A decrease in absolute numbers (abs.) of circulating dendritic cells (DCs) and recruitment into target organs has been reported, but whether the level of proteinuria associates with circulating DC abs. has not been clarified.We conducted a cross-sectional study of 210 patients with kidney disease aged 21–96 years who were admitted to our hospital for kidney biopsy in 2007–2010. For accuracy, the level of proteinuria was thoroughly measured by 24-h urine collection from patients in their admitted condition. The abs. of total DCs (tDCs), myeloid DCs (mDCs) and plasmacytoid DCs (pDCs) was measured by three-color fluorescence-activated cell sorting (FACS). Patients were divided into four groups based upon the quartile of each DC abs. and one-way ANOVA, and multivariable-adjusted regression analyses were performed.2 µL (p = 0.0277), respectively. Multivariate-adjusted regression analysis revealed that the mDC abs. was negatively associated with proteinuria (95% CI − 57.0 to − 8.5) and positively associated with male gender (95% CI 66.2–250.5). Independent associations were also shown between pDCs abs. and estimated glomerular filtration rate (eGFR) (95% CI 0.14–2.67) and C-reactive protein (95% CI − 49.4 to − 9.9) and between tDCs abs. and male gender (95% CI 54.5–253.6) and C-reactive protein (95% CI − 80.5 to − 13.4).Quantile analysis showed that the level of daily proteinuria decreased with increasing blood mDC abs., with mean proteinuria levels (g/day) of 2.45, 1.68, 1.68, 1.10 for those in mDC abs. quartiles ≤ 445, < 686, < 907, ≥ 907 cells/10We first reported that circulating mDC abs. has a negative association with the level of proteinuria. DCs are the major antigen-presenting cells and are distributed throughout the body including kidney . The subSeveral diseases influence circulating DCs, and decreases in circulating DCs and recruitment into target organs have been reported in autoimmune diseases –4, infecIn kidney mDCs and pDCs, both subsets usually locate in the tubulointerstitium often with a high frequency  and incrA reduction in the number of circulating pDCs has been reported in patients with chronic kidney disease (CKD) compared with that in healthy controls. Several reports have shown that the reduction in pDC numbers was more striking than that in mDCs –14. HoweThough the level of proteinuria is a sensitive marker for kidney disease, the association between the circulating DCs abs. and the level of proteinuria has not been clarified to date. If the circulating DC abs. decreases with the level of proteinuria, the measurement of blood DC abs. may be a biomarker for renal disease activity.Considering all the evidence, we evaluated the hypothesis that the circulating DC abs. could be altered by the level of proteinuria, and we found that mDCs abs. was significantly associated with the level of proteinuria negatively, independent of eGFR. We also found that the number of pDCs, but not the number of mDCs, was associated with eGFR by multivariate regression analysis adjusted for proteinuria, C-reactive protein, and gender.This study was a cross-sectional, observational, single-center study and comprised 243 Japanese patients who received kidney biopsy between December 1, 2007, and June 31, 2010. Clinical information and hematological data were collected immediately before the kidney biopsy. This study was performed in accordance with the Declaration of Helsinki. The Ethics Committee for Clinical Research of Jikei University School of Medicine approved this study [permission no. 29-032 (8648)].To explore the association between the abs. of each subset of DCs and clinical data, the following inclusion criteria were selected: 1) adult patients with biopsy-proven kidney disease and (2) patient records with complete data for the following factors: age, gender, body mass index (BMI), glucocorticoid use, level of proteinuria in collected daily urine, and fasting blood samples. As a result, patients were excluded from the analysis due to insufficient laboratory data (33 patients). Therefore, of the 243 patients evaluated, 210 Japanese patients satisfied the inclusion criteria and were included in the analysis  = 194 × Cr−1.094 × age−0.287 (× 0.739 for women).eGFR was calculated using Japanese equation : eGFR  using a combination of fluorochrome-conjugated monoclonal antibodies to different membrane markers. Reagents were obtained from BD Biosciences : 20 µL lineage cocktail 1-fluorescein isothiocyanate (Lin1-FITC), containing a mixture of anti-cluster of differentiation (CD) 3, CD14, CD16, CD19, CD20, and CD56 antibodies; 5 µL anti-CD123-PE; 5 µL anti-CD11c-APC; and 10 µL anti-human leukocyte antigen (HLA)-DR were incubated with 100 µL whole blood for 15 min at room temperature in the dark. Cells were fixed and lysed for 20 min at room temperature using FACS lysing solution (BD Bioscience). Cells were centrifuged at 200×5 cells were measured to obtain a minimum of 1000 HLA-DR-positive DCs. Representative dot plots of the gating strategy for identifying mDCs and pDCs by three-color FACS are shown in Fig. +, Gate R3) or pDCs  [Data acquisition was performed on a three-color FACSCalibur (BD Bioscience) using a well-defined gating strategy in Cell Quest Pro software . At least 5 × 10Gate R4) , 16, 17.p value of < 0.05 was considered significant. Factors associated with DC abs. on univariate analysis were subsequently included in a multivariate model. Multivariate regression analyses were performed to identify the covariates associated with each DC abs. T values and 95% confidence intervals (CIs) were determined using univariate and multivariate logistic regression models for the factors that were significantly associated with each DC abs.Statistical analyses were performed using JMP 9.0 . Data are expressed as the mean ± standard deviation or numbers (percentage) of patients. Comparisons across the various groups were performed using the Pearson Chi square test for categorical data, Dunnett test for continuous data and ANOVA for quartile analysis. All tests were two-tailed, and a n = 67, 31.9%), benign/malignant nephrosclerosis , diabetic nephropathy , membranous nephropathy , minor glomerular abnormalities , crescentic glomerulonephritis , focal segmental glomerulosclerosis , minimal change disease (MCD) , lupus nephritis , IgA vasculitis , obesity-related nephropathy  and others   cells/102 µL, and 415.5 ± 219.7 (0–1192) cells/102 µL, respectively. The relative numbers of tDCs, mDCs, and pDCs (% of leukocytes) were 0.24 ± 0.09 (0.04–0.66)%, 0.12 ± 0.07 (0.01–0.51)%, and 0.07 ± 0.04 (0–0.28)%.The mean abs. of tDCs, mDCs, and pDCs were 1421.4 ± 366.0 (326–2766) cells/102 µL (p = 0.0277), respectively  of 2.45, 1.68, 1.68, 1.10 for the mDC abs. quartiles of ≤ 445, < 686, < 907, and ≥ 907 cells/10ly Table a. Comparly Table a.Compared to patients with the highest pDC abs., patients with the lowest pDC abs. were older and had lower eGFR, higher BUN, higher serum Cr, higher C-reactive protein and lower hemoglobin Table b.Compared to patients with the highest tDC abs., patients with the lowest tDC abs. were older and more often female and had lower eGFR, higher blood urea nitrogen (BUN), higher C-reactive protein and lower hemoglobin Table c.As shown in Table The primary variables were tDCs abs., mDCs abs. and pDCs abs., and the explanatory variables were determined according to their univariate relationship or importance within the explanatory variables as follows: age, gender, albumin, eGFR, level of daily proteinuria, C-reactive protein and hemoglobin.t = − 2.66, 95% CI − 57.0 to − 8.5, p = 0.0084) and a positive association with male gender  after adjusting for age, serum albumin, eGFR, C-reactive protein, and hemoglobin. Although serum albumin, C-reactive protein and hemoglobin were factors associated with mDCs abs. in univariate logistic regression analyses, these factors were not independent risk factors in multivariate-adjusted regression analysis.Multivariate-adjusted regression analysis for mDCs abs. revealed an independent negative association with the level of proteinuria  and a negative association with C-reactive protein  after adjusting for age, male gender, serum albumin, level of proteinuria, and hemoglobin. Although age and hemoglobin were factors associated with pDCs abs. in univariate analyses, these factors were not independent risk factors in multivariate-adjusted logistic regression analysis.Multivariate-adjusted regression analysis for pDCs abs. revealed an independent positive association with eGFR  and a negative association with C-reactive protein  after adjusting for age, serum albumin, eGFR, level of proteinuria, C-reactive protein, and hemoglobin.Again, multivariate-adjusted regression analysis (least-squares method) for tDCs abs. revealed an independent positive association with male gender  independently associated with circulating mDC abs., pDC abs. or tDC abs. by multivariate regression analysis as follows. First, we revealed that the circulating mDC abs. was negatively associated with the level of proteinuria, independent of eGFR. These data suggest that mDCs may recruit from circulating blood to injured kidneys exhibiting heavy proteinuria, independent of the CKD stage. For accuracy, the level of proteinuria was thoroughly measured by 24-h urine collection from patients in their admitted condition, not by spot-measuring g/Cr. This accurate measurement may contribute to the result. Second, the circulation of pDC abs. had a positive association with eGFR and no association with proteinuria. This suggests that pDCs are recruited into the kidney as kidney function declines, independent of the level of proteinuria. Thus, CKD stage and heavy proteinuria may regulate the distribution of each DC subset separately. Third, C-reactive protein was negatively associated with both pDC abs. and tDC abs. Fourth, male gender was positively associated with both mDC abs. and tDC abs.A possible mechanism of the decrease in circulating DCs involves either DC death, a distribution shift from circulating blood to organs, or the other unknown mechanism in some kidney disease without inflammatory cell infiltration. DCs may die following infection, as in malaria  or severp = 0.0638). Similar to our study, several reports have shown a decrease in the mDC or/and pDC count in inflammatory diseases, such as septic shock [In the present study, all DC subsets showed a negative association with C-reactive protein, though a weak association was observed between C-reactive protein and mDCs  than this study (4.5–133.2 ml/min/1.73 m2), so that they could not detect a significant differences between DC number and eGFR. And this study involved only the patients who received kidney biopsy and the frequency of diabetic nephropathy was small (39.7% in Pauls’ vs 12.4% in this study) so that this study might not be able to detect a significant differences between DC number and the presence of diabetes mellitus. These discrepancy of patient characteristics might influence the results.The present multivariate regression analysis (210 patients) supports the proposal that renal dysfunction (decrease in eGFR) could have a negative effect on at least pDCs but not mDCs in patients with predialysis CKD, such as that in a report by Hesselink et al. . SeveralHuman blood mDCs and pDCs express estrogen receptors (ERs) and respond to estrogens, suggesting that estrogens may contribute to the sex differences in immunity by regulating DC biology . Among oOrsini et al. also observed that only the pDC abs. significantly decreased with increasing age by univariate analysis, whereas the mDC abs. did not show any age correlation in the subjects over 20 years . We alsoWe conducted a cross-sectional study of 210 patients with various kidney disease and we could not detect disease specific relationship between proteinuria and circulating DCs in each renal disease.This study included a relatively smaller number of patients with diabetic nephropathy compared to the usual CKD population given their relatively lower indication for kidney biopsy, which may influence the results. Thus, further large-scale study will be needed to clarify the relationship in each renal disease.2), 16 patients with a normal eGFR > 90 ml/min/1.73 m2, and 23 patients with a normal daily proteinuria < 0.15 g, these patient characteristics may influence the results.This study did not involve intact healthy control subjects; however, the range of eGFR was wide (4.5–133.2 ml/min/1.73 mWe did not detect the local mDC by immunostaining nor by histopathological findings of tubulointerstitial inflammation and we did not measure any serum concentrations of IFN, inflammatory cytokines and chemokines, so we could not clarify the mechanism by which proteinuria influences the decrease in circulating mDCs.The measurement of both daily proteinuria and DC count was performed in 1 day simultaneously, although it would be preferable if more than two times simultaneous measurement of both was possible.We first reported that circulating mDC abs. had a negative association with the level of proteinuria and no association with eGFR in CKD patients not receiving dialysis. C-reactive protein was negatively associated with both the pDC abs. and tDC abs., and male gender was positively associated with both the mDC abs. and tDC abs. in these patients.Notably, we observed a strong reduction in mDCs, while the pDCs appears mostly unchanged in patients with heavy proteinuria. This is in line with several diseases where the involvement of DCs as a factor associated with the diseases was highlighted.A decrease in the circulating mDC abs. may reflect the degree of kidney damage in CKD patients, and measurement of the blood mDC abs. may be a biomarker for renal disease activity.


The lack of adherence to prescribed antihypertensive medication occurs in 50% of patients and leads to poor health outcomes and increased medical costs. Consistent use of antihypertensive medications among patients with hypertension is essential to the reduction of short- and long-term cardiovascular complications. Strategies to improve medication adherence include syncing prescription medications in the pharmacy, which allow patients to retrieve chronically prescribed medications in one visit. The adoption of medication synchronization has been shown to improve adherence to medications; however, there is a lack of data showing if the intervention reduces blood pressure and improves long-term health outcomes.This study aims to determine the association between participation in an appointment-based medication synchronization service and blood pressure levels among patients on antihypertensive medications.This longitudinal prospective cohort study will observe changes in blood pressure among individuals in a medication synchronization program and those in a usual care group. Patients on at least two antihypertensive medications and four total medications have been recruited to participate in the study. All participants will be required to have at least a 6-month history of filling prescriptions at the pharmacy prior to enrollment in the study. Based on an estimated standard deviation of 14 mmHg, a sample size of 70 participants provides approximately 80% power with a two-sided .05 significance to detect a difference of 9 mmHg blood pressure between the two cohorts.As of the publication of this paper, patients are completing final blood pressure visits at the pharmacy and medication data are being collected from the pharmacy. Once patients complete all blood pressure visits, data analysis will begin.This study will link medication synchronization and changes in blood pressure levels among individuals with hypertension. This study will provide preliminary data for a randomized clinical trial that will assess the impact of medication synchronization on blood pressure.DERR1-10.2196/12527 Despite the multitude of treatment options for the management of hypertension, heart disease and stroke remain the leading causes of death attributed in part to high blood pressure. Among the 75 million Americans with hypertension, 74.9% of patients are being treated and only 52.5% of patients are under control . AlthougP=.04) [P=.001 and 10 mmHg, P<.001, respectively) [Pharmacy-based strategies to improve medication adherence include the incorporation of synchronization, which enables patients to retrieve multiple medications systematically on one predetermined date -6. CurreP=.04) . Howeverctively) . These sctively) . A reviectively) . Additioctively) . This maP=.04) [Other literature may not specify medication synchronization as the intervention used. Although other studies may not be identically comparable to medication synchronization, there are some similarities to the interventions performed that merit investigation. One such example is the Federal Study of Adherence to Medications in the Elderly (FAME) study, which examined the effect of a pharmacy care program on patient adherence and persistence . This stP=.04) . The phaP=.04) . MedicatThough there are similarities between medication synchronization and the FAME study intervention, the key difference is the use of blister packaging in the study. Thus, the study cannot directly be compared to a medication synchronization intervention due to the inclusion of compliance packaging. Although medication synchronization has been shown to indirectly improve medication adherence, it is not known if utilization leads to reduced blood pressure. Therefore, we aim to determine the association between participation in a medication synchronization service and blood pressure levels among patients on antihypertensive medications.The overall objective of this project is to determine the association between participation in an appointment-based medication synchronization service and blood pressure levels among patients on antihypertensive medications. Other study objectives are to examine whether the effect of an appointment-based medication synchronization service on patients’ blood pressure vary based on time-invariant demographic characteristics and time-variant characteristics of the subjects. Data generated from this study will determine the appropriate study size for a larger clinical trial of the intervention.Aim 1 is to examine the association between enrollment in an appointment-based medication synchronization program and study participants’ blood pressure levels. Hypothesis 1 is as follows: consistent enrollment within a medication synchronization service will be associated with lower blood pressure measurements. This hypothesis will be tested using blood pressure ascertained over a follow-up period of up to 10 months.Aim 2a is to examine whether the effect of an appointment-based medication synchronization program on patients’ blood pressure varies based on time-invariant demographic characteristics of patients, such as gender, age, education, income, and ethnic origin.Aim 2b is to examine whether the effect of an appointment-based medication synchronization program on patients’ blood pressure varies based on time-variant characteristics of patients, such as weight, lifestyle change, adverse health behavior, ethanol use, frequency and intensity of exercise, adverse drug reaction, the number of medications, and adherence rates.Aim 3 is to describe the level of patient satisfaction with enrollment in an appointment-based medication synchronization program.This is a longitudinal prospective study to observe whether enrollment in an appointment-based medication synchronization program will change SBP and DBP levels in patients with hypertension  [Group 1 (medication synchronization group) comprises patients voluntarily participating in the study and who will enroll in the appointment-based medication synchronization program.Group 2  comprises patients voluntarily participating in the study and who refuse to participate in the appointment-based medication synchronization program.All procedures, informed consent protocols, and study documents were approved by the University of Toledo Biomedical Institutional Review Board. Study procedures were all approved by the pharmacy practice site.All research subjects will sign written informed consent forms prior to participating in the study.Recruitment will occur among patients that utilize an independent pharmacy for prescription services in Toledo, Ohio. The pharmacy’s computer software will be screened for patients that potentially meet the inclusion criteria see . Up to 1After the baseline clinic visit, patients that meet the study inclusion and exclusion provisions will be followed by cohort depending on the services they decide to utilize in the pharmacy. Participants will be categorized as participating in medication synchronization or traditional medication pick-up group. Participation in any of these pharmacy services are voluntary and are a part of care offered by the pharmacy. Participants will notify investigators as to which cohort they would like to participate in at the month-1 research visit.The baseline visit procedure includes confirmation that all inclusion and exclusion criteria are satisfied, verification of participant consent and contact information, and completion of baseline data survey, including obtaining blood pressure for analysis. The follow-up visit schedules for data collection do not differ by cohort groups. Proportion of days covered will be utilized to assess medication adherence. Monthly medication data will be collected to evaluate prescription records to assess adherence. Seated blood pressure, weight, sociodemographic data, and smoking and alcohol usage will be collected every 3 months starting from the first month of the study.A multitude of physical measures and medication data will be collected throughout the study via five research pharmacy visits or four phone call surveys. A one-time medical history will be collected at baseline to serve as a screening tool as well as an eligibility and stratification factor. The following physical measures will be collected at baseline: seated blood pressure, weight, height, gender, age, medical history, sociodemographic data, and smoking and alcohol usage. Seated blood pressure, physical measures, and questionnaires will be gathered during pharmacy visits, while all other information, including health status and satisfaction with pharmacy services, will be collected via phone call surveys. Sociodemographic information will reflect age, race, date of diagnosis, ethnicity, gender, level of education, marital status, persons living with participants, and US postal code. This data will be used to identify eligible participants and to characterize the final study population. Baseline medication data include the number of medications, type of medications, and adherence. Inclusion criteria:Subject is an adult, 18 years of age or older.Subject is able to read, understand, and sign a written informed consent form to participate in the study.Subject has systolic blood pressure >140 mmHg or diastolic blood pressure >90 mmHg during the baseline period.Subject has been an established patron of Toledo Family Pharmacy Inc for at least six previous refills of blood pressure medications.Subject has filled more than four prescriptions at Toledo Family Pharmacy each month for the past 3 months.Subject has a history of hypertension and has been prescribed at least two antihypertensive medications.Subject possesses an active phone number where they can be reached.Subject is willing and able to comply with the study protocol.Exclusion criteria:Subject is participating in another clinical trial within four weeks of the baseline period involving an intervention that could affect the outcome measures of this study.Subject has end-stage renal disease and is on dialysis.Subject had a heart attack in the previous six months.Subject had a stroke in the previous six months.Subject is currently enrolled in an auto-delivery program or medication packaging program.Subject has a psychiatric or mental disorder that, in the judgment of the investigator, could interfere with provision of informed consent or completion of survey questionnaires.Patient has a designated power-of-attorney that signs for medications, which renders them not competent to sign the informed consent form.The pharmacy will supply the research team with medication data on each of the participants enrolled in the study each month. Data provided will include the number of medications, names of medications, costs of medications, the date when the prescription was filled, and the date when the prescription was picked up from the pharmacy. For each drug, the quantity filled, days’ supply, and prescription number will be obtained. Protected health information collected in the study will be maintained on a key code sheet. This code sheet will be locked in the principal investigator’s office at the University of Toledo. A code will be on all data collection forms to refer to each participant in the study. These efforts will maintain research participants’ confidentiality throughout the study. Data collected on data forms will be maintained in a locked cabinet in the principal investigator’s office, separate from the key code sheet. Data collected on code sheets will also be transcribed to electronic format using REDCap provided by the University of Toledo.By assuming a medium effect size of 0.75 times standard deviation for the intervention and set power at 0.8 with a two-sided alpha of .05, the sample size needed is 70 participants, with 35 in each group, allowing for up to 20% attrition. Based on the estimated standard deviation of 14 mmHg, the proposed sample size will provide approximately 80% power to detect a difference as small as 9.5 mmHg between intervention and control groups [pseudo-population, where treatment arm and control arm are balanced over the confounder [Given the observational nature of the study and patients’ self-selection of the treatment arms, inverse-probability -of-treatment weighted (IPTW) estimation of marginal structural models (MSMs) will be employed in this study. This estimation addresses both sources of potential bias, such as confounders or covariates  and missing data under the missing-at-random (MAR) assumption, the most popular informative mechanism in practice. MSMs are a class of statistical models used for causal inference in epidemiology and use a multi-step estimation strategy to control for the effect of confounding variables, allowing the investigator to obtain unbiased estimates -12. Amonnfounder . When apFunding was provided on May 12, 2017. As of the publication of this paper, patients are being enrolled in the study and completing follow-up visits in the pharmacy. Also, as of the publication of this paper, investigators are collecting prescription data provided by the pharmacy for all participants in the study. Data collection and analysis will be completed by May 2019.Pharmacy’s Appointment-Based Model Implementation Guidefor Pharmacy Practices [The medication synchronization service will be delivered as described in the American Pharmacists Association Foundation’s ractices . There aOnce a sync day has been assigned, a pharmacy staff member will call the patient at least a week before the sync date. This is to ensure there are no medication changes prior to filling medications and for the pharmacy staff to confirm if there have been any new medications added or medications that have been discontinued. Once the patient comes to pick up medications that have been synced, the pharmacist will briefly counsel the patient.The monthly appointments are times when patients will come in to retrieve their prescriptions. This is also a time the pharmacist can provide some education to the patient on blood pressure management. This encounter is not meant to go beyond the usual patient consultation that occurs when a patient picks up a prescription. The intent of this consultation is to provide a focused means of providing patients with education on hypertension and medication use. A brief topic outline described in Although there are studies that demonstrate medication synchronization has an impact on adherence metrics, there is still need for data to connect it with a clinical measure such as blood pressure. Data generated from this study will provide an association between medication synchronization and blood pressure levels. It will also determine effect size and the appropriate sample size for a future clinical trial of the intervention. Limitations of this study include lack of randomization at the point of entry in the study. Study investigators will use MSMs to overcome self-selection bias.


Context: Huoxin formula is a Traditional Chinese Medicine for coronary heart disease (CHD) treatment.Objective: To explore the therapeutic mechanism of the Huoxin formula on arterial functions in CHD patients.Materials and methods: Fifty-eight CHD patients receiving cardiovascular drugs including β-receptor blocker, statins, and antiplatelet medications or others were randomized into intervention  and control [only cardiovascular drugs (n = 28)] groups. Serum biomarkers , and cardiovascular indicators of the common and internal carotid arteries (ICAs) were monitored before and after the treatments.Results: After 3 months of treatment, the increases of intima-media thicknesses (IMT) of the left and right common carotid arteries (CCAs) as well as of the left and right ICAs and the increases of the left and right cardio-ankle vascular index were all significantly  less in the intervention than in control group . Serum concentrations reductions of hs-CRP, IL-18, IL-17 and MMP9  levels were higher in the intervention compared to the control group, which correlated with the changes of left ICA .Discussion and conclusion: Since the Huoxin formula improved arterial functions and reduced inflammatory factor activities in CHD patients, a large-scale clinical trial is warranted. Coronary heart disease (CHD) is characterized by coronary arterial stenosis or obstruction caused by progressive coronary atherosclerotic lesions, and the resulting myocardial ischemia or necrosis of the involved cardiac muscle  var. mandshuricum (Maxim.) Kitag. (Aristolochiaceae), Dalbergia odorifera T. Chen (Leguminosae), Panax notoginseng (Burk.) F. H. Chen  and Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao (Fabaceae) and other accessory materials, which have been reported to have cardioprotective  drugs and other cardiovascular medications and analyzed the mechanisms of therapeutic benefits in addition to the effects on inflammatory-factors. We hypothesized that the Huoxin formula might have actions on arterial elasticity and inflammatory mediators.From October 2013 to December 2015, 58 patients diagnosed with CHD were included in the study from the outpatient clinic and the ward of the cardiovascular department in Shuguang Hospital, Shanghai University of Traditional Chinese Medicine. The ethical committee of Shuguang Hospital approved the study. Informed consent was obtained from all the subjects included in the study, which was carried out in accordance with approved guidelines. The clinical registration number of the study is ChiCTR-TRC-13004040, and the date of registration 19 December 2013.The medical diagnosis was based on ‘The diagnostic and therapeutic guidelines of chronic stable angina pectoris’ published by the Chinese Society of Cardiology and the editorial board of the Chinese Journal of Cardiology in 2007  and a control group (n = 28). The patients in the control group were treated only with the conventional CHD medicines, while the patients in the intervention group were additionally medicated with the Huoxin formula.According to ‘The diagnostic and therapeutic guidelines of chronic stable angina pectoris’, all the patients included in the study were treated with conventional CHD medicines, mainly including β-receptor blocker, statins, antiplatelet drugs with/without ACEI (ARB) drugs, and fewer nitrates, aspirin, enteric-coated tablets as well as angiotensin-converting enzyme inhibitors when necessary. Patients were randomized into an intervention group (A. heterotropoides (Fr. Schmidt) var. mandshuricum (Maxim.) Kitag. (Aristolochiaceae), D. odorifera T. Chen (Leguminosae), P. notoginseng (Burk.) F. H. Chen  and Ast. membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao (Fabaceae) (The Huoxin formula consists of a mixture from roots of abaceae) .Supplementary Figure 1) and specific chromatograms as well as high performance liquid chromatography-mass spectrometry in order to maintain constant quality of mixture batches .The ready-to-use herbal granules , were dissolved at home and taken by each patient twice a day for 3 months. Each medication consisted of granules from packages with single herbs, which have been poured into a cup and solved in 150 mL of warm water. The producer guaranteed high quality according to the Good Agricultural Practice of Medicinal Plants and Animals advanced quality control, with detection technologies including fingerprint  and effect indexes  were measured before the treatment, as well as 3 months after the initiation of therapy.We collected 4 mL blood from the cubital veins of each fasting patient in the morning, and centrifuged the samples for 10 min at 3000 rpm. The separated serum was stored in a freezer at −70 °C. Factors such as hs-CRP, IL-18, IL-17, TNF-α and MMP-9 were measured using ELISA .Pulse pressure (mmHg) = systolic blood pressure − diastolic blood pressure (mmHg). After resting the patients for 5 min, a mercury sphygmomanometer was used to measure the blood pressure in the right upper arm with the patient lying on a bed. The mercury column was brought down at a rate of 2 mmHg/s, and 2 min after the first measurement was completed, a second reading was taken and the mean blood pressure calculated. Systolic and diastolic blood pressure was recorded when the first and fifth Korotkoff sounds were separately detected.These two indexes were measured with a VaSeraTM VS-1000, which electronically measures blood pressure and the pulse . When patients were resting in a prone position on a bed, the pressure cuff and sensor was applied. The pressure cuffs were set at the same positions as when measuring blood pressure and small cushions were placed around the elbows and heels of patients. The cuffs were kept at the same level as the heart in order to keep the pulse wave stable. Two ECG electrodes were attached to the wrists to record waveforms from lead I. A phonocardiogram (PCG) sensor was added to the right knee with the gasbag pointing at the centre of the back of the knee. The strap was adjusted until the green light on the sensor was illuminated (on). Then, we connected and fixed Velcro beside the kneecap, and made sure the pulse wave was clear and stable. Then the START button was pressed to initiate the measurements, with the PCG and stable pulse waves displayed on a monitor. CAVI and ABI reports were automatically printed when the measurements were completed.IMTs were measured using a HP Sonos 5500 (Hewlett Packard) ultrasound instrument at a frequency of 7.0–10 MHz. Patients were placed in the supine position with a head deviation of 45 degrees to the side, with the neck fully exposed. All measurements of the anterior and posterior walls of the CCA, 1 cm proximal to the bilateral carotid branch and in the ICA were performed by the same senior surgeon and the average value of the measurements computed; local plaques were not included.p < 0.05 considered to be statistically significant. Measurements of normally distributed data are presented as the mean ± standard deviation (x ± SD), while data with a skewed distribution are presented as the median, minimum and maximum values. Enumeration data was described by constituent ratios. A chi-squared test was performed to compare the rates of two samples and variance analysis was conducted to compare the repeated measurements in the follow-up (baseline data was set as the concomitant variable).SPSS Statistics for Windows  was used to perform statistical analyses with p > 0.05), indicating the comparability of the therapeutic effects of the two different treatments  in the intervention group. The reductions of IL-18 (p < 0.001), IL-17 (p < 0.001) and MMP9 (p < 0.001) as well as TNF-α levels were greater in the intervention group, indicating that the combinatory use of traditional chinese medicine was able to suppress systemic inflammatory reactions (As shown in eactions .p = 0.043) as well as with R-CAVI  and L-CAVI . IL-18 correlated with left ICA IMT  and pulse pressure   indicating that inflammatory factors contribute to blood vessel obstructions.Based on a correlation analysis, we found that there were various significant intra and inter correlations between CCA IMTs, ICA IMTs and left ABI as well as between CAVIs and ABIs, whereas pulse pressure correlated with left and right ABI , which i= 0.004) , whereasr = 0.328, p = 0.021)  .Radix astragali, Panax, D. odorifera and Asarum cause liver injury  and P. notoginseng (notoginsenoside R1) have been shown to protect against cardiac hypertrophy  and P. notooginseng (Pseudoginseng). We also analyzed the concentrations of multiple serum inflammatory factors before and after combinatory therapy. The concentration of IL-18 was positively correlated with the incidence of cardiovascular events  in macrophages  and intercellular adhesion molecule 1 (ICAM-1) are expressed by endothelial cells following cytokine signalling and bind lymphocytes through cell surface adhesion molecules (Davies et al. Huoxin formula as an adjunct to conventional stable angina pectoris medication slowed down IMT in the common and ICAs, improved CAVI and ABI values as well as pulse pressure significantly better than conventional CHD medicine alone, which was related with changes of inflammatory factor activities. Thus, the progression of coronary stenosis was significantly slowed down in patients treated with adjunctive Huoxin formula.


Background: Oxygen uptake (VO2) evaluations by cardiopulmonary exercise test is expensive and time-consuming. Estimating VO2 based on a field test would be an alternative.Objective: To develop and validate an equation to predict VO2peak based on the modified shuttle test (MST).Methods: Cross sectional study, with 97 children and adolescents with asthma. Participants were divided in two groups: the equation group (EG), to construct the equation model of VO2peak, and the cross-validation group (VG). Each subject performed the MST twice using a portable gas analyzer. The peak VO2peak during MST was used in the equation model. The patients’ height, weight, gender, and distance walked (DW) during MST were tested as independent variables.Results: The final model [-0.457 + (gender × 0.139) + (weight × 0.025) + (DW × 0.002)] explained 87% of VO2peak variation. The VO2peak predicted was similar to VO2peak measured by gas analyzer  (p = 0.67), and presented significant ICC 0.91 (IC95% 0.77 to 0.96); p < 0.001. The Bland–Altman analysis showed low bias (-0.15 L/min) and limits of agreement (-0.65 to 0.35 L/min). There was no difference in DW between EG (760 ± 209 m) and VG (731 ± 180 m), p = 0.51.Conclusion: The developed equation adequately predicts VO2peak in pediatric patients with asthma. Asthma is a prevalent chronic pulmonary disease in the childhood. It is usually characterized by chronic airway inflammation that results in a decreased aerobic capacity, and its prevalence is increasing in many countries .Cardiorespiratory fitness is considered an important marker of health in youth, since high cardiorespiratory fitness in youth have shown to track do adult ages . In addiO2max) have been considered the gold standard for cardiorespiratory fitness assessment  to develop and validate an equation to predict VO2peak based on MST test.In this context, field tests that includes walking, such as shuttle-walk test  or modifg asthma . HoweverPatients between 6 and 18 years of age were recruited in 2013 through 2015 from the Pediatric Department of the Federal University of São Paulo, Brazil. All patients had been diagnosed by a pediatric physician, according to the GINA steps , as havi2peak, and (2) a cross-validation group (VG), which included patients for preliminary validation of the equation model. Patients were included in the study after their legal guardians read, agreed to, and signed the informed consent form, and the patient had signed the informed consent if they were 12 years of age or older. The study was approved by the Associaçao Educacional Nove de Julho Ethical Committee, São Paulo, Brazil (#738192/2014).Patients were divided into two groups: (1) the equation group (EG), which included patients who were studied to construct the equation model of VOThis is a cross sectional study. All outcome measurements needed for this study were taken on two different days. The outcome measurements that were assessed included an asthma control questionnaire, lung function spirometry (pre- and post-bronchodilator), the MST and the cardiopulmonary exercise test (CPET). The MST and CPET were performed on different days, in random order.The asthma control test (ACT) is a simple questionnaire to assess asthma control based on patient views, used for patients over 12 years old. The children’s ACT (C-ACT) is used for children 4 through 11 years old . Each qu1), FEV1/FVC, and forced expiratory flow at 25 to 75% of FVC (FEF25-75) were expressed in liters and percentage of predict value . The technical procedure, acceptance criteria and reproducibility were according to ATS/ERS statement . The patct value , and z-s2 fell below 82%  were continuously evaluated. The modified Borg scale was used to observe lower limb fatigue and dyspnea at the beginning and end of the test (The modified shuttle test (MST) was performed in a 10-m-long corridor according to the previous description . It is aelow 82% . Each pathe test . The grethe test , was use®, LODE BV Medical Technology, Groningen, Netherlands) was used. After 2 min of unloaded cycling, the load was increased (10 to 20 watts/minute)  , and the/minute) . The hea2000, MedGraphics Corporation®, Saint Paul, MN, United States). The VO2, pulmonary dioxide production (VCO2), the relationship between them (RER: VCO2/VO2), and minute ventilation (VE) were continuously measured.All patients performed the MST and CPET while connected to a validated  system ft-student test was used to compare the characteristics between CPET and MST. The unpaired Student’s t-test or Mann–Whitney U-test was used to compare the characteristics of CPET and MST between EG and VG groups. The Pearson’s coefficient of correlation was performed to study the correlation between VO2peak and age, height, weight, and distance walked. Subsequently, the multiple backward linear regression analysis was performed to identify predictors of the dependent variable (VO2peak) measured during MST. Age, height, weight, sex, and distance walked during MST were tested as independent variables. To validate the equation, the measured VO2peak of the VG patients was compared to the predicted VO2peak. The intraclass correlation coefficient (ICC) and its 95% confidence interval (CI) were calculated using an absolute agreement definition, to evaluate the relative reliability between VO2peak measured and predicted. The agreement between VO2peak measured and predicted was assessed by Bland–Altman analysis. A sample size of at least 15 patients per variable was used in the final equation model developed using linear regression. The post hoc power of the validation group was calculated.The Shapiro–Wilk test was used to determine the normality of the data. The variables were described by mean ± SD or median (IQ 25 to 75%), according to the normality of the data. The paired p-value less than 0.05 was considered statistically significant.SPSS version 20  statistical software was used in the study. A Table 1.A total of 153 children and adolescents with asthma were invited to participate, of which 97 were included . Assuming a ratio 2:1, a total of 68 patients were randomly included in the equation group (56% boys). Another 29 VG patients (61% boys) were evaluated to validate the equation. They also had normal lung function and a median GINA step of 3 (2 to 4), 2peak at MST (1.9 ± 0.6 L/min) than they did at CPET (1.5 ± 0.4 L/min), p < 0.001. A similar result was observed for their heart rate , but not statically difference for RERpeak between tests .Of 84 (86%) of 97 patients performed the CPET according to the recommendations. The variables of CPET and MST were compared to be sure that a field test (the MST) had similar responses to the gold standard test. They had a significantly higher VO2peak, distance walked, age, weight, and height  for the EG . The final model of the linear multiple regression analysis included gender, weight, and distance walked during MST. The standardized coefficient β is shown in Table 2.All patients stopped the MST at maximal effort . There was a significant association between VO2: 0.87; SEE: 0.24;Adjusted RGender: boys = 1; girls = 0;Weight: kg;DW: distance walked at MST, in meters.Table 3). No adverse events were reported during or after the MST. The VO2peak measured during the MST of the validation group (1.9 ± 0.5 L/min) was not different of the VO2peak predicted by the equation (2.0 ± 0.5 L/min), p = 0.67 . The post hoc power between the VO2peak measured and predicted was 83%.There was no difference between the equation and validations groups in the variables at the peak of MST . The Bland–Altman analysis showed a mean bias of -0.15 L/min, with the limits of agreement ranging from -0.65 to 0.35 L/min.The ICC of the VO2peak; (ii) There was no difference between the equation and validations groups in the variables at the peak of MST; (iii) The ICC of the VO2peak measured and predicted in the validation group was highly significant; (iv) MST elicited higher peak VO2 and heart rate than CPET.The main results of the present study were: (i) gender, weight, and DW during MST explained 87% of the variance in VO2peak and heart rate during MST compared to the CPET. This result occurred because the CPET was performed on a cycle ergometer and MST is a walk/run test. The CPET is considered a gold standard assessment for exercise capacity, regardless of whether it is performed on a treadmill or a cycle ergometer , it is similar to the results of 2peak) between VO2peak measured and predicted in a study of patients with adolescents with asthma.This is the first study to determine an equation for prediction of VO2peak in children and adolescents from 6 to 18 years with asthma presented psychometric characteristics similar than other equations in adolescent and adult patients with asthma. The MST seems to be an appropriate test to evaluate pediatric patients, because it is performed at a wide range of speeds, and this avoids floor or ceiling effects , which reduced the external validity for the equation in this subgroup of patients. The CPET was not properly performed in all participants, but more than 80% of them performed it. There was not included a control group of healthy participants, and whether differences exist between healthy and asthmatic youth cannot be determined. However, considering that equations for this specific group was not available until now, the inclusion of only asthmatic still relevant to determine an equation for pulmonary chronic conditions.2peak based in the MST in pediatric patients with asthma. Therefore, this test can be a useful alternative to assess VO2peak in clinical settings.The developed equation indicated high correlation and low bias in the prediction VOFL had substantial contributions to the study including: conceptualization/design; supervision/oversight; funding acquisition; formal analysis; drafting significant parts of the work. MR and RS had substantial contributions to the study including: methodology, investigation, data curation. RR-D, GW, and DS had substantial contributions to the study including: critically revising it so as to contribute to the interpretation. MvB, HH, SDC, and TT had substantial contributions to the study including: analysis and interpretation of research data drafting significant parts of the work or critically revising.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.


Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disease and is characterized by gradual cyst growth and expansion, increase in kidney volume with an ultimate decline in kidney function leading to end stage renal disease (ESRD). Given the decades long period of stable kidney function while cyst growth occurs, it is important to identify those patients who will progress to ESRD. Recent data from our and other laboratories have demonstrated that metabolic reprogramming may play a key role in cystic epithelial proliferation resulting in cyst growth in ADPKD. Height corrected total kidney volume (ht-TKV) accurately reflects cyst burden and predicts future loss of kidney function. We hypothesize that specific plasma metabolites will correlate with eGFR and ht-TKV early in ADPKD, both predictors of disease progression, potentially indicative of early physiologic derangements of renal disease severity.To investigate the predictive role of plasma metabolites on eGFR and/or ht-TKV, we used a non-targeted GC-TOF/MS-based metabolomics approach on hypertensive ADPKD patients in the early course of their disease. Patient data was obtained from the HALT-A randomized clinical trial at baseline including estimated glomerular filtration rate (eGFR) and measured ht-TKV. To identify individual metabolites whose intensities are significantly correlated with eGFR and ht-TKV, association analyses were performed using linear regression with each metabolite signal level as the primary predictor variable and baseline eGFR and ht-TKV as the continuous outcomes of interest, while adjusting for covariates. Significance was determined by Storey’s false discovery rate (FDR) q-values to correct for multiple testing.Twelve metabolites significantly correlated with eGFR and two triglycerides significantly correlated with baseline ht-TKV at FDR q-value < 0.05. Specific significant metabolites, including pseudo-uridine, indole-3-lactate, uric acid, isothreonic acid, and creatinine, have been previously shown to accumulate in plasma and/or urine in both diabetic and cystic renal diseases with advanced renal insufficiency.This study identifies metabolic derangements in early ADPKD which may be prognostic for ADPKD disease progression.www.clinicaltrials.gov identifier: NCT00283686; first posted January 30, 2006, last update posted March 19, 2015.HALT Progression of Polycystic Kidney Disease (HALT PKD) Study A; Clinical The online version of this article (10.1186/s12882-019-1249-6) contains supplementary material, which is available to authorized users. Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disorder and affects 1/1000 individuals. It is characterized by gradual enlargement of numerous cysts in the kidneys over decades, and the disease process begins long before loss of estimated glomerular filtration rate (eGFR) occurs. There are at least three definite genetic causes of ADPKD. The majority of ADPKD cases ~ 75%) are caused by mutations in polycystin 1 . Recentl% are cauThe course of ADPKD is variable depending not only on which gene is mutated  and the With the continuing advent of new therapeutic approaches, identifying patients at risk for progression to ESRD has become paramount. The Consortium for Radiologic Imaging Studies of Polycystic Kidney Disease (CRISP) study established ht-TKV as assessed by magnetic resonance imaging as an accurate and reliable measure of renal cyst burden, which also strongly predicts those patients who will progress to advanced chronic kidney disease (CKD) . Ht-TKV IRBs at each participating center approved the HALT-A clinical trial. These included University of Colorado Health Sciences Center, Denver (Aurora), Colorado; Emory University School of Medicine, Atlanta, Georgia; University of Kansas Medical Center, Kansas City, Kansas; Tufts Medical Center, Boston, Massachusetts; Beth Israel Deaconess, Boston, Massachusetts; Cleveland Clinic, Cleveland, Ohio; Mayo Clinic, Rochester, Minnesota. All subjects provided written consent, and parental consent was required and obtained for all participating subjects between the ages of 15 and 18 years of age at each participating site. Two Data Coordinating Centers were involved in the HALT-A clinical trial: Washington University, St. Louis Missouri and the University of Pittsburgh, Pittsburgh, Pennsylvania. IRB study approval was obtained at each Data Coordinating Center.clinicaltrials.gov) Study A is a double-blind, placebo-controlled trial consisting of 588 hypertensive participants  [This study was approved by the Institutional Review Boards both at the University of California, Davis, and at the University of Chicago. The HALT Progression of Polycystic Kidney Disease  . BrieflySamples and standards were analyzed as described . Peaks wSamples and standards were analyzed as described . The samMetabolomic and lipidomic analyses were performed on all 277 plasma samples by gas chromatography-time of flight mass spectrometry (GC-TOF/MS). Patient samples were analyzed along with 35 (metabolomics) and 36 (lipidomics) splits of the “reference” (pooled) quality control sample placed evenly in between patient samples for quality control assessment. Run order of patient samples was randomly determined. Note that for the lipidomics data, one patient sample showed anomalous behavior and was markedly dispersed from the rest of the patient samples  and thus removed from further analysis.A total of 116 plasma metabolites and 101 lipids were identified, quantitated, and included in downstream statistical analysis. Also 174 unknown plasma metabolites were analyzed separately in the same manner described below. For lipidomic data, lipids quantified in fewer than 50% of samples were discarded from downstream analysis to reduce the bias that could be induced by imputation due to the detection limit. Unobserved values for any remaining undetected lipid below the pre-defined detection limit were imputed as one-half of the lipid-specific minimum of the observed values prior to normalization and statistical inference tests.2 transformed to reduce the influence of extreme values and to meet the homogeneity of variance assumption.For between-sample normalization, the intensity values for each sample were summed, and the median value of the sums across all samples were determined. The intensity values of each sample were then scaled such that the sum of the scaled intensities equaled the median value of all samples. Thus, the sum of the scaled intensities was the same for all samples. Normalized intensity values were then logp-value < 0.05. Multicollinearity was inspected for possible collinearity issues using variance inflation factor (VIF) score with a suggestive threshold of 2.5. We investigated potential effect modifiers such as sex and PKD genotype by adding interaction terms in regression models. When an interaction term reached statistical significance, post-hoc analysis was performed stratified by the effect modifier. Five missing values for BMI were imputed with the mean of the observed values. We used the Storey’s false discovery rate (FDR) q-values to correct for multiple testing [Prior to assessing the association between metabolite intensity levels and disease severity (baseline eGFR and ht-TKV), we assessed the distributions of eGFR and ht-TKV and then log transformed them to meet normality and homoscedasticity of linear regression models. To identify individual metabolites whose intensities are significantly associated with eGFR and ht-TKV, we performed association analyses using linear regression with each metabolite signal level as the primary predictor variable and baseline eGFR and ht-TKV as the two outcomes of interest, analyzed separately. All analyses accounted for the covariates attributed to the HALT-A study design including age, sex, PKD genotype (PKD1 or PKD2), and participating clinical site of sample collection. Other HALT covariates, including baseline systolic and diastolic blood pressure, weight, body mass index (BMI), height-adjusted liver volume, and urine levels for sodium, potassium, and creatinine, were evaluated for inclusion in multiple linear regression models to determine whether any of those covariates are influential in estimating the magnitude of association of metabolite levels with the outcomes. Variable inclusion and exclusion were determined based on Akaike weight-based relative importance (RI) scores , with a  testing . All staDue to the large number of samples  influential in association between metabolites and outcomes, eGFR and ht-TKV  and final variable selection was carried out via likelihood ratio test (LRT). Age, genotype, sex, site of sample collection, and BMI were significantly  above for eGFR and to the model depicted by equation 2) above for ht-TKV. For eGFR, none of the metabolites showed significant effect modification by sex or genotype at FDR q-value< 0.05  for eGFR as previously described, while eGFR was added as an additional covariate to the model depicted by equation 2) for ht-TKV. When controlled for ht-TKV as a covariate, four metabolites were determined to be significantly associated with eGFR at FDR q-value < 0.05, including pseudo-uridine, indole-3-lactate, creatinine, and uric acid . Indole-3-lactate and indole-3-acetate are both present in the urine of a juvenile cystic mouse model [Several metabolites, including indole-3-lactate and likely the unknown compound 191801, are microbial metabolites, the former being the end-product of tryptophan metabolism produced by gut microbes , and these model . MeanwhiWe found significant associations between two large chain length triglycerides and baseline values of ht-TKV in ADPKD patients. This kind of association between specific triglycerides and a marker of progression risk has not been previously described in early ADPKD patients with relatively intact kidney function. Lipid dysregulation is associated with ADPKD in mouse models, including impaired fatty acid oxidation in the kidney  and upreWe also evaluated the metabolites when accounting for sex and genotype as possible effect modifiers in their association with outcome. We found evidence that metabolite-outcome associations might be altered by sex or by genotype. However, none of the metabolites were found to be significant at FDR q < 0.05 when controlling for multiple testing so neither sex nor PKD genotype are likely a differentiating factor in the association of all or most metabolites with those outcomes, but nonetheless further investigation is warranted.In summary, we have shown the presence of metabolites and lipids which highly correlate with eGFR and ht-TKV in ADPKD patients with normal kidney function. These metabolites and lipids can be useful for uncovering metabolic derangements, as well as reprogramming, which appear to occur early in disease. With further evaluation we may find additional predictors of early kidney dysfunction that are potentially less costly than measuring ht-TKV, which is currently the only measurement known to predict the rate of disease progression. Perhaps these metabolites will allow for prognostication and stratification of patients into slow and rapid progressors to ESRD in ADPKD.This study identifies metabolic derangements in early ADPKD which may be prognostic for ADPKD disease progression. Such data will be useful for future studies designed to predict outcome of disease based on such early metabolic changes and will likely lead to new therapeutic paradigms for a disease with quite limited therapeutic options.Additional file 1:Figure S1. Boxplots of the distribution of Metabolite intensity levels  for all 35 reference samples of metabolomics data. Figure S2. Boxplots of the distribution of Lipid intensity levels  for all 36 reference samples of lipidomics data. (PPTX 103 kb)Additional file 2:Table S1. Significant (p < 0.05) effect modification of patient’s sex/genotype on metabolite/lipid-eGFR association. Table S2. Significant (p < 0.05) effect modification of patient’s sex/genotype on metabolite/lipid-ht-TKV association. Table S3. Plasma metabolites and lipids significantly (p < 0.05) associated with (a) eGFR, after controlling for ht-TKV as a covariate and with (b) ht-TKV, after controlling for eGFR as a covariate. (XLSX 19 kb)


T-test, variance analysis, chi-square test, Pearson linear correlation analysis, and logistic regression, there was a good correlation between pulmonary artery systolic pressure and thyroid stimulating hormone (TSH) and receptor antibodies (TRAb)  , but there was no significant correlation between the pulmonary artery systolic pressure and other thyroid-related parameters . Based on variance analysis, PASP rose as the level of TRAb gets higher. What is more, patients with HHD combined with PH showed a significantly higher serum level of TRAb; moreover, serum TRAb concentration was remarkably correlated with the PASP level. Therefore, TRAb participates in the process of pulmonary hypertension caused by hyperthyroid heart disease.Hyperthyroidism is a clinical state that results from increased thyroid hormone levels which has a significant impact on cardiac function and structure. Graves' disease is the most common cause of hyperthyroidism in iodine-replete areas. Hyperthyroid heart disease may be associated with pulmonary hypertension in patients who have overt hyperthyroidism. To investigate the association of pulmonary hypertension induced by hyperthyroid heart disease and autoantibody, one hundred and one cases with hyperthyroid heart disease who were consecutively admitted to the inpatient department of endocrinology and metabolism of the Shandong Provincial Hospital between November 2014 and April 2018 were collected and analyzed statistically. According to the Independent samples  Graves' disease (GD) is one of the most common endocrine diseases with prominent cardiovascular manifestations, which is responsible for 80% of hyperthyroidism in areas of iodine sufficiency. The population is generally susceptible, especially prevalent in women aged 40–60 years .The pathogenesis of GD is complex since both genetic and environmental factors can result in the attack of GD among susceptible populations, but thyrotropin receptor antibody (TRAb) ultimately accounts for hyperthyroidism . The bonIt has been reported that excessive secretion of thyroid hormones could give rise to deleterious clinical consequences for the cardiovascular system. Except for thyroid gland, susceptive receptors exist in both myocardial and vascular endothelial tissues, enabling even slight changes in circulating thyroid hormones to generate infaust influence on multiple organs and systems. Abnormal combination of receptors in the cardiovascular system could create a hyperdynamic state , which iPositive rates of other thyroid autoantibodies, such as anti-thyroid peroxidase antibody (TPOAb) and anti-thyroglobulin antibody (TgAb), are also significantly increased in Graves' disease patients, which demonstrate that Graves' disease is an autoimmune thyroid disease from another point of view.For the past few decades, mounting evidences have demonstrated that hyperthyroid heart disease (HHD) may be associated with pulmonary hypertension (PH) in patients who have overt hyperthyroidism. According to several observational studies, the prevalence of pulmonary hypertension in patients with hyperthyroidism varies between 35 and 47% .Pulmonary hypertension (PH) is an infrequent, progressive, and fatal disease characterized by abnormally sustained elevations in pulmonary artery pressure and progressive increase in pulmonary vascular resistance. In most cases, due to the lack of early-stage clinical manifestations, the patients with pulmonary hypertension once exhibit obvious clinical symptoms and signs, dyspnea on exertion (DOE), for instance, has already developed severe decline in right ventricular function, and this is one of the primary causes of hospitalization and death.Therefore, as for patients with HHD complicated with PH, close monitoring and early intervention of pulmonary arterial systolic pressure (PASP) together with thyroid function can prevent the deterioration of hemodynamics as well as the occurrence and development of heart failure .Previous studies have shown that autoimmunity plays a significant role in the pathogenesis of thyroid disorder; meanwhile, the increase of cardiac output is an essential cause of hyperthyroid heart disease with pulmonary hypertension. Nevertheless, to our knowledge, no study to date has examined the contribution of thyroid hormones and autoantibodies to the development of pulmonary hypertension among hyperthyroidism patients with cardiovascular complications. Hence, the aim of this study was to evaluate thyroid autoimmunity and thyroid function in patients suffering from hyperthyroid heart disease complicated with pulmonary hypertension.One hundred and one cases with hyperthyroid heart disease who were consecutively admitted to the inpatient department of endocrinology and metabolism of the Shandong Provincial Hospital between November 2014 and April 2018 were included in the study. Untreated Graves' disease patients at onset were diagnosed with HHD by at least two experienced physicians, on the basis of common clinical and laboratory criteria.Confirmed hyperthyroidism case according to medical history, clinical symptoms and signs, and test resultsPatients were included if they had one or more of the following: arrhythmia such as paroxysmal or persistent atrial fibrillation, frequent ventricular premature beat, conduction block, paroxysmal supraventricular tachycardia; cardiac dilatation; congestive heart failure; angina pectoris or myocardial infarction; and valve prolapse with pathological murmur in the cardiac auscultation areaThe above abnormalities disappeared or improved significantly after clinical cure or complete remission of hyperthyroidismExcluded other causes of heart diseaseAll patients were diagnosed with hyperthyroidism and hyperthyroid heart disease for the first time, and they did not receive any kind of treatmentPulmonary hypertension (PH) was defined by a mean pulmonary arterial systolic pressure ≥25 mmHg at rest.Inclusion criteria include the following: (1) patients who met the diagnostic criteria for HHD were enrolled in the study and (2) patients with complete data.Exclusion criteria include (1) patients with a medical history of thyroidectomy and (2) patients with a history of untreated hypertension or any disease which would affect the heart condition.For each patient, the following tests and examinations were undertaken: detailed physical examination, blood pressure, heart rate, pulse, weight, routine blood count, routine urine test, liver and kidney functions, electrocardiogram, and chest X-ray films (PA position).Vmax2+10 mmHg.Transthoracic echocardiographic evaluations were performed by experienced technicians, who were blinded to the clinical findings, with a Vivid 7 echocardiographic ultrasound device  to measure the peak flow velocity of the tricuspid valve which was used in the calculation of the pulmonary artery systolic pressure using the modified Bernoulli equation: (PASP)=4 × All measurements were performed using the chemiluminescent immunometric assay (Roche Diagnostics GmbH 2010). The interassay coefficients of variation (CVs) for FT3, FT4, and TSH were 4.3%, 4.6%, and 4.8%. Interassay CVs for FT3, FT4, and TSH were 6.1%, 6.4%, and 6.7%. The patients have no history of iodine exposure, and no drugs affecting the hypothalamic-pituitary-thyroid axis were taken prior to blood collection –17.T-test and chi-square test were used for comparisons of means between two groups of HHD patients. To investigate the contribution of thyroid hormones and autoantibodies to the development of PH among HHD patients, logistic regression analyses were performed for all subjects. Pearson linear correlation was used to evaluate the relationship with various tested markers with PASP levels among HHD patients with PH. Variance analyses were adopted to analyze whether different levels of TRAb have a significant effect on PASP. All analyses were calculated by Statistical Product and Service Solutions 22.0 (SPSS 22.0). P value less than 0.05 was considered as statistically significant.For quantitative variables, data were presented as mean ± standard deviation (SD). The independent samples A total of 101 patients  were enrolled in the final analysis based on the inclusion and exclusion criteria. Average age was 46.9 ± 13.5 years, with age ranging from 17 to 74. Pulmonary hypertension, defined as PASP ≥ 25 mmHg, was present in 72 patients with HHD, accounting for 71.3%. All the cases tested positive for at least two kinds of antibodies, 100 cases tested positive for TRAb, 93 cases tested positive for TPOAb, and 65 cases tested positive for TgAb.P=0.012). Out of the total 101 patients, 13 patients had the case history of hypertension (12.87%); however, all of them took drugs for hypertension by prescription and had their blood pressure under control. 71 patients had varying degrees of goiters. 22 male patients had either a current or former cigarette smoking history of at least 10 pack-years. 30 patients exhibited the symptom of atrial fibrillation.The clinical and biochemical characteristics of the patients with HHD are shown in P=0.026; after adjusting for age, FT3, FT4, and TSH), as illustrated in We also found that serum TRAb levels remained independently associated with pulmonary hypertension among patients suffering from HHD  between TRAb with HHD complicated with PH, as shown in the scatter diagram of PASP and serum TRAb levels (P > 0.05). In the next place, PASP positively correlated with a maximum velocity of blood flow in the artery of the tricuspid valve  and maximum pressure on both sides of the tricuspid valve , respectively.Correlation analyses of pulmonary arterial systolic pressure (PASP) with laboratory parameters were summarized in b levels . HoweverQ1: <11.596, Q2:11.596–40.000, and Q3: ≥40.000 IU/l), and then the levels of PASP were compared by using variance analysis; the results are presented in Due to the absence of the exact number of pulmonary arterial systolic pressure in hyperthyroid heart disease patients without pulmonary hypertension, correlation analyses together with variance analysis were done between the patients suffering from HHD combined with PH instead of all subjects. We arbitrarily divided patients suffering from HHD combined with PH into 3 groups according to tertiles of the serum TRAb level (Q3 was found to have significantly higher mean levels of PASP (37.1 ± 7.6 vs. 39.5 ± 8.3 vs. 44.5 ± 10.1); that is to say, it can be confirmed that PASP rises as the level of TRAb gets higher. In addition, TPOAb, maximum velocity of blood flow in the artery of tricuspid valve, and maximum pressure on both sides of the tricuspid valve increase as the level of TRAb goes up.The comparisons of data obtained from patients who were divided into 3 groups according to the serum TRAb content are shown in While thyroid autoimmunity has been reported to be associated with thyroid hormone disorders as well as cardiovascular complications, to the best of our knowledge, this is the first retrospective study conducted investigating the association between autoantibodies and HHD complicated with PH. This study demonstrated that patients with HHD combined with PH showed a significantly higher serum level of TRAb; moreover, serum TRAb concentration was remarkably correlated with the PASP level.Abnormal increase of circulating thyroid hormone has a close relationship with target organ damage of hyperthyroidism, for instance, the undesirable cardiovascular complications. Elevated levels of thyroid hormone can directly act on myocardium and exert positive chronotropic and inotropic effect by enhancing cardiac excitability and myocardial contraction. In addition to direct effect, high concentrations of thyroid hormone could also stimulate angiogenesis; moreover, it accelerates vascular smooth muscle cell relaxation.Hyperthyroidism heart disease is a kind of cardiomyopathy due to metabolic disorder, which turns out to be the consequence of long-term invalid control or exacerbation of hyperthyroidism, characterized by increased pulmonary vascular resistance. Pulmonary hypertension is a hemodynamic and pathophysiological condition in which pulmonary artery pressure rises and eventually exceeds a certain threshold and may ultimately lead to right heart failure even death if without proper treatment . MultiplThese results aforementioned suggested a significant correlation between TRAb and the pathogenesis of pulmonary hypertension among HHD patients. Furthermore, the rate of HHD combined with PH patients who were tested positive for at least two types of autoantibodies reached up to 100% in our research. In this regard, Chu et al.  have conComparatively speaking, the positive rate of TPOAb and TGAb in the two studies aforementioned was only 54% and 35%, respectively. By contrast, we found a higher occurrence of autoimmune hyperthyroidism (99% for TRAb and 100% for at least two types of thyroid autoantibodies). In addition, there was no significant correlation between PASP and FT3, FT4, TSH, anti-TPO, and anti-TG in the HHD cases we included. However, TRAb strongly correlated with PASP. Therefore, it can be demonstrated that the elevation of pulmonary arterial systolic pressure is related with the increasing level of serum TRAb.Although no association of FT4 with HHD complicated PH was observed in our study, a few recent retrospective cohort studies have arrived at different conclusions. Okosieme et al. observed increased risk of mortality and major adverse cardiovascular events with increasing concentrations of FT4 in serial measurements, but not for increasing concentrations of TSH. Moreover, risks of mortality and major adverse cardiovascular events were higher in older patients than in younger patients . By contOn the contrary, our results showed that all the patients suffering from HHD complicated with PH developed various degrees of tricuspid regurgitation. These findings were consistent with what was known about the pathogenesis of right heart failure –30. TricGraves' disease is an autoimmune thyroid disorder mediated by TRAb, leading to superabundant synthesis and secretion of T3 and T4, which have been verified as the main cause of hyperthyroidism. Consequently, all patients with hyperthyroidism are supposed to reexamine the TRAb level during monthly or quarterly regular follow-up. The persistent elevation of TRAb indicates that hyperthyroidism is in the active stage, while the remission phase is normally accompanied by the decline of TRAb , 33. RobNicolls et al.  revealedBeyond that, multiple research studies have highlighted the value of physical or emotional stress as a trigger of Graves' disease-related hyperthyroidism. It is commonly believed that the deterioration or relapse of Graves' disease is preceded by at least one stressful event. In addition, the more stressed these patients are, the shorter is the interval between stressful events and Graves' disease hyperthyroidism. Above all, the interplay between environmental factors and susceptibility genes eventually leads to disruption of the immune tolerance –43.It has been repeatedly reported that, as long as under standard treatment of hyperthyroidism, a great mass of patients developing HHD accompanied by PH would exhibit different degrees of reduction in PASP after the thyroid function returns to normal , 45.In summary, our findings indicated that TRAb is closely associated with HHD combining with PH, all of which is obvious interrelated to hemodynamic alterations and myocardial damage attributed to changes in the serum thyroid hormone level. At the same time, there might be common immune and genetic mechanisms between them, which needs further investigation. The screening of pulmonary arterial pressure in patients with hyperthyroid heart disease may be able to boost the detection rate of patients with pulmonary hypertension. Furthermore, it is necessary for us to apply the research to the epidemiology and pathogenesis of pulmonary hypertension among hyperthyroidism patients, in order to improve the clinical scientific research level in this field.There are a few limitations of our study: (1) although ultrasound showed no sign of increased PASP, the lack of accurate data of pulmonary arterial systolic pressure among hyperthyroid heart disease patients without pulmonary hypertension is the main limitation of our study. On the contrary, invasive ways such as the Swan–Ganz catheter may lead to a small risk of serious morbidity. Consequently, we used cardiac ultrasound to diagnose PH in the present study; (2) on the one hand, we only enrolled Han nationality individuals in our study, and on the other hand, the cohort of subjects was relatively small. Therefore, our conclusion may not apply to other populations.All in all, except for the effect of thyroid hormone as well as autoantibodies on the cardiovascular system, autoimmune-mediated pulmonary arterial systolic pressure elevation may have a vital function in hyperthyroid heart disorder-linked pulmonary hypertension.


There was no correlation between the ABPM Systolic Blood Pressure (SBP)-lowering effect and the total number of ablated spots in renal arteries (17.7 ± 6.0) either at 12  or at 24 months . However, correlations between systolic BP-lowering effect and the number of ablation spots performed in the distal segment and branches were significant at the 12  and 24 months  follow-up. Our findings indicate a substantial correlation between the numbers of ablated sites in the distal segment and branches of renal arteries and the systolic BP-lowering effect in the long-term.We searched for an association between changes in blood pressure (BP) at 12 and 24 months after renal denervation (RDN) and the different patterns of ablation spots placement along the renal artery vasculature. We performed a post-hoc analysis of a 24-month follow-up evaluation of 30 patients who underwent RDN between 2011 and 2012 using our previous database. Patients who had (i) resistant hypertension, as meticulously described previously, and (ii) Chronic kidney disease (CKD) stages 2, 3 and 4. Correlations were assessed using the Pearson or Spearman correlation tests as appropriate. The mean change in systolic ambulatory BP monitoring (ABPM) compared to baseline was −19.4 ± 12.7 mmHg at the 12th ( Catheter-based renal denervation (RDN), targeting the renal afferent and efferent sympathetic nerves is an established alternative treatment option for patients with hypertension ,2,3,4. TBased predominantly on experimental work in large animal models it has become evident that increasing the number of radiofrequency (RF) lesions in the main renal artery of pigs, although consistently effective in reducing renal norepinephrine (NE) and axon density relative to naive kidneys, this was less reliable in achieving a clear dose-response relationship regarding BP . In contThe SPYRAL HTN-OFF MED  trial, iIn the SPYRAL HTN-OFF MED  trial, oPreviously, our group reported that another different RDN approach was also efficient in reducing BP in resistant hypertensive patients with chronic kidney disease in mid-  and longWe must highlight that this study is a secondary analysis from our previous published data . We cond2 and 89 mL/min/1.73 m2 ; and [We performed a post-hoc analysis of a 24-month follow-up evaluation of 30 patients who underwent RDN from June 2011 to December 2012. Patients were recruited from a university hospital and public health network of the country . The incia); and .Exclusion criteria included pregnancy; valvular heart disease with significant hemodynamic consequences; use of warfarin; stenotic valvular heart disease; acute myocardial infarction, unstable angina, stroke, or transient ischemic attack within the previous six months; renovascular anomalies ; and diabetes mellitus type 1 or other secondary causes of hypertension. Patients underwent a complete medical history and physical examination. Hypertension was diagnosed based on national guidelines in force at that time ,18. PatiThe acute procedures pre- and post-RDN, as well as, anticoagulation, analgesia and anesthesia techniques, were described previously . AngiogrAfter the procedure, patients remained hospitalized for 24 h in an inpatient ward. We followed-up patients weekly for the first month, monthly from the second to the sixth months, bimonthly from the seventh to the 12th months, and quarterly during the second year. Doppler ultrasound was performed to evaluate the anatomy of the renal arteries of the patients at the first and sixth months after the RDN. ABPM was performed  at the 12th and 24th months after the procedure to evaluate the BP control and the effectiveness of RDN.2, respectively. For these three patients, the presented data were collected until their last follow-up visit at 12 months. After these patients began hemodialysis, they were censored from the study. Twenty-seven out of 30 patients completed the 24 months of follow-up, and three patients required chronic renal replacement therapy. One patient-initiated dialysis at the 13th month of follow-up after an acute renal insult associated with pulmonary sepsis. The other two started dialysis at the 14th month, also after acute renal injury episodes: one related to a perforated gastric ulcer and the other following decompensation of heart failure and lung infection. Of note, their eGFR at baseline were 17, 15 and 16 mL/min/1.73 mn) and percentage (%). Continuous variables  were expressed as mean ± SD, 95% CI or median (Q1–Q3). Comparisons between ABPM measurements at the 12th and 24th month follow-up and their respective baseline values, for systolic and diastolic ABPM, were performed using analysis of variance (ANOVA) for repeated measures and complemented by a post hoc test. Differences between 12- and 24-month follow-up values were compared by the unpaired t-test, due to three missing values at the 24th month post-RDN and were expressed as mean and 95% CI. Categorical variable frequencies were compared with the Fisher test. p values < 0.05 were considered significant. Correlation between the number of ablated spots per renal arteries regions and ABPM measurements was assessed by the Pearson correlation test, in the case of a Gaussian distribution, or Spearman’s correlation test as an alternative. All statistical analyses were performed using the program GraphPad Prism v 8.0 .Categorical variables  were expressed as the number of patients  and −21.3 ± 14.1 mmHg (95% CI: −26.9 to −15.7) at the 24th month (p < 0.0001) follow-up , as shown in p = 0.0015) and −8.1 ± 11.7 mmHg (95% CI: −12.8 to −3.5) at the 24th month (p = 0.0013) follow-up , as shown in The mean change in systolic ABPM compared to baseline was −19.4 ± 12.7 mmHg (95% CI: −24.1 to −14.6) at the 12th month  or at 24  months post-procedure . Correlations between SBP fall and the number of treatments in the middle segment (5.4 ± 2.0) at 12  and 24  months follow-up were not significant. However, there were moderate correlations between the numbers of ablated sites in distal portion (5.2 ± 2.1) of renal arteries and SBP-lowering effect at 12  and 24  months follow-up. The same could be observed in branches (1.9 ± 2.9 treated sites) and SBP-lowering effect at 12  and 24  months follow-up. Surprisingly, the correlations between the number of treated sites in the distal section and branches together (7.1 ± 4.6) became stronger to both follow-up time points -lowering effect and the total number (17.7 ± 6.0) of ablated spots in renal arteries either at 12  in renal arteries , and middle segments . Correlation between DBP fall and the number of treatments in the distal segment (5.4 ± 2.0) was significant at 12  and 24  months follow-up. For the number of treatments delivered in branches, correlation between DBP fall and the number of treatments was significant at 12  and 24  months follow-up. A similar correlation was observed for DBP-lowering effect and the number of ablated spots in distal segment summed to branches at 12  and 24  months follow-up as shown in There were no correlations between ABPM diastolic blood pressure (DBP)-lowering effect at 12 (−8.4 ± 11.9 mmHg) and at 24 (−8.1 ± 11.7 mmHg) months post-procedure and the total number of ablated spots  per branch resulted in even greater NE reduction (−82% ± 18%) . FurtherWe also found that the sum of ablations performed in distal segment and branches presented a significant correlation to the SBP-lowering effect at the 12th and 24th months after RDN C,D. ThisAnother crucial point is that our patients presented CKD beyond hypertension, which was not observed in the SYMPLICITY HTN-1, HTN-2 and HTN-3 trials neither in the SPYRAL HTN-OFF and HTN-ON MED trials. As we know, the sympathetic activation is a hallmark of the essential hypertensive state occurring early in the clinical course of the disease ,30. In CThe relatively small sample is a limitation of this study. However, the present post hoc analysis is unique in the literature as it addresses the correlation between the number and location of RDN ablations and the sustained BP-lowering effect in the long-term. Nevertheless, because this analysis was performed from an uncontrolled study, our findings should be interpreted with caution. In addition, this is a post hoc analysis and, as such, the primary research  was not Our findings indicate that RDN in resistant hypertensive CKD patients provided a significant sustained reduction in BP. More importantly, we observed a clear and substantial correlation between the numbers of ablated sites in the distal segment and branches of renal arteries and the systolic BP-lowering effect in the long-term. Although encouraging, our data are from a post hoc analysis and must be validated in a larger and properly designed study to address this specific aim.The positive results from SPYRAL HTN-OFF MED, RADIANCE SOLO and SPYRAL HTN-ON MED signify re-ignited the RDN field. In concordance of these studies, our data corroborate the perspective that the number of treated sites in branches is extremely important for the SBP-lowering effect.