Background: Although brachial-ankle pulse wave velocity (baPWV) is simple and convenient, its usefulness as an initial screening test for hypertensive patients is not well-known. This study aimed to investigate the association of baPWV with left ventricular (LV) geometry and diastolic function in treatment-naive hypertensive patients.Methods: A total of 202 untreated hypertensive patients  without documented cardiovascular diseases were prospectively enrolled. Both baPWV and transthoracic echocardiography were performed on the same day before antihypertensive treatment.Results: In multiple linear regression analysis after adjustment for potential confounders, baPWV had significant correlations with structural measurements of LV including relative wall thickness  and LV mass index , and four diastolic parameters including septal e′ velocity , E/e′ , left atrial volume index , and maximal velocity of tricuspid regurgitation . The baPWV was significantly increased in patients with LV hypertrophy, abnormal LV remodeling, or diastolic dysfunction, compared to those without . In the receiver operating characteristic curve analysis, the discriminant ability of baPWV in predicting LV hypertrophy and diastolic dysfunction had an area under the curve of 0.646  and 0.734 , respectively.Conclusion: baPWV was associated with parameters of LV remodeling and diastolic function in untreated hypertensive patients. The baPWV could be a useful screening tool for the early detection of adverse cardiac features among untreated hypertensive patients. Increased arterial stiffness reflects the vascular damage and arteriosclerosis , 2, and This prospective cohort study was conducted at a general hospital in a big city . Patients who visited or were referred to the outpatient clinic to evaluate hypertension for the first time and not previously diagnosed with hypertension nor receiving antihypertensive medications were consecutively recruited between December 2016 and October 2020. The diagnosis of hypertension was made when the systolic/diastolic blood pressure of ≥140/90 mmHg was confirmed at least twice in the office. The office blood pressure measurement was performed by a single experienced cardiologist (H.L.K) using the oscillometric method on the right upper arm. If the diagnosis of hypertension was not determined through the office blood pressure measurements, 24-h ambulatory blood pressure monitoring was additionally performed with the diagnostic cutoff of systolic/diastolic blood pressure of ≥135/85 mmHg . Hyperte2). Obesity was defined as body mass index ≥25 kg/m2 according to the Asian-Pacific obesity criteria. Blood pressure and heart rate were measured after 5 min of rest. Mean arterial pressure was calculated by the following equation: (1 × systolic blood pressure + 2 × diastolic blood pressure)/3. Pulse pressure was calculated as the systolic blood pressure minus the diastolic blood pressure. Diabetes mellitus was defined by either a previous diagnosis of diabetes mellitus, the current use of oral antihyperglycemic agents or insulin, or fasting glucose ≥126 mg/ml or glycated hemoglobin ≥6.5%. Dyslipidemia was defined by one of the following: a previous diagnosis of dyslipidemia, lipid-lowering medications, or low-density lipoprotein cholesterol of ≥160 mg/dl. After 12 h or more of overnight fasting, we obtained venous blood levels of white blood cell, hemoglobin, glucose, glycated hemoglobin, creatinine, total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, triglyceride, and C-reactive protein. Glomerular filtration rate was calculated using the Modification of Diet in Renal Disease formula.Patients' clinical data were collected at the time of recruitment. Body mass index was calculated as the ratio of weight in kilograms divided by the height in meters squared  , 13, 15.2 in men and >95 g/m2 in women, which was further categorized as concentric  and eccentric  LV hypertrophy (2). A diagnosis of LV diastolic dysfunction was made if more than half of the criteria were met according to the 2016 American Society of Echocardiography/European Association of Cardiovascular Imaging guidelines  curves analysis was performed to evaluate the discriminant ability of baPWV for predicting LV hypertrophy and diastolic dysfunction. The DeLong test was used to compare areas under the curve (AUC) of ROC. A two-tailed P-value of < 0.05 was considered statistically significant. All analyses were performed using R .Categorical variables are presented as frequencies (percentages) and continuous variables as mean ± standard deviation or median with interquartile range (IQR). The differences between the groups were analyzed using the χOur study cohort comprised 202 patients diagnosed with hypertension, none of whom received previous antihypertensive medications. The baseline characteristics of these patients are summarized in Parameters of transthoracic echocardiography and baPWV are demonstrated in r = 0.261, P < 0.001) and LV mass index . The baPWV also had significant correlations with all four diastolic parameters including septal e′ velocity , E/e′ , left atrial volume index , and maximal velocity of tricuspid regurgitation . Correlation plots between baPWV and these LV geometry and diastolic parameters are demonstrated in P < 0.05 for each in both model 1 and model 2) (The result of simple correlations of baPWV with the indices of LV geometry and diastolic function are summarized in model 2) .P = 0.004]. When further stratified into four groups of LV hypertrophic remodeling patterns, the group of patients with concentric remodeling as well as that with concentric and eccentric LV hypertrophy had elevated baPWV compared to patients with normal LV (P = 0.009). Patients with diastolic dysfunction also had higher baPWV values compared to those without .We further examined whether arterial stiffness is associated with adverse LV structure and diastolic dysfunction . PatientP = 0.004], with 67.1% sensitivity and 59.8% specificity for a best cutoff value of 1,719 cm/s (P < 0.001), which had the cutoff value of 1,617 cm/s with 92.0% sensitivity and 44.1% specificity (P = 0.975). The baPWV had a higher AUC for predicting diastolic dysfunction compared to that of the pulse pressure, although not statistically significant .In the ROC analysis, the AUC of baPWV for the prediction of LV hypertrophy was 0.623 [95% confidence interval (CI), 0.544–0.703, 719 cm/s . The AUCcificity . The disP = 0.001), with more frequent LV hypertrophy and elevated E/e' (P < 0.001 for each) (P = 0.056).In our cohort, women were older than men  . Women hThe simple correlations between baPWV and the parameters of LV geometry and diastolic function were significant in both sex , as in tThis study demonstrated that the baPWV had significant associations with relative wall thickness, LV mass index, and four diastolic parameters in untreated hypertensive patients, even after controlling for multiple confounding factors. The baPWV was significantly elevated with patients with LV hypertrophy, abnormal LV remodeling, and diastolic dysfunction and also showed fair discrimination ability, especially for predicting diastolic dysfunction. The result of this study provides additional evidence for a significant contribution of arterial stiffness to LV remodeling and diastolic function. Furthermore, our results highlight the clinical value of baPWV in risk stratification and an initial screening test for newly diagnosed hypertensive patients.Hypertension is highly prevalent and a powerful contributor to fatal cardiovascular events. The key step in managing hypertensive patients is assessing the degree and extent of hypertensive-mediated organ damage (HMOD), which are the subclinical structural and functional end-organ changes under the long-standing exposure to high blood pressure , 23, 25.Multiple lines of evidence suggest that PWV has substantial value in predicting adverse cardiovascular events. A meta-analysis of 17,635 subjects involving the general population or patients with cardiovascular risk factors demonstrated that PWV is significantly associated with the incidence of coronary heart disease, stroke, and cardiovascular mortality, with an approximately 100 cm/s increase in cfPWV corresponding to a 7% increased risk . In addiHowever, the routine use of PWV for estimating the extent of organ damage in hypertensive patients is not well-established. While the European guideline suggested cfPWV as an optional assessment for HMOD , the AmeOur study demonstrated that baPWV is significantly associated with the indices of LV remodeling and diastolic parameters in treatment-naïve hypertensive patients. Although many studies investigated the association of baPWV with cardiac damages previously, the majority of them included patients who are already treated with antihypertensive medications or a mixed population with and without hypertension –19, makiChronic pressure overload in hypertension induces various remodeling of the heart . The priSeveral mechanisms underlying the associations of arterial stiffness with LV remodeling and diastolic function have been suggested. The most plausible explanation is that stiffened arteries reflect the pressure pulse wave more rapidly backward to LV, resulting in increased afterload of LV –16, 43. This study has some limitations. First, this study could not provide casual relationships of baPWV with LV geometry and diastolic parameters due to its cross-sectional design. The impact of increased baPWV on the myocardial damages for the long-term follow up period could be explored in future studies with the longitudinal design. In addition, since the patients enrolled were all Koreans, it should be further investigated whether this result could be generalized to other ethnicities.In treatment-naive hypertensive patients, baPWV is significantly associated with LV geometry and diastolic dysfunction. Because the measurement of baPWV is very simple, it may be useful in the initial examination of hypertensive patients.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 Boramae Medical Center, Seoul; IRB number: 26-2017-22. The patients/participants provided their written informed consent to participate in this study.SK and MI performed the statistical analysis. SK drafted the manuscript. H-LK, MI, W-HL, J-BS, S-HK, J-HZ, and M-AK reviewed/edited the manuscript and contributed to the interpretation of data. H-LK conceptualized the overall study design and supervised all aspects of the study and revised the manuscript critically. All authors have read and approved the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Introduction: Hypertension is one of the most common cardiovascular diseases, and the prevalence of hypertension continues to rise across the globe. National and international guidelines recommend angiotensin-converting enzyme (ACE) inhibitors or angiotensin receptor blockers (ARBs), calcium channel blockers (CCBs), diuretics, and beta-blockers for the management of hypertension. CCBs are among the most used antihypertensive medications and Cilnidipine is a newer dihydropyridine CCB shown to have a prolonged antihypertensive property.Objective: This meta-analysis of comparative randomized and non-randomized clinical trials evaluated the effect of Cilnidipine monotherapy or combination therapy on systolic blood pressure (SBP), diastolic blood pressure (DBP), and pulse rate (PR) over 48 weeks of therapy.Study design: PubMed (MEDLINE) and Google scholar databases were searched to identify studies designed to evaluate the effects of Cilnidipine in the treatment of hypertensive patients. The study criteria for inclusion into the meta-analysis were all prospective, randomized, and non-randomized clinical studies published till March 2021, studies published in a peer-reviewed journal, the inclusion of patients with hypertension, assessment of blood pressure and heart rate, and a follow-up of four weeks or longer. The initial search identified 82 potential articles; of these, 24 met the inclusion criteria. Studies with <4 weeks treatment period and those not having a CCB were excluded.Outcomes: Change in SBP, DBP, and PR from baseline at the end of therapy compared between the Cilnidipine and other CCB's.Results: Cilnidipine caused a significant reduction (p<0.05) in SBP, DBP, and PR at end of therapy, whereas the reduction in SBP, DBP, and PR with Cilnidipine was similar to other CCB's (p>0.05). The results of this meta-analysis revealed that there were no significant differences in the efficacy in the treatment of hypertensive patients with Cilnidipine and the other therapies.Conclusion: Cilnidipine has similar anti-hypertensive effects compared with other first-line antihypertensive drugs commonly used in practice. We recommend Cilnidipine as a novel first-line CCB for the management of hypertension either as a monotherapy or as a combination therapy. Hypertension (HTN) is one of the most common cardiovascular diseases, and the prevalence of hypertension continues to rise across the globe . DespiteNational and international guidelines recommend angiotensin-converting enzyme (ACE) inhibitors or angiotensin receptor blockers (ARBs), calcium channel blockers (CCBs), diuretics, and beta-blockers for the management of hypertension. CCBs are among the most used antihypertensive medications currently in the market, and the use of CCBs is especially effective for the treatment of hypertension in the elderly who frequently have large-vessel stiffness .Calcium antagonists dilate blood vessels to reduce peripheral vascular resistance (PVR) which reduces blood pressure. The calcium blockers block calcium influx into vascular smooth muscle cells, resulting in vasodilatation and reduction of peripheral vascular resistance.Cilnidipine is a newer dihydropyridine calcium antagonist shown to have a prolonged antihypertensive property . CilnidiCilnidipine is an L/N-type calcium channel blocker, which lowers the BP in part by sympathetic nerve inhibition at the peripheral sympathetic nerve endings in vivo . It has Old CCB like Amlodipine and a newer CCB like Cilnidipine have shown equal efficacy in reducing blood pressure in hypertensive individuals. But Cilnidipine being an N‑type and L‑type calcium channel blocker is associated with a lower incidence of pedal edema compared to only the L‑type channel blocked by Amlodipine .Cilnidipine has been reported to have more beneficial effects on proteinuria progression in hypertensive patients than Amlodipine, an L-type CCB . The N-tIn a study conducted by Ramya et al., Cilnidipine was found to be safe and effective in reducing microalbuminuria and blood pressure in Indian mild-to-moderate hypertensive patients with type 2 diabetes mellitus. In this study, Cilnidipine caused a significant reduction in the mean (SD) SBP from 150.07 (5.44) mm Hg at baseline to 123.03 (5.23) mm Hg after six months. Cilnidipine also produced a significant reduction in the microalbuminuria from 66.62 (8.39) mg/L to 38.8 (6.45) mg/L after six months .In another large-scale prospective post-marketing surveillance study of post-stroke hypertensive patients  who were treated with Cilnidipine, the blood pressure control with Cilnidipine treatment was very good .Although several meta-analyses have been published on the use of CCBs in cardiovascular disorders, only one has been reported with Cilnidipine . This meReview methodologyWe performed a meta-analysis of comparative clinical trials (randomized and non-randomized) where Cilnidipine was used as monotherapy or combination therapy in the management of hypertension. This meta-analysis complied with the QUOROM  statement . PubMed The study criteria for inclusion into the meta-analysis were all prospective, randomized, and non-randomized clinical studies published till March 2021, studies published in a peer-reviewed journal, the inclusion of patients with hypertension, assessment of blood pressure and heart rate, and a follow-up of four weeks or longer. Studies of all sample sizes were included. The search strategy was based on the search terms “Cilnidipine,” “Calcium Channel Blocker,” “CCB,” “Hypertension,” “Systolic Blood Pressure (SBP),” “Diastolic Blood Pressure (DBP),” “Heart rate,” “Pulse rate,” and “Ambulatory Blood Pressure Monitoring (ABPM).”Abstracts of studies published in all languages (with English translations) were reviewed, and the full text was reviewed after the study satisfied the inclusion criteria. We excluded studies that were not available in English (translations), and studies that excluded hypertensive subjects.The initial search identified 82 potential articles; of these, 24 met the inclusion criteria Figure . One stuOutcomesThe data were abstracted, and differences were resolved by consensus. Data of change in various parameters from baseline to end of therapy were estimated using standard formulae based on analysis of paired data. Efficacy outcomes included SBP, DBP, and PR. Of the twenty-four studies, two studies present blood pressure using a 24-hour ambulatory recording. For these studies, the daytime values for SBP, DBP, and PR were used for analysis along with other studies' data. There were 18 RCTs and six non-RCTs, three studies had diabetic hypertensive patients, two used fixed-dose therapy of Cilnidipine with some other antihypertensive drug, and 15 studies had Amlodipine as control therapy.Statistical analysesMeta-analysis was performed using windows based ‘MedCalc Statistical Software’ version 19.6.1 (2020). Data computations and imputations did in Stata-IC 13.1 . A meta-analysis was done using the random-effects model for comparisons baseline (pre-treatment) versus post-treatment values for different measurement parameters (continuous measure). The random-effects model tends to give a more conservative estimate , but the results from the two models usually agree where there is no heterogeneity. Since the current data are heterogenous (p<0.05), the random-effects model was used. The standardized mean differences  were calculated for SBP, DBP, and PR using the Hedges g statistic, which is the difference between the two means (Cilnidipine and control) divided by the pooled standard deviation (SD), with a correction for small sample bias. If the value 0 was not inside the 95% CI, then the SMD is statistically significant at the 5% level (P<0.05). Cohen's rule of thumb for interpretation of the SMD statistic is: a value of 0.2 indicates a small effect, a value of 0.5 indicates a medium effect, and a value of 0.8 or more indicates a large effect. Publication bias was estimated using the funnel plot. Heterogeneity of the data is estimated using Cohran's Q and I2 statistics, where I2 values ≥50% were indicators of a substantial level of heterogeneity. Forest plots were also used for visual inspection. Funnel plots of effect estimates against the standard error were examined to assess publication bias. Sub-group analysis was done for all parameters based on study design (RCT and non-RCT), diabetes status, a therapy used (fixed-dose combination therapy or monotherapy), setting of blood pressure recording (ambulatory and office setting), duration of therapy , and comparator used (Amlodipine and other drugs).Systolic blood pressureThere were 24 studies that had evaluable data for SBP. There was a reduction in SBP with both Cilnidipine-based therapy and control (Amlodipine and non-Amlodipine-based therapy) in all patients with hypertension  in all patients with hypertension Figure . The ovePulse rateThere were only 12 studies that provided data of PR which could be analyzed. There was a reduction in PR with both Cilnidipine-based therapy and control (Amlodipine and non-Amlodipine-based therapy) in all patients with hypertension Figure . The oveSensitivity analysesIn the efficacy analysis, there was no difference in the overall response rates between Cilnidipine and the control group for all efficacy parameters assessed. The effect size varied from −0.48 (PR) to 0.66 (DBP) without any statistical significance (p>0.05). Further, no differences were found in the different sub-groups based on study design, diabetes status, therapy used, setting of blood pressure recording, duration of therapy, and comparator used.Summary of the literature qualityIn an analysis of the articles, we found that all trials that were included in the meta-analysis were of high quality. The Jadad score was at least two points for each of the 24 studies used for the meta-analysis. There was evidence of publication bias found for SBP (four studies), DBP (13 studies), and PR (three studies). However, there were significant heterogeneities between studies for SBP , DBP , and PR . Combined, this suggests that the overall quality of the analysis was moderate to good.DiscussionThe current meta-analysis showed that Cilnidipine significantly reduced systolic blood pressure, diastolic blood pressure, and pulse rate in hypertensive patients. Cilnidipine has been extensively studied in the management of hypertension. Cilnidipine has been proved to have a reno, neuro, and cardioprotective effect. It decreases heart rate and proteinuria, apart from its BP-lowering effect . HyperteThere were few methodological insufficiencies that require mention. These include (i) randomization methods for the individual studies may not be rigorous because the methods were not clearly described in a few studies; (ii) possibility of selection bias due to poorly described allocation concealment methods; (iii) possibility of measurement bias because the study design was not described in one study; and (iv) we did not evaluate the clinical and laboratory adverse events in the analysis.The current meta-analysis of 24 clinical trials showed that Cilnidipine significantly reduced systolic blood pressure, diastolic blood pressure, and pulse rate in hypertensive patients. The results of this meta-analysis revealed that there were no significant differences in the efficacy in the treatment of hypertensive patients with Cilnidipine and the other therapies. However, there is the possibility of selection bias in a few studies. We can conclude, therefore, that Cilnidipine has similar anti-hypertensive effects compared with other first-line antihypertensive drugs commonly used in practice.We recommend Cilnidipine as a novel first-line CCB for the management of hypertension either as a monotherapy or as a combination therapy. Cilnidipine is highly lipophilic and shows low BP variability among CCBs. The organ protection, especially the reno-protective effect of Cilnidipine deserves special attention. In earlier trials, Cilnidipine was found to be safe and effective in reducing microalbuminuria in hypertensive patients. We warrant further studies to reinforce the cardio-protection and renoprotection efficacy of Cilnidipine, particularly in hypertensive diabetic patients.


The impact of a provider-driven assessment and treatment algorithm based on remote OptiVol  fluid index levels on hospitalizations for congestive heart failure (CHF) remains unknown. We implemented a physician-guided screening and educational strategy for elevated OptiVol fluid index levels measured on remote implantable cardioverter-defibrillator (ICD) monitoring and assessed clinical outcomes over a five-year period. Patients with CHF and a left ventricular ejection fraction (LVEF) of 40% or less with a previously implanted ICD underwent monthly remote monitoring from January 2015 to November 2019. An OptiVol fluid index of 60 Ω-days or more triggered a protocol-based CHF screening and therapy adjustment according to clinical presentation. Among 279 patients included in the study, 228 (81%) were male and 205 (73%) had ischemic cardiomyopathy. The average LVEF was 29% (± 7.3%). A total of 6,616 monthly transmissions were reviewed over five years; of those, 575 (8.7%) were associated with elevated OptiVol fluid index levels in 178 (64%) patients, and clinical follow-up data were available in 459 of 575 (80%) cases. Following abnormal OptiVol fluid levels on remote monitoring, CHF hospitalization occurred in 10 of 459 (2.2%) patient cases. In conclusion, monthly remote monitoring of OptiVol fluid index levels with a health care provider–guided CHF screening and an educational approach to abnormal OptiVol fluid index levels were associated with a low CHF hospitalization rate. This compared favorably to prior similar studies, and randomized controlled prospective studies evaluating similar algorithms are warranted. Remote monitoring was performed monthly from January 2015 to November 2019 and reviewed by a cardiac electrophysiologist. Low intrathoracic impedance measurements were defined as an OptiVol (Medtronic) fluid index level of 60 Ω-days or more at the time of monthly measurement. Patients with abnormal intrathoracic impedance values were managed as follows Patients were called by a physician assistant trained in CHF management who assessed them for signs and symptoms of CHF and provided education on diet, weight monitoring, and recognition of the signs and symptoms of CHF.Patients with no reported clinical CHF manifestations were instructed to self-monitor for signs and symptoms of CHF. Patients with stable CHF symptoms were instructed to see their cardiologist if their symptoms worsened, whereas patients with new or worsening CHF signs and symptoms were referred to their cardiologist for further evaluation and management. In the latter instance, the electrophysiologist contacted the patient’s cardiologist.Patients were contacted again the following month to reassess their clinical status and document whether they had been evaluated and treated by their cardiologist or had been hospitalized for heart failure. Changes to medications were also noted.This was a retrospective chart review of de-identified patient data.Table 1. Their mean age was 78 years (± 9.1 years), and 227 patients (81%) were male. Two hundred four (73%) patients had ischemic cardiomyopathy, and the average LVEF was 29% (± 7%). One hundred and twenty-nine (46%) patients had class III CHF and 183 patients (64%) had biventricular defibrillators.Two hundred seventy-nine patients over the five-year remote monitoring period met the criteria for analysis; of those, 59 (21%) died over the course of the study. Baseline characteristics are listed in A total of 6,616 monthly transmissions were analyzed over the five years; of those, 575 (8.7%) were associated with an elevated OptiVol fluid index. Abnormal OptiVol fluid index levels were present on at least one monthly remote transmission in 178 (64%) patients. Clinical follow-up from the 575 abnormal OptiVol fluid index level transmissions was available in 459 (80%) patient cases. At the one-month follow-up for these 459 cases, 221 (48%) patients had seen their cardiologist and 90 (20%) had undergone a medication adjustment. The overall 30-day CHF hospitalization rate was 2.2% (10/459 patient cases).The 10 hospitalizations occurred in eight patients, with one patient hospitalized on three separate occasions. Four of the patients were already hospitalized at the time of our initial phone call. Of the remaining six hospitalizations, four patients documented symptoms during our phone conversation and had seen their cardiologist prior to their hospitalization. Two patients reported no symptoms during our phone conversation but subsequently developed symptoms and were hospitalized.1 CHF hospitalizations are further associated with a lower quality of life and increased morbidity and mortality.20 As such, implementation of outpatient modalities and clinical algorithms that allow for early identification of CHF exacerbation and appropriate medical intervention is a crucial component in the long-term strategic approach to patients with cardiomyopathy and CHF.CHF management is a major economic burden on the United States health care system with the majority of health care expenditures resulting from hospitalizations for CHF exacerbation. Specifically, the estimated cost of CHF hospitalizations in the United States was over $11 billion in 2014, and with a growing aging population, the expected yearly cost of CHF management is expected to exceed $65 billion by 2030.17 In the current study, we present 30-day CHF hospitalization rates following an abnormal OptiVol fluid index level on remote monitoring. We analyzed more than 6,500 transmissions in 279 patients over a five-year period. In contrast to our prior report, the present study defined abnormal intrathoracic impedance values as OptiVol fluid index levels of 60 Ω-days or more. All patients with abnormal intrathoracic impedance values were educated on the clinical recognition of CHF. As abnormal intrathoracic impedance values can normalize without any intervention, patients were referred to their cardiologist for further evaluation and management only when they reported clinical signs and symptoms of CHF. Primary cardiologists were then instructed to treat based on clinical and/or laboratory findings of CHF and not solely based on abnormal intrathoracic impedance levels. The remote screening protocol allowed for early clinical referral to a primary cardiology provider for in-person evaluation when needed. However, in the absence of clinical signs or symptoms of CHF, patients benefited by learning how to prevent and recognize clinical manifestations of CHF.We previously showed that a targeted and health care provider–guided OptiVol monitoring and intervention strategy minimized CHF hospitalizations in patients with abnormal remote intrathoracic impedance values, using a cutoff of 80 Ω-days or more for abnormal OptiVol levels.10 Nevertheless, intrathoracic impedance is an early measurement of pulmonary fluid accumulation and can normalize without any intervention. It is therefore difficult to predict progression to clinical CHF and hospitalization. The Program to Access and Review Trending Information and Evaluate Correlation to Symptoms in Patients With Heart Failure (PARTNERS-HF) study analyzed 694 patients with biventricular ICDs, using an algorithmic approach including monitoring of intrathoracic impedance, low patient activity, and atrial fibrillation, and reported a 30-day hospitalization rate of 3.9%.6 Cowie et al. similarly used a monthly remote CHF monitoring algorithm and reported a 30-day hospitalization rate of 6.8% in their higher-risk patient group.3 Similarly, Triage-HF identified a high-risk group utilizing remote monitoring and reported a 30-day hospitalization rate of 6.9%.21 Our study findings compare favorably to these studies and indicate that the implemented direct health care provider–guided approach resulted in very low 30-day hospitalization rates. Certain differences and similarities are notable and likely explain the differences in the overall reported hospitalization rates among studies. Similar to PARTNERS-HF and Cowie et al.’s study, the present study implemented a monthly monitoring period in order to allow for early capture of abnormal impedance values in high-risk patients. As an abnormal intrathoracic impedance triggered a screening and education program rather than direct intervention, patients whose intrathoracic impedance would have normalized regardless of any intervention were not overtreated. It is possible that an even more frequent evaluation period could have prevented hospitalizations, specifically four of the 10 hospitalizations in our study that occurred prior to our phone call. Further studies evaluating shorter monitoring periods are warranted.Prior studies have shown that patients with decreased intrathoracic impedance measurement are at a higher risk of CHF hospitalization in the ensuing weeks.22 The Diagnostic Outcome Trial in Heart Failure (DOT-HF), Lung Impedance Monitoring in Treatment of Chronic Heart Failure (LIMIT-CHF), and the Monitoring Resynchronization Devices and Cardiac Patients (MORE-CARE) trials all analyzed the usefulness of an audible alert to detect abnormal OptiVol fluid index levels. DOT-HF saw an increase in CHF hospitalization, and LIMIT-CHF and MORE-CARE showed no benefit of an audible alert for abnormal OptiVol fluid index levels. In DOT-HF and LIMIT-CHF, the intrathoracic impedance was a main determinant in patient treatment and intervention. While our rate of cardiology visits was similar to that seen in DOT-HF, medication adjustments and hospitalizations were significantly reduced. Although 48% of our patients with decreased intrathoracic impedance measurements saw their cardiologist over the following month, only 20% of patients had a change in their CHF medication, compared to a 50% medication change in the DOT-HF trial12 and 100% medication change in the LIMIT-CHF trial.11Our results are in contrast with prior studies that attempted to use intrathoracic impedance to reduce CHF hospitalizations.22 There was no significant difference in CHF hospitalization between the two groups. In these prior studies, the audible alert may have also triggered patient treatment and intervention even in the absence of clinical signs and symptoms of CHF. In our study, cardiology visits and medication changes were a result of signs, symptoms, or a physical exam consistent with CHF, not solely based on abnormal intrathoracic impedance measurements. The ability in our study to only treat the higher-risk patients resulted in a more targeted intervention.MORE-CARE compared remote monitoring and traditional in-office visits with an audible alert for abnormal intrathoracic impedance.10 we cannot exclude potential exclusion bias. We chose a threshold for OptiVol fluid index level greater than 60 Ω-days, which allowed for increased sensitivity but also decreased specificity.This was a retrospective assessment of a single group’s clinical practice and not necessarily applicable to all practices. Clinical follow-up was not available in 20% of patient cases. While this follow-up is similar to prior studies,We present a five-year single-center study utilizing monthly remote OptiVol fluid index levels combined with a health care provider–guided screening and educational and early referral approach to reduce CHF hospitalizations. The 30-day hospitalization rate following monthly remote monitoring of abnormal OptiVol fluid index levels was low (2.2%) and compared favorably to prior studies. Randomized controlled prospective studies evaluating similar algorithms are warranted.


Hypertension is recognized as an important contributor to high cardiovascular mortality in Russia. A comprehensive analysis of data from Russian studies that measured blood pressure in population-based samples has not been previously undertaken. This study aims to identify trends and patterns in mean blood pressure and the prevalence of hypertension in Russia over the most recent 40 years.We obtained anonymized individual records of blood pressure measurements from 14 surveys conducted in Russia in 1975–2017 relating to a total of 137,687 individuals. For comparative purposes we obtained equivalent data from 4 surveys in the USA and England for 23,864 individuals. A meta-regression on aggregated data adjusted for education was undertaken to estimate time trends in mean systolic and diastolic blood pressure, the prevalence of elevated blood pressure (> 140/90 mmHg), and hypertension (defined as elevated blood pressure and/or the use of blood pressure-lowering) medication. A meta-analysis of pooled individual-level data was used to assess male-female differences in blood pressure and hypertension.p < 0.1) at age 35–54 years and by 0.8 mmHg (p < 0.01) at ages 55 and over. The prevalence of elevated blood pressure also decreased by 0.8% per year (p < 0.01), but the prevalence of hypertension remained stable. Mean blood pressure and prevalence of hypertension were higher in Russia compared to the USA and England at all ages and for both sexes.During the period 1975–2017 mean blood pressure, the prevalence of elevated blood pressure and hypertension remained stable among Russian men. Among Russian women, mean systolic blood pressure decreased at an annual rate of 0.25 mmHg (In contrast to the generally observed downward trend in elevated blood pressure in many other countries, levels in Russia have changed little over the past 40 years, although there are some positive trends among women. Improved strategies to bring down the high levels of mean blood pressure and hypertension in Russia compared to countries such as England and the USA are important to further reduce the high burden of CVD in Russia.The online version contains supplementary material available at 10.1186/s12889-021-12320-4. Blood pressure is a strong predictor of cardiovascular mortality  and elevDuring the last 5 years, several global studies have been published in which mean blood pressure and the prevalence of hypertension have been compared between countries and regions. Based on pooled data from 1479 studies the NCD Risk Factor Collaboration (NCD-RisC)  found thIn Russia since the late 1970s, a number of population-based health surveys have been conducted that measured blood pressure and collected information on the use of anti-hypertensive medications. The earliest estimates of hypertension prevalence in Russia were from the collaborative USSR-USA Lipid Research Clinics Prevalence Study (LRC) conducted in 1975–77 and 1982–84 . SubsequWe obtained from investigators data from studies that had measured blood pressure in population-based samples in Russia with adequate documentation of their design and protocols. To be included studies had to have aimed at being representative of their target population, defined either as the entire country, one or several regions, or defined geographic areas. Although the precise designs and settings varied among studies, our analysis aims at identifying any common patterns that may reflect true underlying characteristics in the general population of Russia. Studies were identified based on our extensive knowledge of Russian population-based research. We supplemented this in January-February 2019 with a Google search for “hypertension AND Russian population” and “blood pressure AND Russian population”. This did not yield any studies that we were not already aware of. We approached investigators of all the studies we considered to be eligible for inclusion. As we discuss later in the paper only one study refused to provide their data.Our analysis is based on individual blood pressure measurements from 14 independent cross-sectional surveys conducted in Russia that jointly included a total of 137,687 participants , “Have you been taking any medications lowering blood pressure? , “Do you take prescribed drugs to control your blood pressure?” (IFS2). In SAGE and NHANES the question about blood pressure medication was asked to those who had been informed by a doctor about blood pressure elevation, in other studies to all respondents. For 4/14 of the studies information about the precise question asked about medication was not available.In all studies, hypertension was defined as measured elevated blood pressure and/or the use of blood pressure-lowering medication.In each study, we aggregated education level reported by respondent into three categories: high, middle, and low education. High education corresponds to a university degree or another type of higher education degree. Middle education corresponds to completed secondary education or vocational secondary education. Low education means less than completed secondary education.Our analysis started with an inspection of the age profiles  of the mean SBP and DBP and the prevalence of elevated blood pressure and hypertension  with cutoffs chosen so that we had adequate numbers in each age group on which to estimate means and proportions. The study-specific outcomes  were regressed on the central year of each survey with adjustment for mean age and the share of high education using a random-effects meta-regression model (metareg command in Stata)  on studyordinary least squares (OLS) meta-analysis regression (ipdmetan command in Stata) [To investigate whether male-female differences in the blood pressure outcomes changed over time we compared the magnitude of the sex difference between and within studies according to central year of survey. We used n Stata)  on indiv All tabulations and regression models were implemented in Stata 14 .Figure In both women and men, the highest SBP means in Russia tended to be found in the earliest studies conducted up until the early 2000s. Among women, the lowest values were found in the most recent study .p < 0.01) and a marginally significant decline of 0.25 mmHg per year in the younger age group 35–54 (p < 0.1). There was little evidence for a decline in either age group among men.The trends over time for blood pressure means by central year of study were examined using meta-regression stratified by sex and age group 35–54 and 55+ years Table . The resFigure Unlike SBP there was no obvious trend of DBP with year of survey. The meta-regression results Table  confirm Figure Among the Russian studies at any age, the highest prevalences were generally found in the earliest studies, although men in the IFS2 study (2008–9) had the highest levels. Among women, the lowest prevalence of elevated blood pressure was for the most recent survey KYH 2015–17). The meta-regressions  had the lowest prevalences. In contrast, there was a more heterogeneous pattern among men across the different Russian studies. The two most recent studies (ESSE 2012–2014 and KYH 2015–2017) showed slightly higher prevalences in middle age than those observed a decade earlier. Consistent with the impression from Fig. 5–17 had The numeric values underlying Figs. p < 0.05) per year among women with hypertension aged 35–54 years and of 1.8% (p < 0.01) per year among hypertensive women aged 55+. Similarly among men, anti-hypertensive medications use also significantly increased by 1.5% (p < 0.05) at age 35–54 years and by 1.9% (p < 0.05) among those aged 55+ years.We analyzed the time trend in the percentage of people with hypertension who reported use of anti-hypertensive medications. Our results Table  show thaWe have already noted that there are some differences between men and women in levels and trends. The majority of studies included participants in the age range 55–64 years, allowing us to analyze gender differences in blood pressure dynamics holding age constant. The results are shown in Supplmentary Figs. Our analyses are the most detailed examination to date of all key dimensions of blood pressure and hypertension in Russia spanning over 40 years from 1975 to 2017. Our clearest finding is that mean blood pressures and the prevalence of elevated blood pressure and hypertension in the Russian surveys are higher than in the comparator surveys from the USA and England even in the most recent years. This is consistent with the continued excess of cardiovascular disease mortality in Russia compared to Western countries and underlines how important raised blood pressure is likely to be as a key driver of the very high burden of cardiovascular disease in Russia.A key aims of this study has been to examine trends over time in Russia. Our results provide some evidence for improvements in mean blood pressure and prevalence of elevated blood pressure from the earliest to the most recent surveys. This evidence is strongest for women aged 55 years and older, with less sizeable and convincing trends apparent for men. This is broadly consistent with the conclusion of the NCD-RisC analysis  that repGiven these trends in mean and elevated blood pressures, it is striking that we found almost no evidence in either men or women of changes over time in the prevalence of hypertension per se defined in terms of elevated blood pressure (140/90) and/or taking antihypertensive medication. This might be explained by increases over time in the use of anti-hypertensives in Russia that were particularly pronounced among women. The higher the proportion of the population taking anti-hypertensive medication the higher the nominal prevalence of hypertension regardless of whether this translates into an impact on mean or elevated blood pressure. Certainly, even in the most recent survey, there are high rates of uncontrolled hypertension especially among men .The decrease in mean blood pressure among women led to the initial gender difference in the prevalence of elevated blood pressure with women having the highest levels was reversed from the 2010s. This positive change among women combined with the growing use of medications among them may indicate more effective hypertension control than among men.Our most unexpected finding concerned contrasts between mean SBP and mean DBP. Firstly, diastolic blood pressures for both men and women showed less variation across the Russian surveys than systolic blood pressure. Moreover, the gap between the Russian studies and those in England and the US was larger and more pronounced for DBP than SBP. Secondly, there were striking differences in the relationship of age to mean DBP in the Russian compared to the US and England. Whereas in the US and English surveys mean DBP showed a clear decline from the fifth decade of age, none of the Russian surveys included in analysis showed this effect. Instead, most of the Russian surveys showed either continued increases with age or a plateauing or small decline in DBP from around age 50 years. This is consistent with the small longitudinal declines seen in DBP in the Russian HAPIEE study .In contrast to what we have noted for Russia, there is abundant other evidence of a pronounced decline in DBP after the age of 50 years in Western high-income countries both in cross-sectional and in longitudinal studies . One expOne of the important influences on blood pressure levels in a population is diet especially salt intake . It is oThe inevitable weakness of our study, and indeed of all other parallel data syntheses including the Global Burden of Disease and NCD-RisC , is thatThe optimal approach to study population trends in a parameter such as blood pressure would be to have data from a single study of a consistently defined geographic location sampled at random which used a consistent recruitment and measurement methodology at successive time points. This is the basic design of the Health Survey for England and the US NHANES we used as international comparators in this study. However, in Russia, there are no equivalent repeated large-scale nationally representative surveys. The Russian Longitudinal Monitoring Survey (RLMS) which we have previously used to study trends in smoking  has not Some of the studies we used recruited participants via health care facilities while others used population lists as a basis for sampling. Only one of the Russian studies explicitly attempted to draw a nationally representative sample, while many of the studies were restricted to one or two regions or cities. This diversity in survey location, design, and size will almost certainly have introduced sources of heterogeneity that will make temporal trends challenging to identify with certainty. To minimize some of the extraneous sources of variation, we used methods of meta-analysis that account for the heterogeneity of data sources. To adjust for inter-survey differences in the socio-demographic composition among surveys, we standardized our outcome measures to reflect the fixed educational distribution of the population at the 2002 Russian Census: roughly the mid-point of the period over which our constituent studies were conducted. This will have also helped reduce variations due to selection bias in the recruitment of participants whereby survey participation tends to be higher among those who are most educated. However, this may have introduced a degree of underestimation of real population trends over time through the elimination of changes caused by a general rise of the educational level of the Russian population.Over the past 4 decades of observation, the prevalence of hypertension in Russia has been stable among men and women. Among women, this was due to two opposing trends: a decrease in mean systolic blood pressure and elevated blood pressure prevalence but an increase in the use of anti-hypertensives, potentially resulting in better control of raised blood pressure. In contrast, among men, there was little change in mean and elevated blood pressure and no improvement in its control. Despite improvements among women, the mean blood pressure (particularly diastolic at older ages) and the prevalence of elevated blood pressure remain high in Russia compared to England and the USA. Insufficient blood pressure control in Russia is an obstacle to further reduction of the CVD risks related to hypertension, especially among men.Additional file 1: Supplementary Table S1. Characteristics of surveys included in the analysis. Supplementary Table S2. Methods of blood pressure measurement. Supplementary Table S3. Mean Systolic Blood Pressure (mm Hg) by age, sex and survey adjusted for education. Supplementary Table S4. Mean Diastolic Blood Pressure (mm Hg) by age, sex, and survey adjusted for education. Supplementary Table S5. Percentage of individuals with elevated blood pressure (> = 140/90 mmHg) by age, sex, and survey adjusted for education. Supplementary Table S6. Percentage of individuals with hypertension (BP > =140/90 mmHg or use of medications) by age, sex, and survey adjusted for education.Additional file 2: Supplementary Figure S1. Sex difference in the mean SBP in the age group 55–64 (SBP(m) minus SBP(w)) in Russian surveys (mm Hg). Supplementary Figure S2. Sex difference in the mean DBP in the age group 55–64 (DBP(m) minus DBP(f)) in Russian surveys (mm Hg). Supplementary Figure S3. Male to female odds ratio for the elevated blood pressure in the age group 55–64 in Russian surveys. Supplementary Figure S4. Male to female odds ratio for hypertension in the age group 55–64 in Russian surveys.


How likely is regression of mild to severe chronic kidney disease (CKD) in adults?In this population-based cohort study of adults with incident CKD, the 5-year probability of regression was similar to that of progression or kidney failure in mild (14.3% vs 14.6%), moderate (18.9% vs 16.5%), and severe (19.3% vs 20.4%) CKD. As the risk of death increased with advancing age, the risk of progression decreased substantially, whereas the probabilities of regression varied to a lesser extent and exceeded the risk of progression in moderate or severe CKD.These findings suggest that CKD regression should be considered in the allocation of health resources and in patient counseling. The burden of chronic kidney disease (CKD) is expected to increase worldwide as the global population ages, potentially increasing the demand for nephrology services. Understanding whether CKD inevitably progresses or may regress can inform clinical decision-making and health policy.To study CKD progression and regression by age in adults with CKD.2 for longer than 3 months, from April 1, 2009, to March 31, 2015. Data were analyzed from July 20 to November 30, 2020.This population-based cohort study used linked administrative and laboratory data to assess adults in the province of Alberta, Canada, with incident mild, moderate, or severe CKD, defined by estimated glomerular filtration rate (eGFR) of 45 to 59, 30 to 44, or 15 to 29 mL/min/1.73 mAge.2 for >3 months), death, or censoring .Time to the earliest of CKD regression or progression , kidney failure (the earlier of kidney replacement initiation or eGFR <15 mL/min/1.73 mStudy participants with CKD (55.2% women and 44.8% men) included 81 320 with mild CKD , 35 929 with moderate CKD , and 12 237 with severe CKD . The annual incidence of CKD increased with advancing age, from 180 per 100 000 population younger than 65 years to 7250 per 100 000 in those 85 years or older. After cohort entry, the 5-year probability of regression was similar to that of progression or kidney failure in mild (14.3% vs 14.6%), moderate (18.9% vs 16.5%), and severe (19.3% vs 20.4%) CKD. As mortality at 5 years increased with advancing age in moderate (from 9.6% for age <65 years to 48.4% for age ≥85 years) and severe (from 10.8% for age <65 years to 60.2% for age ≥85 years) CKD, the risk of progression or kidney failure decreased substantially , whereas the probabilities of regression varied to a lesser extent .This cohort study found that with advancing age, CKD regression and death were more likely than CKD progression or kidney failure. These findings have important implications for patient care and for assessing the potential effect of population aging on the burden of CKD. This population-based cohort study assesses progression and regression of disease among adults with chronic kidney disease in Alberta, Canada. Chronic kidney disease is a worldwide public health challenge associated with significant morbidity, mortality, and high health care costs.3 The expected increasing incidence and prevalence of CKD with global population aging may put additional pressures on current limited capacity for the delivery of kidney health care worldwide.6The number of people 65 years and older is projected to more than double from 703 million in 2019 to more than 1.5 billion in 2050.7 Although referral to a nephrologist is recommended for progressive CKD or for estimated glomerular filtration rate (eGFR) of less than 30 mL/min/1.73 m2, patients with less severe CKD are increasingly referred for specialist nephrology care,10 perhaps in anticipation that such care will eventually be required. When eGFR improves to greater than 30 mL/min/1.73 m2, discharge from nephrology clinics back to primary care occurs infrequently,11 partly because of the limited data that are available on the frequency and outcomes of such improvements,14 which are occasionally referred to as CKD regression.15 Understanding how often CKD regression occurs and its outcomes and whether CKD regression varies by age may inform clinical decision-making and health policy, including referral recommendations.Chronic kidney disease is often considered a condition that tends to progress over time.We studied the outcomes of CKD regression and progression in an incident cohort of adults with mild to severe CKD using population-based data. We quantified the probabilities of sustained kidney function improvement (regression), progression, kidney failure, and death, and how these outcomes varied with age. We also assessed the outcomes of participants who experienced CKD regression. Analyses were designed to minimize regression of CKD after acute kidney injury or owing to biological variability in eGFR measurements.16 More than 99% of Alberta residents are registered with Alberta Health and have universal access to hospital care, laboratory testing, and physician services . The institutional review boards at the Universities of Alberta and Calgary approved the study with a waiver of participant consent because retrospective deidentified data were used. We followed the reporting standards recommended by Reporting of Studies Conducted Using Observational Routinely-Collected Health Data (RECORD)17 and used definitions from Kidney Disease: Improving Global Outcomes18 when reporting kidney function and CKD. Data were analyzed from July 20 to November 30, 2020.We conducted a population-based cohort study using linked administrative and laboratory data from Alberta, Canada. The Alberta Health database contains information on demographics, vital statistics, diagnostics, and procedures for inpatient and outpatient health services.19 We assumed that all participants were White, because data on race were not available and misclassification of eGFR is expected to be minimal in Alberta, where only 3% of the population is Black.20 We used only outpatient serum creatinine measurements to enhance the generalizability of findings to general practices and minimize the inclusion of people with episodes of acute kidney injury. We used the mean value of eGFR when there were multiple measurements on the same day.We calculated eGFR using the Chronic Kidney Disease Epidemiology (CKD-EPI) equation, with serum creatinine values standardized to the isotope-dilution mass spectrometry reference method.2 [stage G3a]), moderate , and severe  CKD. We followed guideline recommendations to define CKD based on confirmed eGFR reduction for longer than 3 months.7 To determine each individual’s eligibility to enter a CKD stage and reduce the effect of regression to the mean,21 we screened all consecutive series of 2 or more eGFR measurements, where the first and the last eGFR were separated by more than 90 days and all intervening measurements were within 90 days. We implemented a sustained eGFR method to select the earliest series  for cohort entry, whereby the last eGFR (index eGFR) in the qualifying period defined the corresponding CKD stage and all eGFR values in that series of measurements were below the upper limit of the defined CKD stage  or eGFR of less than 15 mL/min/1.73 m2 for longer than 3 months. When these 2 events occurred on the same day, we defined the outcome as kidney replacement therapy. We ascertained death from Alberta Vital Statistics.16 For the subset of individuals who experienced CKD regression, we studied the competing outcomes of further regression, progression, kidney failure, and death (eTable 1 in the The primary outcome was time to the earliest of sustained kidney function improvement (regression), CKD progression (progression), kidney failure, or death, which we treated as competing events. We defined kidney outcomes using the same methods we implemented to define stage entry  and in restricted samples  , followed by those with moderate (n = 35 929) and severe (n = 12 237) CKD  and severe CKD  were lower than those for mild CKD but increased more steeply with advancing age .Within each stage, the 5-year probabilities of regression were comparable to the risk of progression or kidney failure . With woRegression was observed in all groups defined by age and CKD severity, with 5-year probabilities ranging from 12% to 22%. With advancing age, the probability of regression decreased to a lesser extent  than the risk of progression or kidney failure  in moderate to severe CKD . With moThe probability of regression was lower in individuals with severe albuminuria, regardless of age and CKD stage, although the pattern of association between outcomes and age was similar. For example, the probability of regression at 5 years for moderate CKD was 32.3% for mild or normal albuminuria vs 7.7% for severe albuminuria in individuals younger than 65 years and 17.2% for mild or normal albuminuria vs 9.2% for severe albuminuria in individuals 85 years or older  and death (25.0%). Further regression occurred in some individuals with regression from moderate  or severe CKD , although most remained in the improved stage, experienced progression, or died  still had sustained eGFR of greater than 60 mL/min/1.73 mTime spent in the initial CKD stage was longer when regression was not considered among the reasons to leave that stage. Without considering regression as a reason to leave a CKD stage, the median time spent in severe CKD increased from 2.3  to 3.2  years and the median time spent in moderate CKD increased from 3.6  to longer than 5.0 years .We found similar results in analyses based on the moving average eGFR method or applying an upper limit of 15 months to the chronicity criterion. Details are provided in eTable 8 and eFigures 16 to 20 in the In this population-based cohort study of adults with incident CKD, regression was observed in all groups defined by age and CKD severity, with 5-year probabilities ranging from 12% to 22%. Three important findings emerged. First, regression was at least as common as progression or kidney failure in mild to severe CKD. Second, as the competing risk of death increased with advancing age, the probability of regression decreased to a lesser extent than the risk of progression or kidney failure in moderate to severe CKD. Finally, after regression, stable CKD or further regression was more likely than progression, regardless of age or CKD severity.7 Stability of kidney function over time is seen as the best-case scenario for a person with CKD. In contrast to this perception, we found that the probability of CKD regression was similar or higher than the risk of progression or kidney failure among adults 65 years or older with mild to severe CKD. Chronic kidney disease regression may be due to regression to the mean, recovery from episodes of acute kidney injury, GFR-independent factors ,24 or true improvement because of risk factor modification by treatment or spontaneous recovery. By using 2 or more eGFR measurements longer than 90 days apart when forming CKD cohorts and identifying outcomes, we attempted to estimate each individual’s true mean kidney function over time by minimizing the effect of regression to the mean.21 Although some participants could have recovered from acute kidney injury rather than experienced regression of CKD, the former is less likely in our study, because we applied a chronicity criterion of longer than 3 months to identify CKD cohorts and kidney outcomes and used only outpatient eGFR measurements. Our findings were confirmed in subgroup analyses that excluded participants at risk for acute kidney injury , illness, or amputation with possible muscle mass reduction 25 or included only those with stable CKD for at least 1 year. Similar probabilities of sustained kidney function improvement across CKD stages, including mild CKD, are consistent with findings from animal models and in patients with diabetes after pancreatic transplant.15Chronic kidney disease is often viewed as a chronic and irreversible condition with a lifelong course.26 whereas CKD regression may still occur in older adults who are less frail. Our finding of reduced probability of regression in people with severe albuminuria supports this hypothesis.Because the competing risk of death increased with advancing age, the risk of progression or kidney failure decreased to a larger extent than the probability of regression. A potential explanation is that in the absence of severe albuminuria, CKD in older adults may result from multiple chronic disease–causing mechanisms leading to accelerated frailty before progression or kidney failure can ensue,4 With advancing age, the risk of progression or kidney failure decreased in moderate to severe CKD, whereas regression was at least as likely as progression or kidney failure in mild to severe CKD stages. Our findings are consistent with a study of mild to moderate CKD in primary care (n = 1741) 12 in which most people (34%) had stable kidney function, only a very small fraction (0.2%) developed kidney failure, 18% had progression, and 19% had no evidence of CKD after 5 years. Other studies of patients with hypertensive CKD or receiving nephrology care14 also reported CKD regression. To our knowledge, our study is the first to assess the incidence and outcomes of incident mild, moderate, and severe CKD across categories of age, as well as the outcomes after CKD regression using population-based data.This study further expands our previous work on the association between age and kidney failure and death in adults with severe CKD.27 Second, for adults who are younger than 65 years or have severe albuminuria, the high risk of progression or kidney failure and the low probability of regression warrant more intensive monitoring and treatment, even in less severe stages. Considering the extremely low incidence of mild to moderate CKD in adults younger than 65 years (165 per 100 000 population per year) and the observation that only 5% to 11% of them have severe albuminuria, nephrology consultation in this patient group is not expected to overwhelm nephrology clinics. Most of the remaining CKD population, which includes older adults (aged ≥65 years) with less severe albuminuria and thus higher probability of regression than progression, includes many candidates for discharge from nephrology clinics to primary care. Finally, our study highlights the importance of considering the outcome of regression, regardless of age and CKD severity, and has implications for research using readily available, community-based laboratory data to study CKD burden and prognosis. Ignoring the possibility of regression will result in overestimation of disease duration and thus disease prevalence in modeling studies.This study has clinical practice, health policy, and research implications. First, CKD regression is an important outcome to discuss along with progression, kidney failure, and death in clinical decision-making and advance care planning. Considerations of both adverse and favorable outcomes will provide patients with a more comprehensive view of the disease trajectory and promote optimism and hope, with potentially positive psychological effects that may enhance patient engagement in CKD management.7 We also used a long washout period to maximize the inclusion of incident cases. Finally, the large sample size and extended follow-up allowed us to explore how the probabilities of favorable and adverse outcomes varied by age and across CKD stages. Although regression to the mean does happen and progression takes time to occur, a 5-year observation period should have been long enough to capture these outcomes at the population level.This study has several strengths, including its community-based setting in a geographically defined population with universal access to health care (including serum creatinine assessment and kidney replacement). Furthermore, we applied a chronicity criterion of longer than 3 months to minimize regression to the mean when forming CKD cohorts and identifying outcomes.Our study has some limitations. First, we used routinely collected laboratory data, and thus many factors could have influenced the frequency of eGFR measurements, including the severity of CKD, age, and prognosis. Generally, older individuals or those at low risk of progression are assessed less often. If kidney outcomes were classified using prespecified prospective data, we would arguably have detected a higher probability of CKD regression, especially in older individuals or in those with less severe CKD, and thus this limitation is unlikely to have threatened our conclusions. Second, the observed CKD regression may be explained by changes in muscle mass or diet. However, we obtained similar results after excluding people with previous hospitalization, including those who may have lost muscle mass because of illness or limb amputation. After CKD regression, mortality was less likely than stable CKD or further regression. Third, the study source population was predominantly White, and this may limit generalization of our findings to populations with different ethnic composition. The study population was based on the CKD-EPI equation and included stages G3a, G3b, and G4 CKD; thus our findings may not apply when other methods are used to estimate GFR or to earlier stages of CKD.According to this population-based cohort study, CKD regression may be at least as common as CKD progression or kidney failure for mild to severe CKD. Although the incidence of CKD increases with advancing age, the probability of CKD regression and mortality far exceeded the risk of progression or kidney failure. Therefore, the aging of the general population may not necessarily translate into increased CKD burden for patients and health services.


Vascular structures and functions were assessed by carotid intima media thickness (CIMT) test and brachial ankle pulse wave velocity (baPWV), respectively. Estimated mean 24h urinary sodium excretion of Tibetans in Luhuo County was 5.26±1.61 g. After adjustment, estimated 24h urinary sodium was positively associated with CSBP  and CPP . Line graph of means across urinary sodium quartiles showed that associations of 24 h urinary sodium excretion with AIx75 and baPWV presented approximate “J” shape after controlling for confounders. Estimated 24 h sodium excretion was independently and positively associated with CSBP and CPP. Moreover, association between urinary sodium excretion and arterial elasticity, as evaluated by baPWV and AIx75, presented “J” shape. Further studies are needed to verify J‐shaped association and “safe” zone of sodium intake.Research reports on associations of urinary sodium excretion with central hemodynamic parameters and vascular changes are quite limited in general or non‐hypertensive population. The purpose of the current study was to explore such associations in Chinese general Tibetans living at high altitude. This cross‐sectional study was conducted in Luhuo County, Ganzi Tibetan Autonomous Prefecture with average elevation of 3800 meters from December 2018 to January 2019. A total of 294 Tibetans were included in the current study. Twenty‐four hour urinary sodium excretion was estimated by second fasting spot urine in the morning using Kawasaki formula. Central hemodynamic parameters, including central systolic blood pressure (CSBP), central diastolic blood pressure (CDBP), central pulse pressure (CPP), central mean arterial pressure (CMAP), augmentation pressure (AP), and augmentation index standardized for heart rate of 75 (AIx In addition, findings of existing studies are inconsistent depending on different populations, average amount of salt intake, and status of blood pressure. In particular, no study focusing on such association has been reported in people living at high altitude areas. Low oxygen availability at high altitude imposes extreme physiological challenges on aerobic organisms. Hypoxic environment leads to impairment of endothelial function  and 24 h urinary sodium was estimated using Kawasaki formula2.3BaPWV is a noninvasive method for detecting arterial stiffness, which has been validated with carotid femoral pulse wave velocity (cfPWV)BaPWV examination was conducted using Omron oscillometry‐based device  in warm and quiet room. After 5‐min rest in supine position, four cuffs were wrapped in bilateral upper arms and ankles of patients, two ECG pads were clipped to both wrists and one heart sound sensor was placed at the corner of sternum. Pulse wave velocity was computed automatically based on estimated distances and time consumed between sampling points. Average values of left and right sides were used for analysis.2.4CIMT test was undertaken by trained ultrasound physician using Philips portable carotid ultrasonography system (Philips CX‐50). Study patients sat in supine positions with their heads deviating towards opposite side to detector during examination. Measurements were made in anterior and posterior wall of common carotid artery at approximate distance of 10 mm from bifurcation on both sides. Mean values of anterior and posterior wall of both sides were computed for analysis.2.575 was computed to standardize for heart rate of 75 beats/min.Parameters of central hemodynamics, including CSBP, central diastolic blood pressure (CDBP), central pulse pressure (CPP), central mean arterial pressure (CMAP), augmentation pressure (AP), and AIx, were noninvasively determined by applying proprietary digital signal processing and transfer function using cuff‐based SphygmoCor system .2.625, P75) and qualitative data were presented as percentages or frequencies. Continuous variables in different groups were compared using analysis of variance. Differences in categorical variables among groups were compared using χ2 tests. Partial correlation analysis was used to analyze correlations of estimated 24 h sodium intake with central hemodynamic parameters, baPWV and CIMT after controlling for age, sex, smoking status, alcohol consumption, total cholesterol (TC), fasting blood glucose (FBG), waist circumference (WC), uric acid (UA), heart rate (HR), antihypertensive therapy, and lipid‐lowering therapy . Variables that presented significant correlation with estimated 24 h sodium intake were included in multiple linear regression model as dependent variables and estimated 24 h sodium intake was analyzed as an independent variable. Adjusted confounders in multiple linear regression model included significant variables in univariate linear regression (p < .1) and that known to be correlated with dependent variables. Line graph of means across urinary sodium excretion quartiles were drawn to visualize relationship between urinary sodium excretion with dependent variables before and after controlling for confounders. p < .05 was considered statistically significant.Quantitative data were presented as mean ± SD or median . Mean age of patients was 42.3 ± 10.8 years, 32.3% were males (mean age: 46.9 ± 13.2 years) and 67.7% were females (mean age: 40.2 ± 8.7 years). SBP/DBP were 129.7 ± 18.1/81.9 ± 13.0 mm Hg, estimated 24 h urinary sodium excretion was 5.26 ± 1.61 g and estimated 24 h salt intake was 13.36 ± 4.09 g. Basic characteristics of study patients across urinary sodium excretion quartiles are presented in Table 3.2p = .005) and office PP  but it was not correlated with office DBP  and office MAP .Partial correlation analysis in the current study showed that estimated 24 h urinary sodium excretion was positively correlated with office SBP . However, correlations of estimated 24h urinary sodium excretion with CDBP, CMAP, and Ln AP  were not significant. Estimated 24h urinary sodium excretion was significantly and negatively correlated with AIx75 (p = .036). These findings are presented in Table p = .008 and β = 0.87, p = .013, respectively). However, associations of estimated 24 h urinary sodium excretion with CDBP, CMAP, and Ln AP were not significant. Findings of the current study also established that 24 h urinary sodium excretion was significantly and negatively associated with AIX75   but it was not correlated with CIMT (p = .168)   Table .75 presented “J” shape as shown in Figure After controlling for confounders, line graph of means across 24 h urinary sodium excretion quartiles showed that there was a positive linear association between 24 h urinary sodium excretion with CSBP and CPP. Associations of 24 h urinary sodium excretion with baPWV and AIx475 presented “J” shape.Findings of the current study established that estimated 24 h urinary sodium excretion of native general Tibetans living in Luhuo County was 5.26±1.61 g and estimated 24 h salt intake was 13.36±4.09 g. In addition, the current study showed that estimated 24 h urinary sodium excretion was independently and positively associated with CSBP and CPP. The current study also established that association of 24 h urinary sodium excretion with baPWV and AIx+/K+ ratio was independently related to CSBP and CPP, but not CDBP, in general population. Similarly but not identically, the current study indicated that 24 h urinary sodium was positively associated with CSBP and CPP in general Tibetans, suggesting that salt restriction intervention might be beneficial for control of central blood pressure. In the current study, 1 g increase in urinary sodium excretion was associated with 1.7 mm Hg increase in CSBP after controlling for age, sex, smoking, alcohol consumption, WC, HR, UA, FBG, TC and 1.2 mm Hg after additionally controlling for peripheral MAP.Central blood pressure was reported to be more closely related to coronary perfusion and CV events compared with peripheral blood pressure. So far, few studies have explored association between 24 h salt intake with central blood pressure. Han and coworkersCurrent medical guidelines lack recommendations for lower limits of salt intake. Furthermore, relevance of “the lower, the better” clarion for salt intake is still debatable. O'Donnell and coworkers75 presented “J” shape, which was similar to relationship of urinary sodium excretion with CV events established by O'Donnell and coworkers Lowest point of J curve was mildly higher compared with previous reported “safe” zone of sodium intake in a study by O'Donnell (3–6 g/day), we speculated that the observed deviation might be attributed to three reasons: First, spot urine samples were only collected once, which did not exclude day‐to‐day variation of urinary sodium excretion. Second, applicability of Kawasaki formula for estimating actual 24 h sodium intake in the studied population is unclear. Third, long‐established high salt diet due to alpine and cold climate as well as special Tibetan genetic profiles might have rendered locals more tolerable to detrimental effects of high salt intake compared with people living in plains. This implies that safe zone of sodium intake might have been mildly higher.Association between urinary sodium excretion with arterial stiffness has rarely been reported and most previous studies focused on patients with hypertension or obesity. Few studies has been undertaken in general population or non‐hypertensive people and yielded quite different results. Seung Ku Lee and coworkers22 This balance may be an essential adaptation of preserved vascular function. Therefore, these complicated adaptations reduce effects of hypoxic environment in the current study on associations of urinary sodium excretion with central blood pressure and vascular changes.In the current study, linear relationship of urinary sodium with CSBP and CPP, as well as J‐shaped associations of urinary sodium with baPWV and AIx75 were not affected by hypoxic environment in Luhuo County, because similar findings have been reported by previous studies conducted in low altitude areas. Permanent settlement at high altitude area may have invoked different physiological adaptations among residents including vascular function. Previous studies showed that healthy Andean highlanders and Sherpa presented similar or only slightly lower vascular functions compared with lowlanders at sea level.Mechanisms of arteriosclerosis caused by high salt intake have been well‐established in previous studies. High salt activates renin‐angiotensin‐aldosterone (RAAS) system and sympathetic system and inhibits synthesis of nitric oxide produced by endothelial cells, which are closely associated with arteriosclerosis.The current study had some limitations. Firstly, although determination of 24 h urinary sodium excretion is the gold standard for assessing salt intake, it is difficult to undertake accurate collection and inconvenient to operate. The current study, therefore, adopted Kawasaki formula to estimate urinary sodium excretion by spot urine, which might have deviated from actual status of salt intake to some extent. However, the current study considered Kawasaki formula applicable in population study. Secondly, hormones, including renin, angiotensin, aldosterone, and catecholamine, were not determined for practical reasons, which should be improved in future studies to explore potential mechanisms. Thirdly, inherent nature of cross‐sectional study confined explanation of causal relationships between urinary sodium excretion and vascular changes. Large randomized controlled studies are needed to overcome these limitations.Based on findings of the current study, dietary salt restriction is essential for control of central blood pressure considering current status of high salt intake in this high‐altitude area. J‐shaped association between urinary sodium with vascular changes should be confirmed in future studies by enlarging sample sizes from more sites in high altitude Tibetans area and collecting 24 h urine samples. A recent study revealed that daily pattern of urinary sodium excretion, aside from the amount of sodium excretion, was associated with central blood pressure and arterial stiffness.On behalf of all authors, the corresponding author states that there is no conflict of interest.Xiaoping Chen, Zhipeng Zhang, Hang Liao and Xin Zhang are responsible for designing the study, selecting the research location, communicating execution details with local CDC staffs. Zhipeng Zhang, Hang Liao, Xin Zhang, Qingtao Meng, Rufeng Shi, Jiayue Feng, Xinran Li, Qiling Gou, Runyu Ye and Xianjin Hu participant in performing the examinations for study patients. Zhipeng Zhang are responsible for writing and editing the manuscript. Hang Liao are responsible for editing the manuscript.Supporting materialClick here for additional data file.


Background: Centhaquine (CQ) (Lyfaquin®) is in late stage clinical development as a safe and effective first-in-class resuscitative agent for hemorrhagic shock patients . Acute kidney injury (AKI) is known to be associated with hemorrhagic shock. Hence, effect of CQ on protection of kidneys from damage due to hemorrhagic shock was investigated.Methods: To assess effect of CQ on AKI in shock, we created a rat model with hemorrhagic shock and AKI. Renal arteries were clamped and de-clamped to induce AKI like ischemia/reperfusion model and hemorrhage was carried out by withdrawing blood for 30 min. Rats were resuscitated with CQ (0.02 mg/kg) for 10 min. MAP, heart rate (HR), and renal blood flow (RBF) were monitored for 120 min.Results: CQ produced a significant improvement in RBF compared to vehicle (p< 0.003) even though MAP and HR was similar in CQ and vehicle groups. Blood lactate level was lower (p = 0.0064) in CQ than vehicle at 120 min post-resuscitation. Histopathological analysis of tissues indicated greater renal damage in vehicle than CQ. Western blots showed higher HIF-1α (p = 0.0152) and lower NGAL (p = 0.01626) levels in CQ vs vehicle. Immunofluorescence in the kidney cortex and medulla showed significantly higher (p< 0.045) expression of HIF-1α and lower expression of Bax (p< 0.044) in CQ. Expression of PHD 3 (p< 0.0001) was higher, while the expression of Cytochrome C (p = 0.01429) was lower in the cortex of CQ than vehicle.Conclusion: Results show CQ (Lyfaquin®) increased renal blood flow, augmented hypoxia response, decreased tissue damage and apoptosis following hemorrhagic shock induced AKI, and may be explored to prevent/treat AKI.Translational Statement: Centhaquine (CQ) is safe for human use and currently in late stage clinical development as a first-in-class resuscitative agent to treat hemorrhagic shock. In the current study, we have explored a novel role of CQ in protection from hemorrhagic shock induced AKI, indicating its potential to treat/prevent AKI. Blood loss in excess may lead to hemorrhagic/hypovolemic shock, due to insufficient blood circulation, insufficient tissue perfusion and poor oxygenation . HemorrhCenthaquin (INN: Centhaquine)  (2-[2-4. The clip = 0.0049), 30 min (p = 0.003), 90 min (p = 0.004) and 120 min (p = 0.0017)  of 250 to 50 after hemorrhage and KI [ 0.0017)  post-resBlood lactate level was equally elevated in both vehicle and CQ rats after hemorrhage, indicating similar extent of hemorrhagic shock in both group of rats. Following resuscitation, lactate levels were significantly lower in rats treated with CQ than vehicle .Effect of CQ on cardiovascular actions in the rat model of hemorrhage and KI/R. A significant decrease in heart rate and mean arterial blood pressure (MAP) was observed in both CQ and NS groups of rats after kidney ischemia and hemorrhage  while a Effect of CQ on the expression of hypoxia responsive survival factors. Western blot analysis was carried out to assess the expression of hypoxia responsive proteins HIF-1α and HIF-1β in kidney tissues  Significp = 0.0272) than sham kidneys, while in CQ it was lower than vehicle; however, the difference was not significant compared to either vehicle (p = 0.0787) or sham  staining and histopathological analysis. Pathology Damage Score (PDS) in vehicle was significantly higher  in CQ treated rats than that of vehicle rats , acid-base imbalance (base deficit), decrease in pH (acidosis), increase in lactate (lactate or metabolic acidosis), change in pO2 and pCO2 levels, etc . It has p = 0.0277) at end of study in control and from 1.16 ± 0.12 at baseline to 0.68 ± 0.05 (p = 0.0021) at end of the study in CQ cohort  of acute injury . To incrThe microcirculatory system consists of complex network of small diameter (<100–150 µm) blood vessels e.g. arterioles, venules and capillaries and plays an important role in regulating oxygen supply to meet the metabolic needs of tissues. In homeostasis, they are systemically regulated by sympathetic and parasympathetic nervous system  However,Our previous studies demonstrated contrasting effects of CQ at high and low doses, and it is used as a hypotensive agent at high dose (0.2–0.45 mg/kg), while as a resuscitative agent at low dose (0.01–0.05 mg/kg) . Based oChange in tissue blood flow after hemorrhage is known to alter oxygen supply in organs, which leads to change in cellular metabolism with partial catabolism of glucose to produce lactate instead of CO2 and H2O. Excessive production of lactate causes lactic acid (metabolic) acidosis, which is an important parameter for determining patients’ condition and known to be correlated with death in shock patients . We obseThe regulation of HIF1 A level is mediated through PHD 1–3 (Prolyl Hydroxylase-4 Domain 1–3). PHDs hydroxylase HIF1 1α in presence of oxygen and facilitate its ubiquitination and degradation . Althougin situ PCR technique in kidney tissues of the rats. Since ROS generation mostly takes place in mitochondria and known to cause mitochondrial DNA damage  generation is also known to affect the stabilization of HIF-1α. Although we have not measured the ROS in our study, yet we assessed the mitochondrial DNA using A damage . We asse studies . We obse studies .We further analyzed kidney tissue for expression of kidney damage markers and apoptotic markers. We observed, a significant decrease in early kidney damage marker, NGAL in CQ than that NS treated rats. While we observed no significant difference in the expression of another early AKI marker, cystatin C among all the groups tested in this study. The comparative study in post-operative AKI patients has shown that change in NGAL and cystatin C could be detected in patients’ serum as early as 2 h after operation, while change in creatinine was not detected in even at 24 h post operation . The stuWe also evaluated expression of apoptotic marker, Bax and cytochrome C in these samples. Both, Bax and cytochrome C are essential components in mitochondria dependent apoptosis pathway. We observed significantly reduced level of Bax in CQ cortical and medullary regions of kidneys than that of NS treated rats. Significantly reduced level of cytochrome C was seen in the cortical region, while in medullary region it decreased but could not reach the level of statistical significance. To examine the renal function, we assessed the levels of creatinine and BUN in the rat plasma at baseline and at 120 min after hemorrhage and resuscitation. We observed significantly higher levels of creatinine and BUN in vehicle and CQ than sham rats; however, similar levels of creatinine and BUN in vehicle and CQ were observed at 120 min . The recThus, the results of the study indicate that resuscitation with CQ helps in protection of kidney tissues by increasing tissue blood perfusion and hypoxia inducible factors mediated anti-apoptotic and survival responses. Based on the results of the study, the potential mechanisms of CQ could be enhancement of blood flow in kidneys after acute injury and anti-apoptotic role mediated via hypoxia responsive factors activation, which would be promoting angiogenesis and metabolic adaptation in injured tissues for their survival and function.Overall, our study has demonstrated a novel role of resuscitation with CQ in improving kidney blood flow, hypoxia survival response and protection against acute kidney injury. Further studies on other kidney damage models  will help to prove its potential to develop as a new drug for acute kidney injury.To explore the role of CQ on kidney perfusion and protection of kidney tissues against hypoxic damage, we performed CQ resuscitation in a rat model with hemorrhagic shock and kidney ischemia/reperfusion (KI/R). To induce AKI in hemorrhagic shock condition, renal arteries of rats were clamped, and hemorrhage was carried out. After shock and renal ischemia, resuscitation with CQ (0.02 mg/kg) or vehicle was carried out to assess the effect of CQ  guideline listed in EQUATOR Network library. Ethical aspects: All animal care and use for experimental procedures were performed in accordance with the guidelines of the Institutional Animal Care and Use Committee (IACUC) of Midwestern University. All biohazards were handled in accordance with Midwestern University Bio-safety Committee standard operating procedures and in compliance with OLAW/OSHA regulations.The animal experiment protocol was approved by the Institutional Animal Care and Use Committee of Midwestern University (IACUC protocol 2502). Male Sprague-Dawley rats with an average weight of 288 g were purchased from Envigo, Indianapolis, IN. All rats were acclimatized to for at least 4 days with proper care and readily available food and water. Animals were anesthetized using urethane. Urethane was administered in two doses of 1.5 g/kg body weight via intraperitoneal injection. Urethane was chosen as the anesthetic agent because it produces long lasting (8–10 h) anesthesia with minimal cardiovascular and respiratory system depression. After anesthesia, surgery was carried out to open the abdominal area and renal arteries were located. Right kidney was exposed to assess blood flow using a Laser Doppler probe . A femoral artery was catheterized and the catheter was attached to the pressure monitoring probe of AD Instruments PowerLab . Continual mean arterial pressure and heart rate measurements were carried out throughout the experiment. The femoral artery in the other leg of the rat was catheterized to draw blood to induce hemorrhage and to collect blood samples for blood gas analyses. A femoral vein was catheterized and was used as an infusion site for resuscitative agents. Following the surgery and catheterization, heparin (1 U/g body wt) was infused through femoral vein and 10 min of stabilization period was provided. After stabilization of vital signs, 0.8 ml of blood was drawn from the femoral artery and 0.3 ml was used for blood gas analysis (first blood gas reading) and 0.5 ml was stored for future plasma assessments . Both renal arteries were clamped using strings and hemorrhage was started, simultaneously. Hemorrhage was induced by manually withdrawing blood from the femoral artery at a rate of approximately 0.5–1 ml/min until a MAP between 35 and 40 mmHg was reached . A little volume of blood was infused back if MAP was fallen below 35 mmHg. A total of approximately 6.4 ml of blood from each rat was drawn, which is about 33% of the total blood volume of an adult rat weighing 280–300 gm. The MAP at 35–40 mmHg was maintained for the entire 30 min of hemorrhage by further withdrawal or infusion of blood if necessary. After completion of the hemorrhage, 0.8 ml of blood was withdrawn (0.3 ml for the second blood gas reading and 0.5 ml stored for the second plasma reading). For the sham treatment, rats were anesthetized similar to test rats and after anesthesia, surgery was carried out to open the abdominal area; however, no clamping of renal arteries was performed. Femoral arteries were catheterized to collect blood samples for blood gas analyses and monitoring mean arterial pressure as well as heart rate; however, no hemorrhage was carried out. The rats were monitored for the same time period of test rats.The blood loss volume was calculated for each animal and the same volume was used for infusion with resuscitative agent. The infusion consisted of centhaquine (CQ) dissolved in saline  or saline (control group). An approximately 6.4 ml of CQ solution was infused in each of the test rats and same volume of saline was infused in vehicle or control rats. The rate of infusion was 1/10th volume of resuscitative agent per minute . Thirty min post infusion (or 30 min after completion of resuscitation), 0.8 ml blood was drawn . At 120 min post infusion, 1.3 ml blood was drawn . At the end of experiment, urine was collected from the bladder and stored in the 4°C. The rats underwent bilateral nephrectomy. The right kidney (imaged) was stored in 10% formalin. And the left kidney was snap frozen in liquid nitrogen and stored in −80°C.The kidney blood flow in the right kidney of each rat was measured using a PeriCam PSI system . The system measures blood perfusion/flow through laser speckle contrast analysis (LSCA) technology in real time. Real time blood flow graph and video recording of the kidney flow both were recorded simultaneously and were analyzed using a software package “PIMSoft”. The whole kidney was selected as the ROI and blood flow at different time points  were analyzed by selecting an area on the graph at respective time points.+, K+, and Ca++ (A portion of blood (0.3 ml) isolated at different time points of the experiments  were used for blood gas analysis. All blood gas samples were analyzed using a GEM Premier 3,000 blood gas analyzer . A number of blood components were analyzed including lactate, pH, pO2, pCO2, TCO2, HCO3, Hct, Hbc, Naand Ca++ .Snap-frozen kidney tissues were homogenized in RIPA buffer . Homogenate was centrifuged at 12,000 RPM at 4°C for 30 min and supernatant was collected in a new tube. Folin-Ciocalteu’s Reagent was used to determine sample protein concentration. About 60 μg of protein from each sample was mixed with Laemmli sample buffer  and was denatured in a boiling water bath for 2 min. Proteins were resolved using a 10% SDS PAGE. The resolved proteins were transferred to a nitrocellulose membrane , using a wet transfer method. 4% fat-free milk solution prepared in 1X TBST was used to block the membrane after completion of the transfer. Blocking was done at RT for 1 h. The membrane was incubated overnight with one of the primary antibodies anti-HIF1 A, HIF1 B, NGAL, HFABP, NAG2, TIM1, and Cystatin C at a dilution of 1:1,000 at 4°C overnight. The primary antibodies were purchased from Abcam, Cambridge, MA, United States. The membrane was washed three times with incubation of 10 min each at RT. The blot was incubated with respective secondary antibodies goat anti-rabbit or goat anti mouse IgG conjugated with horseradish peroxidase (HRP) at dilution of 1:10,000 for 2 h at room temperature. The secondary antibodies were purchased from Santa Cruz Biotech., Santa Cruz, CA, United States. Anti-GAPDH  at dilution 1:10,000 was used to detect GAPDH as a loading control. The HRP chemiluminescence was developed with SuperSignal WestPico Chemiluminescent Substrate  and signals were acquired using the Bio-Rad gel documentation system XR . The protein band intensity was analyzed using ImageJ (NIH) software and graphs were plotted after normalizing with GAPDH expression.Immunofluorescence technique was used to detect expression of hypoxia factors , kidney injury markers and apoptotic markers (Bax and Cytochrome C) in cortical and medullary regions of kidneys. A portion of snap-frozen kidneys was dissected out and submerged in 4% paraformaldehyde (PFA) prepared in 1X PBS and thawed at RT for 30 min. Kidney tissues were sliced into 10 µm using a cryostat  at −20°C. Further steps of immunofluorescence were followed with slight modifications as described in our previous studies . In brieThe right kidneys of rats were used for histopathological analysis. At the end of the experiment (at 120 min after resuscitation) rats were sacrificed and the kidney was dissected out and preserved in 10% neutral buffered formalin solution and stored at room temperature. Formalin fixed kidneys were shipped to IDEXX BioAnalytics, Columbia, MO, for their histopathological analysis. The kidney samples were processed and analyzed by professional technicians at IDEXX BioAnalytics in a blinded manner. Briefly, the kidney tissue was embedded in paraffin wax and sectioned longitudinally with a microtome. Hematoxylin and eosin staining of the sections were carried out after deparaffinization. Coverslips were mounted and tissue sections were examined under the microscope at magnification of 20X. Tissue sections from different regions of each kidney were randomly selected and examined for various parameters including tubular dilatation, intratubular hyaline casts and tubular necrosis. These parameters were used to calculate the pathology damage score of kidneys.In situ PCR was carried out to assess the mitochondrial DNA in the rat kidney tissues, with following our previously developed in situ PCR protocol . Sample size: The sample size in this study was 8 per experimental group (N = 8), which was based on our previous study of resuscitative effect of centhaquine on rats after hemorrhagic shock  115–124), where a power analysis was conducted using GraphPad Instat-2.00 . The power was set to 80% , and the level of significance  used was 0.05. Power analysis indicated that a sample size of at least five per group was required to achieve a power of 80%, when the level of significance alpha is 0.05. Randomization: After hemorrhagic shock and I/R rats were randomized to receive either CQ or normal saline. Blinding: The histopathology analysis of kidney tissues was performed and interpreted by investigators blinded to the treatment groups of rats.Results were expressed as Mean ± SEM. A P value <0.05 was considered to be significant. Statistical analysis was carried out using Unpaired