The aim of the present study was to examine the association between the levels of circulating vascular endothelial growth factor receptor (VEGFR)2 levels, serum lipid composition and plasma receptor for advanced glycation end-products (RAGE) expression in patients undergoing hemodialysis (HD). A total of 50 patients on HD (27 men and 23 women; median age, 66 years; age range 28-88 years; HD mean time, 29.0, 3.9-157.0 months) were enrolled. Age-matched healthy subjects (n=26) were used as the control group. Plasma VEGFR2 and RAGE levels were determined using ELISA. Dyslipidemia (D) in patients on HD was diagnosed according to the Kidney Disease Outcomes Quality Initiative Clinical Practice Guidelines for Managing Dyslipidemias in Chronic Kidney Disease. Circulating VEGFR2, RAGE and serum lipids were compared between dyslipidemic and non-dyslipidemic patients on HD and controls. In patients on HD, the plasma VEGFR2 levels were lower compared with those in the healthy population. D was associated with high plasma VEGFR2 levels. The triglyceride/HDL-cholesterol ratio was strongly associated with plasma VEGFR2 levels. The plasma VEGFR2 concentration was associated with circulating RAGE levels. Therefore, circulating VEGFR2 levels may be partly associated with lipid abnormalities and plasma RAGE levels in patients receiving HD.
Patients with end-stage renal disease (ESRD) on maintenance hemodialysis (HD) are susceptible to changes in their blood vessels. A number of studies have confirmed the high prevalence of atherosclerotic lesions in the arteries of patients on HD (
The protocol used in the present study was performed in accordance with the guidelines described in the Declaration of Helsinki (
A total of 50 Caucasian patients on HD (27 men and 23 women; median age, 66 years; age range, 28-88 years; HD mean time, 29.0, 3.9-157.0 months) who had been treated with maintenance HD for ≥6 months were recruited. All patients underwent HD 3 times for 4-4.5 h per week on low-flux polysulphone-based membranes with a surface area of 1.3-2.1 m2, low-molecular-weight heparin was used as an anticoagulant. The dialysis efficiency was evaluated based on single-pool Kt/V urea nitrogen, according to the National Kidney Foundation-Kidney Disease Outcomes Quality Initiative (K/DOQI) Guidelines (
A total of 26 age-matched self-declared healthy Caucasian volunteers were included in the control group. No substantial health deviations were recorded during the medical interview and physical examinations. The volunteers had not received lipid-lowering drugs, at least in the 3 months prior to the commencement of the study. Control individuals were recruited between June and November 2018, and all controls were students from the University of the Third Age in Nowy Tomyśl and Wolsztyn, Poland. The blood samples were taken on December 10th and 11th, 2018 (according to the protocol).
According to Kouw
Dyslipidaemia (D) in the control group was assessed according to The European Society of Cardiology and the European Atherosclerosis Society guidelines (
Plasma VEGFR2, RAGE, insulin, glycated hemoglobin (HbA1c), glucose levels, lipid profiles, albumin levels and high-sensitivity C-reactive protein (hsCRP) concentration were measured. After a minimum of 8 h of overnight fasting, venous blood was drawn into an EDTA tube and promptly centrifuged at 2,000 x g for 10 min at 4˚C. The obtained plasma was frozen at -80˚C in aliquots until protein analysis was performed. The plasma VEGFR2 levels were measured using the Human VEGFR2 ELISA kit (cat. no. ab213476; Abcam), according to the manufacturer's protocols. No significant cross-reactivity or interference was observed. The limits of VEGFR2 concentration detection were 34.3-25,000 pg/ml. The sensitivity of the assay was <70 pg/ml. The intra-assay coefficient of variation (CV) was 2.5%, and the CV for inter-assay precision was 5.8%. The plasma RAGE levels were measured using the Human RAGE ELISA kit (cat. no. ab190807; Abcam). All measurements were performed in duplicate. Plasma insulin concentrations were determined using the electrochemiluminescence immunoassay method, according to the manufacturer's protocol (Cobas E411; cat. no. 12017547122; Roche Diagnostics GmbH). Homeostasis model assessment of insulin resistance (HOMA-IR) was determined as fasting plasma insulin (µU/ml) x fasting plasma glucose (mmol/l)/22.5(
Participants were categorized as having D when they met the criteria described in the K/DOQI for D (
The normality of distribution of variables was assessed using the Shapiro-Wilk test for each group separately. Numeric variables are expressed as the mean ± standard deviation, or as a median and range, as appropriate; categorical variables are presented as percentages. A Student's t-test was used to compare normally distributed data, or otherwise a Mann-Whitney U test was used. As plasma VEGFR2 levels were not normally distributed, a Spearman's rank correlation was performed to determine the correlation between this variable and the other parameters. Univariate receiver operating characteristic (ROC) curves were evaluated using Medical kit version 4.0.67 (statsoft.pl). Statistical analysis was performed using STATISTICA version 13 (TIBCO Software Inc.). P<0.05 was considered to indicate a statistically significant difference.
The clinical and demographic characteristics of patients in the HD and control groups, with and without D, are summarized in
The laboratory parameters of patients on HD with and without D are presented in
The laboratory parameters of controls with and without D are presented in
There was no difference in plasma VEGFR2 levels between patients with and those without MeS (1.38±0.915 vs. 1.10±0.785 ng/ml, respectively; P=0.338).
In the control group, plasma VEGFR2 levels were significantly positively correlated with TG/HDL-chol ratio and TG levels (
To the best of our knowledge, there are no previous studies showing the association of plasma VEGFR2 levels with lipid abnormalities, MeS, plasma RAGE levels and other biochemical parameters in patients on HD. The following were the major findings of the present study in patients on HD: i) The plasma VEGFR2 levels were lower compared with those in the control subjects; ii) in patients on HD with D, the plasma VEGFR2 levels were higher compared with those in patients on HD without D; iii) there were positive correlations between plasma VEGFR2 levels and lipid abnormalities (Tchol, LDL, TG, Non-HDL, LDL/HDL-chol ratio, TG/HDL-chol ratio and plasma RAGE levels); iv) there was no difference in plasma VEGFR2 levels between patients with and without MeS.
In the present study, lower levels of VEGFR2 were observed in patients on HD compared with the control subjects, which was in line with the results reported by Sepe
There have been several reports of increased carbonyl stress (
MeS commonly occurs in patients on HD, and is closely associated with endothelial cell dysfunction and VEGFR2 expression (
NAFLD is known to be strongly associated with MeS, and is considered as a novel risk factor of cardiovascular events in patients undergoing HD (
D is the primary characteristic of MeS and a common feature amongst patients on HD. In the present study, >50% of the patients on HD were also diagnosed with D, based on the diagnostic recommendations of the K/DOQI guidelines (
The association between LDL-chol and plasma VEGFR2 levels was also examined. Data from previous studies indicated that LDL-chol affects the structure and activity of VEGFR2, but the mechanism underlying this phenomenon is not fully understood. Jin
Recently, the HDL/Tchol ratio was reported to be a more reliable risk factor for atherosclerotic changes compared with Tchol or HDL-chol alone (
The present study has several limitations. First, all patients in this study were Caucasian, and any differences with other ethnicities were not examined. Second, single-center trials are associated with a potential bias. Third, the number of patients and healthy subjects was relatively small.
In conclusion, the findings of the present study demonstrated that circulating VEGFR2 levels were lower in patients on HD compared with those in the healthy population, D was associated with higher plasma VEGFR2 levels, the TG/HDL-chol ratio (an index of atherogenic D) was strongly associated with plasma VEGFR2 levels, and plasma VEGFR2 concentration was associated with circulating RAGE levels. It may be inferred that, in patients on HD, circulating VEGFR2 levels may be partially associated with lipid abnormalities and plasma RAGE levels. However, further studies are required to define the pathophysiological role of circulating VEGFR2 and its precise association with D.
Not applicable.
The datasets used and/or analyzed during the present study are available from the corresponding author on reasonable request.
LN conceived and designed the study, and was involved in data collection and analysis. HD performed the analyses of VEGFR2 and RAGE levels and data analysis. WW was involved in data analysis and performed the statistical analysis. All the authors have read and approved the final version of the manuscript. HD and WW confirm the authenticity of all the raw data.
The protocol used in the present study was performed in accordance with the guidelines described in the Declaration of Helsinki and was approved by the Institutional Review Board of Poznań University of Medical Sciences, Poland. Written informed consent was obtained from all subjects before participation.
Not applicable.
The authors declare that they have no competing interests.
Comparison of plasma VEGFR2 concentration in HD patients and controls. VEGFR2, vascular endothelial growth factor receptor 2; HD, hemodialysis.
ROC curve of VEGFR2 in the prognosis of dyslipidemia in patients receiving hemodialysis. VEGFR2, vascular endothelial growth factor receptor 2; ROC, receiver operating characteristic.
Correlation between plasma VEGFR2 and plasma RAGE expression levels in the entire hemodialysis group. VEGFR2, vascular endothelial growth factor receptor 2; RAGE, receptor for advanced glycation end-products.
Correlation between plasma VEGFR2 and TG/HDL-chol levels in the entire hemodialysis group. VEGFR2, vascular endothelial growth factor receptor 2; TG, triglycerides; HDL-chol, high density lipoprotein cholesterol.
Clinicopathological characteristics of the hemodialysis patients and controls with and without D.
HD patients, n=50 | Controls, n=26 | |||||
---|---|---|---|---|---|---|
Parameter | With D | Without D | P-value | With D | Without D | P-value |
N | 26 | 24 | - | 15 | 11 | - |
Age, years |
65, 47-89 | 66, 28-81 | 0.938 | 72, 60-90 | 68, 61-77 | 0.338 |
Sex | - | |||||
Male | 14 | 13 | 6 | 5 | ||
Female | 11 | 12 | 9 | 6 | ||
DM, n (%) | 11(42) | 15(62) | - | 0 | 0 | - |
Waist to hip ratio |
0.965±0.095 | 0.968±0.096 | 0.933 | 0.918±0.042 | 0.894±0.098 | 0.067 |
Waist to height ratio |
0.615±0.084 | 0.613±0.102 | 0.931 | 0.596±0.076 | 0.531±0.063 | 0.642 |
SBP, mmHg |
123±15 | 125±15 | 0.598 | 130±10 | 128±10 | 0.743 |
DBP, mmHg |
70, 60-90 | 70, 60-80 | 0.825 | 80, 60-90 | 70, 60-100 | 0.442 |
BMI, kg/m2 |
26.6±4.8 | 27.1±5.9 | 0.742 | 27.7±5.42 | 27.3±3.44 | 0.723 |
Period of HD, months |
32.2±20.8 | 41±36.1 | 0.641 | - | - | - |
eKT/V |
1.34±0.23 | 1.36±0.18 | 0.675 | - | - | - |
Residual diuresis, l/24 h, range | 0.5, 0-2.5 | 0.6, 0-2.1 | 0.754 | - | - | - |
aMean ± standard deviation;
bmedian, range. BMI, body mass index; DBP, diastolic blood pressure; DM, diabetes mellitus; D, dyslipidemia; HD, hemodialysis; eKt/V, adequacy of HD; SBP, systolic blood pressure; DBP, diastolic blood pressure.
Values of laboratory parameters in HD patients and controls.
Parameter | HD patients, n=50 | Controls, n=26 | P-value |
---|---|---|---|
VEGFR2, ng/ml |
1.32, 0.330-5.0 | 2.24, 0.700-5.9 | 0.025 |
Albumin, g/dl |
3.65±0.33 | 4.32± 0.21.0 | 0.054 |
Total cholesterol, mg/dl |
175.0±49.0 | 203.0±43.0 | 0.001 |
LDL-cholesterol, mg/dl |
93.3±33.2 | 115.0±36.6 | 0.005 |
HDL-cholesterol, mg/dl |
41.8±12.0 | 68.3±19.2 | 0.005 |
TG, mg/dl |
173.0±83.0 | 94.0±33.0 | <0.0001 |
Non-HDL, mg/dl |
133.0±47.0 | 137.0±38 | 0.676 |
LDL/HDL-cholesterol ratio |
2.35±0.87 | 1.70±0.64 | 0.004 |
HDL/total cholesterol, % |
22.2, 14-50 | 32.9, 55-21 | 0.0005 |
TG/HDL-cholesterol ratio |
4.17, 0.8-8.2 | 1.28, 0.5-3.5 | <0.0005 |
hsCRP, mg/l |
14.1±2.6 | 5.0±1.68 | 0.05 |
RAGE ng/ml |
1.23±1.06 | 0.680±0.048 | 0.035 |
HB, g/dl |
11.4±1.4 | 13.5±0.9 | 0.07 |
aP<0.05;
bP<0.01;
cP<0.001;
dmean ± standard deviation;
emedian, range. HB, hemoglobin; HD, hemodialysis; HDL, high density lipoprotein; hsCRP, high sensitivity C reactive protein; LDL, low density lipoprotein; RAGE, receptor for advanced glycation end products; TG, triglyceride; VEGFR2, vascular endothelial growth factor receptor 2.
Laboratory parameters in HD patients with and without dyslipidemia.
Parameter | HD with D | HD without D | P-value |
---|---|---|---|
VEGFR2, ng/ml |
1.96, 0.330-5.0 | 0.930, 0.390-3.31 | 0.01 |
Albumin, g/dl |
3.74±0.32 | 3.56±0.32 | 0.058 |
Total cholesterol, mg/dl |
194.0, 159.0-349.0 | 136.0, 98.0-178.0 | <0.0001 |
LDL-cholesterol, mg/dl |
118.0±22.3 | 66.3±18.3 | <0.0001 |
HDL-cholesterol, mg/dl |
38.0, 27.0-67.0 | 38.5, 26.0-82.0 | 0.953 |
TG, mg/dl |
220.0±82.9 | 122.0±44.5 | <0.0001 |
Non-HDL, mg/dl |
156.0, 127.0-300.0 | 94.5, 58.0-139 | <0.0001 |
LDL/HDL-cholesterol ratio |
2.95±0.62 | 1.69±0.62 | <0.0001 |
HDL/Total cholesterol, % |
19.0, 14.0-33.5 | 30.2, 17.3-50.0 | <0.0001 |
TG/HDL-cholesterol ratio |
5.2, 1.82-10.1 | 2.76, 0.810-7.08 | 0.0005 |
hsCRP, mg/l |
4.7, 4.0-126.0 | 9.45, 4.0-65.1 | 0.41 |
HB, g/dl |
11.3±1.4 | 11.4±1.4 | 0.772 |
aP<0.01;
bP<0.001;
cmean ± standard deviation;
dmedian, range. D, dyslipidemia; HB, hemoglobin; HD, hemodialysis; HDL, high density lipoprotein; hsCRP, high sensitivity C reactive protein; LDL, low density lipoprotein; RAGE, receptor for advanced glycation end products; TG, triglyceride; VEGFR2, vascular endothelial growth factor receptor 2.
Laboratory parameters in the control cohort with and without D.
Parameter |
Controls with D | Controls without D | P-value |
---|---|---|---|
VEGFR2, ng/ml | 2.75±1.63 | 2.39±1.14 | 0.535 |
Albumin, g/dl | 4.31±0.18 | 4.34±0.24 | 0.724 |
Total cholesterol, mg/dl | 225.0±38.0 | 174.0±30.0 | <0.001 |
LDL-cholesterol, mg/dl | 138.0±31.0 | 72.0±15.0 | <0.00001 |
HDL-cholesterol, mg/dl | 66.0±16.0 | 71.0±23.0 | 0.557 |
TG, mg/dl | 121.0±41.0 | 89.0±22.0 | 0.237 |
Non-HDL, mg/dl | 159.0±31.0 | 103.0±14.0 | <0.0001 |
LDL/HDL-cholesterol ratio | 2.13±0.55 | 1.30±0.39 | 0.0002 |
HDL/Total cholesterol, % | 29.5±5.6 | 40.0±7.9 | 0.0005 |
TG/HDL-cholesterol ratio | 1.61±0.81 | 1.42±0.70 | 0.537 |
hsCRP, mg/l | 4.21±1.68 | 3.66±1.22 | 0.432 |
HB, g/dl | 12.6±1.3 | 12.3±1.30 | 0.621 |
aP<0.001;
bmean ± standard deviation D, dyslipidemia; HB, hemoglobin; HDL, high density lipoprotein; hsCRP, high sensitivity C reactive protein; LDL, low density lipoprotein; RAGE, receptor for advanced glycation end products; TG, triglyceride; VEGFR2, vascular endothelial growth factor receptor 2.
Significant correlations between VEGFR2 and other clinicopathological characteristics in the examined groups treated with HD, with and without dyslipidemia.
A, Whole HD group, n=50 | ||
---|---|---|
Correlated parameter | R | P-value |
VEGFR2 and age | 0.334 | 0.017 |
VEGFR2 and RAGE | 0.686 | <0.00001 |
VEGFR2 and Tchol | 0.297 | 0.036 |
VEGFR2 and LDL-chol | 0.291 | 0.039 |
VEGFR2 and TG | 0.362 | 0.009 |
VEGFR2 and non-HDL | 0.361 | 0.009 |
VEGFR2 and LDL/HDL-chol ratio | 0.402 | 0.004 |
VEGFR2 and HDL/Tchol ratio | -0.403 | 0.004 |
VEGFR2 and TG/HDL-chol ratio | 0.362 | 0.009 |
B, Dyslipidemic HD group, n=26 | ||
Correlated parameter | R | P-value |
VEGFR2 and age | 0.397 | 0.04 |
VEGFR2 and RAGE | 0.737 | 0.0002 |
C, Non-dyslipidemic HD group, n=24 | ||
Correlated parameter | R | P-value |
VEGFR2 and RAGE | 0.677 | 0.00027 |
VEGFR2 and PLT | -0.457 | 0.024 |
aP<0.05;
bP<0.01;
cP<0.001. HDL-chol, high density lipoprotein cholesterol; LDL-chol, low density lipoprotein cholesterol; PLT, platelets; RAGE, receptor for advanced glycation end products; Tchol, total cholesterol; TG, triglyceride; VEGFR2, vascular endothelial growth factor receptor 2.