Serum cardiovascular risk factors in obstructive sleep apnea

Serum Cardiovascular Risk Factors in

Obstructive Sleep Apnea*

Murat Can, MD; S¸erefden Ac¸ikgo¨z, MD; Go¨rkem Mungan, MD;

Taner Bayraktarog˘lu, MD; Erdem Koc¸ak, MD; Berrak Gu¨ven, MD; and

Selda Demı˙rtas, MD

Background: Obstructive sleep apnea (OSA) patients have increased cardiovascular morbidity

and mortality. The cardiovascular markers associated with OSA are currently not defined.

Objectives: The aims of this study were to determine whether OSA is associated with serum

cardiac risk markers and to investigate the relationship between them.

Methods: Sixty-two male patients were classified into two groups with respect to apnea-hypopnea

index (AHI): group 1, sleep apnea (n

ⴝ 30), with AHI > 5; and group 2 (n ⴝ 32), with AHI < 5. We

compared cardiovascular risk factors in both groups with control subjects (n

ⴝ 30) without OSA (AHI

<

1). Serum cholesterol, triglyceride, high-density lipoprotein cholesterol (HDL-C), low-density

lipoprotein cholesterol (LDL-C), apolipoprotein A-I, apolipoprotein B, lipoprotein (a), C-reactive

protein (CRP), and homocysteine were measured. Statistical significance was assessed with analysis of

variance at p < 0.05. In correlation analysis, Pearson correlation was used.

Results: There was no significant difference between group 1 and group 2 in total cholesterol, LDL-C,

HDL-C, triglyceride, apolipoprotein A-I, apolipoprotein B, and lipoprotein (a). All of the M-mode

echocardiographic parameters were in the normal reference range. Serum homocysteine and CRP

levels were significantly increased in group 1 compared to group 2 (p < 0.05). Serum CRP values

were increased in both group 1 and group 2 when compared with control subjects (p < 0.05). Serum

homocysteine values were higher in group 1 than in control subjects (p < 0.05).

Conclusions: Our results show that OSA syndrome is associated not only with slight

hyperteinemia but also with increased CRP concentrations. Increased plasma concentrations of

homocys-teine and CRP can be useful in clinical practice to be predictor of long-term prognosis for

cardiovascular disease and the treatment of OSA.

(CHEST 2006; 129:233–237)

Key words: C-reactive protein; homocysteine; obstructive sleep apnea

Abbreviations: AHI⫽ apnea-hypopnea index; CPAP ⫽ continuous positive airway pressure; CRP ⫽ C-reactive protein; HDL-C⫽ high-density lipoprotein cholesterol; LDL-C ⫽ low-density lipoprotein cholesterol; OSA ⫽ obstructive sleep apnea

O

bstructive sleep apnea (OSA) is a common

chronic respiratory disorder.

_{OSA occurs in}

approximately 4% of men and 2% of woman

⬎ 30

years old.

_{Increase in the ratio with age may depend}

on the role of OSA on the complications of the

disease.

OSA is well-defined syndrome that includes one or

two of the following symptoms: severe snoring,

nocturnal respiratory arrest, repeated nocturnal

awakening, nonrecuperative sleep, diurnal fatigue,

and altered concentration. These clinical findings are

related to the extent of hypoxemia and hypercapnia

that develop as a result of disordered breathing.

*From Faculty Of Medicine (Drs. Can, Ac¸ikgo¨z, Mungan, and Gu¨ven), Department Of Biochemistry, and Faculty Of Medicine (Drs. Bayraktarog˘lu and Koc¸ak), Department Of Internal Medi-cine, Karaelmas University, Zonguldak; and Faculty Of Medicine (Dr. Demı˙rtas), Department Of Biochemistry, Ufuk University, Ankara, Turkey.

Manuscript received April 8, 2005; revision accepted July 16, 2005.

Reproduction of this article is prohibited without written permission from the American College of Chest Physicians (www.chestjournal. org/misc/reprints.shtml).

Correspondence to: Murat Can, MD, Karaelmas University, Faculty of Medicine, Department of Biochemistry, Zonguldak, Turkey; e-mail: drcanmurat@yahoo.com

CHEST

Original Research

Although OSA patients have increased

cardiovas-cular morbidity and mortality,

_{how much of their}

cardiovascular diseases are due to OSA rather than to

other risk factors such as upper-body obesity; insulin

resistance; increased age; alcohol and caffeine

con-sumption; and cigarette smoking is yet unknown.

Therefore, identifying the possible risk factors

in-volved in OSA cardiovascular morbidity and

mortal-ity is of great clinical importance. Several studies

report a strong association between homocysteine,

C-reactive protein (CRP),

_{total cholesterol,}

low-density lipoprotein cholesterol (LDL-C),

high-den-sity lipoprotein cholesterol (HDL-C),

triglycer-ide,

_{apolipoprotein A-I and B,}

_{lipoprotein (a)}

levels,

_{and coronary heart disease.}

For editorial comment see page 218

The pathophysiology of the underlying

mecha-nisms and complications of OSA is complex and

multifactorial. The aims of this study were to

deter-mine whether OSA syndrome is associated with

serum cardiac risk markers, and to investigate the

relationship between serum cardiac risk markers.

Materials and Methods

Patients were recruited for the study based on medical history and written consent. The study protocol was approved by the university ethics committee and was performed in accordance with the current revision of the guidelines in accordance with the Declaration of Helsinki.

Patients

OSA was diagnosed on the basis of the International Classifi-cation of Sleep Disorders.14_{Sixty-two male patients who were}

referred for suspected sleep apnea underwent an overnight sleep study. Patients were classified into two groups with respect to apnea-hypopnea index (AHI); group 1 (n⫽ 30), AHI ⬎ 5; group 2 (n⫽ 32), AHI ⬍ 5. We compared cardiovascular risk factors in both groups with control subjects (n⫽ 30) without OSA (AHI ⬍ 1). All subjects with OSA snored and reported excessive daytime sleepiness or two or more other features such as impaired concentration, unrefreshing sleep, witnessed apneas, restless sleep, and irritability/personality change. Before enroll-ment, the subjects were asked about their regular medications and medical history regarding diabetes mellitus, renal diseases, and ischemic heart disease. The patients were assessed for coronary artery disease with resting and stress 12-lead precordial ECG. Previous myocardial infarction, unstable angina, prior coronary intervention, arrhythmias, conduction abnormalities, heart failure, digoxin therapy, inability to perform tests, hyper-tension (BPⱖ 140/90 mm Hg or receiving medication), chronic renal disease, and diabetes mellitus were the exclusion criteria. Subjects who smoked or had systemic infections at the time of the study were also excluded. Before polysomnography, baseline demographic data, BP, ECG, and echocardiography were as-sessed throughout the day, and blood samples were collected between 8 pm and 9 pm.

Polysomnography

Polysomnography was started at 9 pm and ended at 6:30 am. Surface electrodes were applied using standard techniques to obtain an EEG, an electromyogram of the chin, an ECG, and an electrooculogram. Sleep was defined according to the criteria of Rechtschaffen and Kales.15 _{Ventilation was monitored using}

inductive plethysmography. Airflow was monitored by ther-mistors placed at the nose and mouth, while arterial oxygen saturation was monitored continuously with a pulse oximeter. A polygraph was run continuously at 10 mm/s to record all of the above physiologic data simultaneously throughout the course of the experiment. All parameters were stored in a data recorder for subsequent analysis. Apnea was defined as the cessation of airflow at the nose and mouth lasting for⬎ 10 s. Hypopnea was defined as a decrease of ⱖ 50% in thoracoabdominal motion associated with a fall in the baseline oxygen saturation ofⱖ 4%. All AHI values were calculated to express the number of episodes of apnea and hypopnea per hour of total sleep time.

Echocardiographic Analyses

We performed echocardiography with M mode (GE-VingMed System 5 Ultrasound System; GE-VingMed Sound AB; Horten, Norway). All studies were performed and interpreted by the same operator and recorded on videotape. Left ventricular end-diastolic dimension, left ventricular end-systolic dimension, and thickness of the interventricular septum and posterior wall were measured. The ejection fraction was calculated from area measurements using the area-length method applied to the average apical area. Echocardiographic data were recorded ac-cording to the guidelines of the American Society of Echocardi-ography.

Blood Collection

All blood samples were obtained in the nonfasting state. The subjects did not perform any specific exercise or apply any specific diet during the study period. For homocysteine, serum samples were centrifuged immediately and placed on ice prior to separation. After centrifugation, the serum aliquots were frozen and stored at⫺ 80°C.

Biochemical Analysis

Serum cholesterol, triglyceride, and HDL-C were measured by enzymatic colorimetric methods with commercially available kits (Cobas Integra 800; Roche Diagnostics GmbH; Mannheim, Germany), and LDL-C was calculated according to the Friede-wald formula: total cholesterol⫺ HDL-C ⫺ (0.45 ⫻ triglycer-ide). Apolipoprotein A-I and apolipoprotein B were measured by immunoturbidimetric method, and lipoprotein (a) and CRP were determined by particle-enhanced immunoturbidimetric method on the Roche Integra 800 analyzer. Serum homocysteine was measured by enzyme-linked immunosorbent assay (Axis Homo-cysteine EIA; Axis-Shield Diagnostics; Dundee, Scotland) on a diagnostic instrument (LP 400; Diagnostics Pasteur; Chaska, MN). Vitamin B12and folate levels were measured by

electro-chemiluminescent immunoassay on a Roche Elecsys 2010 ana-lyzer (Vitamin B12and folate kit; Roche Diagnostics).

Erythro-cyte count, hemoglobin concentration, mean cell volume, and mean cell hemoglobin concentration were measured (MAXM; Beckman Coulter; Fullerton, CA), and stained RBC examinations of the patients were studied.

Statistical Analysis

Results are expressed as mean⫾ SE. We used analysis of variance to analyze any differences in demographic and

hemo-dynamic characteristics between the two groups. In correlation analysis, a Pearson correlation was used. All statistical analysis was performed with a statistical program (SPSS, version 11.0; SPSS; Chicago, IL), defining statistical significance as p⬍ 0.05.

Results

When we compared the patients with the healthy

control subjects, there were no significant

differ-ences between group 1, group 2, and the control

group with respect to age, body mass index, and BP

(Table 1). All of the M-mode echocardiographic

parameters (left ventricular end-diastolic dimension

[LVEDD], 4.46

⫾ 0.45 cm; left ventricular

end-systolic dimension [LVESD], 2.93

⫾ 0.41 cm; EF,

64.6

⫾ 2.66%; IVS, 1.28 ⫾ 0.09 cm; and PW,

1.17

⫾ 0.10 cm) were in normal reference range.

With regard to the conventional parameters (total

cholesterol, LDL-C, HDL-C, triglycerides,

apoli-poprotein A-I, apoliapoli-poprotein B, and liapoli-poprotein (a))

there were no significant differences between group

1 and group 2. Total cholesterol, LDL-C,

triglycer-ide, apolipoprotein B, and lipoprotein (a) values

were increased in both group 1 and group 2 when

compared with control group (p

⬍ 0.05). There were

no significant differences between group 1, group 2,

and the control group in apolipoprotein A-I and

HDL-C results (Table 2). Serum CRP values were

increased in both group 1 and group 2 when

com-pared with control group (p

⬍ 0.05). Serum

homo-cysteine values were higher in group 1 than in

control subjects (p

⬍ 0.05). Comparison of serum

homocysteine and CRP levels revealed a significant

difference (p

⬍ 0.05) between group 1 and group 2.

(Table 2). Distribution of plasma homocysteine and

CRP is shown in Figure 1. We did not find a

significant correlation between CRP and

homocys-teine levels (r

⫽ 0.06, p ⬎ 0.05). There was no

rela-tionship between AHI and both CRP and

homocys-teine (r

⫽ 0.12, p ⬎ 0.05; r ⫽ 0.31, p ⬎ 0.05,

respectively). The values of vitamin B

, folate,

erythrocyte count, hemoglobin, mean cell volume,

and mean cell hemoglobin concentration were in

reference range. Stained RBC examination results

were in the normal range.

Discussion

Homocysteine is a thiol-containing amino acid that

is an intermediate substance produced during

intra-cellular demethylation of methionine. Elevated

lev-els of homocysteine are found in patients with

cardiovascular diseases.

_{A clear correlation was}

shown between mildly elevated total blood

homocys-teine concentrations and premature coronary artery

diseases,

_stroke,

_{peripheral artery diseases, or}

venous thrombosis.

Elevated homocysteine levels in OSA patients

were reported only in patients with associated

isch-emic heart disease and/or hypertension.

_{Jordan et}

al

_{reported 30% decrease in homocysteine level}

after long-term continuous positive airway pressure

(CPAP) treatment in a small group of patients mostly

with hypertension or diabetes. However, Svatikova et

Table 1—Clinical Characteristics of OSA Patients and

Control Groups* Characteristics Group 1 (n⫽ 30) Group 2 (n⫽ 32) Control (n⫽ 30) Age, yr 47.14⫾ 1.62 45.31 ⫾ 1.21 42.6 ⫾ 3.2 Body mass index, kg/m² 29.63 ⫾ 0.67 31.12 ⫾ 0.76 20.2 ⫾ 0.82 Systolic BP, mm Hg 124.4⫾ 2.29 125⫾ 2.28 122 ⫾ 4.5 Diastolic BP, mm Hg 76.43⫾ 1.17 76.88 ⫾ 1.03 75.4 ⫾ 2.1

AHI 25.04⫾ 3.85 2.72⫾ 0.28 ⬍1

*Data are presented as mean⫾ SE.

Table 2—Demonstration of the Whole Parameters Detected in Patients and Control Subjects* Parameters Group 1 (n⫽ 30) Group 2 (n⫽ 32) Control (n⫽ 30) Homocysteine,␮mol/L 21.53⫾ 14.2† 7.4⫾ 5.12 6.8⫾ 4.7 CRP, mg/L 5.08⫾ 3.25†‡ 2.7⫾ 0.60§ 1.8⫾ 0.61 Total cholesterol, mg/dL 212.8⫾ 46.0‡ 197.7⫾ 27.0§ 118.4⫾ 32.2 Triglyceride, mg/dL 153.8⫾ 69.0‡ 132.7⫾ 65.0§ 78.0⫾ 18.8 HDL-C, mg/dL 44.0⫾ 10.0 43.2⫾ 10.9 47.1⫾ 9.4 LDL-C, mg/dL 133.3⫾ 33.3‡ 124.6⫾ 27.1§ 56.4⫾ 20.6 Lipoprotein (a), mg/dL 16.6⫾ 11.0‡ 13.0⫾ 9.3§ 8.7⫾ 3.2 Apolipoprotein A-I, mg/dL 126.4⫾ 20.6 121.7⫾ 39.1 132.1⫾ 27.9 Apolipoprotein B, mg/dL 110.8⫾ 33.7‡ 101.8⫾ 23.3‡ 80.4⫾ 16.7

*Data are presented as mean⫾ SE.

†Significant difference between group 1 and group 2 (p⬍ 0.05). ‡Significant difference between group 1 and control group (p⬍ 0.05). §Significant difference between group 2 and control group (p⬍ 0.05).

al

_{have shown that plasma levels of homocysteine}

are not elevated in OSA patients, and neither acute

untreated OSA nor treatment with CPAP and

dis-turbed sleep affect plasma homocysteine levels or

obscure its diurnal variation. This result is in contrast

to the findings of our study, in which we found

slightly enhanced homocysteine levels in OSA

pa-tients. The OSA patients in our study did not have

any heart disease and/or hypertension. Thus, we can

say that OSA might be independently associated with

mildly increased blood homocysteine levels.

A recent epidemiologic study

_{has shown that}

enhanced levels of CRP are a strong independent

predictor of risk of future myocardial infarction,

stroke, peripheral arterial disease, and vascular death

among persons without known cardiovascular

dis-ease. In OSA patients, hypoxia and reoxygenation

episodes can also cause activation of inflammatory

cells, as observed for neutrophils and monocytes,

and ongoing inflammatory responses play important

roles in atherosclerosis.

Although CRP is a nonspecific marker of

inflam-mation, epidemiologic studies

_{suggest that CRP}

is an important risk factor in atherosclerosis and

coronary artery disease. CRP that was found at high

concentrations in the atherosclerotic lesion

_{has a}

direct role on secretion of inflammatory mediators

by vascular endothelium,

_{up-regulates the}

expres-sion of adheexpres-sion molecules in endothelial cells, and

increases low-density lipoprotein uptake into

macro-phages.

_{The development of systemic}

inflamma-tion, characterized by elevated levels of certain

potent proinflammatory mediator such as CRP, may

have an important and direct role in the

develop-ment of atherosclerotic lesions and in promoting

cardiovascular morbidity.

Population-based cross-sectional studies

_have

shown that plasma CRP concentrations are elevated

in obesity.

_{In this study, CRP levels of both groups}

were found to be significantly higher than those of

the control group. Chronic subclinical inflammation

effects may be one pathophysiologic mechanism

explaining the enhancement of CRP levels in OSA

patients. Shamsuzzaman et al

_{reported higher CRP}

values in patients with moderate-to-severe OSA than

in control subjects. Yokoe et al

_{observed elevated}

CRP values in patients with moderate-to-severe OSA

as well, and they noted a decrease in CRP levels by

treatment with nasal CPAP. In agreement with these

studies, we found high levels of CRP in OSA

pa-tients, but this enhancement did not have any

cor-relation with the severity of OSA. Similarly, we did

not find any correlation between CRP and

homocys-teine. Both of the cited authors

_reported

signif-icant positive relationship between CRP and AHI;

however, our results disagree with these authors.

The lack of correlation between AHI and CRP levels

is explained by the fact that apnea-related hypoxia

was not sufficient in patients with mild-to-moderate

OSA. This finding shows that CRP may be an

independent risk factor in patients with

mild-to-moderate OSA for future cardiovascular events.

Our results show that OSA syndrome is associated

not only with slight hyperhomocysteinemia but also

with increased CRP concentrations. The lack of

Figure 1. Distribution of plasma homocysteine (top, A) and CRP (bottom, B) in patients and control subjects.

correlation between homocysteine and CRP

sup-ports the possibility that homocysteine and CRP are

independent risk factors for cardiovascular disease in

OSA patients. Increased plasma concentrations of

homocysteine and CRP can be useful in clinical

practice to be predictor of long-term prognosis for

cardiovascular disease and the treatment of OSA,

providing many benefits to the patients and society.

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