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HDL lipids in close relatives of coronary heart disease patients
419
34 (1979) 419-429 @ Elsevier/North-Holland Scientific Publishers, Ltd.
Atherosclerosis,
HDL LIPIDS IN CLOSE RELATIVES PATIENTS Environmental
and Genetic
OF CORONARY
HEART DISEASE
Influences
DANIEL POME’M’A, HORACE MICHELI, ALAN SUENRAM and CONSTANTIN JORNOT
Divisions de Diabktologie et d’znformatique, Geneva, Geneva (Switzerland)
Department
of Medicine of the University of
(Received 21 May, 1979) (Revised, received 10 July, 1979) (Accepted 11 July, 1979)
Female fiistdegree relatives of CHD patients differed, after the age of forty, from the normal control population by their low HDL cholesterol. Between the ages of 20 and 40 years a slight but significant increase in HDL triglycerides was observed. Except for hormonal contraception which induces significant lowering of HDL cholesterol in the first degree relatives, the observed differences in HDL lipids did not seem related to environmental factors but rather to be genetically determined. In contrast, the low HDL cholesterol observed in the wives of the CHD patients appeared to be related to differences in alcohol intake. Key words:
Coronary heart disease - Enuironmental factors -Female relatives of CHD patients - HDL cholesterol triglycerides - Wives of CHD patients
Introduction The high incidence and prevalence of coronary heart disease (CHD) today represents a major medical threat. Familial clustering of coronary artery disease has been demonstrated but the reasons for the increased familial prevalence are not firmly established and genetic or environmental factors may play a role.
Supported by a grant (6.13000.74) of the Swiss National Science Foundation. Reprint requests to: Prof. D. Porn&a. Division de MabCtolode. HOpital Canton& Switzerland.
CH-1211 Geneva 4.
420
The influence of serum cholesterol and triglycerides on the development of coronary artery disease has been studied extensively. More recently the interest has been focusing on the association of HDL cholesterol with coronary heart disease. Clinical and epidemiological studies have established that there is a strong negative correlation between plasma HDL cholesterol and the existence of coronary heart disease [ 1,3]. It has been shown retrospectively in patients older than 40 years that the power of the HDL cholesterol as a negative predictor for myocardial infarction is greater than the power of the LDL cholesterol as a positive predictor [4]. We have been able to demonstrate that male relatives of patients suffering from myocardial infarction have low HDL cholesterol even in the absence of cardio-vascular disease [5], an observation which may be of significant importance for explaining the development of coronary heart disease in families and may suggest genetic determinism. Myocardial infarction occurs less frequently and at a later age in women than in men, therefore it is of interest to determine whether the lipoprotein lipid pattern, and particularly the HDL cholesterol content in female relatives (sisters, daughters, mothers) of patients suffering from acute myocardial infarction, show a pattern similar to that observed in male relatives of CHD patients or if they differ. In order to distinguish between the role of genetic and environmental factors on HDL cholesterol, the wives of the CHD patients who share similar environments but have different genetic background from the probands were examined. Material and Methods The female firstdegree relatives of 117 consecutive male and female patients with acute myocardial infarction hospitalized in the Department of Medicine were admitted into the study as well as the wives of the male patients. The relatives and wives of male probands 55 years and older and female probands 70 years and over were excluded from the study. The difference in age for the men and the women was chosen because of the lower incidence of myocardial infarction among women below the age of 55. Families with fewer than two first-degree relatives were not considered for the study. However, if fewer than two first-degree relatives of the eligible families attended the medical examination, the probands, wives or relatives already examined were not excluded from the study. The control group previously examined for the four European Cities Survey [6] consisted of 132 female Genevese civil servants without clinical or electrocardiographic evidence of ischaemic heart disease, peripheral vascular disease or any chronic or acute illness. Subjects taking drugs affecting serum lipids were excluded from the study. However, considering the wide use of hormonal contraception in young women, the relatives on “the pill” were investigated and included as a separate group in the present study. Medical examination was performed in all subjects with special emphasis on atherosclerosis (history of angina pectoris, intermittent claudication, peripheral pulses, skin temperature and color, xanthoma). It included blood pressure
421
measurement and ECG recording. Smoking and drinking habits and physical activity were assessed by direct questioning. The alcohol consumption was expressed as liters of wine (10’) per week as in Geneva the use of wine is more widespread than other alcohols. Beer and hard liquors consumed were therefore transformed in wine “equivalent” on the basis of pure alcohol content. Serum and lipoprotein cholesterol was determined by a modification of Abell and Kendall’s method [ 71 as described previously [ 51. Triglycerides were measured according to Soloni [ 81. VLDL, LDL and HDL were separated by quantitative preparative ultracentrifugation in two steps at densities 1.006 and 1.063 as previously described [ 51. The recovery was 93% for cholesterol and 92% for triglycerides. Chemical and ultracentrifugation procedures were previously standardized by sample exchange between laboratories [6]. The statistical analysis of the differences between means of two samples was performed using the Student t-test, the Mann-Whitney U Test and the comparison of regression lines. Before calculation, serum and VLDL triglycerides and VLDL cholesterol were transformed to their logarithms. In order to take into account the influence of age on lipoprotein lipids, the results for the first-degree relatives are reported separately for the age groups 20-39 years, 40-49, and 50 and over. Because of the smaller number of wives and their limited age range, they were all considered together and compared to the controls in the same age range. Results One hundred and seventeen families of 76 male and 41 female probands entered the study. There were 231 living female firstdegree relatives (sisters, daughters, mothers) aged 20 and over eligible for the study. The percentage of dead relatives at the time of the study differs according to the generation. Among the probands’ mothers, 56% were dead as compared with 1% among the daughters. The mortality among the probands’ sisters was 7 percent. 147 (64%) of the living relatives were examined. Most of the relatives who did not attend the medical examination were living too far away from Geneva. In the younger age group (20-39 years) 15 of the 49 relatives (31%) were currently taking “the pill”. Their results were considered separately. In addition, 66 spouses of male probands were examined. Six of them using hormonal contraception were not considered in the present study. Firs t-degree relatives Serum lipids Serum cholesterol was elevated (5.80 mmol/l, P< 0.05) among relatives of patients with myocardial infarction in the age group 40-49 years as compared to the controls (5.27 mmol/l) but no difference was found in serum cholesterol in the younger and older age groups (Table 1). Serum triglycerides did not differ between relatives of patients with ischaemic heart disease and the control group even among the women using hormonal contraception.
422 TABLE 1 SERUM CHOLESTEROL AND TIENTS AND CONTROLS
TRIGLYCERIDES
(mmol/l)
IN FEMALE
4049
Age (years)
20-39
Groups Hormonal contraception Number
relatives
relatives
controls
relatives
yes 15
no 34
no 45
no 36
SWll
RELATIVES
OF CHD PA-
50 and over controls
relatives
controls
no 62
no 50
5.27 0.94 -0.1029 0.2155 0.79
6.15 1.08 0.0583 0.1947 1.14
6.50 1.11 0.0362 0.1940 1.09
??I
Cholesterol SD Triglycerides (log) SD antilog
4.70 0.80 -0.0975 0.1850 0.80
4.72 0.65 -0.1252 0.1968 0.75
4.91 0.79 -0.1735 0.1756 0.67
5.80 * 1.02 0.0200 0.2533 0.96
HDL Cholesterol SD Triglycerides SD
1.39 * 0.32 0.17 * 0.06
1.44 0.35 0.14 * 0.06
1.58 0.32 0.11 0.04
1.42 * 0.34 0.14 0.06
1.65 0.44 0.14 0.08
1.45 * 0.31 0.15 0.05
1.58 0.33 0.14 0.06
LDL Cholesterol SD Triglycerides SD
2.67 0.70 0.29 *J 0.08
2.68 0.52 0.23 0.11
2.78 0.63 0.22 0.07
3.51 ** 0.88 0.29 0.08
2.94 0.85 0.25 0.10
3.76 0.95 0.33 0.10
4.06 0.88 0.31 0.08
-0.1768 0.2129 0.67 -0.4609 0.2838 0.35
-6.2727 0.2952 0.53 -6.5538 0.2717 0.28
-6.0845 0.2494 0.82 -0.4762 0.3181 0.33
-9.503 0.2442 1.12 -9.2599 0.2855 0.55
-0.0201 0.2609 1.05 -0.2627 0.3053 0.55
VLDL Cholesterol SD antilog Triglycerides (log) SD antilog
-0.2037 0.3030 0.63 -0.5302 0.2881 0.30
-0.0133 0.3176 0.97 -9.3611 0.3810 0.44
Comparison relatives vs controls * P < 0.05, ** P < 0.01. 1 Comparison relatives using the pill vs relatives not using the pill.P < 0.06,
and vs the controls,
P < 0.01.
Serum lipoproteins HDL. At age 40 years and above, the HDL cholesterol (Fig. 1) was significantly lower, 1.42 mmol/l, P < 0.05 (age group 40-49 years) and 1.45 mmol/l, P < 0.05 (50 years and above), in the close relative8 of patient8 with myocardial infarction than in the controls (1.65 mmol/l and 1.58 mmol/l, respectively). HDL triglycerides did not differ in relatives and controls after the age of 40 year8 (Table 1). In the younger age group (20-39 years) the HDL cholesterol had a tendency to be lower among the relatives but the difference was not significant. In contrast the HDL triglyceride8 were higher (0.14 mmol/l, P< 0.05) among the relatives than in the control8 (0.11 mmol/l) (Fig. 2). When the CHD patients’ relatives using hormonal contraception were considered, it appeared that the difference8 observed between relatives of coronary patients and controls were more marked. HDL triglycerides were increased (0.17 mmol/l, P < 0.05) a8 compared to the controls (0.11 mmol/l) and their HDL cholesterol wa8 significantly lower (1.39 mmol/l, P < 0.05) when compared to the HDL cholesterol of the controls (1.58 mmol/l).
423
Cholesterol
-
(mmolll)
rrletivcs lno drugs) controls
0
1.8
??
peo.05
1.6
1.4
,I
C age
LO- 49
groups
“=
31
Fig. 1. HDL over).
36
cholesterol
years
X0 50
in female
62
first-degree
relatives
of CHD patients
and controls
(age 40 years and
LDL. In the age group 40-49 years, the LDL cholesterol was increased in the relatives of CHD patients (3.51 mmol/l, P< 0.001) as compared to the controls (2.94 mmol/l) (Table 1). There was no increase in LDL cholesterol in younger or older women related to CHD patients. LDL triglycerides did not differ between relatives and controls except in relatives using the “pill” who had higher LDL triglycerides (0.29 mmol/l, P< 0.05) than the controls (0.22 mmol/l). VLDL. No difference in VLDL cholesterol and triglycerides was found in any age group between the controls and the relatives, whether or not on the “Pill”. Q
sgr20-39 yrsrs
Imrsn f SEMI
Trlglycerldes IrnrnolllI
Cholesterol
0.20.
~p
-p
0
controls
(L5)
Fig. 2. HDL contraception
m
relatives (no drug) (3L1
cholesterol and triglycerides or not and in controls.
m
relatwes on the”p~ll” in young
(151
female first-degree
?e.htives
either
using hormonal
424 TABLE 2 SERUM
AND
PATIENTS
LIPOPROTEIN
CHOLESTEROL
AND TRIGLYCERIDES
AND CONTROLS Wives (n = 60)
Serum Cholesterol SD Triglycerides (log) SD antilog
4.97 0.88 -0.1129 0.0851 0.77
Controls (n = 76)
5.34 P < 0.05 0.98 -0.0864 0.2227 0.82
Wives (n = 58)
Controls (n = 76)
1.44 0.29 0.15 0.05
1.59 0.40 0.13 0.07
2.85 0.72 0.24 0.09
3.05 0.81 0.26 0.10
-0.5600 0.2964 0.28 -0.0902 0.3397 0.81
4.5422 0.2813 0.29 a.4330 0.3231 0.37
HDL Cholesterol SD Triglycerides SD LDL Cholesterol SD Triglycerides SD VLDL Cholesterol (log) SD antilog Triglycerides (log) SD antilog
Cholesterol (mmolll) 1.8
1.6 p-zo.02
“=
controls
wives
91
56
Fig. 3. HDL cholesterol in wives of CHD patients.
P < 0.02
(mmolfl)
IN WIVES OF CHD
425
Wives of CHD patients The age of the 60 wives ranged from 26 to 58 years. Their mean age was 43.8 years and the median age 43.0 years. They were compared with the women of the control group in the same age range who had a mean and median age of 43.2 years. The serum cholesterol of the patients’ wives was 4.97 mmol/l and triglycerides 0.7 7 mmol/l. Due to technical difficulties serum ultracentrifugation was not performed in two wives. The mean HDL cholesterol of the 58 wives was lower (1.44 mmol/l, P < 0.02) than HDL cholesterol of the controls (1.59) (Table 2 and Fig. 3). Environmental factors Cigarette smoking The percentage of smokers was 33% among the relatives (average 18 cigarettes daily), 32% among the probands’ wives (18 cigarettes daily) and 28% among the controls (10 cigarettes daily). Physical activity 3% of the relatives and 3% of the probands’ wives had high physical activity at work and/or at leisure as compared to 5% of the controls. The other subjects had medium or low physical activity. Alcohol in take 46% of the firstdegree relatives and 49% of the controls drank alcohol as opposed to 7% of the probands’ wives. The average weekly alcohol intake was 1.5 liter wine equivalent in all 3 groups. Discussion Since all patients with acute myocardial infarction in the Geneva area were referred to the same hospital, they and their relatives are representative of the local population. Only when death occurred before or shortly after hospital admission, was it not possible on practical and psychological grounds to contact and investigate the patients’ relatives. The results show that female first-degree relatives of patients hospitalized for acute myocardial infarction differ from women of a normal control population by their HDL lipids. Before the age of 40 years, the HDL triglycerides were increased in female relatives of CHD patients without significant lowering of HDL cholesterol. None of these women had clinical evidence of cardiovascular disease. Similar increases in HDL triglycerides without changes in HDL cholesterol have been observed by Bergstrand [9] in myocardial infarction patients. In contrast, the HDL cholesterol was lower in the older relatives (age 40 years and over) than the control group, but HDL triglycerides did not differ. The low HDL cholesterol in the female relatives aged 40-49 years, as in male relatives of CHD patients [ 51, does not appear to be secondary to extensive atherosclerosis since only one out of the 36 female relatives in this age group
426
had clinical evidence of atherosclerosis. Her HDL cholesterol was low (1.16 mmoi/l). When this one value was removed from the group, the mean HDL cholesterol of the relatives without evidence of cardio-vascular disease still remained lower than the controls (P < 0.05). Similar to male relatives of CHD patients [ 51, HDL lipid changes in female relatives do not appear to be secondary to clinically overt cardiovascular disease but precede its occurrence. Eight of the 62 relatives aged 50 and over had clinical manifestations of cardiovascular disease. The HDL cholesterol of these 8 women was lower (1.32 mmol/l) than the controls (1.58 mmol/l). In contrast, the HDL cholesterol of the relatives without clinical evidence of cardiovascular disease in the same age group was 1.47 mmol/l, not different either from HDL cholesterol of the controls or of the atherosclerotic relatives. Atherosclerosis may be present without clinical manifestations. It is likely that the absence of differences in HDL cholesterol between the relatives with and without evidence of atherosclerosis may be due to older women having various degrees of atherosclerosis in the absence of symptoms. It is, however, apparent that the relatives with the lowest HDL cholesterol had developed clinically more advanced atherosclerosis than the others. It appears from this study that in female relatives of CHD patients HDL lipid changes occur in the absence of clinical occurrence of cardiovascular disease. In contrast to young male relatives of CHD patients [ 51, young female relatives had high HDL triglycerides without significant lowering of HDL cholesterol. The lack of significance of the low HDL cholesterol is possibly related to the small number of young relatives who were examined. However, it is known that females develop coronary vascular disease with a 15- to 20-year delay as compared to males. It is therefore possible that the low HDL cholesterol which is already significant at the age of 20-29 years in male relatives [5] becomes apparent at a later age in female relative of CHD patients. Another factor which may influence HDL cholesterol at a younger age in male than in female relatives are the VLDL triglycerides. They are more elevated and show a wider range of values in males than in females in the younger age group [5]. Higher VLDL triglycerides may further lower the HDL cholesterol in young male relatives as compared to women. The inverse relationship between HDL cholesterol and VLDL triglycerides has been observed both in the female relatives and controls. The regression analysis showed no difference in the slope and intercepts for either group (Fig. 4). This was in contrast to the previous finding that the male close relatives of the CHD patients [5] had similar slopes to, but different intercepts from, the controls, which means that at a given VLDL triglycerides concentration, the male relatives had a significantly lower HDL cholesterol level than the controls. The lack of differences between female relatives and controls could be related to the lower VLDL triglycerides measured in female as compared to male relatives. The effect of hormonal contraception on HDL was evident in female relatives of CHD patients using “the pill” who had higher LDL and HDL triglycerides and lower HDL cholesterol than the controls. Hormonal contraception has been shown to favor the development of cardiovascular disease [lo, 111 possibly by lipoprotein changes [12-141.
427
HDL
-
Controls
(l-l = 132)
-------
Relatives
(n i 167)
VLDL
Triglycerides
Cholesterol ~mmollll
Immolll)
Fig. 4. Regression lines with their 95% confident limits for ordinate of HDL cholesterol versus VLDL triglycerides.
Due to the wide variety of pills in use it was not possible to evaluate the specific effect of oestrogens or progestin. The HDL lipid changes observed in the families of the CHD patients may be due to genetic or environmental factors or both. In order to assess further the influence of familial environment on HDL, the HDL cholesterol and triglycerides of probands’ wives were analysed. They share the same environment as their husband but have a different genetic background. If genetic influence is to be expected as being one of the causes of the low HDL cholesterol observed in the female and male first-degree relatives of CHD patients, one would anticipate that patients’ wives would have a HDL cholesterol content similar to the control population. On the other hand, if environmental factors play a major role on the HDL cholesterol content of the close relatives of coronary patients, the patients’ wives would be more likely to have low HDL cholesterol. In order to minimize the possible effect of age on serum lipids, the wives of the CHD patients were compared to the women of the control group in the same age range. The spouses of the CHD patients had a lower serum cholesterol (4.97 mmol/l, P< 0.05) than the controls (5.34 mmol/l). The low serum cholesterol was due in part to lower (but not significantly) LDL cholesterol but mainly to low HDL cholesterol and triglycerides (Table 2). In view of this observation one may suspect that environmental factors may play an important role in the lowering of HDL cholesterol in the families of CHD patients. Among environmental factors which may influence HDL cholesterol, alcohol intake, cigarette smoking and physical activity both at work and at leisure have been studied. Hormonal contraception has already been discussed. Alcohol increases HDL cholesterol [ 151. The low HDL cholesterol of the first-degree relatives of CHD patients was not related to differences in alcohol consumption as the percentage of alcohol consumers and their average intake was not different between the first degree relatives and the controls. In contrast there were 7% only of the wives of CHD patients who used to drink alcohol as compared to 49% of the controls. This suggests that the low HDL cholesterol in
428
probands’ wives could be related to differences in alcohol intake. Cigarette smoking which may lower HDL cholesterol [16] cannot explain the HDL cholesterol differences between the relatives, the wives and the controls as the percentage of smokers and the daily cigarette consumption was not different between the probands’ wives, the relatives and the controls. Heavy physical activity, which has been shown to increase HDL cholesterol [ 161, could not account for HDL cholesterol differences since less than 5% of the women in each group had high physical activity. It appears from this study that the low HDL cholesterol observed in female first-degree relatives was not explained by increased VLDL triglyceride or by known environmental factors, like smoking and drinking habits or physical activity. As most of the firstdegree relatives aged 20 years and over do not live in the probands’ house the low HDL cholesterol is likely to be a genetic trend possibly related to apoprotein composition. Conclusion The HDL cholesterol and triglycerides content differ in female first-degree relatives of patients with acute myocardial infarction from those observed in a normal control population. After the age of 40 years, HDL cholesterol was significantly lower in the relatives of CHD patients that in normals. It was not related to differences in physical exercise, alcohol intake or cigarette smoking. As most of the close relatives do not share the proband’s household, it appears more likely that the low HDL cholesterol in the close relatives of CHD patients was genetically determined and possibly related to the apoprotein composition [ 171. In the younger age group (20-39 years) there was a slight but significant increase in HDL triglycerides and a trend (not significant) toward low HDL cholesterol which was significant among the relatives using hormonal contraception. Probands’ wives who have a different genetic background than the probands but share the same households have a low HDL cholesterol level as compared to controls. Lower alcohol intake in probands’ wives could account for the difference. Acknowledgements We wish to thank Mrs. B. Kalix, F. Ruinard and B. Von Arx, technicians, Mrs. M. Jaillot, nurse, for their great help during this study.
and
References 1 Medalie, J.H.. Kahn, H.A.. Neufeld. N.H., Riss. E. and Goldbourt, U.. Five-year myocardial infarction incidence, Part 2 (Association of single variable to age and birth place), J. Chron. Dis., 26 (1973) 329. 2 Micheli. H.. Pometta, D. et Bloch. A.. CholestCrol et txiglyc&ides des lipoprot&nes driques isolees par ultracentrifugation - Relation avec I’ath~roscldrose coronarienne, Schweiz. Med. Wschr., 104 (1974) 1794. 3 Miller, G.J. and Miller, N.E., Plasma high-density lipoprotein concentration and development of ischaemic heart-disease, Lancet, 1 (1975) 16.
429
4 Gordon, T., C&eIIi, W.P.. Hjortland, M.C., Kannel, W.B. and Dawber, T.R.. High density lipoprotein as a protective factor against coronary heart disease -The Framingham Study. Amer. J. Med., 62 (1977) 707. 5 MicheIi, H., Pometta. 6
7 8 9
10
D., Jornot,
C. and Scherrer.
J.R., High density lipoproteins
cholesterol
in Inale
relatives of coronary patients, Atherosclerosis, 32 (1979) 269. Lewis, B., Chait. A., Sigurdsson, G.. Mancini. M., Farinaro, E.. Oriente, P.. Carlson. L.A.. Ericsson, M.. MicheIi, H. and Pometta. D., Serum lipoproteins in four European communities -A quantitative comparison, Europ. J. CIin. Invest., 8 (1978) 165. Abe& L.L.. Levy, B.B.. Brodie, B.B. and Kendall, F.E.. Simplified method for the estimation of total cholesterol in serum and demonstration of its specificity, J. Biol. Chem., 195 (1952) 357. for determination of serum triglycerides, Clin. Chem.. Soloni, F.G.. Simplified manual micromethod 17 (1971) 529. Bergstrand, R., WikIund, 0. and Helm, G., Glucose tolerance, plasma insulin and alpha-lipoproteins in young male myocardial infarction patients compared with control matched on serum cholesterol level. In: 0. Wiklund. Alpha Lipoproteins in Relation to Myocardial Infarction. Thesis University of G&eberg, 1978, PP. l-12. Mann. J.I., Vessy. M.P.. Thorogood. M. and Doll. R., Myocardial infarction in young women with special reference to oral contraceptive practice, Brit. Med. J., 2 (1975) 241.
11 Stem, M.P., Brown, B.W., HaskeII. W.L., Farquhar, J.W.. Wehrle. C.L. and Wood, P.D.S., Cardiovascular risk and use of estrogens or estrogen-progestagen combinations - Stanford Three-Community Study. Amer. J. Med., 235 (1976) 811. 12 Bradley. D.D., Wingerd, J.. Petitti, D.B., Krauss. R.M. and Ramcharan. S., Serum high-density lipoprotein - Cholesterol in women using oral contraceptives. estrogens and progestins. New EngI. J. Med.. 299 (1978) 17. 13 Arntzenius. A.C., Van Gent, C.M.. Van Der Voort, H., Stegerhoek, C.I. and Styblo. K., Reduced highdensity lipoprotein in women aged 40 to 41 using oral contraceptives, Lancet, 1 (1978) 1221. 14 Pometta. D. and MicheIi. H., Influence de la contraception hormonale sur Ies fiPoProt6ineSS6riWeS. Schweiz. Med. Wschr., 108 (1978) 2012. 15 CasteIIi. W.P., Doyle. J.T., Gordon, T.. Hames. C.G.. Hjortland, M.C.. Hulley, S.B., Kagan. A. and Zukel. W.J.. Alcohol and blood lipids -The cooperative lipoprotein phenotyping study, Lancet. 2 (1977) 153. 16 Enger, S.C., Herbj@rnsen, K.. Erickssen, K. and Fretland. A.. High density 1ipoProteins (HDL) and physical activity - The influence of physical excercise, age and smoking on HDL cholesterol and the HDL/totaI cholesterol ratio, Stand. J. CIIn. Lab. Invest., 37 (1977) 251. 17 Albers, J.J., Cheung, M.C. and Hazard. W.R.. High-density lipoproteins in myocardiai infarction survivors. Metabolism. 27 (1978) 479.
34 (1979) 419-429 @ Elsevier/North-Holland Scientific Publishers, Ltd.
Atherosclerosis,
HDL LIPIDS IN CLOSE RELATIVES PATIENTS Environmental
and Genetic
OF CORONARY
HEART DISEASE
Influences
DANIEL POME’M’A, HORACE MICHELI, ALAN SUENRAM and CONSTANTIN JORNOT
Divisions de Diabktologie et d’znformatique, Geneva, Geneva (Switzerland)
Department
of Medicine of the University of
(Received 21 May, 1979) (Revised, received 10 July, 1979) (Accepted 11 July, 1979)
Female fiistdegree relatives of CHD patients differed, after the age of forty, from the normal control population by their low HDL cholesterol. Between the ages of 20 and 40 years a slight but significant increase in HDL triglycerides was observed. Except for hormonal contraception which induces significant lowering of HDL cholesterol in the first degree relatives, the observed differences in HDL lipids did not seem related to environmental factors but rather to be genetically determined. In contrast, the low HDL cholesterol observed in the wives of the CHD patients appeared to be related to differences in alcohol intake. Key words:
Coronary heart disease - Enuironmental factors -Female relatives of CHD patients - HDL cholesterol triglycerides - Wives of CHD patients
Introduction The high incidence and prevalence of coronary heart disease (CHD) today represents a major medical threat. Familial clustering of coronary artery disease has been demonstrated but the reasons for the increased familial prevalence are not firmly established and genetic or environmental factors may play a role.
Supported by a grant (6.13000.74) of the Swiss National Science Foundation. Reprint requests to: Prof. D. Porn&a. Division de MabCtolode. HOpital Canton& Switzerland.
CH-1211 Geneva 4.
420
The influence of serum cholesterol and triglycerides on the development of coronary artery disease has been studied extensively. More recently the interest has been focusing on the association of HDL cholesterol with coronary heart disease. Clinical and epidemiological studies have established that there is a strong negative correlation between plasma HDL cholesterol and the existence of coronary heart disease [ 1,3]. It has been shown retrospectively in patients older than 40 years that the power of the HDL cholesterol as a negative predictor for myocardial infarction is greater than the power of the LDL cholesterol as a positive predictor [4]. We have been able to demonstrate that male relatives of patients suffering from myocardial infarction have low HDL cholesterol even in the absence of cardio-vascular disease [5], an observation which may be of significant importance for explaining the development of coronary heart disease in families and may suggest genetic determinism. Myocardial infarction occurs less frequently and at a later age in women than in men, therefore it is of interest to determine whether the lipoprotein lipid pattern, and particularly the HDL cholesterol content in female relatives (sisters, daughters, mothers) of patients suffering from acute myocardial infarction, show a pattern similar to that observed in male relatives of CHD patients or if they differ. In order to distinguish between the role of genetic and environmental factors on HDL cholesterol, the wives of the CHD patients who share similar environments but have different genetic background from the probands were examined. Material and Methods The female firstdegree relatives of 117 consecutive male and female patients with acute myocardial infarction hospitalized in the Department of Medicine were admitted into the study as well as the wives of the male patients. The relatives and wives of male probands 55 years and older and female probands 70 years and over were excluded from the study. The difference in age for the men and the women was chosen because of the lower incidence of myocardial infarction among women below the age of 55. Families with fewer than two first-degree relatives were not considered for the study. However, if fewer than two first-degree relatives of the eligible families attended the medical examination, the probands, wives or relatives already examined were not excluded from the study. The control group previously examined for the four European Cities Survey [6] consisted of 132 female Genevese civil servants without clinical or electrocardiographic evidence of ischaemic heart disease, peripheral vascular disease or any chronic or acute illness. Subjects taking drugs affecting serum lipids were excluded from the study. However, considering the wide use of hormonal contraception in young women, the relatives on “the pill” were investigated and included as a separate group in the present study. Medical examination was performed in all subjects with special emphasis on atherosclerosis (history of angina pectoris, intermittent claudication, peripheral pulses, skin temperature and color, xanthoma). It included blood pressure
421
measurement and ECG recording. Smoking and drinking habits and physical activity were assessed by direct questioning. The alcohol consumption was expressed as liters of wine (10’) per week as in Geneva the use of wine is more widespread than other alcohols. Beer and hard liquors consumed were therefore transformed in wine “equivalent” on the basis of pure alcohol content. Serum and lipoprotein cholesterol was determined by a modification of Abell and Kendall’s method [ 71 as described previously [ 51. Triglycerides were measured according to Soloni [ 81. VLDL, LDL and HDL were separated by quantitative preparative ultracentrifugation in two steps at densities 1.006 and 1.063 as previously described [ 51. The recovery was 93% for cholesterol and 92% for triglycerides. Chemical and ultracentrifugation procedures were previously standardized by sample exchange between laboratories [6]. The statistical analysis of the differences between means of two samples was performed using the Student t-test, the Mann-Whitney U Test and the comparison of regression lines. Before calculation, serum and VLDL triglycerides and VLDL cholesterol were transformed to their logarithms. In order to take into account the influence of age on lipoprotein lipids, the results for the first-degree relatives are reported separately for the age groups 20-39 years, 40-49, and 50 and over. Because of the smaller number of wives and their limited age range, they were all considered together and compared to the controls in the same age range. Results One hundred and seventeen families of 76 male and 41 female probands entered the study. There were 231 living female firstdegree relatives (sisters, daughters, mothers) aged 20 and over eligible for the study. The percentage of dead relatives at the time of the study differs according to the generation. Among the probands’ mothers, 56% were dead as compared with 1% among the daughters. The mortality among the probands’ sisters was 7 percent. 147 (64%) of the living relatives were examined. Most of the relatives who did not attend the medical examination were living too far away from Geneva. In the younger age group (20-39 years) 15 of the 49 relatives (31%) were currently taking “the pill”. Their results were considered separately. In addition, 66 spouses of male probands were examined. Six of them using hormonal contraception were not considered in the present study. Firs t-degree relatives Serum lipids Serum cholesterol was elevated (5.80 mmol/l, P< 0.05) among relatives of patients with myocardial infarction in the age group 40-49 years as compared to the controls (5.27 mmol/l) but no difference was found in serum cholesterol in the younger and older age groups (Table 1). Serum triglycerides did not differ between relatives of patients with ischaemic heart disease and the control group even among the women using hormonal contraception.
422 TABLE 1 SERUM CHOLESTEROL AND TIENTS AND CONTROLS
TRIGLYCERIDES
(mmol/l)
IN FEMALE
4049
Age (years)
20-39
Groups Hormonal contraception Number
relatives
relatives
controls
relatives
yes 15
no 34
no 45
no 36
SWll
RELATIVES
OF CHD PA-
50 and over controls
relatives
controls
no 62
no 50
5.27 0.94 -0.1029 0.2155 0.79
6.15 1.08 0.0583 0.1947 1.14
6.50 1.11 0.0362 0.1940 1.09
??I
Cholesterol SD Triglycerides (log) SD antilog
4.70 0.80 -0.0975 0.1850 0.80
4.72 0.65 -0.1252 0.1968 0.75
4.91 0.79 -0.1735 0.1756 0.67
5.80 * 1.02 0.0200 0.2533 0.96
HDL Cholesterol SD Triglycerides SD
1.39 * 0.32 0.17 * 0.06
1.44 0.35 0.14 * 0.06
1.58 0.32 0.11 0.04
1.42 * 0.34 0.14 0.06
1.65 0.44 0.14 0.08
1.45 * 0.31 0.15 0.05
1.58 0.33 0.14 0.06
LDL Cholesterol SD Triglycerides SD
2.67 0.70 0.29 *J 0.08
2.68 0.52 0.23 0.11
2.78 0.63 0.22 0.07
3.51 ** 0.88 0.29 0.08
2.94 0.85 0.25 0.10
3.76 0.95 0.33 0.10
4.06 0.88 0.31 0.08
-0.1768 0.2129 0.67 -0.4609 0.2838 0.35
-6.2727 0.2952 0.53 -6.5538 0.2717 0.28
-6.0845 0.2494 0.82 -0.4762 0.3181 0.33
-9.503 0.2442 1.12 -9.2599 0.2855 0.55
-0.0201 0.2609 1.05 -0.2627 0.3053 0.55
VLDL Cholesterol SD antilog Triglycerides (log) SD antilog
-0.2037 0.3030 0.63 -0.5302 0.2881 0.30
-0.0133 0.3176 0.97 -9.3611 0.3810 0.44
Comparison relatives vs controls * P < 0.05, ** P < 0.01. 1 Comparison relatives using the pill vs relatives not using the pill.P < 0.06,
and vs the controls,
P < 0.01.
Serum lipoproteins HDL. At age 40 years and above, the HDL cholesterol (Fig. 1) was significantly lower, 1.42 mmol/l, P < 0.05 (age group 40-49 years) and 1.45 mmol/l, P < 0.05 (50 years and above), in the close relative8 of patient8 with myocardial infarction than in the controls (1.65 mmol/l and 1.58 mmol/l, respectively). HDL triglycerides did not differ in relatives and controls after the age of 40 year8 (Table 1). In the younger age group (20-39 years) the HDL cholesterol had a tendency to be lower among the relatives but the difference was not significant. In contrast the HDL triglyceride8 were higher (0.14 mmol/l, P< 0.05) among the relatives than in the control8 (0.11 mmol/l) (Fig. 2). When the CHD patients’ relatives using hormonal contraception were considered, it appeared that the difference8 observed between relatives of coronary patients and controls were more marked. HDL triglycerides were increased (0.17 mmol/l, P < 0.05) a8 compared to the controls (0.11 mmol/l) and their HDL cholesterol wa8 significantly lower (1.39 mmol/l, P < 0.05) when compared to the HDL cholesterol of the controls (1.58 mmol/l).
423
Cholesterol
-
(mmolll)
rrletivcs lno drugs) controls
0
1.8
??
peo.05
1.6
1.4
,I
C age
LO- 49
groups
“=
31
Fig. 1. HDL over).
36
cholesterol
years
X0 50
in female
62
first-degree
relatives
of CHD patients
and controls
(age 40 years and
LDL. In the age group 40-49 years, the LDL cholesterol was increased in the relatives of CHD patients (3.51 mmol/l, P< 0.001) as compared to the controls (2.94 mmol/l) (Table 1). There was no increase in LDL cholesterol in younger or older women related to CHD patients. LDL triglycerides did not differ between relatives and controls except in relatives using the “pill” who had higher LDL triglycerides (0.29 mmol/l, P< 0.05) than the controls (0.22 mmol/l). VLDL. No difference in VLDL cholesterol and triglycerides was found in any age group between the controls and the relatives, whether or not on the “Pill”. Q
sgr20-39 yrsrs
Imrsn f SEMI
Trlglycerldes IrnrnolllI
Cholesterol
0.20.
~p
-p
0
controls
(L5)
Fig. 2. HDL contraception
m
relatives (no drug) (3L1
cholesterol and triglycerides or not and in controls.
m
relatwes on the”p~ll” in young
(151
female first-degree
?e.htives
either
using hormonal
424 TABLE 2 SERUM
AND
PATIENTS
LIPOPROTEIN
CHOLESTEROL
AND TRIGLYCERIDES
AND CONTROLS Wives (n = 60)
Serum Cholesterol SD Triglycerides (log) SD antilog
4.97 0.88 -0.1129 0.0851 0.77
Controls (n = 76)
5.34 P < 0.05 0.98 -0.0864 0.2227 0.82
Wives (n = 58)
Controls (n = 76)
1.44 0.29 0.15 0.05
1.59 0.40 0.13 0.07
2.85 0.72 0.24 0.09
3.05 0.81 0.26 0.10
-0.5600 0.2964 0.28 -0.0902 0.3397 0.81
4.5422 0.2813 0.29 a.4330 0.3231 0.37
HDL Cholesterol SD Triglycerides SD LDL Cholesterol SD Triglycerides SD VLDL Cholesterol (log) SD antilog Triglycerides (log) SD antilog
Cholesterol (mmolll) 1.8
1.6 p-zo.02
“=
controls
wives
91
56
Fig. 3. HDL cholesterol in wives of CHD patients.
P < 0.02
(mmolfl)
IN WIVES OF CHD
425
Wives of CHD patients The age of the 60 wives ranged from 26 to 58 years. Their mean age was 43.8 years and the median age 43.0 years. They were compared with the women of the control group in the same age range who had a mean and median age of 43.2 years. The serum cholesterol of the patients’ wives was 4.97 mmol/l and triglycerides 0.7 7 mmol/l. Due to technical difficulties serum ultracentrifugation was not performed in two wives. The mean HDL cholesterol of the 58 wives was lower (1.44 mmol/l, P < 0.02) than HDL cholesterol of the controls (1.59) (Table 2 and Fig. 3). Environmental factors Cigarette smoking The percentage of smokers was 33% among the relatives (average 18 cigarettes daily), 32% among the probands’ wives (18 cigarettes daily) and 28% among the controls (10 cigarettes daily). Physical activity 3% of the relatives and 3% of the probands’ wives had high physical activity at work and/or at leisure as compared to 5% of the controls. The other subjects had medium or low physical activity. Alcohol in take 46% of the firstdegree relatives and 49% of the controls drank alcohol as opposed to 7% of the probands’ wives. The average weekly alcohol intake was 1.5 liter wine equivalent in all 3 groups. Discussion Since all patients with acute myocardial infarction in the Geneva area were referred to the same hospital, they and their relatives are representative of the local population. Only when death occurred before or shortly after hospital admission, was it not possible on practical and psychological grounds to contact and investigate the patients’ relatives. The results show that female first-degree relatives of patients hospitalized for acute myocardial infarction differ from women of a normal control population by their HDL lipids. Before the age of 40 years, the HDL triglycerides were increased in female relatives of CHD patients without significant lowering of HDL cholesterol. None of these women had clinical evidence of cardiovascular disease. Similar increases in HDL triglycerides without changes in HDL cholesterol have been observed by Bergstrand [9] in myocardial infarction patients. In contrast, the HDL cholesterol was lower in the older relatives (age 40 years and over) than the control group, but HDL triglycerides did not differ. The low HDL cholesterol in the female relatives aged 40-49 years, as in male relatives of CHD patients [ 51, does not appear to be secondary to extensive atherosclerosis since only one out of the 36 female relatives in this age group
426
had clinical evidence of atherosclerosis. Her HDL cholesterol was low (1.16 mmoi/l). When this one value was removed from the group, the mean HDL cholesterol of the relatives without evidence of cardio-vascular disease still remained lower than the controls (P < 0.05). Similar to male relatives of CHD patients [ 51, HDL lipid changes in female relatives do not appear to be secondary to clinically overt cardiovascular disease but precede its occurrence. Eight of the 62 relatives aged 50 and over had clinical manifestations of cardiovascular disease. The HDL cholesterol of these 8 women was lower (1.32 mmol/l) than the controls (1.58 mmol/l). In contrast, the HDL cholesterol of the relatives without clinical evidence of cardiovascular disease in the same age group was 1.47 mmol/l, not different either from HDL cholesterol of the controls or of the atherosclerotic relatives. Atherosclerosis may be present without clinical manifestations. It is likely that the absence of differences in HDL cholesterol between the relatives with and without evidence of atherosclerosis may be due to older women having various degrees of atherosclerosis in the absence of symptoms. It is, however, apparent that the relatives with the lowest HDL cholesterol had developed clinically more advanced atherosclerosis than the others. It appears from this study that in female relatives of CHD patients HDL lipid changes occur in the absence of clinical occurrence of cardiovascular disease. In contrast to young male relatives of CHD patients [ 51, young female relatives had high HDL triglycerides without significant lowering of HDL cholesterol. The lack of significance of the low HDL cholesterol is possibly related to the small number of young relatives who were examined. However, it is known that females develop coronary vascular disease with a 15- to 20-year delay as compared to males. It is therefore possible that the low HDL cholesterol which is already significant at the age of 20-29 years in male relatives [5] becomes apparent at a later age in female relative of CHD patients. Another factor which may influence HDL cholesterol at a younger age in male than in female relatives are the VLDL triglycerides. They are more elevated and show a wider range of values in males than in females in the younger age group [5]. Higher VLDL triglycerides may further lower the HDL cholesterol in young male relatives as compared to women. The inverse relationship between HDL cholesterol and VLDL triglycerides has been observed both in the female relatives and controls. The regression analysis showed no difference in the slope and intercepts for either group (Fig. 4). This was in contrast to the previous finding that the male close relatives of the CHD patients [5] had similar slopes to, but different intercepts from, the controls, which means that at a given VLDL triglycerides concentration, the male relatives had a significantly lower HDL cholesterol level than the controls. The lack of differences between female relatives and controls could be related to the lower VLDL triglycerides measured in female as compared to male relatives. The effect of hormonal contraception on HDL was evident in female relatives of CHD patients using “the pill” who had higher LDL and HDL triglycerides and lower HDL cholesterol than the controls. Hormonal contraception has been shown to favor the development of cardiovascular disease [lo, 111 possibly by lipoprotein changes [12-141.
427
HDL
-
Controls
(l-l = 132)
-------
Relatives
(n i 167)
VLDL
Triglycerides
Cholesterol ~mmollll
Immolll)
Fig. 4. Regression lines with their 95% confident limits for ordinate of HDL cholesterol versus VLDL triglycerides.
Due to the wide variety of pills in use it was not possible to evaluate the specific effect of oestrogens or progestin. The HDL lipid changes observed in the families of the CHD patients may be due to genetic or environmental factors or both. In order to assess further the influence of familial environment on HDL, the HDL cholesterol and triglycerides of probands’ wives were analysed. They share the same environment as their husband but have a different genetic background. If genetic influence is to be expected as being one of the causes of the low HDL cholesterol observed in the female and male first-degree relatives of CHD patients, one would anticipate that patients’ wives would have a HDL cholesterol content similar to the control population. On the other hand, if environmental factors play a major role on the HDL cholesterol content of the close relatives of coronary patients, the patients’ wives would be more likely to have low HDL cholesterol. In order to minimize the possible effect of age on serum lipids, the wives of the CHD patients were compared to the women of the control group in the same age range. The spouses of the CHD patients had a lower serum cholesterol (4.97 mmol/l, P< 0.05) than the controls (5.34 mmol/l). The low serum cholesterol was due in part to lower (but not significantly) LDL cholesterol but mainly to low HDL cholesterol and triglycerides (Table 2). In view of this observation one may suspect that environmental factors may play an important role in the lowering of HDL cholesterol in the families of CHD patients. Among environmental factors which may influence HDL cholesterol, alcohol intake, cigarette smoking and physical activity both at work and at leisure have been studied. Hormonal contraception has already been discussed. Alcohol increases HDL cholesterol [ 151. The low HDL cholesterol of the first-degree relatives of CHD patients was not related to differences in alcohol consumption as the percentage of alcohol consumers and their average intake was not different between the first degree relatives and the controls. In contrast there were 7% only of the wives of CHD patients who used to drink alcohol as compared to 49% of the controls. This suggests that the low HDL cholesterol in
428
probands’ wives could be related to differences in alcohol intake. Cigarette smoking which may lower HDL cholesterol [16] cannot explain the HDL cholesterol differences between the relatives, the wives and the controls as the percentage of smokers and the daily cigarette consumption was not different between the probands’ wives, the relatives and the controls. Heavy physical activity, which has been shown to increase HDL cholesterol [ 161, could not account for HDL cholesterol differences since less than 5% of the women in each group had high physical activity. It appears from this study that the low HDL cholesterol observed in female first-degree relatives was not explained by increased VLDL triglyceride or by known environmental factors, like smoking and drinking habits or physical activity. As most of the firstdegree relatives aged 20 years and over do not live in the probands’ house the low HDL cholesterol is likely to be a genetic trend possibly related to apoprotein composition. Conclusion The HDL cholesterol and triglycerides content differ in female first-degree relatives of patients with acute myocardial infarction from those observed in a normal control population. After the age of 40 years, HDL cholesterol was significantly lower in the relatives of CHD patients that in normals. It was not related to differences in physical exercise, alcohol intake or cigarette smoking. As most of the close relatives do not share the proband’s household, it appears more likely that the low HDL cholesterol in the close relatives of CHD patients was genetically determined and possibly related to the apoprotein composition [ 171. In the younger age group (20-39 years) there was a slight but significant increase in HDL triglycerides and a trend (not significant) toward low HDL cholesterol which was significant among the relatives using hormonal contraception. Probands’ wives who have a different genetic background than the probands but share the same households have a low HDL cholesterol level as compared to controls. Lower alcohol intake in probands’ wives could account for the difference. Acknowledgements We wish to thank Mrs. B. Kalix, F. Ruinard and B. Von Arx, technicians, Mrs. M. Jaillot, nurse, for their great help during this study.
and
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