Serum osteoprotegerin is an independent marker of central arterial stiffness as assessed using carotid–femoral pulse wave velocity in hemodialysis patients: a cross sectional study

The cross-sectional study was conducted at a medical center in Hualien between March and July in 2015 and was approved by the Institutional Review Board of Tzu-Chi Hospital. Patients aged more than 20 years and who had undergone HD for at least 3 months with standard 4-h dialysis three times per week were included. The same type of high-flux polysulfone disposable artificial kidney (FX class dialyzer, Fresenius Medical Care, Bad Homburg, Germany) was used by all patients. Patients who had any infection, acute myocardial infarction, stroke, peripheral arterial occlusive diseases, pulmonary edema, arrhythmias and atrial fibrillation at the time of enrollment or who refused to participate were excluded from the study. DM was defined based on a history of antidiabetic drug use, and hypertension was defined based on a history of or receiving antihypertensive agents. Coronary artery disease (CAD) was diagnosed through coronary angiography. The drug use history was collected by a review of medical records, which included the use of angiotensin-converting enzyme inhibitors (ACEIs), angiotensin receptor blockers (ARBs), calcium channel blockers (CCBs), β-blockers, and statins.

Body weights were measured in light clothing after HD, and height was measured to the nearest half centimeter without shoes. Body mass index (BMI) was calculated as body weight (kilograms) divided by height squared (meters). Blood pressure was measured before the HD session using standard mercury sphygmomanometers.

PWV was evaluated from the time taken for the arterial pulse waveform to propagate from the carotid to the femoral artery, using applanation tonometry (SphygmoCor system, AtCor Medical, Australia). A transcutaneous recording of the pressure pulse waveform in the underlying artery was performed in the supine position after a minimum 10-min rest before the HD session. The cfPWV was calculated using the distance and mean time difference between the two superficial artery sites. A detailed explanation of this device has been provided elsewhere [12]. Patients with cfPWV > 10 m/s were defined as the high arterial stiffness group, according to the European Society of Hypertension and the European Society of Cardiology (ESH-ESC) 2013 Guidelines [10].

Blood samples were collected from the patients for analysis before the HD session. Fasting blood samples (approximately 5 mL) were immediately centrifuged at 3000 g for 10 min. The obtained serum samples were stored at 4 °C and used for biochemical analyses within 1 h of collection. Serum levels of blood urea nitrogen (BUN), creatinine, glucose, total cholesterol, calcium, phosphorus, and C-reactive protein were measured using an autoanalyzer (Siemens Advia 1800, Siemens Healthcare GmbH, Henkestr, Germany). Corrected serum calcium levels were calculated as follows: corrected calcium (mg/dL) = serum calcium (mg/dL) + 0.8 [4 – serum albumin (mg/dL)], and the upper normal limit was chosen as the reference value according to the K/DOQI recommendations [13]. The levels of intact parathyroid hormone (i-PTH) were measured by an autoanalyzer (Diagnostic Systems Laboratories, Webster, Texas, USA), and levels less than two times the upper limit and more than nine times the upper normal limit were defined as low and high parathyroid hormone levels, which are 130 and 595 pg/mL, respectively [14]. Serum OPG levels were measured using a commercially available enzyme-linked immunosorbent assays (ELISA) (eBioscience Inc., San Diego, CA, USA). The fractional clearance index for urea (Kt/V) was measured before and immediately after dialysis using a formal, single-compartment dialysis urea kinetic model.

The normality of the distributions of continuous variables was assessed using the Kolmogorov–Smirnov test. Variables with normal distribution were expressed as mean ± standard deviation and analyzed by the Student’s independent t-test, whereas those not normally distributed were expressed as medians and interquartile ranges and analyzed by the Mann–Whitney U test. Categorical variables were expressed as absolute (n) and relative frequency (%) and analyzed by the χ² test or Fisher’s exact test. Serum OPG levels were analyzed categorically into tertiles, and characteristics of the study population in different OPG groups were evaluated for linear trend using one-way analysis of variance (ANOVA) or the Cochran–Armitage test for trend. The values of cfPWV, HD duration, glucose levels, iPTH levels, and serum OPG levels showed skewed distribution and were logarithmically transformed to achieve normality. Univariable correlations between cfPWV and potential explanatory variables were assessed by Pearson’s correlation coefficient. Multivariable linear and logistic regression analyses were conducted using cfPWV values and the presence of high arterial stiffness (defined as cfPWV > 10 m/s) as dependent variables and potential predictors (age, sex, DM, systolic blood pressure, corrected serum calcium, phosphorus, iPTH, and OPG levels) as independent variables. Data were analyzed using SPSS for Windows (version 19.0; SPSS Inc., Chicago, IL, USA). A p value < 0.05 was considered as statistically significant.

A total of 120 chronic HD patients were enrolled in our study. The mean age of the patients was 63.3 ± 13.7 years, and 53 (44.2%) of them had high arterial stiffness. The clinical and laboratory characteristics of the 120 chronic HD patients with low or high arterial stiffness are presented in Table 1. Compared with the low arterial stiffness group, patients in the high arterial stiffness group had higher systolic blood pressure (p = 0.038), cfPWV (p <  0.001), corrected serum calcium (p = 0.007), and OPG levels (p <  0.001). In addition, the proportion of DM was higher in the high arterial stiffness group (p = 0.001).

The clinical characteristics of the study population were stratified by tertiles of serum OPG levels (Table 2). Increasing tertiles of serum OPG levels showed a significant association with greater height (p = 0.011), male gender (p = 0.008), higher cfPWV (p = 0.020), and lower iPTH levels (p = 0.049).

The continuous variables were found to correlate with cfPWV values among the 120 HD patients, as shown in Table 3. The cfPWV values showed a positive correlation with systolic blood pressure (r = 0.196, p = 0.032), serum glucose (r = 0.182, p = 0.047), and OPG levels (r = 0.281, p = 0.002).

The factors associated with cfPWV values were analyzed among the 120 HD patients. After multivariable adjustment, increasing tertiles of serum OPG levels (β = 0.89 and 1.63 for tertile 2 and tertile 3, respectively, p for trend = 0.035) and the presence of DM (β = 1.83, p = 0.008) were the two independent factors that were associated with cfPWV values. Notably, low iPTH levels were also marginally associated with cfPWV values (β = 1.29, p = 0.055), compared to iPTH levels in the normal range (Table 4).

Table 5 delineates the results of the multivariable logistic regression analysis regarding the factors associated with high arterial stiffness. Age (odds ratio, [OR] = 1.04, 95% confidence interval [95% CI] = 1.01–1.08, p = 0.026), increasing tertiles of serum OPG levels (OR = 5.34 for tertile 2; OR = 7.06 for tertile 3; p for trend = 0.002), DM (OR = 3.65, 95% CI = 1.34–9.98, p = 0.012), low iPTH levels (OR = 2.83, 95% CI = 1.06–7.53, p = 0.038), and high corrected calcium levels (OR = 3.73, 95% CI = 1.16–11.98, p = 0.027) were independently associated with high arterial stiffness.

Furthermore, we analyzed the correlation between corrected calcium levels and cfPWV values based on the tertiles of serum OPG levels, as shown in Fig. 1. The corrected calcium levels showed a positive correlation with cfPWV values in the tertile 3 group (r = 0.408, p = 0.009), but not in the tertile 1 or 2 group.