Prehypertension in dyslipidemic individuals ; relationship to metabolic parameters and intima-media thickness

Introduction. Like hypertension, prehypertension is associated with cardiovascular disease. Aims. The aim of this study was to evaluate: a) the prevalence of prehypertension/hypertension in individuals with various dyslipidemic phenotypes; b) the relation between blood pressure (BP) and other risk factors for atherosclerosis; c) atherogenic potential of prehypertension by the assessment of intima-media thickness of the arteria carotis communis (IMT). Methods. 667 clinically asymptomatic subjects were divided into four dyslipidemic phenotypes (DLP) according to apolipoprotein B (apoB) and triglycerides (TG): DLP1 (n=198, normo-apoB/normo-TG), DLP2 (n=179, normo-apoB/ hyper-TG), DLP3 (n=87, hyper-apoB/normo-TG), DLP4 (n=203, hyper-apoB/hyper-TG). DLP1 served as a control group. Results. There was significantly higher prevalence of prehypertension and hypertension in subjects with dyslipidemia (DLP2 43.0%, 41.3%; DLP3 42.5%, 29.9%; DLP4 42.4%, 47.8%) than in normolipidemic individuals (DLP1 32.8%, 20.2%). Systolic and diastolic blood pressure (SBP + DBP) correlated with age, total cholesterol, TG, non-HDL-cholesterol, body mass index and waist circumference; SBP additionally with C-peptide, fasting glycemia; DBP additionally with apoB, homeostasis model assessment (HOMA) and plasminogen activator inhibitor-1. The IMT of hypertensive and of prehypertensive subjects was higher than that of subjects with normal BP in all DLPs. Conclusions. The prevalence of prehypertension was higher in all dyslipidemic patients. The common prevalence of prehypertension/hypertension was highest in the hypertriglyceridemic subjects. Prehypertensive and hypertensive patients had higher IMT than normotensive individuals in all DLPs.


INTRODUCTION
Prehypertension has been defined as systolic blood pressure (SBP) 120 to 139 or diastolic blood pressure (DBP) 80 to 89 mm Hg, based on two or more properly measured seated blood pressure (BP) readings on each of two or more office visits 1 .Prehypertension tends to increase in severity over time 2 and associates with increased incidence of cardiovascular disease (CVD), particularly in those with BP levels in the 130 to 139/85 to 89 mm Hg range 3,4 .Compared with normal BP, prehypertension has been associated with a 27% increase in all-cause and 66% increase in CVD mortality 5 .A study of 1999 to 2000 NHANES data has suggested that 64% of prehypertensive subjects have ≥1 other abnormal CVD risk factors, the percentage increased to 94% in those ≥60 years of age 6 .The risk ratios for obesity, dyslipidemia, metabolic syndrome, and diabetes are all greater in prehypertensive than normotensive subjects 7,8 .The dyslipidemias represent heterogenic metabolic disorders.Sniderman suggested an algorithm of dyslipidemia classification based on apolipoprotein B (apoB) and triglyceride (TG) concentrations 9 .The classification divides dyslipidemias into four dyslipidemic phenotypes (DLP) on the basis of apoB and TG concentrations: DLP 1 (normo-apoB / normo-TG), DLP 2 (normo-apoB / hyper-TG), DLP 3 (hyper-apoB / normo-TG) and DLP 4 (hyper-apoB / hyper-TG).Assessment of ApoB used instead of LDL cholesterol (LDL-C) in this diagnostic algorithm is rational and fully justified.ApoB is a better predictor for the risk of coronary artery disease than total cholesterol (TC), LDL-C and non-HDL cholesterol (non-HDL-C).This has repeatedly been shown in a large number of prospective epidemiological studies dealing with lipids and apoB (ref. 10).
The aims of the present study were to investigate: the prevalence of prehypertension and hypertension in dyslipidemic individuals without clinical manifestation of atherosclerosis; the relation between blood pressure and other risk factors for atherosclerosis and the atherogenic capacity of prehypertension by assessment of the intima-media thickness of the arteria carotis communis (IMT), as a surrogate marker of subclinical atherosclerosis manifestation.

Study design and subjects
The study was carried out as a cross-sectional study on subjects who had been examined at the Lipid Center of the 3 rd Department of Internal Medicine, University Hospital Olomouc, Czech Republic in the period October 2005 to June 2011.All subjects (patients with dyslipidemia, their spouses, relatives and healthy volunteers; n=828) filled out a questionnaire on their medical history, especially cardiovascular status, medication and smoking habits.All subjects were tested for an underlying cause of secondary hyperlipidemia: diabetes mellitus, hypothyroidism, hepatic or renal impairment and nephrotic syndrome.Subjects with these diagnoses were not enrolled in the study.Other exclusion criteria were history of clinically manifest atherosclerosis (coronary artery disease, cerebrovascular ischemic disease and peripheral arterial disease), hypolipidemic treatment in the previous 8 weeks, hormone therapy with estrogens and clinical presence of acute infectious disease or trauma.The study was reviewed and approved by the institutional Ethics Committee of the Faculty of Medicine and Dentistry and University Hospital and informed consent was obtained from all participants.Individuals who met the above mentioned criteria (n=667), were divided into four dyslipidemic phenotypes (DLP) based on apolipoprotein B and triglycerides: DLP 1 (n=198, apoB<1.2g/L and TG<1.5 mmol/L), DLP 2 (n=179, apoB<1.2g/L and TG≥1.5 mmol/L), DLP 3 (n=87, apoB≥1.2g/L and TG<1.5 mmol/L), DLP 4 (n=203, apoB≥1.2g/L and TG≥1.5 mmol/L).The use of TG value in this algorithm is justified as well.).A value 1.2 g/L for apoB was chosen, because at this level the cardiovascular risk rapidly increased 9,12 .This value corresponds to the 75 th percentile of the North American population, the level chosen to define an elevated LDL-C (ref. 13,14).Subjects with DLP 1 -who were normolipidemic (normo-apoB / normo-TG) -served as a control group.The presence of metabolic syndrome was assessed according to National Cholesterol Education Program Adult Panel III (ref. 15).The ten-year risk of having a heart attack using risk assessment tool based on information from Framingham Heart Study was estimated in all participants 16 .

Blood pressure measurement
Body mass index (BMI), waist circumference, SBP and DBP were determined.The auscultatory method of BP measurement with a properly calibrated and validated mercury sphygmomanometer was used.Persons were seated quietly for at least 5 min in a chair, with feet on the floor, and arm supported at heart level.An appropriate-sized cuff was used to ensure accuracy.The first and fifth Korotkoff sounds were used to identify SBP and DBP (ref. 1 ).At least three sitting BP measurement with a 30-second interval were taken and the mean from the last two was calculated.Subjects with SBP ≥ 120 and < 140 or DBP ≥ 80 and < 90 mmHg were defined as prehypertensive.Patients treated by antihypertensive drugs, or with SBP ≥ 140 or DBP ≥ 90 mmHg were taken as hypertensive 1 .There was a substantial part of treated patients within the hypertensive group.Hypertensive patients with DLP 1 (n=40) were treated by ACE inhibitors (ACE-I) or angiotensin II receptor blockers (ARBs) in 37.5%, by calcium channel blockers (CaCBs) in 5.0%, by beta blockers (BBs) in 35

Biochemical analyses
Venous blood samples were drawn in the morning after a 12 h fast.Total cholesterol, triglycerides and HDLcholesterol were determined enzymatically on Modular SWA analyzer (Roche, Basel, Switzerland) using commercially available kits (Cholesterol SYS 917, Triglycerides GPO-PAP and HDL cholesterol plus, 3 rd generation kits, Roche, Basel, Switzerland).Determination of HDL-C was made by a direct method without precipitation of apo B containing lipoproteins.LDL-C levels were calculated according to the Friedewald formula (for TG < 4.5 mmol/L).We also calculated non-HDL cholesterol (non-HDL-C = TC -HDL-C).Concentration of apoB and apolipoprotein A1 (apoA1) was determined immunoturbidimetrically on Modular SWA analyzer (TinaQuant Apo A1, TinaQuant Apo B kits, all Roche, Basel, Switzerland).High-sensitivity C-reactive protein (hs-CRP) was assessed by means of an ultra sensitive latex immunoturbidimetric method on Modular SWA analyzer (CRP latex TinaQuant kit, all Roche, Basel, Switzerland).Glycemia was determined by enzymatic-colorimetric method (Glucose GOD-PAP kit) on Modular SWA analyzer.Insulin was determined using the commercially available kits -Insuline (Immunotech, Marseille, France) using specific antibodies by IRMA (immunoradiometric assay) method.The result obtained was then used for calculation of the parameter of insulin resistance HOMA [homeostasis model assessment, described by Matthews et al.: fasting glycemia (mmol/L) * fasting insulin (mU/L) / 22.5] (ref. 17).C-peptide and intact proinsulin were determined using the commercially available kits -C-peptide (Immunotech, Marseille, France) and Proinsulin (DRG Instruments GmbH, Marburg, Germany) using specific antibodies by IRMA method for C-peptide and radioimmune assay (RIA) method for proinsulin.Concentrations of insulin, proinsulin, C-peptide and hs-CRP were measured in the serum stored at -80° C. Serum levels of the soluble intercellular cell adhesion molecule-1 (s-ICAM-1) and plas-minogen activator inhibitor-1 (PAI-1) were measured by enzyme-linked immunosorbent assay (ELISA) using commercially available standard kits s-ICAM-1 (Immunotech, Marseille, France) and PAI-1 (Technoclone, Vienna, Austria).These markers of endothelial damage were used because we previously found significant differences in their levels between individual DLPs (ref. 18,19).

IMT measurement
Ultrasound scanning of IMT was performed with a 10 MHz linear array transducer (Philips Sonos 5500, 2004).All measurements were performed with subjects in a supine position.The head was tilted to the one side at an angle of 45°.The longitudinal B-mode image of the common carotid artery (CCA) was displayed just before the widening of the bulb.When an optimal longitudinal image of the far wall of the CCA in the region of 1 cm proximally from the bulb was obtained, it was frozen on the R wave according to a simultaneous ECG and videotaped.Three video records were made on both CCA.IMT measurements were processed off-line using the software Image-Pro Plus (Version 4.0, Media-Cybernetics, Silver Spring, USA).The region under evaluation was the CCA wall 1-2 cm distant proximally from the mentioned border.The average of all mean IMT of three frozen images of both sides was chosen as the outcome variable.The measurement of IMT was made without knowledge of laboratory results.

Statistical analyses
All values are expressed as means ± SD or as median with 25 and 75 percentiles for variables with non-normal distribution.The Kolmogorov-Smirnov test was used to test for normal distribution.Variables with non-normal distribution (TG, proinsulin, insulin, C-peptide, HOMA, hs-CRP, PAI-1) were log transformed to normalize their distribution before statistical analysis.Differences in means between groups were analyzed using ANOVA after adjustment for age, sex, BMI, and smoking.Differences in categorical variables were analyzed by Χ 2 test.For statistical evaluation of correlations between parameters we used a univariate Pearson correlation analysis.Multivariate regression analysis was used for testing for an independent association between dependent and independent variables.Statistical analysis was performed using SPSS for Windows version 12.0 (Chicago, Illinois, USA).Probability values of P<0.05 were considered as statistically significant.

Basic characteristics
Table 1 shows the biochemical and anthropometric characteristics in individual DLPs.Individuals with hyper-apoB DLPs (DLP 3 and 4) had significantly increased total cholesterol and LDL-C; individuals with hyper-TG DLPs (DLP 2 and 4) had decreased HDL-C and apoA1 compared to normolipidemic subjects (DLP 1).Non-HDL-C and apoB/apoA1 significantly rose from DLP 1 to DLP 4. Subjects with hyper-TG DLPs (DLP 2 and 4) showed increased markers of insulin resistance (HOMA, C-peptide, insulin, proinsulin), obesity (BMI, waist circumference) and also increased prevalence of metabolic syndrome.They had higher blood pressure (SBP, DBP) and levels of PAI-1.Patient with DLP4 had significantly increased IMT and s-ICAM-1 levels.

IMT according to blood pressure in individual dyslipidemic phenotypes
IMT rose from DLP 1 to DLP 4 (see Table 1).The highest elevation of IMT was observed in patients with simultaneously elevated levels of TG and of apoB (DLP 4).In the whole group of all participants IMT was thicker in prehypertensive (IMT=0.67±0.14;P<0.05) and in hypertensive (IMT=0.73±0.12;P<0.05) patients respectively than in normotensive individuals (IMT=0.57±0.10)after adjustment for age, sex, BMI and smoking.Fig. 2 shows differences of IMT in various groups according to BP and the presence of various DLP.IMT of hypertensive and of prehypertensive subjects respectively was higher than of subjects with normal BP within all DLPs.By comparison between DLPs the significant differences were found between IMT of normotensive (DLP 1 versus DLP 3, and DLP 4 respectively) and prehypertensive subjects (DLP 1 versus DLP 4).Because anti-hypertensive drugs could influence BP and IMT, only untreated subjects were enrolled in the further analysis.IMT correlated with age (r=0.69;

DISCUSSION
The results showed significantly higher prevalence of prehypertension/hypertension in clinically asymptomatic subjects with dyslipidemia (DLP 2, 3, and 4) than in normolipidemic individuals (DLP 1) even after adjustment for age, sex, smoking and body mass index.Cumulative prevalence of prehypertension or hypertension was the highest in the hypertriglyceridemic subjects (DLP 2 and 4).They had significantly increased blood pressure.SBP correlated with age, total cholesterol, TG, non-HDL-C, C-peptide, fasting glycemia, BMI and waist circumference.DBP correlated with age, total cholesterol, TG, non-HDL-C, apoB, HOMA, BMI, waist circumference and PAI-1.
Many cross-sectional studies of different populations demonstrated that prehypertensive individuals had higher total cholesterol, LDL-C, TG and lower HDL-C than normotensive subjects [6][7][8][20][21][22][23][24][25][26][27][28][29][30] . Some pospective studies also showed that dyslipidemia could predict occurrence of incident hypertension by years [31][32][33] .The mechanism by which dyslipidemia leads to BP elevation is only partially understood.Dyslipidemia causes endothelial dysfunction, the loss of vasomotor reactivity and arterial stiffness 34 .We found total cholesterol, and non-HDL-C independent predictors for BP elevation. PAI-1as a maker of endothelial dysfunction was independently associated with DBP in all dyslipidemic patients of this study, too.These findings may be explained by the suggesting pathogenetic mechanism.Lipid particles could activate recently identified liver X receptor, which is a potential regulator of renin expression 32,35 .The activation of renin-angiotensin-aldosterone system (RAAS) increases BP and also leads (via angiotensin II) to PAI-1 expression 36 .PAI-1 can promote fibrosis, vascular remodeling, arterial stiffness and BP increase by preventing the activation of matrix metalloproteinases and the degradation of extracellular matrix by plasminogen activators and plasmin 37 .The significant increase of blood pressure and the highest cumulative prevalence of prehypertension/hypertension were detected in hypertriglyceridemic subjects and the levels of PAI-1 were significantly elevated in these patients too.
Hypertriglyceridemic subjects also showed increased markers of insulin resistance and of abdominal obesity.Many papers found the association between insulin resistance and prehypertension 8,38,39 or hypertension 40 .Hyperinsulinemia directly increases sodium re-absorption in renal tubules and may lead to fluid retention 41,42 .4][45] ).However, not insulinemia alone but insulin resistance is usually related to blood pressure in prehypertensive and hypertensive patients and it is able to predict incident hypertension [46][47][48] .There was not found correlation between insulin levels and BP in the present study, too.Only C-peptide and fasting glycemia correlated with SBP, HOMA with DBP.However, in multivariate regression analysis no more specific marker of insulin resistance (except fasting glycemia) rested to predict BP levels.SDP and DBP BP were independently predicted by waist circumference.Abdominal obesity seems to play an independent role in BP elevation of all dyslipidemic patients.Among various anthropometric measurements waist circumference was detected the best predictor for incidence of hypertension 49 and waist circumference was also independent predictor for progression of prehypertension to hypertension 50 .The possible mechanism linking abdominal obesity with BP elevation is production of adipokines.Visceral fat produces more angiotensinogen and PAI-1 than subcutaneous fat 51 .The activation of RAAS leading to vasoconstriction and increase sodium re-absorption, together with chronic inflammation and endothelium dysfunction are probably the main causes of prehypertension and hypertension in patients with abdominal obesity.
The present study also showed increased IMT of prehypertensive, hypertensive patients respectively compared to normotensive individuals in all dyslipidemic phenotypes.IMT was independently predicted by age, total cholesterol, SBP, and waist circumference.It was closely related to 10-year coronary heart disease risk estimated using Framingham Risk Score 15 .Several papers also found IMT of the common carotid artery in patients with prehypertension or borderline hypertension thicker compared to normotensive controls 52,53 .Many studies detected the increase of IMT in patients with dyslipidemia.Our findings showed cumulative effect of dyslipidemia and BP increase on IMT.We found significant differences between IMT of prehypertensive subjects according to DLPs after adjustment for age, sex, smoking and BMI.Prehypertensive patients with DLP 4 had the same IMT as normolipidemic patients with hypertension.While the second group should be treated by antihypertensive drugs, the pharmacological treatment for prehypertension has been still recommended only in patients with diabetes mellitus, chronic kidney disease or coronary artery disease 1 .Also assessment of global cardiovascular risk in clinically asymptomatic dyslipidemic middle-aged patients with prehypertension only by SCORE(ref. 54) or by Framingham Risk Score 15 may not result to recommendation for hypolipidemic treatment.And thus measurement of IMT could constitute useful diagnostic tool for cardiovascular risk prediction and making decision on pharmacological treatment in these people.This noninvasive examination should be considered especially in asymptomatic dyslipidemic individuals with hyperapoB/ hyperTG phenotype, who have not reached the level of risk for initiation of therapy based on clinical and laboratory data only.

CONCLUSION
The prevalence of prehypertension was significantly higher in all dyslipidemic patients compared to normolipidemic subjects.Common prevalence of prehypertension and hypertension was the highest in the hypertriglyceridemic subjects.Prehypertensive and hypertensive patients had higher IMT than normotensive individuals in all dyslipidemic phenotypes.Dyslipidemia and blood pressure have cumulative effect on IMT.IMT measurement should be considered in asymptomatic individuals with hyper-apoB/hyperTG phenotype to improve estimation of their cardiovascular risk.

Table 3 .
Predictive values of independent predictors on the levels of systolic and of diastolic blood pressure by multivariate backward stepwise regression analysis in dyslipidemic individuals.