Effects of smoking on serum lipid and lipoprotein concen-trations and lecithin:cholesterol acyltransferase activity

Effects of smoking on serum lipid and lipoprotein concen-trations and lecithin:cholesterol acyltransferase activity

Melahat Dirican^a, Emre Sarandol^a, Engin Ulukaya^b, Hatice Asuman Tokullugil^a

Melahat Dirican^a, Emre Sarandol^a, Engin Ulukaya^b, Hatice Asuman Tokullugil^a

Abstract:Cigarette smoking is one of the major risk factors for cardiovascular disease.The mechanism responsible for this association is still unknown. We measured the activity of lecithin:cholesterol acyltransferase (LCAT), a key factor in the esterification of plasma cholesterol and reverse cholesterol transport, and the levels of lipids and apolipoproteins in the serum of 27 cigarette smoking and 31 non-smoking (control) men. We could not find any significant difference among these parameters between the groups. Serum LCAT activity was lower in smokers, but the difference was statistically nonsignificant. We also classified the two groups in respect to their serum lipid levels as hyper- and normolipidemic, we observed that normolipidemic-smokers had lower (p<0.05) high density lipoprotein-cholesterol (HDL-C) and HDL-ester cholesterol levels compared to the normolipidemic-nonsmokers. While there were no any significant differences between hyperlipidemic-smokers and nonsmokers with respect to any of the parameters.
In the end we have got the idea that smoking seems to affect HDL-C and HDL-ester cholesterol levels in the normolipidemic-smokers group, only, Also, LCAT activity tended to be lower in smokers compared to nonsmokers. J. Med. Invest. 46:169-172, 1999

Keywords:Lecithin:cholesterol acyltransferase, smoking, lipids

INTRODUCTION
Cigarette smoking is accepted as a major risk factor for ischemic heart disease (IHD) as well as hypertension and hyperlipidemia (1). Certain compo-nents of cigarette smoke, such as nicotine and carbon monoxide, have been reporterd to be responsible for the development of IHD by increasing plasma catecholamine levels and producing hypoxia (2). On the other hand, cigarette smoking alters plasma lipoprotein levels (3-5) and increases the susceptibil-ity of low density lipoprotein (LDL) to oxidation (6). However, the exact mechanism responsible for the harmful effect of smoking on IHD is not fully under-stood (7).
High density lipoprotein-cholesterol (HDL-C) is known to be an important protective factor for IHD. Several epidemiological studies have indicated that low serum levels of HDL-C are associated with an increased risk of IHD (8-10). Reverse cholesterol transport is also an antiatherogenic process in which excess cholesterol is transported from peripheral tissues back to the liver or to other peripheral tissues that in need cholesterol. The excess cholersterol is taken by HDL and esterified by lecithin:cholesterol acyltransferase (LCAT, EC 2. 3. 1. 43) and cholesteryl esters are transfered from HDL towards very-low and low-density lipoproteins (VLDL and LDL) by cholesterol ester transfer protein (CETP) (11).
Conflicting data on the influence of smoking on LCAT activity have been published In this respect we investigated the effects of smoking on the levels of plasma lipids, lipoproteins, apolipoproteins and LCAT activity in fasting blood from healthy smokers and nonsmokers, matched for age and body mass index (BMI).

MATERIALS AND METHODS
Twenty-seven cigarette smokers and thirty-one non-smokers were investigated. All subjects were men. The smokers smoked at least 10 cigarettes per day (mean, 18 ± 6cigarettes;range 10 to 30) for at least 7 years (mean, 15 ± 6 years, range7to28). The non-smokers had never smoked in the past. The ages of the subjects were 29 to 53. Subjects with hepatic, renal or cardiovascular disease, and diabetes mellitus were excluded. None of them were taking any drugs, such as beta-blockers, diuretics or lipid-lowering agents. Informed consent was ob-tained from each participant in the study.
Blood was drawn from an antecubital vein into vacutainer tubes after 10-12 hours of fasting. Blood was centrifuged at 1500g for 15 minutes. Sera were stored at -20°C and analyzed within 30 days. Serum triglyceride (TG) level was determined enzymatically (Biosystems, Spain). Total cholesterol (TC) was measured with the Lieberman-Burchard method. Free cholesterol (FC) was determined in serum and HDL fraction with the Lieberman-Burchard method after digitonin precipitation (12). HDL-C was assayed after the precipitation of apolipopro-tein (apo) B-containing lipoproteins with dextran sulphate-magnesium chloride (13). LDL-C was deter-mined according to Friedewald et al (14). Esterified cholesterol was deduced from the difference be-tween total and free cholesterol. Apo A-I and apo B were measured by nephelometry (Sanofi Pasteur, France). Lipoprotein (a) was analyzed by ELISA (Chromogenix, Sweden). Serum LCAT activity was determined as a function of the decrease of free cholesterol which is esterified during incubation at37°C, as described by Hitz et al (15).
Results are expressed as mean±SD. The Student's t-test was used to compare the smoker and non-smoker subjects. A p value of <0.05 was considered to be significant.

RESULTS
The study groups comprised of 27 male smokers (mean age ± SD, 41.9 ± 6.7 years) and 31 male non-smokers (41.5 ± 6.9 years). The body mass indexes were 25.6 ± 2.8 and 26.4 ± 3.5kg/m2 in smokers and non-smokers, respectively (Table1).There were no significant differences in serum TC, TG, lipoprotein (a), apo A-I and B, LDL-C and HDL- and serum-free and- ester cholesterol levels between smokers and non-smokers. Serum LCAT activity was lower in smokers than in non-smokers, although this difference was statistically nonsignifi-cant (Table2and 3).
The smoker and non-smoker subjects were grouped into normolipidemics (n:30) and hyperlipidemics [n:28, isolated hypercholesterolemia (n:8) and mixed hypertriglyceridemia (n : 20)] HDL-C and HDL-esterified cholesterol were lower in normolipidemic-smokers than in normolipidemic-nonsmokers (Table4).

DISCUSSION
Smoking is a significant risk factor for IHD. There are several studies showing associations between cigarette smoking and altered serum lipid and lipo-protein concentrations. These alterations, which are associated with smoking, are higher serum levels of TC, TG, VLDL-C, LDL-C and lower serum levels of HDL-C and Apo A-I. Many of these variables have been associated with an increased risk of IHD (5).
Several studies have reported that serum HDL-C level is found to be lower in smokers than in non-smokers (16-18). However Siekmeier et al. (19) reported that smoker and non-smoker subjects had similar HDL-C levels;this result agrees with our result. In this study, concentrations of TC, TG, LDL-C, HDL-C, Lp (a), apo A-I and B did not differ between smokers and non-smokers.
There are several studies showing lower serum LCAT concentrations or activities in smokers com-pared to non-smokers (20, 21). Haffner et al. reported a negative correlation between smoking and LCAT mass (21). The positive correlation between LCAT concentration and TC and LDL-C was also reported in these studies (20-22). These findings are in ac-cordance with the suggestion that LCAT plays an important role in the reverse transport of cholesterol. In addition to in vivo studies, in vitro studies have shown that cigarette smoke inhibited LCAT activity (23). In another study, it was reported that rats given nicotine had lower LCAT activity (24). However, in several studies, LCAT activity was found not to be different in smokers and non-smokers (7, 17, 22, 25). In our study, serum LCAT activity was found to be 15.8% lower in smokers than in non-smokers, but this difference was not statistically significant.
The data indicate that cigarette smoking does not affect lipid, lipoprotein and apolipoprotein levels. LCAT activity tended to be lower in smokers compared to non-smokers. So, it may be considered that the reverse cholesterol transport can be diminished and cholesterol can accumulate in the peripheral tissues, more in the smokers than non-smokers.

REFERENCES
1. Kannel WB:Update on the role of cigarette smoking in coronary artery disease. Am Heart J101 (3):319-328, 1981
2. Yokode M, Kita T, Arai H, Kawai C, Narumiya S, Fujiwara M:Cholesteryl ester accumulation in macrophages incubated with low density lipoprotein pretreated with cigarette smoke extract. Proc Natl Acad Sci 85:2344-2348, 1988
3. McGill HC:Potential mechanisms for the aug-mentation of atherosclerosis and atherosclerotic disease by cigarette smoking. Prev Med 8:390-403, 1979
4. Topping DL:Effects of tobacco smoke and its constituents on lipid and carbohydrate metabo-lism. In:Clemens MJ, Ashwell M, eds. Bio-chemistry of cellular regulation. Boca Raton, Florida, CRC Press, 1980, Vol2, pp. 165-183
5. Craig WY, Palomaki GE, Haddow JE:Ciga-rette smoking and serum lipid and lipoprotein concentrations:an analysis of published data. Br Med J 298:784-788, 1989
6. Scheffler E, Wiest E, Woehrle J, Otto I, Schulz I, Huber L:Smoking influences the atherogenic potential of low-density lipoprotein. Clin Invest 70:263-268, 1992
7. Moriguchi EH, Fusegawa Y, Tamachi H, Goto Y:Effects of smoking on HDL subfractions in myocardial infarction patients:effects on leci-thin:cholesterol acyltransferase and hepatic lipase. Clin Chim Acta195:139-144, 1990
8. Miller GJ, Miller NE:Plasma high density lipoprotein concentration and development of ischaemic heart disease. Lancet 1:16-19, 1975
9. Goldbourt V, Holtzman E, Neufeld HN:Total and high density lipoprotein cholesterol in the serum and risk of mortality:evidence of a threshold effect. Br Med J 290:1239-1243, 1985
10. Castelli WP, Garrison RJ, Wilson PWF, Abbott RD, Kalousaian S, Kannel WB:Incidence of coronary heart disease and lipoprotein cho-lesterol levels. J Am Med Assoc 256:2835-2836, 1986
11. Franceschini G, Maderna P, Sirtori C:Reverse cholesterol transport:physiology and pharma-cology. Atherosclerosis 88:99-107, 1991
12. Bauer JD.:Clinical laboratory methods. St. Louis, C.V. Mosby Co, 9th ed., 1982
13. Demacker PNM, Vos-Janssen HE, Hijmans AGM, Van't Laar A, Jansen AP:Measurement of high-density lipoprotein cholesterol in serum:Comparison of six isolation methods combined with enzymic cholesterol analysis. Clin Chem26 (13):1780-1786, 1980
14. Friedewald WT, Levy RI, Fredrickson DS: Estimation of the concentration of low-density lipoprotein cholesterol in plasma without use of the ultracentrifuge. Clin Chem18:449-502, 1972
15. Hitz J, Steinmetz J, Siest G:Plasma lecithin:cholesterol acyltransferase-reference values and effects of xenobiotics. Clin Chim Acta133:85-96, 1983
16. Criqui MH, Wallace RB, Heiss G, Mishkel M, Schnfeld G, Jones GTL:Cigarette smoking and plasma high-density lipoprotein cholesterol. The Lipid Research Clinics Program Prevalence Study. Circulation62 (suppl4):70-76, 1980
17. Ito T, Nishiwaki M, Ishikawa T, Nakamura H:CETP and LCAT activities are unrelated to smoking and moderate alcohol consumption in healthy normolipidemic men. Jpn Circ J 59(8):541-546, 1995
18. Siekmeier R, Marz W, Kronenberger H, Seiffert UB, Groβ W:Effects of cigarette smoking on plasma lipids, apolipoproteins and lipoprotein (a) in healthy subjects. Clin Chem 40 (7):1350-1351,1994
19. Siekmeier R, Wulfroth P, Wieland H, Groβ W, Marz W:Low-density susceptibility to in vitro oxidation in healthy smokers and non-smokers. Clin Chem 42 (4):524-530, 1996
20. Albers JJ, Bergelin RO, Adolphson JL, Wahl PW:Population-based reference values for lecithin-cholesterol acyltransferase (LCAT). Atherosclerosis43 (2-3):369-379, 1982
21. Haffner SM, Applebaum-Bowden D, Wahl PW:Epidemiological correlates of high density lipo-protein subfractions, apolipoproteins A-I, A-II, and D, and lecithin cholesterol acyltransferase. Effects of smoking, alcohol, and adiposity. Ar-teriosclerosis 5:169-177, 1985
22. Ronnemaa T, Viikari J, Marniemi J. Lecithin:cholesterol acyltransferase activity in children and young adults. Atherosclerosis 77:7-13, 1989
23. McCall MR, van den Berg JJ, Kuypers FA, Tribble DL, Krauss RM, Knoff LJ:Modifi-cation of LCAT activity and HDL structure. New links between cigarette smoke and coronary heart disease risk. Arterioscler Thromb 14 (2):248-253, 1994
24. Latha MS, Vijayammal PL, Kurup PA:Effect of nicotine administration on lipid metabolism in rats. Indian J Med Res 98:44-49, 1993
25. Dullaart RP, Hoogenberg K, Dikkeschei BD, Van Tol A:Higher plasma lipid transfer protein activities and unfavorable lipoprotein changes in cigarette-smoking men. Arterioscler Thromb14 (10):1581-1585, 1994

Received for publication December 8, 1998;accepted May 11, 1999.

Address correspondence and reprint requests to Dr. Melahat Dirican, Uludag University Medical Faculty Department of Biochemistry 16059 Gorukle-Bursa, TURKEY and Fax:+90.224. 4428018.