Rv)22.MCFP is the pressure measured throughout the circulatory system when there is no blood motion and depends on the venous compliance and the intravascular stressed volume , which in turn depends on the total intravascular volume. Rv is a complex component that depends on the different parts of the venous circulation24.Moderate hemorrhage (BW) determined a decrease of IVC flow with a parallel leftward shift of the venous return curve by lowering total and stressed volume without a change in unstressed volume resulting in a decrease in MCFP without significantly changing the Rv Fig. 18,23,24.18. In an experimental hemorrhage model, Cannesson et al. observed that in anaesthetized hypovolemic pigs, PHE boluses (from 0.5 to 4 mcg/kg) induces a dose-related increase in cardiac output secondary to an increase of IVC flow mediated by an unloading of the splanchnic reservoir and vice-versa, when it is preload independent, PHE boluses induce a decrease in CO associated with a decrease of IVC flow25. However, in a rabbit hemorrhage model, we have shown that PHE infusion (15 mcg/kg/min) induced a decrease in aortic flow with a significant SV decrease after BW compared with normovolemia. The different hemodynamic response to PHE may be probably related to the different doses of the α-adrenergic agonist 20. We propose that PHE infusion had a predominant effect on the Rv (decreased slope of the VR curve without a change of MCFP), which tends to decrease VR. However, the concomitant increase in the relative proportion of stressed volume to unstressed volume, tend to offset some of the decrease in VR (shifting the VR intercept with the abscissa (MCFP) to the right). Finally, PHE infusion increases the left ventricular afterload, shifting the ventricular function curve downward  at the time of administration18.The current study has several limitations. Pulmonary hypertension and right ventricular dysfunction can mask hypovolemia and normalize the values of dynamic preload indicesEEOT and ∆AoFEEOT keep the ability to predict fluid responsiveness during PHE infusion in a rabbit model of hemorrhage. This result may suggest the advantage of the EEO test in predicting fluid responsiveness during hypovolemia and vasopressor drug administration.We show that ∆SVThis study was approved by the Ethics Committee on the Use of Animals (CEUA), Facultad de Medicina, Universidad de la República, Montevideo-Uruguay in October 2013 . We strictly complied with the Guide for the Care and Use of Laboratory Animals , prepared by the National Academy of Sciences’ Institute for Laboratory Animal Research.28. The animals were tracheotomized and mechanically ventilated  via an endotracheal tube (ID 2.5 mm). The ventilator was set in the volume-controlled ventilation mode with a tidal volume of 8 mL/kg, end-expiratory pressure of 5 cmH2O, a respiratory rate 38 ± 6 breaths/min, and inspired fractional oxygen of 60%. Peripheral oxygen saturation (Rainbow 7 LNOP newborn sensor) was monitored and maintained above 96% . The respiratory rate was adjusted to maintain an end-tidal CO2 tension between 30–38 mmHg . The arterial partial pressure of oxygen and carbon dioxide were monitored by serial blood gas evaluation . The heart rate/respiratory rate ratio was >3.6. Anesthesia was maintained with a continuous infusion of midazolam (0.5–1 mg/kg/h) and rocuronium bromide (0.6 mg/kg/h). We used a heating pad to keep the animal normothermia.Ten female New Zealand rabbits (body weight 3.1 ± 0.6 kg) were premedicated with acepromazine (0.3 mg/kg i.m.) and meperidine (10 mg/kg i.m.). We placed a fluid-filled catheter (24-gauge) through an ear vein to induced anesthesia (midazolam 0.5 mg/kg i.v.) and to administrate saline solution (7 mL/kg/h) as maintenance requirementsWe placed a 4.5 F triple-lumen central venous catheter (Paediatric Multicath 3-Vygon) in the left jugular vein for measuring central venous pressure, blood withdrawal, and drug infusion. After making a right lumbar incision (extra-peritoneum approach), two non-constricting ultrasonic perivascular flow probe  were positioned one around the infra-diaphragmatic aorta and the other around the inferior vena cava. Both sensors were connected to a Doppler flowmeter  to measure instantaneous aortic and IVC flow, respectively. We monitored systemic arterial blood pressure placing a fluid-filled catheter (20-gauge) through the right femoral artery up to the infra-diaphragmatic aorta, just distal to the aorta flow probe. All pressure transducers (P23Db Gould Statham) were zeroed to atmospheric pressure at the midthoracic level.11.Once the instrumentation was completed, the animals were allowed to stabilize for 30 min, and all the variables were obtained during basal load (BL). After BL measurement, a second set of hemodynamic data were obtained after increasing the vasomotor tone by PHE infusion  during 30 min (BL + PHE). Twenty minutes after stopping PHE infusion, we removed blood progressively by stepwise cumulative volume of 5 mL/kg with a total of 15 mL/kg of body weight (15% of volemia) and a third set of hemodynamic measurements was obtained (BW). After thirty minutes of blood withdrawal, a fourth set of data was obtained during PHE administration (BW + PHE), and finally, a fifth set of data was obtained after volume replacement  with HAES-6% . We used the PHE to increase the vasomotor toneOnce we completed the experimental protocol, the animals were killed with intravenous potassium chloride under deep anesthesia.11.We monitored all signals in real-time and stored digitally at the end of expiration . Hemodynamic values were averaged over a 5-sec period. SV was estimated offline from the integral of the systolic portion of the aortic flow curve for each cardiac cycle5. We calculated the change of SV (∆SVEEOT), and aortic flow (∆AoFEEOT) from the value before the expiratory hold to the value corresponding with the maximal aortic flow reached during the EEO. The hemodynamic changes were expressed as percent changes from the value before the expiratory hold.We performed one EEO test in each experimental condition by interrupting the ventilator at end-expiration over 10–15 sec Fig. 5. We calmax − Qmin)/[(Qmax + Qmin)/2], where Qmax and Qmin are the maximum and minimum values for SV and PP during the same respiratory cycle, respectively17. We averaged the measurements of three consecutive respiratory cycles for statistical analysis.SVV and PPV were assessed using the standard formulae (100 × (Qdyn = PPV/SVV)29.The vasomotor tone was assessed by the dynamic arterial elastance (EaEEOT and ∆AoFEEOT (standardized difference of 1.25: target difference/standard deviation) for a level of significance of 0.05 and a power of 80%30. Normal distribution was tested with the Shapiro-Wilk test. Data are expressed as mean ± SD, except other thing was stated. SVV, PPV, and EEO test were analyzed at the different experimental conditions using a one-way analysis of variance (ANOVA). Post-hoc testing was performed using the Bonferroni test. Paired Student’s t-test was used to compare the different values of Eadyn under PHE infusion (BL + PHE and BW + PHE) versus without PHE infusion . All statistical tests were performed using SPSS software . P-value < 0.05 was considered statistically significant.Preliminary experiments were employed to calculate the sample size. A sample size of ten animals allows detecting a 20% effect in ∆SV


The current gold standard for the diagnosis of white-coat effect is by the 24-h ambulatory blood pressure monitoring (ABPM) which may not be readily available in every primary care setting. Previous studies had shown that deep breathing, through modulating the baroreceptor reflex sensitivity to vagal stimulation over 30 to 60 s, was useful in detection of the white-coat effect. The aim of our study was to evaluate the diagnostic accuracy of the deep breathing test (DBT) as compared with the gold standard of ABPM in the diagnosis of hypertension with white-coat effect in Chinese patients in primary care.This cross sectional study recruited 178 consecutive, eligible, consented, hypertensive patients receiving treatment at a local public primary care Hypertension Clinic.The diagnostic accuracy of the DBT in all recruited patients, patients not taking beta-adrenergic blockers and patients with different clinic SBP cut-off before the DBT by means of area under the receiver operating characteristic (ROC) curve, sensitivity, specificity, positive and negative predictive values was evaluated.p = 0.04). The corresponding sensitivity and specificity of the DBT were 40.9 and 90.9% respectively if SBP drop was > 30 mmHg.The results for the ROC curves for systolic and diastolic BP changes after the DBT were statistically insignificant. The ROC curve was statistically significant for SBP change in patients not taking beta-adrenergic blockers and with pre-DBT clinic SBP ≥ 165 mmHg (ROC curve area of 0.719, 95% CI 0.53 to 0.91, The DBT, even though could not be clinically applied to all patients, was proven to be a potential screening and diagnostic test for white-coat effect in Chinese hypertensive patients with a pre-test SBP of ≥165 mmHg and who were not taking beta-adrenergic blockers.This study was approved by Kowloon East Cluster/ Kowloon Central Cluster Research Ethics Committee/Institutional Review Board of Hospital Authority of Hong Kong under the registration KC/KE-16-0084/ER-3. Hypertension, being a major constituent to the global burden of non-communicable diseases , was repWhite-coat hypertension is defined as persistently raised office blood pressure in ambulatory normotensive patients . It accoThe detection of white-coat effect is important as failure to recognise the condition may lead to unnecessary treatment causing undesirable side effects for the patients and also inflating the cost of treatment –24. Currp < 0.001) among patients with or without white-coat effect respectively [p < 0.26) between the hypertensive patients with or without white-coat effect when using the first criterion but was able to identify a significant difference (p < 0.003) using the second criterion when applied to patients with office SBP < 160/100 mmHg [Deep breathing was shown to be able to lower blood pressure by increasing the baroreceptor reflex sensitivity to vagal stimulation , 28. Stuectively . In the ectively . The Jos100 mmHg . The app100 mmHg .p < 0.001) [Previous evidence also supported a positive correlation between office SBP and white-coat effect , 21. In < 0.001) . In anotThere was also evidence showing that beta-adrenergic blockers might potentiate the baroreflex, possibly through enhancing heart-rate variability and increasing the vagal tone while reducing the sympathetic beta-receptor stimulation and therefore might affect the DBT results , 34. HowThis was a cross sectional study conducted in a public primary care clinic serving more than 10,000 hypertensive patients in year 2015. When patient failed to reach target clinic blood pressure during consecutive follow up visits, they would be asked to bring their home blood pressure machine for validation along with their home blood pressure record. If there was a significant discrepancy between the clinic blood pressure and home blood pressure as measured by a validated home blood pressure machine, they would be referred to the Hypertension Clinic for further evaluation by family medicine specialists. Consecutive patients on anti-hypertensives, attending the Hypertension Clinic and fulfilled the inclusion criteria i.e. with clinic SBP ≥ 140 mmHg and/or DBP ≥ 90 mmHg at latest two clinic visits during the study period were included. Only those who aged 18 years or above and agreed to give consent in participating in the study were recruited until the required sample size was reached. Non-Chinese patients, patients with suspected secondary hypertension or atrial fibrillation and pregnant patients were excluded. The study was conducted from 1st August 2016 to 30th September 2017. This study was approved by Kowloon East Cluster/ Kowloon Central Cluster Research Ethics Committee/Institutional Review Board of Hospital Authority of Hong Kong.All clinic blood pressure readings were obtained in sitting position with the measuring cuff at heart level, using the calibrated manual sphygmomanometers  and appropriate cuff sizes. The patient, after resting for at least 5 min, would have his or her blood pressure measured at both arms. The second blood pressure, spaced 2 min apart would then be taken at the arm with higher measured blood pressure . The higSubsequently, a monitor  would be installed to perform the 24-h ABPM with an appropriate cuff placed on the non-dominant arm and programmed to measure blood pressure every 30 min while patients were awaked and 60 min while patients were asleep. The awake and sleeping time would be programmed individually based on the history given by patients. Patients with ABPM reports showing more than 80% successful readings would be included and their results would be interpreted by family medicine specialists who were blinded to the DBT results. There was no internationally standardized definition on the interpretation of white-coat effect by using the 24-h ABPM. In this study, it would be defined as a decrease of 10 mmHg or more in mean daytime ambulatory SBP when compared with the clinic SBP .Patients’ demographic data including age, gender, body mass index, smoking status, presence of diabetes and cardiovascular diseases and number and types of anti-hypertensives taken were retrieved from computerized record.Previous studies showed that the sensitivity of using 10 mmHg difference as cut-off to define white-coat effect after DBT was 0.8 . The preThe main outcome was to evaluate the diagnostic accuracy of the DBT for white-coat effect by means of area under the receiver operating characteristic (ROC) curve, sensitivity, specificity, positive and negative predictive values in Chinese hypertensive patients. Sub-analysis on the effect of beta-adrenergic blockers and different clinic SBP cut-off on the diagnostic accuracy of the DBT would also be evaluated. Blood pressure responses to DBT in all patients and different sub-groups with or without white-coat effect would be presented in terms of mean blood pressure drop after the test.p < 0.05. SPSS version 21 was used for statistical analysis.Central tendencies and distributions of continuous variables were presented as means and standard deviations respectively. The means of continuous variables were compared with independent samples t test. Categorical variables were presented as proportions and percentages. They were compared with Chi-square test (with Yate’s correction for 2 × 2 comparisons) or Fisher’s exact test. Areas under the receiver operating characteristic (ROC) curve, sensitivity, specificity, positive and negative predictive values were calculated to assess the diagnostic accuracy of the DBT, with respective 95% confidence interval. Differences were considered statistically significant when Five Hundred forty-eight patients attended the Hypertension Clinic during the study period and 209 patients fulfilled the inclusion criteria. 30 14.4%) patients refused to participate and 1 patient was excluded due to insufficient ABPM data resulting in 178 patients being recruited into the study. The mean age of the subjects was 65.0 and more were female patients (64%). More than half of the subjects (54.5%) were receiving monotherapy. Calcium channel blockers were the commonest (74.2%) class of anti-hypertensives being used by the patients, followed by angiotensin-converting enzyme inhibitors (27.5%), beta-adrenergic blockers (21.3%), angiotensin receptor blockers (14.6%), alpha-blockers (9.0%), diuretics (8.4%), hydralazine (7.9%) and methyldopa (5.1%). Other clinical characteristics of the subjects were summarized in Table 4.4% patip < 0.001) (Table p = 0.611) and 0.53  respectively, showing that they were uninformative  Table . 68.5% (p = 0.042). The ROC curve for both SBP and DBP change did not yield any significant results. However, the discriminating power of the DBT by using SBP change was improved with higher pre-test SBP cut-off, with an area under ROC curve of 0.68 (95% CI 0.51–0.84) and p value reaching 0.07 when patients with SBP < 165 mmHg excluded  and ≥ 165 mmHg (p = 0.035).  if the post-test SBP drop was more than 30 mmHg. The relatively high PPV and low NPV implied that DBT was most useful to rule in the existence of white-coat effect.Our study was one of the studies with larger sample size as compared to previous studies and probably the first to examine the deep breathing test’s applicability in Chinese patients. Furthermore, our study was the first to have taken into consideration the practicality of performing the DBT in actual clinical setting where the patients were more likely to be continued on their usual drug regime while performing the DBT. Neither the previous studies conducted by Federico et al. and Marion et al. took into account for the types of anti-hypertensives the patients were taking while they were performing the DBT, while in the Jose et al. study, the study subjects had anti-hypertensive withdrawn for 2–3 weeks before attempting the DBT, which might be less practical in a non-research setting.Our study added information that beta-adrenergic blockers and different clinic SBP cut-off would affect the DBT performance. Since higher office SBP was positively correlated with white-coat effect, we postulated that patients with higher pre-test SBP would exhibit significant difference in BP responses after DBT between the white-coat and non white-coat groups. Though the mean SBP drop between the two groups did not show increasing significance as we moved towards higher SBP cut-off, it reached statistical significance across both groups with SBP ≥145 mmHg and ≥ 165 mmHg and patients not on beta-adrenergic blockers. The findings also supported the evidence that beta-adrenergic blockers could modify the baroreflex and hence affect the DBT results.Previous studies showed conflicting evidence on whether the drop of SBP or DBP post-DBT could be used to evaluate the presence of white-coat effect –32. Our Our study showed a significant result after exclusion of patients on beta-adrenergic blockers and those with pre-test SBP < 165 mmHg. However, the sample size by such exclusivity became smaller and hence affected the statistical accuracy to find a precise SBP cut-off for distinguishing between the white-coat and non white-coat effect. Further research is suggested to specifically investigate this sub-group with a larger sample size. Moreover, the study was conducted in only one single clinic which was specialized on assessing hypertensive patients with suspected white-coat effect. The clinic had exceptionally more patients with white-coat effect diagnosed (68.5%) and the results might not be able to be generalised to all primary care clinic settings in Hong Kong.Throughout the study, there were several occasions where the post DBT blood pressures were higher than the pre-test values. As mentioned earlier, the device guided breathing cycle had its advantages, nonetheless, it also increased the anxiety in some of our patients during the attempt to synchronise their breathing rate to that of the counter on the mobile device. More training and attempts might be needed to allow the patients more adapt to the deep breathing exercise and produce a more pronounced blood pressure lowering effect. For those patients who showed a paradoxically increase in blood pressure after DBT, further evaluation with a standard ABPM should be performed.The DBT could not be clinically applied to all Chinese patients with suspected white-coat effect. However, the study proved that it would be a potential screening and diagnostic test for white-coat effect under certain selection criteria; i.e. in patients with a pre-test SBP of ≥165 mmHg and who were not taking beta-adrenergic blockers. Future larger scale studies should be conducted to enhance the statistical accuracy and generalisability of the results.


P=0.023) and comorbidities such as diabetes mellitus (DM) (P=0.013), chronic kidney disease (CKD) (P=0.007), and risk of developing obstructive sleep apnoea (OSA) (P=0.04) showed significant variation. Medication adherence to prescribed hypertensive medication was a major issue even though significant association was not found with high VVV (P=0.536). The SD of SBP was then classified into high and low VVV groups by means of a cutoff point at the 50th percentile. Bivariate analysis by using Chi-squared test revealed comorbidities such as DM, CKD, and physical activity (P=0.044) were significantly associated with high VVV. Further multivariate regression analysis revealed that comorbidities such as DM and CKD have 1.561 times and 5.999 times more risk to show high variability, respectively. In conclusion, we recommend simple practical measures to achieve sustainable BP control among hypertensive patients with DM and CKD to minimize the VVV and improve their cardiovascular outcome.Visit-to-visit variability (VVV) is a relatively new concept in the hypertensive arena. Data regarding VVV are lacking in our region, and factors associated with VVV are rarely examined in previous studies. This study was conducted among 406 patients attended to the cardiology outpatient department of Teaching Hospital, Jaffna, Sri Lanka, in 2018 to assess the long-term variability of blood pressure (BP) by reviewing last six consecutive BP readings from the records retrospectively. Data regarding sociodemographic variables and behavioural factors such as medication adherence, physical activity, smoking, alcohol consumption, and relevant comorbidities were taken through an interviewer-administered questionnaire. Data were analysed by using SPSS version 25 and VVV of systolic blood pressure (SBP) matrix expressed as mean of SD and association were examined with various factors and VVV of SBP. SBP showed high VVV among the participants as expressed by mean of SD which was 13.06 ± 5.64. When comparing mean SD among the categories of different variables, female sex ( Blood pressure fluctuation is a well-known phenomenon among patients with hypertension known as visit-to-visit variability (VVV). In the past, VVV of blood pressure often dismissed as random fluctuation and is thought to be a limitation of measuring blood pressure at clinic settings . One of n = 406) to assess the long-term variability by looking into their outpatient clinic records. Short-term variability in blood pressure is affected by behavioural, emotional, and postural influences on cardiovascular physiology and cardiac rhythm [VVV is an important independent prognostic marker for stroke, cardiovascular disease, and all-cause mortality –5. The mc rhythm . Arteriac rhythm , 9.Antihypertensive nonadherence would explain the adverse prognosis found in individuals with high VVV of blood pressure and may be a modifiable cause of VVV of blood pressure, and efforts to reduce nonadherence have the potential to decrease the VVV of blood pressure . AdditioAs described above in the past, VVV is thought to be a nuisance factor and considered as not related to future risk of development of cardiovascular diseases. But recent studies primarily from Europe found strong association between higher levels of variability of blood pressure and increased risks of coronary heart disease and stroke, and evidences also revealed that VVV is not a random phenomenon , 13–15. This cross-sectional study was conducted among 406 patients attended to the cardiology outpatient clinic of Teaching Hospital, Jaffna, Sri Lanka, in 2018. Patients with hypertension of more than one year duration who are 18 years of age and more were included in the study. Data were collected over the period of two months by using an interviewer-administered questionnaire. Last six consecutive BP recordings were retrieved after obtaining informed consent from participants. This study was evaluated and approved by the Ethics Review Committee, Faculty of Medicine, University of Jaffna, Sri Lanka.Data regarding BP were retrospectively retrieved from medical records. BP was measured in office-based settings with an interval of 8 weeks by medical officers attached to the cardiology department with the conventional cuff method using a mercury sphygmomanometer. In this study, we used the standard deviation (SD) of BP of six visits as the main metric for VVV of SBP regardless of their being many other metrics available ) as previous study found them to be equally significant. The SD of SBP was then classified into two groups by means of a cutoff point at median: values below the median were defined as low variability of SBP, and values equal to or greater than median are defined as high variability of SBP . We alsoPatient's clinic records were used to get information including year of diagnosis of hypertension and comorbidities such as DM, CKD, and hypertensive medication.Quantity of alcohol was calculated in units by using the validated questionnaire used in different studies in Sri Lanka. Physical activity level was calculated using International Physical Activity Questionnaire (IPAQ score). Both questionnaires were used in many studies of Sri Lanka, and validity was established already.Weight, height, neck circumference, and midarm circumference were done by standard weighing scale and nonstretchable plastic tape.The neck circumference was measured in the midway of the neck, between the midcervical spine and midanterior neck using a nonstretchable plastic tape with the subjects standing upright. The midarm circumference of the upper arm is measured at the point of half way between the olecranon process of the ulna and the acromion process of scapula using the nonstretchable plastic tape.Sleep apnoea risk was assessed using the STOP-Bang questionnaire. This questionnaire was translated in to Tamil and was retranslated to English to check that the meaning was not altered, and also consensus was obtained among three different physicians. Risk was categorised as low, medium, and high.Medication adherence was classified as low, medium, and high adherence based on a standard modified medication adherence questionnaire. This questionnaire also was translated in to Tamil and was retranslated to English to check that the meaning was not altered, and also consensus was obtained among three different physicians.Statistical Analysis. Statistical analysis was performed with IBM SPSS Statistics version 25 [β-coefficient (standard error) and adjusted exp β (odds ratio (OR), 95.0% confidence interval (CI)).rsion 25 . ParticiWe also examined comparison of the mean of SD of SBP between groups of different variables. Comparisons of the mean values between three or four groups were analysed by the one-way analysis of variance.Most of the participants were males (62.6%) and belong to 50–64 age group (82.3%). Majority studied up to secondary education (78.6%), and 71.2% of the participants were housewives or unemployed. Interestingly, most of them belong to poorest income group (63.5%) and 34.4% reported as middle income. Considerable amount of the participants (26.6% CI: 22.5–31.1) were noted to have insufficient (sedentary) physical activity level and 14.3% (CI: 11.1–17.9) were obese. As shown in When considering the associated comorbidities , majoritn = 34).When considering the antihypertensive medications among the participants , commonlP=0.023), DM (P=0.013), CKD (P=0.007), and risk of developing OSA (P=0.04) showed significant variation. No other variables showed significant variation. The average means of SBP and DBP were 127.4 ± 8.62 mm Hg and 78.40 ± 3.64 mm Hg, respectively and were obtained from 6 measurements. The mean average real variability (ARV) of SBP was 14.10 mm Hg ± 7.79. The range of ARV of SBP was 2–48 mm Hg. The average mean of SD was 13.06 ± 5.64, and we examined mean SD comparison among the categories of different variables and found that female sex  and CKD (P=0.019). Physical activity has become nonsignificant (P=0.098). After adjustment for covariates such as age, sex, economic status, education, and occupation, DM and CKD was significantly associated with high VVV of SBP  with high VVV while comparing means of SD with counterpart. No other sociodemographic variables showed significant variation. A previous study also found sodium intake was significantly higher in the high VVV group than in the low VVV group [Another study revealed being older, female gender, and certain comorbid conditions significantly raised VVV . This stVV group , but we P=0.013) and CKD (P=0.007) have showed significant variation when comparing mean standard deviation. Further analysis  by comparing low VVV and high variability found that having DM and CKD as comorbidities have 1.561 times and 5.999 times more risk to show high variability, respectively. This fact is well supported by the study carried out among type 2 diabetics which concluded that VVV in SBP could be a novel risk factor for development and progression of nephropathy [Several studies found that the VVV was associated with progression of vascular disease such as dementia , cognitihropathy . In the hropathy . It was hropathy . Same auhropathy . We beliP=0.04). But we were unable to establish a significant association between risks of developing OSA with high variability during further analysis, and the reason might be that the STOP-bang questionnaire was not validated to our own population. But a study on sleep apnoea and VVV concluded that OSA is associated with abnormal visit-to-visit variability, and sympathetic activation seems to be related in some way [We used STOP-Bang questionnaire to assess the risk of developing OSA, and 65.3% (60.5%–69.8%) found to have high risk to develop OSA. We also compared mean of SD of SBP among low-, medium-, and high-risk OSA groups  which shsome way . A screeP=0.044) showed significant association with high variability in bivariate analysis and multivariate analysis did not show significant association (P=0.098). But we could argue the role of physical activity in control of VVV of SBP and its importance to encourage patients to involve more physical activity.A study at a cardiac rehabilitation setting measured SBP and DBP before exercise training at each visit and determined VVV in BP expressed as the standard deviation of the average BP. Patients who had uncontrolled BP at baseline and who did not change their antihypertensive drugs throughout the study period showed a significant reduction of both SBP and DBP after 3 months. Patients who did not change their antihypertensive drugs were divided into high and low VVV in the SBP groups according to the average value of VVV in SBP or DBP and found that VVV in SBP and DBP in the 1st month was significantly decreased than the 3rd month in both groups . Our stuP=0.151) not showed significant difference with VVV of SBP.It is reasonable to predict class of antihypertensive drugs also may have impact in the long-term variability. A study evaluated whether the antihypertensive medication class differentially affected blood pressure VVV among hypertensive individuals in clinical real-world setting as well as the association between VVV and characteristics. It showed diuretics significantly lowered VVV and alpha/beta blockers resulted in highest VVV , 19. ButP=0.536) between the two groups. The reasons could be medication adherence is low regardless of different categories of VVV and also be due to the adherence questionnaire is not validated among our population. None of the participants of the study reported high adherence, and 22.7% (CI: 18.8–26.9) reported low adherence to medication. A previous study in the same setting also revealed poor medication adherence was prevalent due to several reasons [Similarly a study on medication adherence and VVV of systolic blood pressure in African Americans with chronic kidney disease (CKD) has found lower medication adherence is associated with higher systolic VVV of blood pressure in CKD patients. Efforts to improve medication adherence in this population may reduce systolic VVV of blood pressure , 31. We As ample evidence available for VVV and all-cause mortality , it coulThis study clearly demonstrates that VVV of BP is high among our population and high VVV independently associated with comorbidities DM and CKD. Hence, the clinicians should not consider the adequacy of blood pressure control at a single visit as it is of marginal clinical importance. VVV of BP is considerably high among the study population, and addressing this may reduce patients' cardiovascular morbidity and mortality in poor resource setting. A most stringent BP control is recommended in particular among hypertensive patients with comorbidities of DM and CKD. As medication adherence plays a key role in treating hypertensives, clinicians should adhere to simple prescription pattern to ensure compliance and consequently minimize VVV. Identifying and treating sleep apnoea would also have a great impact on VVV, and it is always desirable to conduct studies related to sleep apnoea in selected patients. Encouraging physical activity could be an important strategy to ameliorate VVV of SBP among hypertensives. In conclusion, VVV of BP is an important target for the treating physician to reduce the cardiovascular and renal outcome of diabetic patients with hypertension.


Asthma and chronic obstructive pulmonary disease (COPD) can present as unique conditions or as a combination known as asthma-chronic obstructive pulmonary disease overlap syndrome (ACOS). These condition(s) can be categorized as obstructive conditions, causing inflammation of small airways leading to decrease airflow, mucus production, and bronchoconstriction. Asthma and COPD affect every age, gender, ethnicity, and socioeconomic status, thus increasing mortality and morbidity burden in our society. Fractional exhaled nitric oxide (FeNO) is an endogenous gaseous molecule which can be measured in the human breath test because of airway inflammation. It has been studied extensively as a marker of inflammation and has been incorporated into an algorithm for asthma management. The purpose of this study was to investigate whether FeNO testing can lead to a change in the diagnosis. A retrospective chart review of 95 patients with asthma, COPD, and ACOS was performed, and FeNO levels were recorded. Out of 95 patients, 36%, 24%, and 22% of the patients had an initial diagnosis of asthma, COPD, and ACOS, respectively. After the FeNO testing, the number of patients with the final diagnosis of asthma and ACOS increased, and COPD decreased. Our results support the utility of FeNO as a viable marker in diagnosing and managing complex cases of asthma, COPD, and ACOS. Asthma and chronic obstructive pulmonary disease (COPD) are respiratory conditions which commonly present with airflow limitation; the patient often complains of experiencing shortness of breath. Asthma and COPD exacerbation present with increased airway inflammation, mucus production, and air trapping. Air trapping results from not being to exhale completely, leading to residual oxygen in the lungs. Data suggests that COPD is the third- and fourth-most cause of deaths in the United States and the world, respectively -3. Some Fractional exhaled nitric oxide (FeNO) is a gaseous molecule produced in response to an inflammatory process and may aid in differentiating asthma from other lung conditions . This paWe obtained records of all FeNO tests (160 patients) and results performed in the outpatient setting at our private practice affiliated with Jersey Shore University Medical Center (JSUMC). We focused on patients with the diagnosis of asthma and COPD who underwent FeNO testing between November 1, 2016, and February 28, 2017  FeNO levels, diagnosis before and after the test, presenting symptoms, smoking history, and past medical history were recorded into an electronic spreadsheet . Next, one of the authors calculated the mean and range of FeNO (ppb) levels along with identifying the change of diagnosis after the FeNO test.Our analysis focused on patients who underwent FeNO testing starting November 1, 2016, to February 28th, 2017. We categorized diagnosis into asthma, chronic obstructive pulmonary disease (COPD), and asthma-COPD overlap syndrome (ACOS). For this study, diagnosis of COPD included patients who had chronic bronchitis and emphysema; bronchiectasis was not included in the diagnosis of COPD.A total of 95 patients were included in this study Table . Before After FeNO testing, our cohort was characterized as Table : 45 asthIn our cohort, one patient diagnosis changed from asthma to ACOS, nine patients with COPD were changed to the diagnosis of ACOS, two patients changed from COPD to asthma, and two patients were changed to having no final diagnosis despite having an initial diagnosis Table .Fractional exhaled nitric oxide (FeNO) is a marker of endogenous inflammation which can be used to monitor inflammatory changes in the airway. Asthma and COPD usually consist of airway inflammation and hyperresponsiveness. FeNO levels correlate with the presence of inflammation in the lungs; however, the precise role in diagnosis is not well defined. Despite lacking robust association, FeNO levels in conjunction with other laboratory markers and clinical signs aid in reaching a diagnosis .In our cohort, 36% of the patients were initially diagnosed as asthmatics. However, this percentage increased to 47% after FeNO testing. Conversely, the number of patients initially diagnosed with COPD decreased following the FeNO testing (24% to 13%). Furthermore, the number of patients without an initial diagnosis decreased from 17 patients to 11 with five patients being diagnosed as asthmatics, and one patient as ACOS after FeNO testing  were reviewed, and the result suggests that FeNO is a reliable test to diagnose and manage asthma even though reported sensitivity and specificity was 0.65 and 0.82, respectively . In a re were revAlthough the clinical application of FeNO has increased over the years, it is important to evaluate all the available patient data before reaching a diagnosis. In Table Limitation of this study can be attributed to the FeNO test itself because of false positive results, although rare. This can occur with viral illness due to its ability to induce the production of nitric oxide in an attempt to utilize its antiviral properties . Data suFractional exhaled nitric oxide (FeNO) is a simple, reproducible, and a noninvasive marker of inflammation which may be used as one piece of data to help the clinician better assess the patients true underlying pulmonary disorder and potentially change the treatment plan. Although the primary purpose of this study was to help more accurately diagnose the patient, the true value of FeNO may be in regards to treatment. Therefore, further studies are needed to better understand FeNO and its applicability across the clinical spectrum.


Nephrotoxicity of intravenous (IV) colistin has impeded its clinical use; aerosolized (AS) colistin may be an alternative, but safety data are lacking. Therefore, this study aimed to evaluate the incidence of acute kidney injury (AKI) and risk factors associated with IV and AS colistin administration.A retrospective study was performed in a tertiary referral hospital. Data were collected before and after colistin administration between October 2012 and April 2016. Exclusion criteria were as follows: age less than 18 years, previous colistin administration, concurrent use of IV and AS colistin, dialysis before colistin use, and colistin use for less than 3 days. We compared AKI incidence following administration of IV versus AS colistin and analyzed risk factors for colistin-associated nephrotoxicity.A total of 464 patients were enrolled . Incidence of AKI was significantly higher in the IV group . Duration of colistin use  and presence of chronic kidney disease  were associated with nephrotoxicity. There were no significant risk factors associated with AS colistin.Although AS colistin was not associated with any significant risk factors for nephrotoxicity, duration of colistin use and baseline kidney function may affect AS colistin-associated nephrotoxicity. Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, and Enterobacter, has risen. These infections represent a threat to public health because these gram-negative bacteria are resistant to commonly used antibiotics such as antipseudomonal penicillins and cephalosporins, aminoglycosides, tetracyclines, fluoroquinolones, and carbapenems  [The estimated glomerular filtration rate (eGFR) was calculated to assess kidney function by using the abbreviated MDRD study equation [eGFR = 175 × SCrnblack)] .2 , and the OR of CKD was high at 2.710 .2, p-value <0.001; baseline BUN 41.69 ± 27.41 vs 29.61 ± 20.18 mg/dL, p-value < 0.001) and more common in CKD patients . There was no difference in the dose between the two groups, but the accumulated dose was higher in the AKI group because colistin was used for a longer duration . Comparison of AKI and non-AKI patients treated with AS colistin yielded similar results to the comparison between AKI and non-AKI without AS colistin . Factors associated with kidney function were worse in the AKI group , according to KDIGO 2012 guidelines [2 [2 because factors that affect renal function could not be precisely controlled because this was a retrospective study. In addition, we felt that simply increasing the SCr threshold by 50% in patients with normal kidney function would be unlikely to be representative of true nephrotoxicity. Despite a 1.5-times increase in SCr in the normal kidney function group, many patients presented with low Scr. Individuals with low SCr were not considered to have impaired renal function and were not treated in clinical practice. According to these standards, the results of our study demonstrated that nephrotoxicity occurred twice as frequently with IV versus AS colistin treatment , meaning that patients with CKD have a high risk of nephrotoxicity. As such, colistin should be used with caution in CKD patients. Contrary to our findings, previous studies have described old age, longer duration of colistin use, diabetes mellitus, high dose per IBW, and septic shock as risk factors ,32.Several studies have examined association of IV colistin-associated AKI with nephrotoxins. Use of vancomycin, glycopeptides, NSAIDs, loop diuretics, and rifampin were reported to induce IV colistin-associated AKI ,10,31–33et al. [et al. [Several studies have examined dose-dependence of IV colistin-induced nephrotoxicity. Kwon et al.  reported [et al.  reported [et al. ,32. Howe [et al. ,31,32,34Comparison of the AKI group and non-AKI group showed that the AKI group used colistin for one week longer. From these results, it could be inferred that duration of colistin use affects nephrotoxicity. Accumulated dose is also significantly higher in the AKI group, but this may be due to the longer use of colistin in AKI. These results showed the same trend with AS colistin administration . ColistiOur study has some limitations that merit discussion. As this was a retrospective study, inaccurate patient information may have been provided. If data were missing, data collected most closely in time to the missing data were used. Furthermore, the mean value was used if there were multiple values obtained for the same factor on the same day. In addition, Patient clinical status was not evaluated using an APACHE II score, which would have been helpful in assessing kidney function and risk of AKI. Despite these limitations, this study, to our knowledge, is the first to attempt to determine risk factors associated AS colistin-induced nephrotoxicity. Furthermore, this is the first study to evaluate nephrotoxicity risks between IV and AS colistin treatment in a large number of patients.From the analysis, duration of colistin use and baseline kidney function may be associated with AS colistin-associated nephrotoxicity. Therefore, to reduce nephrotoxicity, short term use of AS colistin and evaluation of baseline kidney function prior to treatment are recommended.S1 DatasetAll data for this study were included and personal information were anonymous.(SAV)Click here for additional data file.


The cause of reduced exercise capacity (ExCap) in chronic kidney disease (CKD) is multifactorial. The aim of this study was to investigate determinants of aerobic ExCap in patients with mild to severe CKD not undergoing dialysis.We included 52 individuals with CKD stage 2–3, 47 with stage 4–5, and 54 healthy controls. Peak workload and peak heart rate (HR) were assessed by a maximal cycle exercise test. Cardiac function including stroke volume (SV) and vascular stiffness were evaluated by ultrasound at rest. Handgrip strength, body composition, haemoglobin level and self-reported physical activity were assessed.2 explained 82% of variation in peak workload. Peak HR contributed most to the variation; the peripheral variables handgrip strength and vascular stiffness did not improve the explanatory value in regression analysis.Peak workload , peak HR  and haemoglobin level  were all three significantly lower in CKD 2–3 than in controls,  and were even lower in stages 4–5 CKD than in CKD 2–3 . Resting SV and lean body mass did not differ between groups and handgrip strength was significantly lower only in CKD 4–5 compared to controls (p = 0.02). Peak workload was strongly associated with the systemic oxygen delivery factors: SV, peak HR and haemoglobin level. These three factors along with age, sex and heightIn this cross-sectional study of CKD patients not on dialysis, aerobic ExCap decreased gradually with disease severity. ExCap was associated mainly with systemic oxygen delivery factors, in particular peak HR. Neither muscle function and mass, nor vascular stiffness were independent determinants of aerobic ExCap in this group of CKD patients. The exclusion criteria for all participants were current malignancy, kidney transplantation or kidney donation, or blood-transmitted disease. The inclusion criteria for the controls were absence of kidney disease, cardiovascular disease, diabetes or any chronic medication. After inclusion, all participants underwent clinical investigation, anthropometric measurement, exercise stress test, handgrip strength test, laboratory testing, carotid ultrasound, transthoracic echocardiography and body composition scan.Ninety-nine patients with non-dialysis CKD (aged 18–65 years) and 54 healthy controls who had completed an exercise test were included . Glomeruundation . 54 contThe study protocol was reviewed and approved by the Local Ethics Committee and Institutional Review Board of the Karolinska Institutet at the Karolinska University Hospital. All participants gave their written informed consent.A symptom-limited incremental cycle ergometer test on an electronically braked cycle ergometer  was administered according to clinical standards. The initial workload and workload increase/minute  were individualized with the goal of achieving symptom limitation within 6 to 10 minutes. Participants were instructed to cycle at a speed of 60 rpm and were encouraged to continue cycling until exhaustion. Perceived exertion was reported as the highest rating  was used to measure maximum voluntary isometric contraction as a measure of handgrip strength. The test was performed with the participant in a standing position. The measurement was repeated three times using the dominant arm, and the highest value was recorded as handgrip strength.A handheld TakeiEchocardiography was performed only at rest, with a patient in the supine left decubitus position and according to current guidelines . The imaBody composition was measured with a whole-body dual-energy X-ray absorptiometry scan to determine LBM and body fat percentage .Physical activity level was rated by the participants using a four-point scale modified from the Saltin–Grimby Physical Activity Level Scale  as folloRegular exercise: running, swimming, tennis, badminton, gymnastics or similar activity on three or more occasions per week; every session should last at least 30 min and cause sweating.Level 1 = Moderate amount of regular exercise: running, swimming, tennis, badminton, gymnastics or similar activity on 1–2 occasions per week; every session should last at least 30 minutes and cause sweating.Level 2 = Light exercise: walking or cycling or other physical activity during at least 2 h per week, usually without sweating; this includes walking or cycling to/from work, Sunday walks, gardening, fishing, table tennis and bowling or similar activity.Level 3 = Sedentary: mostly reading, watching television, movies or other sedentary activities, or walking, cycling or light exercise for less than 2 h per week.Level 4 = Haemoglobin concentration was measured in venous blood obtained using routine laboratory methods at the Karolinska University Laboratory at the inclusion visit before the exercise test.The rate of systemic oxygen delivery (cardiac output x oxygen content) during peak exercise, expressed as millilitres of oxygen per minute, was estimated by the following equation:Peak systemic oxygen delivery = peak SV × peak HR x ((1.34 × haemoglobin concentration × oxygen saturation) + (0.03 × oxygen tension)).Assuming a normal blood oxygen saturation of 100% and a normal blood oxygen tension of 100 mmHg, the equation may be simplified as follows:Peak systemic oxygen delivery = peak SV × peak HR × (1.34 × haemoglobin concentration + 3).The factor of 1.34 equals the oxygen-binding capacity of haemoglobin, expressed as millilitres of oxygen per gram of haemoglobin (per g/L blood). The factor of 0.03 equals the oxygen solubility coefficient, expressed as millilitres of oxygen per litre of blood at an oxygen tension of 100 mmHg.SV measured at rest in the supine position was used as an approximation of peak SV in the calculation , 36.Continuous variables are presented as mean and standard deviation, and categorical variables as number and percentage of the study sample. Skewed variables are presented as median and interquartile range. Logarithmic transformation was used for skewed variables when analysed in multiple regression models.Group comparisons were performed using one-way analysis of variance, with Tukey’s honest significant difference post hoc test for further between-group analysis; Kruskal–Wallis or chi-square tests were used when appropriate. Bonferroni post hoc testing was performed to adjust p-values between groups when the Kruskal–Wallis test was used. For relevant variables, subgroup analysis was performed in patients not taking beta-blockers. Statistical significance was defined as a p-value <0.05 for a two-tailed test. Partial correlations were calculated to investigate the relationships between two variables while controlling for the influence of a third variable.To explore the determinants of aerobic ExCap in CKD, the two CKD groups were merged into one group (CKD stage 2–5). Peak workload was used as the dependent variable in multiple linear regression analyses following four different strategies (1–4 below). Standardized regression coefficients (RCs) are reported. Adjustment for age, sex and height squared was performed in all analyses because these have been identified as non-disease-related determinants of peak workload in reference materials , 31. HeiSingle determinants—The increase in the explanatory value (R2) for peak workload was analysed after adding single variables to a model with age, sex and height in both CKD 2–5 and controls. This purpose of this analysis was to better understand the contribution of individual variables to ExCap in CKD compared to controls.Multiple determinants—The multiple regression analysis performed in CKD 2–5 with peak workload as the dependent variable included the systemic oxygen delivery factors  and the peripheral factors (handgrip strength and vascular stiffness) after adjustment for age, sex and height.Stepwise multiple determinants—Manual forward regression was used to analyse the stepwise increase in R2 for peak workload in CKD 2–5 by adding the significant independent variables from the regression analysis described above (multiple determinants) after adjustment for age, sex and height.Statistical analyses were performed using IBM SPSS Statistics for Windows .The clinical characteristics for each group  are presented in Peak workload, exercise time, peak HR, HR reserve, estimated peak systemic oxygen delivery and HR recovery at 2 min after the end of exercise were all significantly lower in CKD 2–3 than in controls, and these values were even lower in CKD 4–5 . A simil2 the most. In the controls, only SV (but not peak HR or haemoglobin level) was associated with peak workload after adjusting for age, sex and height. LBM, handgrip strength and physical activity were also associated with peak workload in the controls.Tested as single determinants, the systemic oxygen delivery factors , HR reserve, E/é, vascular stiffness, LBM, handgrip strength and physical activity level were all associated with peak workload in CKD 2–5 after adjusting for age, sex and height . Among tTested as multiple determinants, in CKD 2–5 after adjusting for age, sex and height, systemic oxygen delivery factors  contributed significantly to explain the individual variation in peak workload . Vascula2 after adding significant physiological variables from the multiple regression model shown in 2 produced an R2 of 0.83; peak HR made the strongest contribution to R2. Age was no longer a significant determinant when peak HR was included in the model.2, SV, peak HR and haemoglobin level in patients not taking beta-blockers (n = 65) rendered an R2 of 0.81. Peak HR, SV and haemoglobin level still explained the significant variation in peak workload A separate multiple regression analysis with age, sex, height2, explained 83% of the individual variation in aerobic ExCap in the current CKD sample and peak HR was the main determinant of aerobic ExCap. The peripheral factors handgrip strength and vascular stiffness differed significantly only between CKD 4–5 compared to controls and LBM did not differ between groups. Moreover, after inclusion of the systemic oxygen delivery variables, these peripheral factors did not provide further explanation of the variance in ExCap and seem to be of less importance for aerobic ExCap in this group of CKD patients.In mild to severe non-dialysis CKD, we found that aerobic ExCap, peak HR, haemoglobin level and the estimated peak systemic oxygen delivery decreased gradually with increasing disease severity. ExCap was strongly associated with the systemic oxygen delivery factors SV, peak HR and haemoglobin level. These factors, along with age, sex and height2 for peak workload in these CKD patients Click here for additional data file.


Marinobufagenin (MBG) is an endogenous cardiotonic steroid (CTS) that inhibits the Na+/K+-ATPase. Human MBG is significantly increased in end-stage renal disease and immunization against MBG attenuates cardiovascular fibrosis in a rat model of uremic cardiomyopathy. Mineralocorticoid antagonists (MRA) block MBG binding sites and decrease proteinuria in chronic kidney disease (CKD) patients. We therefore aimed to investigate the association of MBG and albuminuria, as a marker of renal damage, as well as MBG and decline of glomerular filtration rate (GFR).The Graz endocrine causes of hypertension (GECOH) study is a single center study of adults routinely referred for screening of endocrine hypertension. Plasma MBG was measured by an enzyme-linked immunoassay, and in a post-hoc analysis, follow-up creatinine levels were obtained. Patients with proteinuria >3.5g/day at baseline were excluded from further evaluation.ρ = .357; p = .028) and proteinuria . In linear regression analysis, the association remained significant after adjustment for age, sex, and BMI , and for mean systolic blood pressure . In follow-up analyses (N = 30), MBG was significantly associated with decline in GFR after adjustment for time-to-follow-up .We measured MBG concentrations in 40 hypertensive subjects and excluded one patient due to pre-existing proteinuria. Plasma MBG was significantly correlated with albuminuria (Spearman The findings suggest that MBG plasma concentrations were associated with albuminuria as well as decline in kidney function. Whether MBG predicts hard renal endpoints warrants further investigations. Chronic kidney disease (CKD) is one of the most burdensome and frequent medical conditions. In general populations, CKD prevalence of all five KDIGO stages is 13.4%, and of KDIGO stages three to five is 10.6%. However,Mineralocorticoid receptor antagonist (MRA) therapy has been suggested to mitigate renal fibrosis.,6 MRA deMarinobufagenin (MBG) is an endogenous cardiotonic steroid (CTS), all of which are inhibitors of the sodium-potassium adenosine triphosphatase (Na+/K+-ATPase), also called digitalis-like factors. By chemical structure, MBG belongs to bufadienolides. First deThe circulating concentrations of MBG are significantly increased in virtually all patients undergoing dialysis for ESRD.–21 HigheWe therefore evaluated the relation of plasma MBG and albuminuria, as a clinical marker of kidney damage, in patients with arterial hypertension in a post-hoc analysis of the Graz endocrine causes of hypertension (GECOH) study. In addition, we assessed the association of plasma MBG concentrations and decline of estimated GFR at follow-up.The Graz endocrine causes of hypertension (GECOH) study is a single center study at a tertiary care center that comprises adult patients (aged ≥18 years), who have routinely been referred to the outpatient clinic for screening for endocrine hypertension at the Division of Endocrinology and Diabetology, Medical University of Graz, Austria. The study protocol has previously been published elsewhere.  The primIn a first baseline visit, arterial blood pressure (BP) was measured according to the method of Korotkoff after five minutes at rest and a sphygmomanometer with an appropriate cuff. Systolic and diastolic BP values were measured on both arms and the mean values were recorded. Height and weight were determined wearing light clothes and no shoes. Detailed methods about diagnostic criteria of primary aldosteronism (PA) have been described previously.At a first baseline visit, venous blood sampling was performed according to current guidelines. Blood waAfter a maximum of five-year storage at -80°C in aliquots of 1 mL without additional freeze-thaw cycle, plasma MBG was measured at the Department of Nephrology, Transplantation and Internal Medicine, Medical University of Silesia, Katowice, Poland, following extraction with C-18 columns . DetailedPlasma creatinine was measured at the Clinical Institute of Medical and Chemical Laboratory Diagnostics Medical, University of Graz, Graz, Austria, using the Jaffe method, with a calibration traceable to an isotope dilution mass spectrometry (IDMS) reference measurement procedure .2) at baseline and follow-up was determined using the CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration) formula.[In January 2018, we retrospectively obtained available plasma creatinine concentrations of participants from the local hospital information system, where only the single most recent follow-up value was used for the present analysis. Estimated glomerular filtration rate  were excluded from further analyses.r coefficients were computed to assess the association between parametric variables, Spearman’s ρ coefficients for non-parametric variables. Linear regression analyses were further used to account for the influence of potential confounding variables. To lower risk of over-fitting, in cox regression analyses, MBG concentrations were allocated into three equally sized groups and the hazard of grouped plasma MBG concentrations on falling into a lower CKD class according to KDIGO guidelines [Depending on their distribution, continuous data are either presented as means ± SD  or as medians with interquartile ranges (skewed variables). Categorical variables are presented as percentages. To test for normality we used Shapiro-Wilk tests. Group comparisons were performed by analysis of variance (ANOVA), Kruskal-Wallis test by ranks or chi-squared test for linear trends, where appropriate. Pearson’s idelines  at folloBetween February 2009 and December 2012, 233 white study participants were enrolled at the outpatient clinic of the Division of Endocrinology and Diabetology, Medical University of Graz, Austria. Plasma marinobufagenin (MBG) concentrations were available in 40 patients, of which 11 patients were diagnosed with primary aldosteronism (PA) and 29 patients with essential hypertension (after exclusion of PA). One patient was excluded from the present analysis due to pre-existing diabetic nephropathy and excessive proteinuria >3.5g/day at baseline.Overall and disease-specific baseline characteristics of remaining study participants (N = 39) are shown in ρ = .357; p = .028) and proteinuria , but not with GFR at baseline, or systolic BP at baseline. In linear regression analysis, the association remained significant after adjustment for possible confounders, including age, sex, and BMI , and for mean systolic blood pressure . However, after adjustment with GFR at baseline, significance was lost .Plasma MBG concentrations were significantly correlated with albuminuria , but not in patients with PA .In a sensitivity analysis, the correlation between plasma MBG concentrations and albuminuria at baseline was still evident in patients with essential hypertension . In linear regression analysis, the association turned significant after adjustment for time-to-follow-up . MBG correlated with spot urine sodium , but not with spot urine sodium-to-creatinine ratio , nor 24-hour sodium excretion .In sensitivity analyses, significance of the correlation of plasma MBG and decline in GFR was lost for both patients with essential hypertension  and PA . No association was found for albuminuria at baseline and decline in GFR  per year during follow-up.In cox regression, the hazard ratio of increasing plasma MBG concentrations for the risk of falling into a lower CKD class at follow-up according to KDIGO guidelines was 3.6 . Eventually, an increase in 0.1 nmol/L plasma MBG at baseline was associated with a decrease of 2.75 ml/min/1.73mIn this study, we demonstrated a significant association between plasma marinobufagenin (MBG) concentrations and albuminuria, a marker of renal damage. In addition, we could demonstrate that elevated plasma MBG concentrations correlate with a decline in estimated glomerular filtration rate (GFR) at follow-up. The results indicate that, for hypertensive patients, MBG could serve as a potential marker for kidney impairment at follow-up. In subgroup analyses, the association was evident in patients with essential hypertension, although we showed that MBG concentrations are higher in PA patients than in patients with essential hypertension. The seleMBG concentrations are elevated in patients with end-stage renal disease (ESRD) and MBG was suggested to mediate cardiovascular remodeling in uremic cardiomyopathy. In rats with experimental renal disease, MBG was associated with renal fibrosis and passive immunization against MBG attenuated progression of fibrosis as well as improved renal function. To our kMBG usually binds to Na+/K+-ATPase and dissociates very slowly, so that the cell induces endocytosis of the blocked Na+/K+-ATPase. The Na+/K+-ATPase resides ubiquitously at the plasmamembrane barrier of various cell types including human cardiomyocytes, vascular smooth muscle cells, cells of the adrenal gland and epithelial cells of the proximal tubule. Besides the basic function of Na+/K+-ATPase to maintain a gradient of sodium and potassium concentrations across the plasmamembrane barrier, Na+/K+-ATPases take part in signaling pathways of various cell lines. The Na+/MRA therapy was shown to delay CKD progression over the long term. Still, MMBG may not only mediate renal fibrosis but may help to detect individuals at risk for fibrosis and progression of CKD. As MRA were shown to antagonize MBG effects, increased MBG concentrations may further help to identify individuals benefitting most from MRA treatment.We have to acknowledge that in our study the relatively small sample size of patients with MBG measurement and follow-up as well as restricted clinical data at follow-up decrease the power of or study and limit further discrimination of results. Our findings should therefore be rather regarded as hypothesis generating warranting confirmation in larger well characterized cohorts. To some extent, our results could be influenced by increased prevalence of PA with increasing MBG concentrations, as we have already showed that MBG is higher in patients with PA. AlthoughWhile our study suggests increased MBG concentration as a predictor of GFR decline, further studies are necessary to validate our results. Additional investigations are warranted to see whether MRA therapy can prevent GFR decline in association with increased MBG concentrations.Our data support the hypothesis that a high MBG concentration is associated with markers of kidney impairment. Patients with highest MBG concentrations may be at increased risk of future GFR decline. Whether elevated MBG concentrations predict hard renal endpoints warrants further investigations.S1 DatasetData from baseline visit(s) and follow-up.(XLSX)Click here for additional data file.


The purpose of this study was the assessment of volumetric CT-perfusion (CTP) of the lower leg musculature in patients with symptomatic peripheral arterial disease (PAD) of the lower extremities, comparing it with established angiographic and hemodynamic parameters.Thirty-five consecutive patients with symptomatic PAD of the lower extremities requiring interventional revascularization were assessed prospectively. All patients underwent a CTP scan of the lower leg, and hemodynamic and angiographic assessment. Hemodynamic parameters, specifically ankle-brachial pressure index (ABI), ankle blood pressure (ABP), peak systolic velocity (PSV), and segmental pulse oscillography (SPO) level, were determined. Lesion length and degree of collateralization were assessed by interventional angiography. CTP parameters were calculated with a perfusion software, acting on a no outflow assumption. A sequential two-compartment model was used. Differences in CTP parameters and correlations between CTP, hemodynamic and angiographic parameters were assessed with non-parametric tests.-1, mean blood volume (BV) 0.71 ± 0.33 ml/100ml, and mean mean transit time (MTT) 7.22 ± 2.66 s. BF and BV were higher in subjects with longer lesions, and BV was higher in subjects with lower ABI. Significant correlations were found between lesion length and BV (r = 0.65) and BF (r = 0.52). Significant inverse correlations were found between BV and ABI and between BV and ABP .The cohort consisted of 27 subjects with an occlusion, and eight with a high-grade stenosis. The mean blood flow (BF) was 7.71 ± 2.96 ml/100ml*minIn our study, we have shown the feasibility of CTP for the assessment of PAD. In the future, this quantitative method might serve as a non-invasive method, possibly complementing the diagnostic workup of patients with peripheral arterial disease. Z-axis coverage [Peripheral arterial disease (PAD) is one of the major cost-driving factors in public healthcare . Similarcoverage . Beam-hacoverage . Radiaticoverage . Furthercoverage . In contcoverage . Neverthcoverage . This iscoverage , 22–24. Despite several potential benefits, such as fast and non-invasive quantitation of tissue ischemia, the clinical application of CTP of the limbs has so far been challenged by the aforementioned technical difficulties, and larger patient studies are still missing.Therefore, the purpose of our study was the assessment of volumetric CTP of the lower leg musculature in patients with symptomatic PAD, and the comparison of CTP parameters with state-of-the-art angiographic and hemodynamic parameters, in order to further characterize the potential role of this non-invasive method for the quantitative in-vivo assessment of limb ischemia.S1 File.This prospective study was approved by the institutional review board and by the cantonal ethics committee . All patients provided informed signed consent prior to the examinations. Thirty-five consecutive patients  with symptomatic PAD of the legs were evaluated. All patients were referred for angioplasty for unilateral interventional revascularization of arteries that provide blood supply to the lower leg. Symptomatic PAD was defined as prevalence of intermittent claudication, ischemic rest pain and ischemic tissue loss such as gangrene or non-healing ischemic ulcers, and ultrasonographic evidence of a hemodynamically significant obstruction of the common iliac, external iliac, superficial femoral or popliteal artery. Medical history was recorded. Further details are given in All subjects underwent clinical assessment of hemodynamic parameters within 24 hours prior to revascularization (see below). Angiographic parameters were assessed at the beginning of the interventional revascularization. Immediately before the revascularization procedure, all patients underwent a CTP scan of the lower leg.Hemodynamic assessment was performed in all patients prior to the planned revascularization. Evidence of hemodynamically significant obstruction was established by non-invasive vascular testing, pulse volume recordings and duplex ultrasound imaging studies.S1 File. Arteries in subjects with an ABI ≥ 1.40 were considered incompressible. ABI and ABP values of these subjects were not considered for further analysis, because such values indicate the presence of media sclerosis. An ABI of 1.00 to 1.40 was considered as normal, borderline from 0.91 to 0.99, and pathologic if < 0.91 [For the assessment of the maximum ankle-brachial pressure index (ABI) and the systolic ankle blood pressure , patients were placed in supine position at rest for 5–10 minutes in a room with ambient temperature 22–25°C). A digital oscillometric automatic blood pressure device with appropriately sized blood pressure cuffs  was used to measure systolic, diastolic and mean arterial blood pressure on both upper arms simultaneously. Less than 12 mmHg inter-arm systolic pressure gradient was regarded as normal [°C. A digf < 0.91 . The ABISegmental pulse oscillography (SPO) was performed with a two-channel device . For this study, only the pulse curves of the lower calf were taken into account. Pulse curves were rated as normal if the wave morphology was dichotomous, mildly pathologic with blunt wave morphology and absence of dichotomy, and severely pathologic with flat waves. Maximum impairment of arterial perfusion in a given segment was characterized by a zero baseline. Ultrasonographic imaging was performed with a clinical ultrasound system . Clinical standard criteria were applied to quantify the severity of a stenosis. In subjects with vessel stenosis, the ratios of peak systolic velocity  in the stenotic segment and prestenotic PSV were calculated.All CTP examinations were performed with a 128-slice CT scanner  in shuttle mode, allowing for 28.2 cm of axial coverage and 2 s temporal resolution, which included almost all muscles of both lower legs in every subject. After placement on the gantry table, the patients’ feet were fixated using tape to prevent spontaneous movement. The tube current was set to 100 mAs, the tube voltage to 100 kV(p). The duration of the CT-perfusion scan was 60 s, with a rotation time of 2 s. CTP was started 15 s after injection of 60 ml of CM  at 5 ml/s. CM was injected into an antecubital vein by a dual-head pump injection device , followed by a flush of 50 ml of NaCl at 5 ml/s. The collimation was 64 x 0.6 mm. The reconstruction increment was 10 mm at 10 mm slice width. Image reconstruction was performed with a 512 x 512 pixel matrix and medium smooth B30f kernel. For image post-processing and analysis, the reconstructed images of both legs were transferred to a commercially available computer workstation .-1), blood volume , and mean transit time  were calculated with a perfusion software , based on the Patlak analysis. Since veins did not show sufficient CM enhancement in the late phase of the dynamic series and since venous enhancement was furthermore often degraded by pathologic flow within varicose veins in our patient cohort, no outflow curve was obtained. Thus, as previously described in the literature, we acted on a no outflow assumption [CTP parameters blood flow . The lesion length was rated as short (1) if < 4 cm, medium (2) if 4–10 cm, and long (3) if > 10 cm. The degree of collateralization was assessed by visual estimation on angiographic images. It was rated as good (1) if the estimated cross-sectional area of collaterals added up to > 50% of the pre-lesion diameter of the collateralized vessel, medium (2) if < 50%, and poor (3) if only faint or no collaterals were visible. The performing interventional radiologist had 20 years of experience in the field of revascularization procedures.Demographic, hemodynamic, angiographic and CTP parameters in patients with stenosis and occlusion were compared by Mann-Whitney U-test. Differences of CTP parameters between symptomatic and non-symptomatic limbs were analysed by Wilcoxon signed ranks test. CTP parameters were stratified by lesion length, collateralization, ABI and SPO, and were then compared by Kruskal-Wallis test. The correlation between CTP, hemodynamic and angiographic parameters was analysed using Spearman’s correlation. Results were interpreted as almost perfect correlation between ±0.81 and ±1.00, substantial between ±0.61 and ±0.80, moderate between ±0.41 and ±0.60, fair between ±0.21 and ±0.40, there under no correlation . A p-valFig 1A), while the degree of collateralization in both types of lesions was not significantly different. Patients with medium and long lesions had a higher degree of collateralization than patients with short lesions, (p = 0.041) (S1 Table).Twenty-seven patients (77%) were referred for interventional revascularization of an occlusion, eight patients (23%) for a high-grade stenosis. All lesions were located unilaterally in the common iliac, external iliac or superficial femoral artery. Occluded vessel segments were significantly longer than stenotic segments . ABP and ABI values of these subjects were hence excluded from analysis. The majority of the remaining patients had mildly pathologic ABI values (n = 22), five had severely pathologic values, and one a normal value. For a comparison of CTP parameters stratified by ABI, subjects were assigned to three equally distributed groups .Table 1.PSV could not be determined in 6 subjects . Upstream PSV was significantly higher in stenotic than in occluded vessels. Detailed results are given in -1, the mean BV was 0.71 ± 0.33 ml/100ml tissue, and the mean MTT was 7.22 ± 2.66 seconds. Below we provide data of the asymptomatic limb as well, whenever statistical significance is claimed for CT-parameters in the symptomatic limb, in order to identify potential inter-group differences against intra-individual, inter-limb differences.The mean BF of the symptomatic limb was 7.71 ± 2.96 ml/100ml tissue * minTables Fig 1B). However, in the subgroup with short lesions, higher CTP parameters were found in the asymptomatic than in the symptomatic limb (Table 2).Overall, CTP parameters were neither significantly different in patients with stenosis or occlusion, nor in the symptomatic and asymptomatic limb . For MTT no such difference was found.BF and BV of the symptomatic leg were significantly different depending on lesion length, with lowest values obtained in short lesions, and highest values in long lesions (Table 3).CTP parameters were not different when stratified by the degree of collateralization (There was no significant difference in CTP parameters when stratified by the SPO level.p = 0.006), with higher BV in patients with lower ABI . In the asymptomatic limb, there was no significant difference of BV among the ABI groups (p = 0.18).BV of the symptomatic limb was significantly different among ABI groups , and a moderate correlation between BF and lesion length . BV showed a moderate inverse correlation with ABI  and ABP . Similar correlations were obtained in the subgroup analysis of subjects with occlusions, although by trend with lower coefficients (Table 4).A substantial correlation was found between BV and lesion length  without suspicion of media sclerosis had a normal ABI value . HoweverIn patients with a pathologic ABI, further diagnostic tests are necessary in order to localize lesions and assess the grade of stenosis. Several different modalities provide detailed characteristics of the clinically relevant lesion. To date, invasive diagnostic tests are the first line recommendation for patients eligible for revascularization . HoweverIn a CTP study by Barfett et al., a significant difference of foot perfusion values between the side of simulated vascular obstruction and the non-obstructed side were shown . In our Since PAD is considered a generalized disease, changes of the microvascular density might alter the vascular supply of the muscles significantly on both the symptomatic and the asymptomatic side. On one hand, the microvascular occlusion might reduce the perfusion of the muscles. On the other hand, it has been shown that in limbs with a severe stenosis, ischemia induces neo-angiogenesis . This maThe assessment of CT perfusion in calf muscles, as performed in our study, might have advantages over the perfusion measurement in the skin or muscles of the foot. We expect perfusion values obtained in muscle tissue be more reliable compared to skin-derived values, since they are more independent of ambient temperature and resistant to movement artefacts if patients are positioned properly , 13, 16.Besides stenosis diameter and length, BF is also determined by partly microscopic parameters that are more difficult to quantify exactly, such as microvascular obstruction and collateralization. Our study cannot identify which one of these parameters is the dominant factor determining BF. This might be the reason for not finding a strong correlation between CTP and different parameters.It should be mentioned that our study has some limitations. We did perform consensus reading instead interreader evaluation of results . There aAsymptomatic limbs in our cohort are inherent parts of the systemic disease as well, and as shown in our study, cannot serve as standard of reference for absolute CTP values. Future work might analyse CTP parameters in patients with critical limb ischemia at rest and at stress, and before and after revascularization. Also, we did not compare CTP with other quantifiable perfusion imaging, such as PET perfusion. Although an accurate technique, it is only available in a few centers worldwide, owing to financial, technical and logistic issues , 41. FinOverall, since there was a difference in CTP parameters in patients with short stenosis, one possible explanation might be that using CTP-values in direct intra-individual comparison in this indication works best in a specific time window .In our study we showed that BF and BV are higher in subjects with longer lesions and that BV is higher in subjects with lower ABI. In a subgroup with short lesions, BF and BV were lower compared to the asymptomatic limb. Furthermore, we found a positive correlation between CTP parameters BF and BV and the lesion length, and an inverse correlation between BV and ABI. In the future, this quantitative method might serve as a non-invasive method, which might complement the diagnostic workup of patients with peripheral arterial disease.S1 File(DOCX)Click here for additional data file.S1 TablePatients with medium and long lesions had a higher degree of collateralization than patients with short lesions, (p = 0.041).(DOCX)Click here for additional data file.


NS). Studies support the “overfill” theory of NS with pathophysiological proteolytic activation of the epithelial sodium channel (ENaC) which explains the common observation of suppressed renin –angiotensin system and poor therapeutic response to ACE inhibitors. Blockade of ENaC by the diuretic amiloride would be a rational intervention compared to the traditionally used loop diuretics. We describe a 38‐year‐old male patient with type1 diabetes who developed severe hypertension (200/140 mmHg), progressive edema (of at least 10 L), and overt proteinuria (18.5 g/24 h), despite combined administration of five antihypertensive drugs. Addition of amiloride (5 mg/day) to treatment resulted in resolution of edema, weight loss of 7 kg, reduction in blood pressure (150/100–125/81 mmHg), increased 24 h urinary sodium excretion (127–165 mmol/day), decreased eGFR (41–29 mL/min), and increased plasma potassium concentration (4.6–7.8 mmol/L). Blocking of ENaC mobilizes nephrotic edema and lowers blood pressure in NS. However, acute kidney injury and dangerous hyperkalemia is a potential risk if amiloride is added to multiple other antihypertensive medications as ACEi and spironolactone. The findings support that ENaC is active in NS and is a relevant target in adult NS patients.Sodium and fluid retention is a hallmark and a therapeutic challenge of the nephrotic syndrome ( Thus the “diuretic resistance” associated with NS is not absolute and could be related to active ENaC. This is in agreement with mouse studies with segmental knock out of ENaC (Perrier et al. μmol/L are achieved (Andersen et al. The strategy to overcome volume expansion and hypertension in this patient was initially to increase the doses of antihypertensive medication including thiazide/loop diuretic combination. Despite titrating antihypertensives to include five different drugs and notably with high doses of loop diuretics, there was only a very modest reduction in blood pressure and no significant effect on edema. Despite ACE‐inhibition and aldosterone blockade, the powerful action of low‐dose amiloride showed that unopposed ENaC‐mediated NaDespite initial dual blockade of RAAS with ACE inhibitors and spironolactone, hyperkalemia did not develop until supplementation with amiloride. This indicates that hyperkalemia was not the result of RAAS inhibition, but rather a result of blocking ENaC. In the current case, hyperkalemia was likely aggravated by the decrease in kidney function which also could explain the reduction in proteinuria. The decline in eGFR was reversible. It could be speculated that the prompt drop in blood pressure in a patient with possible impairment of the autoregulation of renal blood flow combined with a rapid loss of estimated 7L of extracellular fluid might have worsened kidney function. Acute kidney injury and hyperkalemia have previously been reported in a controlled clinical trial in proteinuric patients given amiloride, and the trial was stopped early (Unruh et al. In summary, the present case shows that the addition of the ENaC blocker amiloride to multidrug regimen reduces edema, body weight, and blood pressure in a severe NS case. It also illustrates that careful monitoring and stepwise titration is necessary to minimize the risk for acute injury and hyperkalemia.Written informed consent was obtained from the patient for publication.The authors declare that they have no competing interests.


Older adults with sarcopenia may be at risk for unstable postural blood pressure due to diminished lean mass that plays a role in maintaining fluid volume. Males have greater lean mass, so risk may be mediated by gender. We compared postural blood pressure changes in older men  and women  with sarcopenia before and after an overnight fast. Sarcopenia was defined using the Lean Mass Index . Body composition was measured using multi-frequency bioelectrical impedance, and blood pressure was measured lying, sitting, and standing. On Day 1  there were significant drops in systolic blood pressure, with an overall decrease of -9.1 ± 2.2 mmHg (p < 0.001) between lying and standing. On Day 2 (overnight fast), postural changes were more profound, with an overall decrease of -14.1 ± 2.8 mmHg (p < 0.001). However, when compared by gender, postural changes between lying and standing remained significant but did not differ between men and women . On both days diastolic blood pressure remained stable. In this group of older adults, significant decreases in postural systolic blood pressure were observed in the early morning fasted condition, increasing the risk for orthostatic hypotension (drop in systolic blood pressure -20.0 mmHg). Interestingly, gender did not influence risk.