The Impact of Selected Factors on the Cardiovascular System in Chronic Kidney Disease

Cardiovascular Diseases Clinical Trial

Official title:

Assessment of the Impact of Selected Factors on the Cardiovascular System in Patients With Different Chronic Kidney Disease Stages

Clinical Trial Details — Status: Completed

Administrative data

• NCT number: NCT05214872
• Other study ID #: PoznanUMS_DF2
• Status: Completed
• Start date: March 25, 2016
• Est. completion date: September 30, 2020

Study information

• Verified date: January 2022
• Source: Poznan University of Medical Sciences
• Contact: n/a
• Is FDA regulated: No
• Study type: Observational

Clinical Trial Summary

Chronic kidney disease (CKD), is characterized by accelerated development of atherosclerosis and advanced remodelling of vessels and the heart. It is associated with many factors, including inflammation, arterial hypertension, hyperlipidemia, hyperhomocysteinemia, secondary hyperparathyroidism, and oxidative stress. Hypertension is one of the most critical risk factors for cardiovascular complications. It leads to the formation of structural changes in the vascular system: it impairs the activity of the endothelium, causes hypertrophy and remodelling of the vascular wall, reduces the susceptibility of the vessels and accelerates the development of atherosclerosis. This study aimed to identify the processes and their representative markers, the concentration of which in the serum may reflect the cardiovascular system status and can predict the increased mortality in HD patients.

Description:

Chronic Kidney Disease has a significant impact on the cardiovascular system. From many different complications of CKD, one to mention is arterial stiffness. This disorder results from many pathologies, including inflammation, arterial hypertension, carbohydrate metabolic disorders, lipid disorders, vascular calcification, chronic inflammation, and oxidative stress.

The main goal of this study was to analyze the mechanisms leading to the increased tendency to cardiovascular disturbances in CKD, with particular focus on the parameters of oxidative stress, inflammation and the results of imaging examinations (intima-media thickness (IMT) assessments) and other non-invasive cardiological examinations based on the results using the Portapres device (Finapres Medical Systems (FMS), the Netherlands), the SphygmoCor tonometer (AtCor Medical), the Colin blood pressure monitor (BMP)-7000 (Japan) Pulse Trace 2000 (Micro Medical Ltd., Rochester, Kent, United Kingdom) The Accuson CV 70 system (Siemens) with a 10 megahertz (Mhz) transducer.

Besides, studied participants were followed 2 years after enrollment to study for recording cardiovascular-related death.

Recruitment information / eligibility

• Status: Completed
• Enrollment: 252
• Est. completion date: September 30, 2020
• Est. primary completion date: September 10, 2020
• Accepts healthy volunteers: Accepts Healthy Volunteers
• Gender: All
• Age group: 18 Years and older

Eligibility

Criteria:

The following criteria of qualifying for the study were adopted for all respondents:

• 18 years of age or older,

• written consent to participate in the study,

• no active inflammatory process,

• no neoplastic disease or a neoplastic disease whose treatment was stopped at least 10 years ago,

• no history of immunosuppressive treatment,

• stable liver function (not more than two times increased activity of transaminases), HBs antigen and anti-HCV negative antibodies,anti-HIV negative antibodies.

In addition, for CKD patients (CKD1-2) and PREDIALYSIS GROUP, the following additional inclusion conditions were applied:

• no acute cardiovascular complications, ie acute heart failure, hypertensive crisis, acute coronary syndrome, at the time of study entry.

At the same time, depending on the technique of renal replacement therapy used, additional inclusion criteria were established for each of the subgroups:

in group HD:

• a minimum of 6 months of treatment with repeated hemodialysis, 3 times a week, for a minimum of 10 hours a week,

• arteriovenous fistula as a vascular access for hemodialysis,

• Estimated dialysis adequacy ratio (eKt / V) of at least 1.2. in the PD group:

• treatment duration UP to a minimum of 6 months, Kt / V =1.8 l / week / 1.73 m2.

For CARD patients, additional conditions include:

• no obvious evidence of renal impairment in the history and at the time of study entry, renal function assessed on the basis of eGFR and urine albumin/creatinine ratio,

• history of angina pectoris,

• documented history of at least one acute coronary syndrome,

• admission to the Department of Intensive Care of Cardiology and Internal Diseases in order to perform a planned coronary angiography, on the day of admission to the study without signs of the acute coronary syndrome, no additional comorbidities, ie those that do not result directly or indirectly from coronary heart disease.

In turn, for the HV group (control group), additional conditions include:

• no obvious evidence of renal impairment in the history and at the time of study entry, renal function assessed on the basis of eGFR and urine albumin/creatinine ratio,

• no obvious signs of cardiovascular impairment in the history and at the time of study entry, estimated on the basis of normal blood pressure (<140/90 mmHg), no abnormalities in the medical history and physical examination,

• not taking any medications on a regular basis.

Related Conditions & MeSH terms

• Atherosclerosis
• Atherosclerosis of Artery
• Cardiovascular Diseases
• Chronic Kidney Diseases
• Dialysis; Complications
• Inflammation
• Kidney Diseases
• Renal Insufficiency, Chronic

Intervention

Diagnostic Test:
• laboratory parameters - complete blood count
the complete blood count was analyzed using Sysmex K-4500 Automated Hematology Analyzer (by GMI Inc., USA): hemoglobin (HGB) [g/dl]; red blood count (RBC) [10^12/l]; hematocrit (HCT) [l/l]; white blood cells (WBC) [10^9/l]; platelet count (PLT) [10^9/l]

Other:
• body mass index (BMI) [kg/m^2] calculation
body mass index (BMI) [kg/m^2] was calculated by dividing a person's weight (post-HD weight in HD group) [kg] by the squared their body height [m]

Diagnostic Test:
• selected parameters of oxidative stress (1)
Serum concentration of: advanced glycation ends products (AGE) [µg/mg protein]; 3-nitrotyrosine (3-NT) [µmol/mg protein]; advanced oxidation protein products (AOPP) [µmol/mg protein]; carboxymethyle(lysine) (CML) [µg/mg protein] were determined with the enzyme immunoassay methods (ELISA) using Shanghai Sunred Biological Technology Co kits, China.
• metalloproteinases
metalloproteinases in the serum [ng/ml]: metalloproteinase 9 (MMP-9) in the serum was determined by the ELISA method using the Quantikine Human MMP-9 (total) kit, by R&D Systems, Canada; tissue inhibitor of metalloproteinase 1 (TIMP-1) in the serum - was determined by the ELISA method using the Quantikine Human TIMP-1 kit, manufactured by R&D Systems, Canada; the MMP-9/TIMP-1 ratio was calculated by the quotient of the MMP-9 and the TIMP-1 concentration.
• parameters of lipids metabolism in the serum
total cholesterol (T-C) [mg/dl] - was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA; low-density lipoprotein cholesterol (LDL-C) [mg/dl] - was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA; high-density lipoprotein cholesterol (HDL-C) [mg/dl] - was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA; triglycerides (TG) [mg/dl] - were assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA; the concentration of low-density lipoprotein (LDL-C) cholesterol - was determined from Friedewalds' equation (LDL-C [mg/dl] = total cholesterol (T-C) [mg/dl]- HDL-C [mg/dl]- TG[mg/dl]/5).
• parameters of iron metabolism
iron concentration [mg/dl] - was analyzed with the Cobas Integra 400 plus biochemical analyzer from Roche Diagnostics, USA; total iron-binding capacity (TIBC) [mg/dl] - was analyzed with the Cobas Integra 400 plus biochemical analyzer from Roche Diagnostics, USA; the unsaturated iron-binding capacity (UIBC) [mg/dl] was determined by an equation in which iron concentration in plasma is subtracted from TIBC [mg/dl]; ferritin [ng/ml] concentration was determined with the Modular E-170 biochemical analyzer from Roche Diagnostics, USA.
• selected inflammatory markers
high-sensitivity C-reactive protein (hsCRP) [mg/l] was measured using DADE Behring, USA and the DADE nephelometer Behring Analyzer II; neopterin [nmol/l] was determined by using the Neopterin ELISA kit, DRG International, Inc., USA; interleukin 18 (IL-18) [pg/ml] concentration was determined by Colorimetric Sandwich ELISA, Quantikine Human IL-18 R&D Inc., USA.

Device:
• carotid intima-media thickness (IMT)
carotid intima-media thickness (IMT) [mm] was measured by The Accuson CV 70 system (Siemens) with a 10 megahertz (Mhz) transducer. Two longitudinal projections were assessed (anterolateral and posterolateral). The distal 1 cm of the common carotid artery just proximal to the bulb was measured by means of a computer analysis system (Medical Imaging Applications, LLC).
• non-invasive cardiological examinations
For non-invasive cardiological examinations, the Portapres device (Finapres Medical Systems (FMS), the Netherlands), the SphygmoCor tonometer (AtCor Medical), the Colin blood pressure monitor (BMP)-7000 (Japan) were used. Main assessed variables: heart rate (HR) [beats per minute [bpm]]; ejection duration (ED) [millisecons]; peripheral systolic (pSBP) and diastolic blood pressure (pDBP) [mm Hg]; peripheral mean arterial pressure (pMAP) [mm Hg]; peripheral end-systolic pressure (pESP) [mm Hg]; central systolic (cSBP) and diastolic blood pressure (cDBP) [mm Hg]; central mean arterial pressure (cMAP) [mm Hg]; central augmented pressure (cAP) [mmHg]; central mean pressure of diastole (cMPD)[mm Hg]; central mean pressure of systole (cMPS) [mm Hg]; central end-systolic pressure (cESP) [mm Hg].
• vessel stiffness assessment
The following parameters of vessel stiffness were assessed by Pulse Trace 2000 (Micro Medical Ltd., Rochester, Kent, United Kingdom): reflection index (RI) [in percentages [%]]; vascular stiffness index (SI) [m/s]; peripheral pulse pressure (pPP) [mm Hg]; central puls pressure (cPP) [mm Hg] peripheral pulse pressure/central pulse pressure (pPP/cPP ratio).

Other:
• cardiovascular (CV)-related death recording during 2-year follow-up
During a 2-year follow-up from the enrollment to this study, CV-related fatal incidents history has been recorded for each subject separately. The primary endpoint was fatal acute myocardial infarction (AMI) or acute ischemic stroke or any unexpected or sudden death only if autopsy proved CV-related. If there was doubt about the cause of death or there was no contact with the patient during the two years from study enrollment, that patient was excluded and not considered further.

Diagnostic Test:
• glucose (Glu)
glucose (Glu) [mg/dl] was assessed in the serum by a routine technique using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.
• klotho
klotho [ng/ml] - was analyzed in the serum by Human KL(Klotho) [ng/ml] ELISA Kit, Shanghai Sunred Biological Technology Co kit, China.
• fibroblast growth factor 23 (FGF-23)
FGF-23 [pg/ml] - was analyzed in rhe serum using Human FGF-23 ELISA Kit, Sigma-Aldrich, USA.
• parameters of calcium and phosphate metabolism
total and ionized calcium [mg/dl], phosphate [mg/dl], intact parathormone (iPTH) [mg/dl] were assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.
• liver enzymes activity assessment
activity of: alanine transaminase (ALT) [U/l]; aspartate transaminase (AST) [U/l]; alkaline phosphatase (ALP) [U/l] were assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.
• total protein and albumin
total protein (TP) [g/dl]; albumin (ALB) [g/dl] were assessed in the serum by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.
• creatinine and urea
creatinine in the serum [mg/dl] - the assay is based on the reaction of creatinine with sodium picrate as described by Jaffe (Jaffes' colorimetric method) - was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA; urea [mg/dl] in the serum - was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.
• selected parameters of oxidative stress (2)
myeloperoxidase (MPO) [ng/ml] in the serum- was determined by the ELISA method using the Quantikine Human MPO test by R&D Systems kit, Canada.
• selected parameters of oxidative stress (3) sRAGE
soluble receptor for advanced glycation end products (sRAGE) [µg/mg protein] in the serum was tested with enzymatic immunoassay (Quantikine ELISA) using R&D Systems kit, Canada.
• selected parameters of oxidative stress (4) MG, CEL, carbamyl protein groups
methylglyoxal (MG) [µg/mg protein]; carboxyethyle(lysine) (CEL) [µg/mg protein]; carbamyl protein groups [µg/mg protein] were assessed in the serum by competitive enzyme immunoassay (competitive ELISA) using kits from Cell Biolabs Inc, USA.
• selected electrolytes assessment
potassium (K) [mmol/l]; sodium (Na) [mmol/l]; magnesium (Mg) [mg/dL] were assessed in the serum by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.
• NT-pro-brain natriuretic peptide (NT-proBNP)
NT-proBNP [fmol/ml] - was analyzed in the serum by enzyme immunoassay using the Nt-proBNP kit from Biomedica, Slovakia.

Other:
• estimated glomerular filtration rate (eGFR) calculation
eGFR [ml/min/1.73m^2] - according to the Kidney Disease: Improving Global Outcomes (KDIGO) 2012 recommendations was calculated based on the Modification of Diet in Renal Disease (MDRD) formula: eGFR = 186 x [creatinine concentration in mg/dl] - 1.154 x [age in years] - 0.203 x [0.724] for the female gender.

Locations

Country	Name	City	State
Poland	Poznan University of Medical Sciences	Poznan

Sponsors (1)

Lead Sponsor	Collaborator
Poznan University of Medical Sciences

Country where clinical trial is conducted

Poland, 

Outcome

Type	Measure	Description	Time frame	Safety issue
Primary	Diagnostic test: basic biochemical parameters: complete blood count - hemoglobin (HGB)	hemoglobin (HGB) [g/dl] was analyzed using Sysmex K-4500 Automated Hematology Analyzer (by GMI Inc., USA).	3 years
Primary	Diagnostic test: basic biochemical parameters: complete blood count - red blood cell count (RBC)	red blood cell count (RBC) [10^12/l] was analyzed using Sysmex K-4500 Automated Hematology Analyzer (by GMI Inc., USA).	3 years
Primary	Diagnostic test: basic biochemical parameters: complete blood count - hematocrit (HCT)	hematocrit (HCT) [l/l] was analyzed using Sysmex K-4500 Automated Hematology Analyzer (by GMI Inc., USA).	3 years
Primary	Diagnostic test: basic biochemical parameters: complete blood count - white blood cell count (WBC)	white blood cells (WBC) [10^9/l] was analyzed using Sysmex K-4500 Automated Hematology Analyzer (by GMI Inc., USA).	3 years
Primary	Diagnostic test: basic biochemical parameters: complete blood count - platelet count (PLT)	platelet count (PLT) [10^9/l] was analyzed using Sysmex K-4500 Automated Hematology Analyzer (by GMI Inc., USA).	3 years
Primary	Diagnostic test: glucose (Glu)	glucose (Glu) [mg/dl] concentration in the serum was assessed by the routine technique using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: urea	urea [mg/dl] concentration in the serum was assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: creatinine	creatinine [mg/dl] concentration in the serum was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA (based on Jaffes' colorimetric method - the assay is based on the reaction of creatinine with sodium picrate as described by Jaffe).	3 years
Primary	Estimated glomerular filtration rate (eGFR) [ml/min/1.73m^2] calculation	eGFR - according to the Kidney Disease: Improving Global Outcomes (KDIGO) 2012 recommendations was calculated based on the Modification of Diet in Renal Disease (MDRD) formula: eGFR = 186 x [creatinine concentration in mg/dl] - 1.154 x [age in years] - 0.203 x [0.724] for the female gender.	3 years
Primary	Body mass index (BMI) [kg/m^2] calculation	Body mass index (BMI) - [kg/m^2] was calculated by dividing a person's weight (post-HD weight in HD group) [kg] by the squared their body height [m].	3 years
Primary	Diagnostic test: parameters of lipids metabolism in the serum - total cholesterol (T-C)	total cholesterol (T-C) [mg/dl] concentration in the serum was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: parameters of lipids metabolism in the serum - high-density lipoprotein cholesterol (HDL-C)	high-density lipoprotein cholesterol (HDL-C) [mg/dl] concentration in the serum was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: parameters of lipids metabolism in the serum low-density lipoprotein cholesterol (LDL-C).	low-density lipoprotein (LDL-C) cholesterol concentration in the serum was determined from Friedewals' equation (LDL-C [mg/dl] = total cholesterol (T-C) [mg/dl] - HDL-C [mg/dl] - TG[mg/dl]/5).; It was assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: parameters of lipids metabolism in the serum - triglycerides (TG)	triglycerides (TG) [mg/dl] concentration in the serum was assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: liver enzymes activity assessment - aspartate transaminase (AST)	activity of aspartate transaminase (AST) [U/l]; was assessed in the serum by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: liver enzymes activity assessment - alanine transaminase (ALT)	activity of alanine transaminase (ALT) [U/l] was assessed in the serum by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: liver enzymes activity assessment - alkaline phosphatase (ALP) [U/l]	activity of alkaline phosphatase (ALP) [U/l] was assessed in the serum by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: total protein (TP)	total protein (TP) [g/dl] concentration in the serum was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: albumin(ALB)	albumin (ALB) [g/dl] concentration in the serum was assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: parameters of iron metabolism - iron	iron concentration [mg/dl] in the serum - was assessed with the Cobas Integra 400 plus biochemical analyzer from Roche Diagnostics, USA;	3 years
Primary	Diagnostic test: parameters of iron metabolism - total iron-binding capacity (TIBC)	total iron-binding capacity (TIBC) [mg/dl] - was determined with the Cobas Integra 400 plus biochemical analyzer from Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: parameters of iron metabolism - the unsaturated iron-binding capacity (UIBC)	unsaturated iron-binding capacity (UIBC) [mg/dl] was determined by an equation in which iron [mg/dl] concentration in plasma is subtracted from TIBC [mg/dl].	3 years
Primary	Diagnostic test: parameters of iron metabolism - ferritin	ferritin [ng/ml] concentration in the serum was determined with the Modular E-170 biochemical analyzer from Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: total and ionized calcium	total and ionized calcium [mg/dl] serum concentrations were assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: phosphate	phosphate [mg/dl] serum concentration was assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: intact parathormone (iPTH)	intact parathormone (iPTH) [mg/dl] serum concentration was assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: klotho (KL)	klotho (KL) [ng/ml] serum concentration was analyzed by Human KL(Klotho) [ng/ml] ELISA Kit, Shanghai Sunred Biological Technology Co kit, China.	3 years
Primary	Diagnostic test: fibroblast growth factor 23 (FGF-23)	fibroblast growth factor 23 (FGF-23) [pg/ml] serum concentration was analyzed using Human FGF-23 ELISA Kit, Sigma-Aldrich, USA.	3 years
Primary	Diagnostic test: selected electrolytes assessment in the serum: potassium (K) and sodium (Na)	Electrolytes: potassium (K) [mmol/l] and sodium (Na) [mmol/l] serum concentrations were assessed by the routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: selected electrolytes assessment in the serum: magnesium	magnesium (Mg) [mg/dl] serum concentration was assessed by routine techniques using Cobas Integra 400 plus biochemical analyzer by Roche Diagnostics, USA.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - 3-nitrotyrosine (3-NT)	Serum concentration of 3-nitrotyrosine (3-NT) [µmol/mg protein] was determined with the enzyme immunoassay method (ELISA) for 3NT using Shanghai Sunred Biological Technology Co kits, China.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - advanced glycation ends products (AGE)	Serum concentration of advanced glycation ends products (AGE) [µg/mg protein] was determined with the enzyme immunoassay method (ELISA) for AGE using Shanghai Sunred Biological Technology Co kits, China.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - carboxymethyle(lysine) (CML)	Serum concentration of carboxymethyle(lysine) (CML) [µg/mg protein] was determined with the enzyme immunoassay method (ELISA) for CML using Shanghai Sunred Biological Technology Co kits, China.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - advanced oxidation protein products (AOPP)	Serum concentration of advanced oxidation protein products (AOPP) [µmol/mg protein] was determined with the enzyme immunoassay method (ELISA) for AOPP using Shanghai Sunred Biological Technology Co kits, China.	3 years
Primary	Diagnostic test: metalloproteinases - metalloproteinase 9 (MMP-9)	metalloproteinase 9 (MMP-9) [ng/ml] concentration in the serum was determined by the ELISA method using the Quantikine Human MMP-9 (total) kit, by R&D Systems, Canada.	3 years
Primary	Diagnostic test: metalloproteinases - tissue inhibitor of metalloproteinase 1 (TIMP-1)	tissue inhibitor of metalloproteinase 1 (TIMP-1) [ng/ml] concentration in the serum - was determined by the ELISA method using the Quantikine Human TIMP-1 kit, manufactured by R&D Systems, Canada.	3 years
Primary	The MMP-9/TIMP-1 ratio assessment	the MMP-9/TIMP-1 ratio was calculated by the quotient of the MMP-9 [ng/ml] and the TIMP-1 [ng/ml] concentration.	3 years
Primary	Diagnostic test: selected inflammatory markers - high-sensivity C-reactive protein (hsCRP)	high-sensitivity C-reactive protein (hsCRP) [mg/l] concentration in the serum was measured using DADE Behring, USA, and the DADE nephelometer Behring Analyzer II.	3 years
Primary	Diagnostic test: selected inflammatory markers - neopterin	neopterin [nmol/l] serum concentration was determined by using the Neopterin ELISA kit, DRG International, Inc., USA.	3 years
Primary	Diagnostic Test: selected inflammatory markers - interleukin 18 (IL-18)	interleukin 18 (IL-18) [pg/ml] concentration in the serum was determined by Colorimetric Sandwich ELISA, Quantikine Human IL-18 R&D Inc., USA.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - myeloperoxidase (MPO)	myeloperoxidase (MPO) [ng/ml] in the serum - was determined by the ELISA method using the Quantikine Human MPO test by R&D Systems kit, Canada.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - methylglyoxal (MG)	methylglyoxal (MG) [µg/mg protein] concentration in the serum was assessed by competitive enzyme immunoassay (competitive ELISA) using MG kits from Cell Biolabs Inc, USA.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - carboxyethyle(lysine) (CEL) [µg/mg protein]	carboxyethyle(lysine) (CEL) [µg/mg protein] concentration in the serum was assessed by competitive enzyme immunoassay (competitive ELISA) using CEL kits from Cell Biolabs Inc, USA.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - carbamyl protein groups [µg/mg protein]	carbamyl protein groups [µg/mg protein] concentration in the serum were assessed by competitive enzyme immunoassay (competitive ELISA) using carbamyl protein groups kits from Cell Biolabs Inc, USA.	3 years
Primary	Diagnostic test: selected parameters of oxidative stress - soluble receptor for advanced glycation end products (sRAGE)	soluble receptor for advanced glycation end products (sRAGE) [µg/mg protein] concentration in the serum was tested with enzymatic immunoassay (Quantikine ELISA) using R&D Systems sRAGE kit, Canada.	3 years
Primary	Non-invasive cardiological examinations (1) with the use of Portapres device (Finapres Medical Systems (FMS), the Netherlands), the SphygmoCor tonometer (AtCor Medical), the Colin blood pressure monitor (BMP)-7000 (Japan) - blood pressures	Blood pressure was measured using the Colin BPM 7000 on both arms (participants were seated). Next, a piezoelectric tonometer Colin BPM was placed over the radial artery for the acquisition of the radial arterial pressure waveform in a supine position. This signal was sent to the SphygmoCor and after averaging various parameters have been assessed and recorded: peripheral systolic blood pressure (sSBP) [mmHg]; peripheral diastolic blood pressure (pDBP)[mm Hg]; peripheral mean arterial pressure (pMAP) [mm Hg]; peripheral end-systolic pressure (pESP) [mm Hg]; central systolic blood pressure (cSBP) [mm Hg]; central diastolic blood pressure (cDBP) [mm Hg]; central mean arterial pressure (cMAP) [mm Hg]; entral augmented pressure (cAP) [mm Hg]; central mean pressure of diastole (cMPD) [mm Hg]; central mean pressure of systole (cMPS)[mm Hg]; central end-systolic pressure (cESP)[mm Hg]	3 years
Primary	Non-invasive cardiological examinations (2) with the use of Portapres device (Finapres Medical Systems (FMS), the Netherlands), the SphygmoCor tonometer (AtCor Medical), the Colin blood pressure monitor (BMP)-7000 (Japan) - heart rate (HR)	Blood pressure was measured using the Colin BPM 7000 on both arms (participants were seated). Next, a piezoelectric tonometer Colin BPM was placed over the radial artery for the acquisition of the radial arterial pressure waveform in a supine position. This signal was sent to the SphygmoCor and after averaging various parameters have been assessed and recorded: - heart rate (HR) in beats per minute [bpm]	3 years
Primary	Non-invasive cardiological examinations (3) with the use of Portapres device (Finapres Medical Systems (FMS), the Netherlands), the SphygmoCor tonometer (AtCor Medical), the Colin blood pressure monitor (BMP)-7000 (Japan) - ejection duration (ED)	Blood pressure was measured using the Colin BPM 7000 on both arms (participants were seated). Next, a piezoelectric tonometer Colin BPM was placed over the radial artery for the acquisition of the radial arterial pressure waveform in a supine position. This signal was sent to the SphygmoCor and after averaging various parameters have been assessed and recorded: - ejection duration (ED) in milliseconds [msec]	3 years
Primary	Device: carotid intima-media thickness (IMT)	Carotid intima-media thickness (IMT) [mm] was measured by The Accuson CV 70 system (Siemens) with a 10 megahertz (Mhz) transducer.; Two longitudinal projections were assessed (anterolateral and posterolateral). The distal 1 cm of the common carotid artery just proximal to the bulb was measured by means of a computer analysis system (Medical Imaging Applications, LLC).	3 years
Primary	Device: vessel stiffness assessments - reflection index (RI)	The following parameter of vessel stiffness was assessed by Pulse Trace 2000 (Micro Medical Ltd., Rochester, Kent, United Kingdom): - reflection index (RI) in percentages [%].	3 years
Primary	Device: vessel stiffness assessments - vascular stiffness index (SI)	The following parameter of vessel stiffness was assessed by Pulse Trace 2000 (Micro Medical Ltd., Rochester, Kent, United Kingdom): -vascular stiffness index (SI) [m/s].	3 years
Primary	Device: vessel stiffness assessments - peripheral (pPP) and central pulse pressure (cPP) [mm Hg]	The following parameters of vessel stiffness were assessed by Pulse Trace 2000 (Micro Medical Ltd., Rochester, Kent, United Kingdom): peripheral pulse pressure (pPP) [mm Hg]; central pulse pressure (cPP) [mm Hg]	3 years
Primary	Device: vessel stiffness assessments - peripheral pulse pressure/central pulse pressure (pPP/cPP) ratio	Peripheral pulse pressure/central pulse pressure (pPP/cPP) ratio was assessed by dividing peripheral pulse pressure (pPP) [mm Hg] by central pulse pressure (cPP) [mm Hg].	3 years
Primary	Cardiovascular (CV)-related death recording during 2-year follow-up	During a 2-year follow-up from the enrollment to this study, CV-related fatal incidents history has been recorded for each subject separately. The primary endpoint was fatal acute myocardial infarction (AMI) or acute ischemic stroke or any unexpected or sudden death only if autopsy proved CV-related. If there was doubt about the cause of death or there was no contact with the patient during the two years from study enrollment, that patient was excluded and not considered further.	2 years for each person qualified for the study
Primary	Diagnostic test: N-terminal pro-B-type natriuretic peptide (NT-proBNP)	N-terminal pro-B-type natriuretic peptide (NT-proBNP) [fmol/ml] concentration in the serum was analyzed by enzyme immunoassay using the Nt-proBNP kit from Biomedica, Slovakia.	3 years