There are limited data on the clinical value of routine postdilatation using noncompliant balloons after contemporary drug‐eluting stent implantation.Additional postdilatation using noncompliant balloons after everolimus‐eluting stent implantation could provide better clinical outcomes.We randomly assigned 1774 patients with coronary artery disease to undergo additional high‐pressure postdilatation using noncompliant balloons and moderate‐pressure dilatation using stent balloons after everolimus‐eluting stent implantation. The primary endpoint was a composite of death, myocardial infarction (MI), stent thrombosis, and target vessel revascularization (TVR) 2 years after randomization.P = .537). In addition, no significant differences were observed between the two groups in the occurrence of an individual end point of death , MI , stent thrombosis , or TVR .The study was discontinued early owing to slow enrollment. In total, 810 patients (406 patients in the high pressure group and 404 in the moderate pressure group) were finally enrolled. At 2 years, the primary endpoint occurred in 3.6% of patients in the high pressure group and in 4.4% of those in the moderate pressure group (The strategy of routine postdilatation using noncompliant balloons after everolimus‐eluting stent implantation did not provide incremental clinical benefits. This study involved 15 cardiac centers in Korea between February 2012 and October 2015. Patients were considered eligible if they had either stable angina or acute coronary syndrome and had at least one de novo coronary lesion  suitable for stent implantation. Patients were excluded if they had contraindications to aspirin and clopidogrel, unprotected left main disease , graft vessel disease, in‐stent restenotic lesion, bifurcation lesion requiring stent implantation for both main and side branches, history of bleeding diathesis or coagulopathy, hepatic dysfunction with aspartate aminotransferase or alanine aminotransferase level ≥3 times the upper normal reference limit, history of renal dysfunction or serum creatinine level of ≥2.0 mg/dL, serious noncardiac comorbid disease with a life expectancy of <2 years, ST‐elevation acute myocardial infarction (MI) within 2 weeks, planned major surgery within the next 6 months with the need to discontinue antiplatelet therapy, or inability to follow the protocol. In patients with multiple lesions who fulfilled the inclusion and exclusion criteria, the first stented lesion was considered the target lesion. The institutional review board at each participating center approved the protocol. All patients provided written informed consent.2.2Patients who met the inclusion and exclusion criteria were randomly assigned in a 1:1 ratio to receive additional high‐pressure postdilatation using a noncompliant balloon and moderate‐pressure dilatation using a stent balloon with an interactive web response system. The allocation sequence was computer generated and stratified according to participating center and blocked with block sizes of four and six varying randomly.The procedure was performed using standard techniques. The stents were deployed at 10 to 14 atm with their stent balloon system. If an acceptable result was achieved, randomization was performed. An acceptable result was defined as a quantitative angiographic residual diameter stenosis of <30% with TIMI  grade 3 flow and absence of major stent edge dissection (type C‐F) and major side branch occlusion (TIMI flow 0 or 1 in the side branches with a reference diameter of ≥2.0 mm). In the moderate pressure group, the procedure was finished. In the high pressure group, postdilatation with at least the same or larger size noncompliant balloons at 18 to 22 atm was followed  and clopidogrel . Heparin was administered throughout the procedure to maintain an activated clotting time of ≥250 seconds. The use of IVUS and administration of glycoprotein IIb/IIIa inhibitors were at the discretion of the operator.2.3The primary endpoint was defined as the occurrence of major adverse cardiac events, including all‐cause death, MI, stent thrombosis, and target vessel revascularization (TVR) 2 years after the procedure. The secondary endpoints included the occurrence of an individual endpoint of death, MI, stent thrombosis, and TVR at 2 years. At 2 years, a composite of death and MI was also assessed.Death was considered to be cardiac in etiology unless an unequivocal noncardiac cause was established. MI was diagnosed on the basis of an increase in the creatine kinase MB  fraction or troponin level greater than the 99th percentile of the upper normal limit with at least one of the following aspects: ischemic symptoms, electrocardiographic changes, and abnormal imaging findings of MI.2.5On the basis of results from previous studies,t test or Mann‐Whitney test. Categorical variables were presented as numbers and percentages and compared using the χ2 test or Fisher's exact test. Survival was assessed using the Kaplan‐Meier method and compared using the log‐rank test. In the patients with multiple clinical events, the first event was considered to be the component of composite outcome. Statistical analyses were performed using the time of first event from randomization. A P value of <.05 was considered statistically significant. All statistical analyses were performed using the SPSS software .All analyses of the two groups were performed according to the intention‐to‐treat principle. Continuous variables are presented as means ± SD or medians (interquartile ranges) and compared using the 33.1Between February 2012 and October 2015, 810 patients were randomly assigned to the high pressure group (n = 406) and moderate pressure group (n = 404) . Consequently, on postprocedural quantitative angiographic analysis, the in‐stent acute gain was higher and the in‐stent diameter stenosis was smaller in the high pressure group than in the moderate pressure group. However, the incidence of edge tear or perforation and no reflow or distal embolization was low and comparable between the two groups (Table The lesion type B2 or C was seen in 568 patients (71%). However, moderate to severe calcification was observed in 44 patients (5.4%). Seventy‐seven patients (9.5%) were treated with overlapping stents. The mean stented length of the target lesions was 28.7 ± 12.6 mm. Although the mean final balloon size was similar between the two groups, the maximal inflation pressure was higher in the high pressure group than in the moderate pressure group  , MI , stent thrombosis , TVR , and death/MI   Figure A. In add4The main findings of this study were as follows: (a) in the patients who underwent contemporary everolimus‐eluting stent implantation, postdilatation using noncompliant balloons was safely performed without increasing incidence of adverse events; (b) postdilatation resulted in higher in‐stent acute gain and smaller in‐stent diameter stenosis; (c) however, the routine postdilatation strategy did not improve the 2‐year clinical outcomes; and (d) therefore, the routine postdilatation approach should be carefully considered in contemporary DES implantation.Optimal stent deployment during PCI has an important effect on short‐ and long‐term clinical outcomes.While reducing the risk of in‐stent restenosis and stent thrombosis, there are potential adverse effects from postdilatation. Postdilatation using noncompliant balloons at high pressures could be associated with a risk of edge tear and vessel rupture.In the bare‐metal stent era, deployment of such stents was often associated with suboptimal stent expansion.In our study, there are possibilities why the routine strategy of postdilatation using noncompliant balloons did not provide incremental clinical benefits. Complex lesion subsets, such as heavy calcification, large plaque burden, small vessel, in‐stent restenosis, or long lesion, are associated with a lower success rate and require more attention and specialized devices to obtain an optimal stent deployment.Our study has also several limitations. First, in South Korea, the registration of clinical studies was not mandatory until March 2018. Since the present study has been conducted in 15 cardiac centers of South Korea between February 2012 and October 2015, our study was possible by approvals of the institutional review board at each participating center without legal restrictions. Second, the decision of enrollment of study was made by the attending operators. Unfortunately, we have no reliable data for patients screened and excluded from the current study. Third, the overall clinical event rate was lower than anticipated. Moreover, the present study was discontinued early due to slow enrollment. Therefore, the sample size could be insufficient for evaluating whether the routine postdilatation strategy benefited all subgroups. Fourth, the use of IVUS was at the discretion of operating physicians. In previous and our studies, routine use of IVUS or postdilatation during PCI did not show incremental clinical benefits.In conclusion, the strategy of routine application of postdilatation using noncompliant balloons after everolimus‐eluting stent implantation did not improve clinical outcomes at 2 years. However, the present study did not have enough statistical power to evaluate the clinical effects of additional postdilatation. Therefore, these findings should be confirmed in further randomized clinical trials with larger populations.The authors declare no potential conflict of interests.Supplementary Figure 1 Flow of Study ParticipantsClick here for additional data file.


When stratified by age and gender, the correlations were observed in women and patients ≤60 years. Furthermore, among patients using calcium channel blocker (CCB) and angiotensin receptor blocker (ARB), those with telomeres lengthening showed a significantly lower level of ∆SBP and ∆PP. There was no correlation between telomere attrition and incidence of cardiovascular events. Our data indicated that increased telomere length of leukocytes was associated with decreased SBP and PP, particularly for patients who received CCB and ARB, supporting that telomere attrition may provide new sight in clinical intervention for hypertension.Leukocytes telomere length has been associated with hypertension, but, whether longitudinal telomeres change could serve as a useful predictive tool in hypertension remains uncertain. This study aimed to examine the longitudinal trajectory of leukocytes telomere length in a population-based prospective study of 1,108 individuals with hypertension. Leukocytes telomere length were measured at baseline and again after a median 2.2 (range 1.5-2.4) years of follow-up. Age as an independent predictor was inversely associated with baseline telomeres and follow-up telomeres. Annual telomere attrition rate was calculated as (follow-up telomeres-baseline telomeres)/follow-up years, and participants were categorized into the shorten and the lengthen groups. Results showed that telomere lengthening was significantly correlated with decreased systolic blood pressure (SBP) ( For triglycerides and relative mean telomere length of leukocytes, the non-parametric Mann-Whitney U test was used due to their sewed distribution. In this study, values of telomere length were logarithm (Lg) transformed so as to reach a normal distribution for analyzing the relation between telomere attrition and blood pressure lowering. Annual telomere attrition rate was calculated using the formula: (follow-up telomere length-baseline telomere length)/follow-up years. All participants were categorized into two groups, of those who experienced telomere length shortening  were categorized as “shorten group”, and those who experienced an increase of telomere length  as “lengthen group”. Here, according to difference in means of telomere length (Lg-transformed) between the shorten  and the lengthen group , the power would achieve >98% to detect a difference in blood pressure lowering with a two-sided α level of 0.05 in 1,108 participants by using the PASS software program (www.ncss.com).Clinical characteristics of participants were compared between groups by the chi-square test for categorical variables (expressed as numbers [percentages]) and theLinear regression analysis was used to examine the relation between annual telomere attrition rate and baseline telomere length and age. The correlations of annual telomere attrition rate with SBP change (ΔSBP), DBP change (ΔDBP) and pulse pressure change (ΔPP) (ΔPP=ΔSBP-ΔDBP) were examined by multivariate linear regression models which first adjusted for age, gender, medical history, baseline features of smoking status , alcohol intake , BMI, waist-to-hip, the stage of baseline blood pressures, fasting glucose, TC, triglycerides, HDL-C, LDL-C, and telomere length (model I), and then further adjusted for changes in BMI, waist-to-hip, fasting glucose, total cholesterol, triglycerides, HDL-C, and LDL-C from 2014 to 2016 during the follow-up. Generalized linear regression model was used to compare the differences in blood pressure change between the shorten and the lengthen group which adjusted for age, gender and covariates mentioned above. The effects of antihypertensive drug treatment on the correlation of annual telomere attrition rate with blood pressure lowering were examined in further stratified analysis by age and gender.The Cox proportional hazards regression model was used to examine the association between annual telomere attrition rate and cardiovascular outcome events. Person-years of follow-up started from the date of recruitment until the occurring date of cardiovascular outcomes, death, or the end of follow-up , whichever came first. A two-tailed probability value of ≤0.05 was considered significant. Analyses were performed with SPSS Statistics 20.0 .β= -0.14; P<0.001) and follow-up telomere length   with blood samples both at baseline and at follow-up were included in this study for assessing the telomeres change, as shown in the flowchart   who had a decrease of telomere length, and the other is the lengthen group (n=722) who had an increase of telomere length . The cliP<0.001) . Aging wP<0.001) .P<0.05). There were no remarkable differences among the stages of hypertension for other traditional vascular risk factors including BMI, lipid profiles, smoking status, alcohol intake, and medical history of cardiovascular diseases as well as diabetes mellitus. In the further analysis of association between telomere attrition rate and blood pressure lowering, we further adjusted for the stage of baseline blood pressure as a covariate in the multiple linear regression models.Considering that the stage of baseline blood pressures might affect the relation between telomere attrition rate and blood pressure lowering, we also compared the characteristics and annual telomere attrition rate for hypertensive patients of different stages . PatientP=0.03) (P=0.04) (interactionP=0.05 by gender) and patients aged≤60 years (interactionP<0.05 by age)  for the lengthen group and -6.9 mm Hg  for the shorten group (P=0.03) ; the ∆PP(P=0.04) . Next, w by age) . There wβ was -3.28 (P=0.02) for ∆SBP and -2.53 (P=0.02) for ∆PP, respectively  for ∆SBP and 3.6 mm Hg  for ∆PP; for ARB users, the difference was 4.7 mm Hg  for ∆SBP and 3.5 mm Hg  for ∆PP .During a median follow-up of 2.2 (range 1.5-2.4) years, there were 70 cardiovascular events and 40 stroke events. The results showed that there was no significant correlation between annual telomere attrition rate and cardiovascular events, and the antihypertensive drugs also did not affect this correlation  from hypertensive rats and patients with essential hypertension, which may contribute to premature cell senescence . MoreoveIn this study, younger patients (≤60 years) with lengthened telomeres at follow-up had significant blood pressure lowering in ΔSBP, ΔDBP and ΔPP. Elevated blood pressures, considered as clinical markers of large artery stiffness, usually aggravate with aging ,37. The A number of studies have found leukocytes telomere length to be longer in women than in men, and the rate of telomere length shortening was slower in women -45. Men Interestingly, we found that annual telomere attrition rate was associated with the heterogeneity in blood pressure lowering in response to antihypertensive drugs, and telomere lengthening had significantly beneficial effects in ΔSBP and ΔPP among patients treated with CCB or ARB. Antihypertensive drugs may affect cell senescence and telomere length shortening. The CCB has been shown to possess antioxidant properties and reduce the oxidative stress in cardiovascular structures, in which suggested mechanisms are to prevent the inactivation of telomerase and increase eNOS activity during the process of vascular endothelial cell senescence . Data frEmerging evidence has demonstrated that leukocytes telomere length shortening is associated with higher risk of cardiovascular diseases; however, to date, the results of systematic review and meta-analysis are inconsistent in the literatures on the relation between telomeres and cerebrovascular diseases such as stroke, due to the between-studies heterogeneity. Moreover, these meta-analyses are based on observational studies reflecting the measurement of telomere length at a single time point at baseline. For example, Haycock et al reviewed 24 studies involving 43,725 participants and 8,400 patients with cardiovascular disease  and showed that shorter leukocyte telomeres are associated with coronary heart disease, but the association with cerebrovascular disease is less certain . A recenLongitudinal studies demonstrate that accelerated telomere attrition rate might show stronger association with increased risk of mortality from cardiovascular disease in high-risk population, such as older people aged ≥70 years  or patieIn the present study, our data showed that there was no significant relationship between telomere attrition rate and incidence of cardiovascular events in hypertensive patients during the follow-up. This might be explained that all hypertensive participants in our cohort had received the antihypertensive treatment, which would reduce the occurrence of cardiovascular events, and thus, the potential effects of accelerated telomere attrition may be attenuated. However, in our study, the limited follow-up period, a relatively smaller person-years, and a short-term exposure duration of telomere attrition, may affect the association between annual telomere attrition rate and cardiovascular events. Further studies with a larger sample size and long-term follow-up will support to identify the relation of telomere attrition with future cardiovascular risk among the hypertensive patients.P=0.89), respectively, their use may not affect the correlation of telomere attrition rate with antihypertensive drugs.Some potential limitations in this study should be mentioned. First, the measurements were restricted to telomere length in circulating leukocytes which do not necessarily reflect telomere attrition in other cell types such as myocardium, endothelium, or atherosclerotic plaque. However, studies by Wilson et al  and ZhanIn summary, this study for the first time showed an association between annual telomere attrition rate and blood pressure lowering in a prospective, longitudinal cohort of primary hypertensive patients. Telomere lengthening was independently associated with decrease in SBP and pulse pressure during the follow-up period of 2 years, and the differences in ∆SBP and ∆PP were more significant in women and younger patients aged≤60 years. Moreover, in patients using CCB or ARB therapy, those having lengthen telomeres showed a significantly lower level of ∆SBP and ∆PP. Our data supported that telomere attrition may provide new sight on telomere biology in clinical intervention for hypertension. Future randomized controlled studies will be helpful to clarify the role of antihypertensive interventions in telomeres lengthening.www.aginganddisease.org/EN/10.14336/AD.2019.0721.The Supplemenantry data can be found online at:


It is now well established that besides being the most common sustained arrhythmia, atrial fibrillation (AF) is a major healthcare burden. Risk of debilitating stroke is increased in AF patients, but even in the absence of stroke, this population is at heightened risk of cognitive decline, depression, and dementia. The reasons for this are complex, multifactorial, and incompletely understood. One potential contributing mechanism is cerebrovascular dysfunction. Cerebral blood flow is regulated by chemical, metabolic, autoregulatory, neurogenic, and systemic factors. The dysfunction in one or more of these mechanisms may contribute to the elevated risk of cognitive decline and cerebrovascular events in AF. This short review presents the evidence for diminished cerebral blood flow, cerebrovascular carbon dioxide reactivity , cerebral autoregulation, and neurovascular coupling in AF patients when compared to control participants in sinus rhythm. Further work is needed to understand the physiological mechanisms underpinning these observations and their clinical significance in atrial fibrillation patients. Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia characterized by an irregularly irregular cardiac output that leads to disrupted peripheral blood flow kinetics. Recognized as a major healthcare burden , incidenAF is often accompanied by structural heart disease, vascular endothelial damage/dysfunction , and abnCerebral blood flow is guided by the careful interplay of chemical, metabolic, autoregulatory, neurogenic, and systemic factors. The dysfunction in one or more of these mechanisms may contribute to the adverse cerebral events associated with AF. This mini-review will present evidence that AF modifies these aspects of cerebral blood flow regulation and the potential underlying mechanisms will be briefly discussed.n = 31, 69  years, median (interquartile range); 51.0 (12.9) cm s−1, mean (standard deviation)] when compared to healthy controls in sinus rhythm . To assess AF irrespective of cardiac rhythm, comparisons were also made of AF patients diagnosed with paroxysmal (transient episodes that resolve spontaneously within 48 h) vs. persistent (untreated episodes last longer than 7 days) AF. Notably, fibrillating patients  exhibit a lower cerebral perfusion [44.4 (10.9) cm s−1] than non-fibrillating AF patients . CVRCO2 was assessed using the slope of MCA Vm  vs. partial pressure of end-tidal CO2  by two 4-min step-increases in inspired CO2 fraction . Strikingly, CVRCO2 was ~31% lower in AF patients [1.90 (1.13) cm s−1 mmHg−1] compared to healthy  as a “disease” control group (CO2 was documented to be ~34% lower in AF patients compared to hypertension patients  and non-fibrillating AF  patients , and PETCO2 may underestimate arterial CO2 , thereby mitigating the risk of ischemia or hemorrhage by preventing under‐ or over-perfusion, respectively. Cerebral autoregulation functions as a high-pass filter whereby slower changes in perfusion pressure (>0.02 Hz) appear to pass unhindered, but more rapid pressure oscillation (<0.02 Hz) are dampened more effectively .n = 30, 69  years], primary hypertensives , and healthy controls . Cerebral autoregulation was determined using transfer-function analysis of the MCA Vm and blood pressure (finger photoplethysmography) responses to repeated squat-to-stand maneuver. AF patients exhibited greater changes in MCA Vm for a given change in blood pressure  compared to hypertensives [1.13 (1.00–1.45)% mmHg−1] and healthy controls , revealing impaired autoregulation. However, unexpectedly, sub-group comparison between AF patients showed that fibrillating AF patients  were better able to delay blood pressure oscillations from transmitting into brain blood flow  compared to non-fibrillating AF patients , and more effective at damping blood pressure driven changes to absolute measures of cerebral perfusion . However importantly, normalized gain failed to show any group differences between fibrillating [1.39 (1.11–1.80)% mmHg−1] and non-fibrillating  patients.Impaired autoregulation may result from a number of interactive mechanisms, including mechanosensitive myogenic ion channels , neurogeCollectively, these observations suggest that cerebral vasculature in AF patients is less able to buffer blood pressure driven fluctuations in brain blood flow  in comparison with primary hypertensives and healthy individuals in sinus rhythm. The apparently conflicting finding of improved absolute gain in fibrillating AF patients may just reflect fibrillation itself, rather than an improvement in cerebrovascular health during fibrillation.The regional metabolic needs of neuronal activation share a close spatial and temporal fidelity with local blood flow, as such ensuring commensurate functional perfusion within the brain . This phn = 12, 71  years] compared to primary hypertensives  and healthy controls . Beat-to-beat posterior cerebral artery (PCA), MCA Vm , and vascular conductance  responses to repeated visual-stimuli  were spline interpolated and then averages and percentage changes calculated %] compared to hypertensives [6.6 (9.4)%] and healthy controls .A blunted peak PCA conductance was observed in AF [18 (8)%] and hypertensive patients [17 (8)%] compared to healthy controls [26 (9)%; icipants . HoweverThese results indicate reduced neurovascular coupling responses in AF patients; however, the underlying mechanisms remain unclear. Neurovascular coupling is mediated by a complex array of feed-forward and feedback mechanisms,  controls and healthy controls in sinus rhythm. These findings may be important in explaining the severity of ischemic strokes, morbidity, and mortality risk from such events, cognitive decline, and cerebral dysfunction in AF. Further cross-sectional and longitudinal studies are needed to better understand the pathophysiological underpinnings and clinical significance of these findings.RJ drafted the manuscript which was critically revised by GL and JF. All authors contributed to the article and approved the submitted version.GL is a consultant for Bayer/Janssen, BMS/Pfizer, Medtronic, Boehringer Ingelheim, Novartis, Verseon, and Daiichi-Sankyo and speaker for Bayer, BMS/Pfizer, Medtronic, Boehringer Ingelheim, and DaiichiSankyo. No fees are directly received personally. JF has received funding from BMS/Pfizer for an investigator-led and competitively reviewed research project. RJ was previously employed by University of Birmingham as a research fellow on BMS/Pfizer funded, investigator-led and competitively reviewed research project which he managed. This required regular contact with funders for continued project grant support. No fees/funds were directly received personally.


HbA1c, renal parameters , and serum biomarkers were collected at intervals of two months. Tocovid supplementation significantly reduced serum creatinine levels (MD: −4.28 ± 14.92 vs. 9.18 ± 24.96), p = 0.029, and significantly improved eGFR (MD: 1.90 ± 5.76 vs. −3.29 ± 9.24), p = 0.011 after eight months. Subgroup analysis of 37 patients with stage 3 CKD demonstrated persistent renoprotective effects over 12 months; Tocovid improved eGFR (MD: 4.83 ± 6.78 vs. −1.45 ± 9.18), p = 0.022 and serum creatinine (MD: −7.85(20.75) vs. 0.84(26.03), p = 0.042) but not UACR. After six months post washout, there was no improvement in serum creatinine and eGFR. There were no significant changes in the serum biomarkers, TGF-β1 and VEGF-A. Our findings verified the results from the pilot phase study where tocotrienol-rich vitamin E supplementation at two and three months improved kidney function as assessed by serum creatinine and eGFR but not UACR.Diabetic kidney disease (DKD) is a debilitating complication of diabetes, which develops in 40% of the diabetic population and is responsible for up to 50% of end-stage renal disease (ESRD). Tocotrienols have shown to be a potent antioxidant, anti-inflammatory, and antifibrotic agent in animal and clinical studies. This study evaluated the effects of 400 mg tocotrienol-rich vitamin E supplementation daily on 59 DKD patients over a 12-month period. Patients with stage 3 chronic kidney disease (CKD) or positive urine microalbuminuria  were recruited into a randomized, double-blind, placebo-controlled trial. Patients were randomized into either intervention group ( Diabetic kidney disease (DKD) is a debilitating microvascular complication of diabetes progressing to end-stage renal failure (ESRF). To date, the only management available for ESRF is restricted to hemodialysis and renal replacement therapy . Hence, The pathophysiology of DKD is complex, multifactorial, and is not fully elucidated. As a result of chronic hyperglycemia, four major metabolic pathways such as protein kinase C, the polyol pathway, advanced glycation end products (AGEs), and the hexosamine pathway are activated. The overproduction of these end-products and by-products of these pathways results in microvascular and macrovascular complications of diabetes . Specifiα, β, γ, δ), that are naturally found in a number of vegetable oils, wheat germ, barley, and certain types of nuts and grains [Vitamin E tocopherols and tocotrienols have potent antioxidant and anti-inflammatory properties . Both cld grains . Tocotrid grains ,6,7. Thed grains  The MICRd grains . A few sd grains ,10,11. Hd grains .Current perspectives regarding the mechanism of action of tocotrienol on the kidneys were mainly on oxidative and inflammatory stress, but this may not reflect the whole picture. This was supported by our pilot study; while there was a significant improvement in renal function after 3 months of supplementation, no correlation was found between renal function and oxidative stress or inflammatory stress markers. Furthermore, this renal improvement was not related to changes in HbA1c, blood pressure, or BMI ,13.There is growing evidence that transforming growth factor beta 1 (TGF-β1), a fibrogenic cytokine, is crucial in mediating the cascade of events leading to DKD. However, its modulation by tocotrienol in DKD remains unexplored . SiddiquThe primary aim of this study was to determine the effect of high-dose tocotrienol-rich vitamin E on DKD, as assessed by serum creatinine and estimated glomerular filtration rate (eGFR) and urine to albumin ratio (UACR). The secondary aim was to investigate the effects of tocotrienol-rich vitamin E on antifibrotic biomarkers, namely TGF-β1.This is a multicenter, placebo-controlled, double-blind, prospective randomized controlled trial to evaluate the effects of tocotrienol-rich vitamin E on diabetic kidney disease patients. This trial commenced in 2019 and was conducted at two clinical research centers in Malaysia, namely the Clinical Research Centres (CRC) of Monash University Sunway and at Monash University Clinical School Johor Bahru (CSJB). The patients were followed up closely at intervals of two months over a year until March 2020 followed by a six-month washout period until September 2020.Study candidates were recruited from a pool of diabetic patients who are reviewed on a regular basis. Potential patients with renal complications were notified to attend for possible inclusion into the study, where all study-related procedures and blood investigations were taken and sent to a designated national certified pathology laboratory. Prior to every visit, patients were reminded to fast for at least 8 h, omit their morning dose of antidiabetic medications, and avoid strenuous exercise the night before. Premenopausal patients with ongoing menses or those who had fallen ill during the scheduled visits were permitted to reschedule their appointment dates.2) or microalbuminuria, UACR > 20–200 mg/mmol. This criterion of stage 3 CKD is outlined by the KDOQI guidelines. Patients with stage 5 CKD (eGFR < 15 mL/min/1.73 m2) were excluded due to the lack of information on drug safety issues.The inclusion criteria for patients included an age range of 18 to 75 years old with stable glycemic control assessed by HbA1c between 6 and 9% (not more than 10% fluctuation over the last 2 months). Resting blood pressure must be less than 150/90 mmHg. Eligible patients must have chronic kidney disease secondary to T2DM defined by either a reduced eGFR , unstable eye disease, or pregnancy. Patients who had consumed any form of water-soluble supplements such as vitamin B, C, and glutathione, or fat-soluble antioxidants such as vitamin A, D, E, and K during the past month were told to return after one month of washout.The study was carried out in accordance with the Declaration of Helsinki and the study protocol was approved by Monash University Human Research Ethics Committee  (Project number: 12090). This trial was registered in the Australian New Zealand Clinical Trials Registry , with the trial registration number ACTRN12619001568101.Written informed consent was obtained from all patients before any procedures were performed during the visit. A thorough history-taking and physical examination were performed, followed by anthropometric measurements . Blood pressure and pulse were measured thrice to obtain an average reading. Blood and urine samples were collected for HbA1c, fasting blood glucose, renal profile, lipid profile, liver function tests, and full blood count. Urine analysis by dipstick test, urine pregnancy test, and UACR were conducted as well. An electrocardiogram (ECG) was also taken to ensure subject eligibility. Patients that fulfilled the criteria were notified and invited to return for randomization two to four weeks after screening.TM; Hovid Berhad, Ipoh, Malaysia) twice daily, while the control group were given identical looking capsules  twice daily. Tocovid SupraBioTM and the placebo capsules were sponsored by ExcelVite, Malaysia and manufactured by Hovid Pharmaceuticals Berhad, Malaysia.Patients were randomized into either the intervention or control group in a 1:1 ratio by an independent party using a computer-generated random sequence. Stratification was performed according to gender, duration of diabetes, and HbA1c levels. Patients in the intervention group were given 200 mg tocotrienol-rich vitamin E  compared to placebo, assuming a 5% standard deviation (SD) of effect (α = 0.05 and 1 − β = 0.8) and an anticipated dropout rate of 10%. To fulfill these specifications, a total of 58 patients were needed.The power calculations were based on the ability to detect a 30% reduction in UACR in the primary analysis of tocotrienol-rich vitamin E , safety tests (lipid profile and liver profile), measurement of vitamin E levels, and biomarkers. ECG was performed as part of the safety tests. Patients were advised to discontinue participating in the study if they experienced serious adverse events or became pregnant during the course of the study.TM) while drug B was the placebo.As per local guidelines, patients in both groups received the standard of care and were required to be reviewed by their regular physician or endocrinologist, alongside the routine review by a research doctor during the follow-up visits. Any changes in medication during the study were documented. The identity of the study drugs was revealed by ExcelVite at the end of study. Drug A was identified as tocotrienol-rich vitamin E  with a measuring range of 4.3–18.8% with coefficient variances (CV) of less than 5%.During the screening visit and every follow up visit, 18mL of fasting blood samples were collected from each patient. These blood samples were collected in serum-separating tubes (SST) and were centrifuged  at 3600 rpm for 15 min to obtain serum. Sera was transferred into 1mL Eppendorf tubes. A portion of these serum samples were then sent to a national certified pathology lab for analysis of serum creatinine, eGFR, liver function, and lipid profile  on the same day. The coefficient variances for these tests were below 6%. eGFR was calculated using the CKD-EPI formula and presented as milliliters per minute per 1.73 mThe remaining serum samples kept in the Eppendorf tubes were stored in a −80 °C fridge on the same day. To minimize inter-assay variation, serum biomarker analysis was performed at the end of the study, on a batch-to-batch basis. These biomarkers were measured using enzyme-linked immunosorbent assay (ELISA) in duplicates and quantified by calorimetric method. An ELISA plate reader  was used. The ELISA kits include Elabscience E-EL-0162  for serum TGF-β1 and Elabscience E-EL-H0111  for serum VEGF. These kits had intra-assay coefficient variances of 4% and inter-assay coefficient variances of 8%. Blood samples collected in EDTA tubes were centrifuged at 3600 rpm for 15 min to obtain plasma samples. Plasma tocotrienols and tocopherol were measured for all patients using high-performance liquid chromatography  with a fluorescence detector (FLD) based on a method by Che et al.  with modCollected urine samples were sent to the same pathology lab on the same day to assess for UACR. The UACR kit  has a coefficient variance of less than 5% for urine creatinine and less than 6% for microalbuminuria. To obtain an average reading of UACR at baseline, two UACR readings were taken on two different visits (screening and visit 1).® Statistics, Version 26.0, Illinois (USA), IBM; 2016. Two-sided p values were calculated; the probability value of ≤0.05 was used to reject the null hypothesis.All statistical analysis was performed using SPSSt-test for continuous variables, while a Chi-square test was performed for categorical variables.Baseline measurements such as sociodemographics, status of diabetes, and renal parameters between two groups were assessed using an independent t-test. The value of d signifies the magnitude of effect size; values > 0.8 depict a “large” effect size which simply means the results are influential.The mean differences of each parameter were calculated between follow-up visits and baseline visit and recorded as continuous variables. Subsequently, these variables were then evaluated for normality using the Shapiro–Wilk test. Parametric continuous variables were evaluated using an independent t-test, while the Mann–Whitney test was used for non-parametric continuous variables. Descriptive statistics of normally distributed data were reported as mean and standard deviations, and median (interquartile) for non-normally distributed data. Categorical data were presented as percentages. The effect size of the study was analyzed using Cohen’s d method and calculated based on the independent To depict real-life situations, a modified intention-to-treat analysis approach was used in this trial. Missing secondary data lost to follow-up were imputed based on the last observation carried forward method, while missing values at random were imputed using the series mean function in SPSS. This form of imputation is valid, as the proportion of missing data constitutes less than 10% of the total sample size.A total of 160 patients with T2DM were screened over a period of 2 months. Of these, 60 patients met the inclusion criteria and were invited to participate in the clinical trial. Fifty-nine patients turned up for randomization—31 in the intervention group and 28 in the placebo group.At the end of the trial, 9 out of the 59 patients did not return for follow up, 5 of which were unable to attend the last follow up visit due to the COVID-19 pandemic, 3 withdrawn due to adverse events unrelated intervention, and 1 moved/migrated from the study area. However, all 59 patients were retained and analyzed per modified intention-to-treat analysis.A summary of the patient flow diagram is shown in 2. The average systolic blood pressure (SBP) and diastolic blood pressure (DBP) readings were 132/76 mmHg, respectively. Based on the Asian criteria BMI by WHO, the average BMI of the patients was 28.54 kg/m2 which is classified as obese [The baseline characteristics of 59 patients are shown in as obese .2) compared to the placebo. Although UACR levels in the control group were much reduced compared to the intervention group, this change was not significant. There were no significant changes seen in the serum biomarker TGF-β1 nor in VEGF-A. No significant changes were seen in HbA1c levels, blood pressure, urea, and uric acid.At the end of six months, tocotrienol-rich vitamin E supplementation significantly reduced serum creatinine  and increased eGFR (MD = 6.0 mL/min/1.73 m2) and reduction in serum creatinine (MD = 13.4 umol/L) compared to the placebo. However, at twelve months supplementation, there were no changes in these renal parameters. This result suggests that the renoprotective effects of tocotrienol-rich vitamin E (Tocovid SupraBioTM) were only sustained until eight months post supplementation. There was a significant change in urea at twelve months of supplementation when compared to the placebo. There were no significant changes in other parameters, nor in TGF-β1 nor in VEGF-A, which are important markers that mediate DKD when comparing both groups.Based on 2 and stage 3 CKD at baseline were stratified into a subgroup. Based on these selection criteria, 20 patients were in the treatment group and 17 in the placebo group.Patients with a glomerular filtration rate of 30–60 mL/min/1.73 mp = 0.05 *) in this subgroup. There was a 7.23 umol/L decrease in serum creatinine levels in the intervention group, while there was an increment of 5.82 umol/L in the placebo. Moreover, there were no significant changes in blood pressure and HbA1c.2 increase in eGFR in the intervention group.Based on 2 over a year which corresponds to the natural history and progression of DKD. More often, the status of eGFR will be used to assess the progression of DKD. Patients with declined eGFR of ≥2 mL/min/1.73 m2 per year will be identified as progressors and have higher chances to eventually progress to ESRF [ to ESRF .The post washout section of the clinical trial protocol was disrupted by the nationwide implementation of the movement control order (MCO) as a public health measure to contain the COVID-19 pandemic. The MCO was executed from March until September 2020. During this time, patients were kept in contact via telephone to assess how they were doing. At 6 months post washout, which coincided with the temporary lifting of the MCO, patients were called to return to the clinic, where they were reviewed and blood and urine samples were collected for post washout analysis. The table below shows the comparison of renal parameters at 12 months Tocovid supplementation and at 6 months post washout between intervention and placebo groups.Post washout, the intervention group showed a statistically significant reduction in UACR by 19.3 mg/mmol in contrast to the placebo group, which showed an increase in UACR by 4.6 mg/mmol. However, there were no significant differences in terms of HbA1c, serum creatinine, eGFR, and uric acid between the groups.2 [Diabetic kidney disease is clinically diagnosed with the presence of persistent high urine albumin-to-creatinine ratio of ≥30 mg/g and/or sustained declined in eGFR below 60 mL/min/1.73 m2 . Classic2 . However2 . Anyanwa2 . This fi2 . As a reThis clinical trial demonstrated that oral supplementation of tocotrienol-rich vitamin E significantly improved renal function as assessed by serum creatinine and eGFR but not UACR . However2 as they persistently benefited from tocotrienol-rich vitamin E supplementation throughout the 12 months study period although there was no improvement in UACR  were supplemented with tocotrienol-rich vitamin E, they appeared to benefit from a sustained increase in eGFR over one year.Furthermore, it is unlikely that the changes in eGFR are secondary to drug-induced hyperfiltration . InterveOur earlier pilot phase study had reported a sustain improvement in creatinine clearance even 6 to 9 months after the intervention had been withdrawn . FurtherOur study demonstrated no significant changes in serum TGF-β1 levels with tocotrienol-rich vitamin E supplementation . HoweverNevertheless, tocotrienol-rich vitamin E significantly reduced serum creatinine and improved eGFR compared to the placebo, indicating there are different pathways or molecular mediators that were modulated by tocotrienol-rich vitamin E that are yet to be discovered. One potential limitation of the study is the sample size. Based on the power calculations performed, the sample size of 59 patients was sufficient to detect statistically significant changes. Novel biomarkers of diabetic kidney disease should be explored further. Future studies could apply proteomics to study protein expression profiling and targeted protein quantification. This may bridge the knowledge gap to exploring other possible molecular mechanisms that are modulated by tocotrienol-rich vitamin E . Further2). Tocotrienol-rich vitamin E supplementation improved renal function, as assessed by a significant reduction in serum creatinine and significant increment in eGFR when compared to placebo, but not urine albumin excretion. At 6 months post washout, there was no improvement in serum creatinine and eGFR. Given the complexity of tocotrienol and the pathophysiology of diabetic kidney disease, the pathway in which tocotrienols exert on the renal function remains elusive. Further studies are warranted to determine the mechanism of action of tocotrienols.In conclusion, this is the first clinical trial that demonstrated that tocotrienol-rich vitamin E supplementation for 12 months was able to ameliorate the progression of DKD, especially in patients with stage 3 CKD (eGFR 30–60 mL/min/1.73 m


A major complication of hypertension is microvascular damage and capillary rarefaction is a known complication of hypertensive end-organ damage which confers a higher risk of systemic disease such as stroke and cardiovascular events. Our aim was to study the effect of hypertension on the retinal microvasculature using non-invasive optical coherence tomography angiography (OCTA). We performed a case-control study of 94 eyes of 94 participants with systemic hypertension and 46 normal control eyes from the Singapore Chinese Eye Study using a standardized protocol to collect data on past medical history of hypertension, including the number and type of hypertensive medications and assessed mean arterial pressure. Retinal vascular parameters were measured in all eyes using OCTA. In the multivariate analysis adjusting for confounders, compared to controls, eyes of hypertensive patients showed a decrease in the macular vessel density at the level of the superficial  and deep venous plexuses  and an increase in the deep foveal avascular zone. This shows that hypertension is associated with reduced retinal vessel density and an increased foveal avascular zone, especially in the deep venous plexus, as seen on OCTA and there is a potential role in using OCTA as a clinical tool to monitor hypertensive damage and identifying at risk patients A major complication of hypertension is microvascular damage, related partly to abnormal vasomotor tone and increased wall-to-lumen ratio in relation to higher blood pressure5. It has also been suggested that vascular rarefaction may due to either functional alterations such as microvessel constriction resulting in non-perfusion or anatomical alterations resulting in actual non-perfusion and vessel loss4.Hypertension is a major cause of morbidity and mortality globally6. These changes have been shown to confer a higher risk of systemic disease such as stroke and cardiovascular events9. Other studies have also looked at larger retinal vessels (200–300 µm) using color fundus photos and demonstrated a correlation between narrower arteriolar diameter in hypertension13.There have been many previous studies on the association between hypertension and retinal vasculature. Retinal fundus photo changes seen in response to hypertension include classic hypertensive retinopathy signs such as arteriovenous (AV) nicking, generalized or focal arteriolar narrowing, microaneurysms, intraretinal hemorrhages, cotton wool spots and papilloedema14. However, FFA is not suitable for use in large cohorts because it is invasive, requiring intravenous access, and has well-known side effects like nausea, vomiting and in rare instances anaphylactic reactions.The impact of hypertension on capillary microvasculature is less clear. Historically, invasive fundus fluorescein angiography (FFA) was needed to evaluate perfusion and the structure of capillary microvasculature. Previous studies using FFA have shown that in patients with hypertension, there is an increase in perifoveal inter-capillary area and decrease in capillary blood velocity17. OCTA has the advantage over conventional FFA as it can image retinal vasculature in 3-dimensions with higher resolution, delineate the foveal avascular zone (FAZ) more accurately and can produce depth resolved images of the superficial and deep vascular plexi17. There are a few OCTA studies that have been published about retinal microvasculature changes related to systemic hypertension. Lee et al. examined only the superficial capillary plexus and showed that in patients with chronic hypertension and hypertensive retinopathy, there was a reduction in the foveal vessel density, perfusion density, mean ganglion cell-inner plexiform layer(GC-IPL) thickness, central foveal thickness (CFT) and retinal nerve fibre layer thickness (RNFL) and an increase in the FAZ, compared to their normotensive control group. Another study by Chua et al. examining the superficial vascular plexus (SVP) and deep vascular plexus (DVP) showed that in patients with poorly controlled blood pressure (BP) ≥ 140/90 mmHg, higher systolic BP and mean arterial pressure and a lower estimated glomerular filtration rate had a reduced DVP capillary density18.Optical coherence tomography angiography (OCTA) is a new, quick, non-invasive imaging modality that allows both the microvasculature within the retinal and choroidal layers to be studied in large populations of patientsThe aim of our current study was to measure retinal vascular density and perfusion using OCTA in individuals with chronic treated hypertension. We hypothesize that OCTA will show subclinical alterations in retinal microvasculature parameters in both the superficial and deep plexi when compared to normotensive individuals.19. In summary, between 2009–2011, SCES-1 recruited 3353 individuals, aged 40–80 years old. Of these, 2661 (87.7%) participated in the SCES-2. A detailed interviewer-administered questionnaire was carried out to collect relevant information about the participants’ medical history along with a physical exam where blood pressure, height and weight were taken.This was a case-control study nested within participants from the population-based Singapore Chinese Eye Study 2 (SCES-2), the 6-year follow up study from the Singapore Chinese Eye Study 1 (SCES-1). The methodology and recruitment details of the study have been previously reportedFor the purpose of this study, we selected (1) hypertension cases, defined as participants with hypertension alone and no other significant co-morbidities that could affect retinal vasculature and (2) normal controls with no past medical history of hypertension or other systemic disease and did not meet the criteria for hypertension as outlined below in both the SCES-1 and SCES-2 studies.20. The age, sex, logMAR visual acuity (VA), intraocular pressure, spherical equivalent, number and type of antihypertensive medications of the patient were also noted. Classes of hypertensive medication in this cohort included angiotensin-converting-enzyme inhibitors, angiotensin II receptor blockers, calcium channel blockers, beta-adrenoreceptor blocking agents, diuretics and alpha-adrenergic agonists. Patients with any systemic illnesses including diabetic mellitus or medications that might affect the retina or had previous cataract surgery, vitrectomies or any obvious retinal pathology that would confound the analysis as determined by trained graders were excluded.Cases were defined to be hypertensive if their systolic blood pressure (BP) was more than or equal to 140 mmHg or diastolic BP more than or equal to 90 mmHg or there was a self-reported history of hypertension or hypertensive medication usage. BP was taken with an automatic blood pressure monitor . Two readings were taken 5 minutes apart and a third reading was taken if the difference between the first 2 readings was greater than 10 mmHg (systolic) and or 5 mmHg (diastolic)This study adhered to the Declaration of Helsinki. Institutional review board of the Singapore Eye Research Institute approved the ethical aspect of the study and written informed consent was obtained from all eligible participants.22. Built-in software upgrades have improved the signal-to-noise ratio for flow detection and minimized artifacts.OCTA of the macula was subsequently performed on cases and controls. The AngioVue OCTA system  used in this study is a spectral domain OCTA which uses motion contrast and a split-spectrum amplitude-decorrelation angiography (SSADA) algorithm that allows for the depth-resolved visualisation of the vasculature within the retinal layers and the choroidTM software was used to analyze the FAZ, macula flow, foveal flow density, parafoveal flow density and total flow density at each level. These parameters were derived from an en face angiogram of the SVP and DVP  automatically segmented scans into 4 cross sectional layers but for the purpose of this study only superficial and deep vascular plexuses were studied. Using anatomical landmarks, the level of the SVP was from the inner limiting membrane to the inner plexiform layer and the level of the DVP was from the inner nuclear layer to the outer plexiform layer. The AngioVue AngioAnalyticsThree trained graders who were masked to the participant’s characteristics, ensured the images were of suitable quality to be graded, marked out the vascular area of interest for analysis and ensured that automated segmentation was accurate, manually adjusting it if necessary. As the automated measurement of the deep FAZ was noted to be inaccurate in a large number of scans, manual measurements using ImageJ software were performed for all gradable images by a single grader who was masked to hypertensive status. Anatomically the FAZ is defined as the area within the fovea that is devoid of any retinal vessels. On the OCTA images, this was defined as an area in the fovea region that did not have clear distinct flow signals demarcated by the border between an area of significant flow from retinal vasculature and areas with no flow. Eyes that were found to have any form of retinal pathology that might confound evaluation of the macular architecture including, but not limited to, pigment epithelial detachments, epiretinal membranes, vitreomacular traction and cystoid macula oedema were excluded. In addition, the poor quality images with: (1) very poor signal strength, (2) significant artifact that either obscured the vascular area of interest or obscured more than half the area of the image for analysis and (3) scans with segmentation failure that could not be manually corrected. To establish inter-observer reliability, the 3 graders independently graded the automated superficial and deep macular flow of the same 10 randomly selected subset of participants and a masked second grader repeated 20  of the manual deep FAZ measurements. The values collected were analyzed to determine the intraclass correlation coefficient.25. Hence to detect a significant difference in our study with a power of 80% and a level of significance of 5% (two sided), a minimum of 31 patients in each group  is required.Based on a previous study, comparing eyes with chronic hypertension and normal controls, the mean difference of 0.05 mm2 in the superficial FAZ was reported as significant with a maximum standard deviation of 0.7Differences between case and control study patients were tested using independent sample t-tests and Fisher exact tests for continuous and categorical data, respectively. Analysis of covariance (ANCOVA) linear regression models were used to assess the association between hypertensive status and the SVP and DVP vessel flow and flow density, FAZ and parafoveal retinal thickness with odds ratios adjusted for age, sex, intra-ocular pressure, visual acuity, spherical equivalent and mean arterial pressure in multivariate analyses. Linear association between the parafoveal inner retina thickness and other the SVP and DVP vessel flow and density, FAZ were measured using Pearson’s correlation coefficients and partial Pearson’s correlation adjusted for age, sex, intra-ocular pressure, visual acuity, spherical equivalent and mean arterial pressure. Intraclass correlation coefficients (ICC) using a 2 way random effects model were also calculated to determine inter-observer reliability between the 3 graders for the automated measurements and between the 2 graders for the manual deep FAZ measurements. All statistical analyses were performed using R3.3.1 statistical computing language .26.We recruited 117 cases with hypertension and 64 controls. Of these, 23 hypertension cases and 18 controls were excluded due to poor quality images or retinal pathology. This left 94 (80%) patients in the hypertensive group and 46 (72%) in the control group. For the automated measurements, calculated ICC showed good inter-observer reliability with an ICC of 0.99  and 0.94  for the superficial and deep macular flow respectively. Similarly for the manual deep FAZ measurements, there was good inter-observer reliability with an ICC of 0.92 The clinical characteristics of cases and controls are shown in Table In the univariate analysis as shown in Table 30. Thus we adjusted for age, sex, intraocular pressure (IOP), logMAR visual acuity, spherical equivalent and MAP. The multivariate analysis . However, of note in our study, outer retinal changes, which may show better correlation to visual function were not specifically graded. More studies need to be done to establish the effect of retinal microvascular changes seen in hypertensive eyes and its impact on the outer retina, photoreceptors and ultimately visual function. Similar to a previous study by Kong et al. on OCT findings in hypertensive eyes, a decrease in peri-foveal macular thickness was observed in the univariate analysis however this was no longer significant in the between the hypertensive and control groups in our multivariate analysis56. In contrast to another cross-sectional cohort study by Lee et al. which found a reduction in the foveal vessel density, perfusion density, mean GC-IPL thickness, CFT and RNFL and an increase in the FAZ at the level of the SVP in the hypertensive group compared to controls, our paper did not find any significant differences between the 2 groups at the level of the SVP. However, their paper may not be comparable to ours because no data on the DVP was presented in their paper and the OCTA macular scan area used was 3 × 3 mm on the Zeiss Angioplex System compared to 6 × 6 mm scans Optovue Angiovue scans used in our study25. Also, the definition of hypertension differed in their study with their 2 comparative groups having either (1) chronic hypertension >10 years and (2) relieved grade IV hypertensive retinopathy for at least 1 year. They also did not adjust their analysis for variables such as age, sex and refractive error and also did not take into account anti-hypertensive medication treatment. Our findings were similar to the study done by Chua et al., in that both our univariate analysis’ found a decrease in the deep capillary density. However, the difference is that in our study after adjusting for IOP, visual acuity, spherical equivalent and MAP in the multivariate analysis, there was no longer a significant difference in this parameter. In addition, our study also looked at the macula flow and FAZ in the SVP and DVP which was not analyzed in the study by Chua et al.Outer retinal losses have been reported previously shown to be associated a with an increase FAZ17. Apart from blood pressure reduction, targeting the microcirculation itself could be beneficial in preventing or reducing end organ damage and thus reducing morbidity and mortality4. Other current methods of microvasculature imaging available for other organs such as nailbed capiloroscopy, contrast based cardiovascular angiography or cerebrovascular angiography have their limitations and are associated with higher risks. So far no other widely available imaging modality has been able to measure microvasculature with such ease, reproducibility and resolution compared to OCTA of the retina57. Hence, studying changes in the microvasculature on OCTA in relation to systemic vascular disease may have the potential to risk stratify, detect early disease, monitor disease progression and detect improvements in vasculature rarefaction in response to treatment.Hence, OCTA allows for a non-invasive, efficient and increasingly widely available method of imaging method the retinal microvasculature with advantages compared to conventional injectable dye angiography, which is unsuitable for use in large cohort studies and for consecutive follow up visits59. However, there are a few limitations to this study. Firstly, hypertension was defined as a patient reported history of hypertension, the use of anti-hypertensives or a single setting reading of increased BP, ambulatory blood pressure was not measured in this study and hence a small number of patients who may have hypertension could have been missed and blood pressure control was not assessed. Secondly, our changes in retinal vasculature was not correlated with other peripheral microvasculature parameters, renal function, body mass index or other end organ damage measurements. Our current OCTA imaging depends on motion contrast from blood flow to visualize the retinal vasculature, hence vessels with flow below the detectable threshold may not be able to be imaged60. We acknowledge that our delimited FAZ may contain some background speckles of flow but this was assumed to be insignificant background noise artifact. In addition, the image artifacts seen on OCTA that obstruct an accurate analysis of the microvasculature had to be excluded, however these only involved a small proportion of the overall cohort and is unlikely to significantly affect our results. Also, our cohort only included participants who were ethnically Chinese and these results may not be applicable to other races.Our study strengths are that this study, to our knowledge, is the largest cohort of both hypertensive and control eyes with standardized imaging protocols and automated quantitative analysis of retinal microvascular parameters which have previously proven to have good reproducibility as well as good inter-observer agreement between graders44.While we noted the number and classes of hypertensive agents, we did not do any subgroup analysis correlating type and number of medications with OCTA parameters due to insufficient numbers. This is a possible area for future research as improvement in vascular rarefaction has been reported with the use of various anti-hypertensive agents and it would be interesting to note if there were differences in improvement in rarefaction between classes of anti-hypertensivesIn conclusion, we have shown that there is an association between systemic hypertension and decreased retinal flow and increased FAZ area and perimeter in the DVP seen on OCTA. However, further studies are needed verify these findings and to study its potential clinical implications as a clinical tool for monitoring hypertensive end organ damage and for risk stratification of normotensive patients.Supplementary Information.Supplementary Information 2.Supplementary Information 3.


Introduction: There is substantial clinical evidence that monotherapy with beta-blockers are less effective in reducing blood pressure among hypertensive Black patients compared to Whites. The highly selective beta-1 agents like nebivolol and bisoprolol have, however, been reported to be effective in reducing blood pressure in African Americans. However, results in African Americans cannot be extrapolated to native Africans because of genetic admixture and gene-environment interaction. There is, therefore, the need for us to generate data that are applicable to Africans residing in sub-Saharan Africa. We therefore decided to evaluate the efficacy and tolerability of highly selective beta-1 agent nebivolol in hypertensive Black patients residing in sub-Saharan Africa.Materials and Methods: The nebivolol study was a multicenter, prospective, observational program among hypertensive patients with 4- and 8-week follow up which was conducted in 5 cities in Nigeria of Abuja, Calabar, Enugu, Oghara, and Port Harcourt. Dosages of nebivolol used in keeping with local prescribing information were 5 and 10 mg once daily each. The effectiveness of treatment was assessed by change from baseline in mean office systolic and diastolic blood pressures, and the proportion of patients achieving the therapeutic goal of <140/90 mmHg. Safety and tolerability of this medication were also assessed.Results: We report the results of the 140 patients studied. The mean age and body mass index were 46.9 ± 7.3 years and 22.3 ± 5.8 kg/m2, respectively, and 57.1% were female. Nebivolol reduced SBP and DBP by 7.6 and 6.6 mmHg, respectively, in 4 weeks, and by 11.1 and 8.0 mm Hg, respectively, in 8 weeks. Blood pressure control was achieved in 54.8% of the patients in 4 weeks and increased to 60.4% in 8 weeks. There was no change in metabolic profile between randomization and at 8 weeks, and erectile dysfunction occurred in 1.3% of the study population.Conclusions: Nebivolol 5 and 10 mg appear efficacious in Nigerian Africans with no negative metabolic effect and minimal side effect profile.Clinical Trial Registration:www.ClinicalTrials.gov, Study Identification: NCT 03598673. It was conducted as a non-interventional study, and therefore study-specific patient visits, tests and monitoring were not imposed, and only data originating from routine clinical practice were collected. Therapy was prescribed according to clinician preference and on the prescribing pattern in Nigeria.Male or female patients aged 30–60 years of age with sitting office systolic BP of 140 mmHg or above and <160 mmHg, and diastolic BP of 90 mm Hg and above but less 100 mmHg and on no antihypertensive treatment for a minimum of six (6) months were recruited into the study. Patients with congestive heart failure, renal failure, coronary heart disease including chronic stable angina, myocardial infarction or acute coronary syndrome, stroke or transient ischemic attack, suspected secondary hypertension, concomitant illness, physical or mental impairment that could interfere with the effective conduct of the study and those who are pregnant or of child-bearing age who are not taking reliable contraception were excluded from the study.The primary outcome measure was a change in office BP value from baseline to 2 months which was calculated as the difference between the mean office BP at randomization and at the end of follow up.Other outcome measures include: the proportion of patients who achieved BP <140 mmHg systolic and <90 mmHg diastolic in 2 months, the proportion of patients who have adverse events, change in plasma fasting blood glucose and fasting lipid profile from baseline to 2 months and proportion of male patients who complain of erectile dysfunction at both the fourth and eighth weeks.Patients were commenced on 5 mg of nebivolol at a starting dose of 5 mg once daily if the BP is not controlled (140/90 mmHg or above) after 4 weeks, the dose is doubled to 10 mg once daily.Office BP was measured after five (5) min of rest in the sitting position using semi-automated blood pressure devices (semi-automated blood pressure monitors (Omron MIT5 Connect). Three BP measurements were taken, and the last two values were used for the calculation of the mean. Patients had their fasting blood glucose and lipid profile assessed at baseline and the 8th week.Each patient was followed up for 2 months at the fourth and eighth weeks. Patients were still followed up till the 8th week even when study medication was discontinued. In the case of discontinuation of study medications, the reason(s) for discontinuation was recorded and reported. The study was completed when the last enrolled patient completed two (2) months of observation.The study was conducted according to the guidelines laid down by the International Conference on Harmonization for Good Clinical Practice . All invp-values was used either as an aid to evaluating a specific difference of interest or as a “flagging” device applied to a large number of safety and tolerability—efficacy variables to highlight differences worth further attention. This was particularly useful for laboratory data which were subjected to quantitative analysis. Patients enrolled in the program with at least one follow-up visit, or a documented adverse event were considered analyzed.Due to the observational nature of the study, descriptive statistical methods were used and supplemented by calculation of confidence intervals wherever this aids interpretation. The calculation of Systolic and diastolic BP were modeled using linear mixed models fitted with restricted maximum-likelihood method which include adjustment for baseline BP, age (<45 or ≥45 years), gender, smoking status, body mass index (BMI) and we considered subject and time as random effects. Site level was not considered in the random effect because it was not adding significant value to the model, and in addition, some sites had very small sample which could bias our results if included in the model.All other patients were included in the evaluation even if they had partially missing data. We investigated existence of interaction within model predictors, and in case of significant interaction between statistically significant predictors in the final model, we explored specific difference between means of blood pressure amongst the groups using pairwise test with Tukey adjustment.Moreover, we presented the adjusted mean reductions in SBP and DBP in interesting subgroups.Firstly, we constructed all models without accounting for missing data and secondly performed a sensitivity analysis, in which we performed several logistic regressions with binary variable (missing and not missing) and added other variables in the models as covariates in order to eliminate the “Missing Completely at Random” hypothesis. With the assumption that all missing data that were missing at random (MAR), we performed multiple-imputation analysis using chained equations. We generated 20 amputated data sets with a maximum of 20 iterations. Variables included in the imputation model were systolic BP, diastolic BP, age, gender, weight, height, BMI, smoking status, site including individuals. All analyses were performed with R Software 4.0.0 .The Trial Steering Committee consisted of the Principal Investigator, the site Investigators, the Statistician, and the representative of Micronova Pharmaceutical team. This committee was chaired by the Principal Investigator. The various site investigators were responsible for entering patients' data into paper case report forms which were subsequently entered into Microsoft data spreadsheet.One hundred and eighty-three (183) patients were screened, but one hundred and forty (140) met the inclusion criteria and these consist of sixty-four (64) patients from University of Abuja Teaching Hospital, thirty (30) patients from Delta State University Teaching Hospital, twelve (12) patients from University of Nigeria Teaching Hospital, seventeen (17) patients from University of Port Harcourt Teaching Hospital and another seventeen (17) patients from University of Calabar Teaching Hospital. The first patient was recruited 2nd February 2018 at the University of Abuja Teaching Hospital, and the last patient was followed up 3rd September 2019 at the University of Calabar Teaching Hospital.2 ± 13.6 with 38 (28.4%) overweight and 40 (29.9%) obese, with a female predominance (57.1%). Duration of hypertension was <5 years in majority of them (81.4%), and the mean systolic and diastolic blood pressures were at 148.2 mmHg ± 6.8 and 94.8 mmHg ± 5.3, respectively.p <0.0001) at 4 weeks and by 4.8 beats per minute  at 8 weeks. There was however no significant difference in pulse rate between the fourth and 8 weeks.Pulse rate was reduced by 4.02 beats per minute , overweight and gender . Therefore, the effect of BMI on systolic BP depend significantly on gender, but gender alone was not significantly associated with systolic BP (p = 0.087). A post-hoc test, exploring specific difference between male with normal weight, male with overweight, male with obesity and respective female subgroups revealed that the changes in SBP in male with overweight compared to female with overweight, in male with obesity compared to female with obesity and in male with obesity compared to female with overweight were significantly different. The adjusted mean reduction of SBP from baseline to 8 weeks follow-up was not statistically different in male regardless of the BMI status and these results were similar female regardless of their BMI status (p = 0.044) but there was no interaction with gender  or age above 45 years . The adjusted mean reduction of SBP from baseline to 8 weeks follow-up for male was 8.2 mmHg  and 8.1 mmHg  for female (In a linear mixed model as shown in I status . Similarr female .A sensitivity analysis that included all patients who were treated in all study sites after multiple imputations confirmed these patterns in treatment effects on systolic BP and diastolic BP.This study has demonstrated that nebivolol which is a cardio-selective beta-1 blocker with vasodilating effects given once-daily as 5 or 10 mg effectively reduced both systolic and diastolic blood pressure in Blacks with stage I hypertension who are residing in sub-Saharan Africa. In addition, 5 or 10 mg nebivolol resulted in response rates of >55%. These findings are of clinical importance, especially because of the excessive burden of hypertension and its complications and the low rates of blood pressure control in sub-Saharan Africa , 16. TheThe findings in this study are also very important as Physicians both in sub-Saharan Africa, Europe and the Americas are less likely to prescribe beta-blockers as monotherapy to Blacks compared to white patients in the general community , 20. ThiAt 8 weeks, nebivolol reduced systolic blood pressure by 11.1 mmHg and diastolic blood pressure by 8.03 mmHg from baseline similar to values recorded in earlier studies in African Americans , 23. ThiApart from being at higher risk for cardiovascular and renal disease, it has been shown that black patients also generally report poorer antihypertensive medication adherence compared with the general hypertensive population , 32, andUnlike traditional beta-blockers, nebivolol had no negative metabolic impact on glucose and lipid profile as demonstrated in this study. LDL cholesterol was however found to be significantly lower at 8 weeks in our study. The mechanism of this reduction is not clear and has to be explored further.One important strength of this trial is that the results were adjusted to account for the heterogeneity of the study population and potential confounding variables such as age, sex, diabetes, and obesity, which are known to determine BP response to treatment within racial groups .The main limitations of this study include the apparently small sample size and the low proportion of diabetic patients who were enrolled in the study, making the extrapolation of results to this population rather difficult. In addition, plasma renin activity and adrenaline levels were not measured making it difficult to fully define the mechanism of action of nebivolol in this population.In conclusion, nebivolol monotherapy is safe and effective in lowering BP in hypertensive black patients residing in sub-Saharan Africa. Nebivolol also has favorable metabolic effect in this population. Studies in a larger population is however needed to confirm these findings.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by University of Abuja Teaching Hospital, Human Ethics Committee. The patients/participants provided their written informed consent to participate in this study.DO was involved in the design and the management of the study and drafted the initial manuscript. BA was involved in the analyzing the data and critical review of the manuscript. LS, EU, EE, CAl, HE, CAg, UN, and VA were involved in data collection and critical review of the manuscript. All authors contributed to the article and approved the submitted version.BA was employed by the company Holo Healthcare limited, Nairobi. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


No relationship was found, with a multivariate analysis, between OCTA parameters and kidney function and neuroradiological signs of central nervous system involvement. OCTA could be a new vascular biomarker in FD, revealing a strong correlation between retinal capillary damage and myocardial impairment, possibly preceding both renal dysfunction and cerebrovascular involvement.In this study, we evaluated the possible relationship between the changes in retinal vessel density (VD) by optical coherence tomography angiography (OCTA) and the vascular alterations involving renal, cardiovascular and central nervous systems in patients affected by Fabry disease (FD). In 50 FD patients, the retinal superficial capillary plexus (SCP) and deep capillary plexus (DCP) in macular region were evaluated by OCTA examination. The patients also underwent a brain magnetic resonance imaging scan, renal and echocardiographic examinations with quantification of systolic pulmonary arterial pressure (PAPs) and left atrial volume index (LAVi). The VD of SCP and DCP was inversely related with E/e’ ratio, LAVi, interventricular septal thickness, global longitudinal strain (GLS) and PAPs ( Fabry disease (FD) is a rare X-linked lysosomal storage disorder caused by a deficit in the enzyme α-galactosidase A, with the progressive accumulation of globotriaosylceramide Gb3) in different cells, such as vascular endothelium, smooth muscle cells, renal cells, cardiomyocytes, and neuronal cells ,2,3,4,5. in diffeThe main causes of vascular abnormalities in FD have been considered the endothelial damage due to Gb3 deposition and the hypertrophy of the medium-caliber arteries due to increased intima-media thickness and vascular smooth muscle cell proliferation .Independently from its pathophysiology, this vascular involvement leads to a progressive severe multiorgan dysfunction, as represented by cardiac involvement (with arrhythmias and left ventricular (LV) hypertrophy), central nervous system (CNS) involvement (mainly represented by the occurrence of cerebrovascular events) and renal dysfunction (causing proteinuria and impairment in glomerular filtration rate—GFR); all factors ultimately leading to premature death ,7,8.Being a systemic disorder, FD also affects the ocular system. Its involvement is mainly represented by the appearance of cornea verticillata, lens opacity and vascular abnormalities. With particular reference to the latter, the pathological deposition of Gb3 in both the retinal vascular endothelial cells and in the tunica media of small vessels is known to induce a decreased resistance to hydrostatic pressure, as well as an impaired vascular remodeling, leading to increased vascular tortuosity, aneurysmal dilatation and occlusive events ,10,11,12The introduction of a non-invasive imaging techniques, such as optical coherence tomography angiography (OCTA), has allowed for a detailed and quantitative analysis of the retinal vascular networks, identifying microvascular changes in different retinal disorders ,14.Although previous OCTA studies have demonstrated significant alterations of the vessel density of superficial capillary plexus (SCP) and deep capillary plexus (DCP) in FD patients, there is currently no information about the possible association with other biomarkers of different organ involvement ,16,17.The aim of this observational study was to correlate OCTA parameters with other imaging biomarkers, to evaluate a possible relationship between the retinal perfusion changes and the vascular alterations involving different systems  in FD patients at diagnosis.2 and/or proteinuria >1 gr/die), and three subjects were excluded because of overt diastolic heart failure, leading to a final number of 50 FD patients were included in this study .Data from 59 FD patients were retrospectively analyzed in subjects evaluated from July 2017 to May 2018. Inclusion criteria were: genetically confirmed diagnosis of FD, 18 years of age, naïve for FD treatment. Exclusion criteria were: evidence of ocular and systemic diseases unrelated to Fabry disease, current or previous macular and retinal vascular diseases, diagnosis of glaucoma, congenital eye disease, high myopia (>6 dioptres), significant lens opacification and reduced-quality OCTA images. FD patients with advanced organ involvement were excluded from the study for possible confounding interaction. In particular, six patients were excluded because of significant renal impairment  according to the Early Treatment of Diabetic Retinopathy Study (ETDRS), slit-lamp biomicroscopy, intraocular pressure measurement, fundus examination, and OCTA.The study was approved by the Institutional Review Board of the University of Naples “Federico II” (protocol number: 109/05) and all investigations adhered to the tenets of the Declaration of Helsinki. Signed informed consent was obtained from each patient.OCTA was performed by the Optovue Angiovue System  that is based on a split-spectrum amplitude de-correlation algorithm (SSADA) and which uses blood flow as intrinsic contrast [The evaluation of the macular capillary network was performed in a 6 × 6 mm scan centered on the fovea. The OCTA software automatically analyzed the macular region divided into whole image, fovea and parafovea in each vascular network of the retina: SCP and DCP, according to the ETDRS classification of diabetic retinopathy.The AngioAnalytics™ software automatically calculated the vessel density that represents the percentage area occupied by the vessels in the analyzed region .Images with a signal strength index less than 40 or residual motion artefacts, incorrect segmentation or low centration and focus were excluded.Within 2 weeks of the ophthalmological evaluation, patients underwent two-dimensional transthoracic echocardiography including determination of LV global longitudinal strain (GLS) with the use of commercially available equipment  equipped with a 2.5 MHz phased-array transducer and provided with a software-based beamforming algorithm. Blood pressure and heart rate were measured at the end of the echo exam. Cardiac chambers quantitative analysis was performed in agreement with 2015 recommendations . LV massSystolic pulmonary artery pressure (PAPs) was computed by measuring the tricuspid regurgitation (TR) peak velocity, summing an estimate of right atrial pressure (RAP) referred to the size and respiratory reactivity of the inferior vena cava (IVC): (a) normal RAP (≈5 mmHg); normal IVC size (IVC diameter < 2.1 cm) with normal inspiratory collapse (>50% decrease in IVC diameter); (b) RAP ≈ 10 mmHg: dilated IVC (diameter > 2.1 cm) or <50% collapse; (c) RAP ≈ 15 mmHg: both dilated IVC and <50% collapse; (d) RAP ≈ 20 mmHg: dilated IVC without visible collapse, as recommended .All statistical data were processed using standard statistical software SPSS . Continuous variables were confirmed for normal distribution by the Kolmogorov–Smirnov test and expressed as mean values ± standard deviation (SD). Differences between the two groups in continuous variables were analyzed using independent t-test for normal distribution and Mann–Whitney U test for non-normal distribution.p < 0.05.From the enrolled subject, one eye was randomly selected for the subsequent analysis, leaving a total number of 50 eyes. Pearson’s correlation was used to evaluate univariate correlates of a given variable. Multivariable linear regression analyses were performed to examine the independent correlates between superficial capillary plexus (SCP) and of the deep capillary plexus (DCP) values and biomarkers of other systems involvement. The null hypothesis was rejected at Demographic data and clinical parameters of the FD are shown in Most patients had normal renal function and proteinuria levels, as shown in All patients were free from cardiac symptoms/signs. LV standard echocardiographic and STE assessment are reported in p < 0.05).In the pooled FD population, the vessel density of SCP and DCP in whole image was inversely related with E/e’ ratio, LAVi, interventricular septal thickness, GLS and PAPs . These cp < 0.001).Patients with neuroradiological signs of CNS involvement (n = 6/11.5%) showed lower vessel density of SCP compared to those without  and proteinuria .No relationships were found between both SCP and DCP parameters and markers of kidney function such as GFR  and DCP in whole image   .In this study, we found that the retinal microvascular alterations, as evaluated by a non-invasive technique such as the OCTA, correlate with microvasculature damages in other organs, suggesting a possible role of this measure as an early biomarker of tissue damage in FD.Indeed, when assessing correlations between OCTA values and other systemic findings, we found a significant negative correlation between the reduced retinal vascular networks and the increased PAPs, LAVi and E/e’ ratio. These echocardiographic parameters are known to represent sensible biomarkers of LV diastolic dysfunction and LV diastolic filling pressure, an early index of myocardial damage ,27. IndeOn the other hand, the lack of correlation of retinal vessel density of SCP and DCP with parameters of kidney function is, although disappointing, not surprising. Indeed, renal vessel disease is a recognized key feature of FD-related nephropathy. However, renal involvement is characterized by the accumulation of Gb3 in virtually all cell types, including endothelial and epithelial glomerular cells, podocytes and tubular cells. Interstitial fibrosis with interstitial infiltrate of inflammatory cells is known to play a role in kidney damage development. Tøndel et al. , by perfSimilarly, the overall absence of significant correlation between the different neuroradiological parameters here investigated and OCTA values can be also interpreted as a further indication of the role of the latter as an early biomarker of the disease. Indeed, the presence of neuroradiological signs suggestive of cerebrovascular involvement, either WMH, stroke or microbleeds, are known to be irreversible manifestations of macrovascular CNS damage, occurring at a late phase of the disease . On the These alterations of the retinal vessel density of SCP and DCP at OCTA have been hypothesized to reflect vessel walls remodeling due to Gb3 accumulation in smooth muscle and endothelial cells . In lineThe retinal and cerebral small vessels share similar embryologic origins, anatomical features, and physiological properties; therefore, the retinal vessels may serve as a research model to detect subclinical cerebrovascular changes associated with FD ,44,45,46The main limitations of this study are represented by the small sample size and the absence of patients with advanced FD stages.Although characterized by these limitations, our study shows the presence of a correlation between the retinal perfusion values and echocardiographic parameters of diastolic dysfunction in FD patients, while other vascular parameters known to be late findings in this condition showed no correlation with OCTA values. This parallel impairment of both vessel density in the macular region and myocardial function supports the hypothesis that OCTA could represent a valid biomarker to predict the early cardiovascular damages in FD. This technique could identify patients with subclinical changes, allowing early interventions and prevent the development of more severe clinical consequences.