Somogyi JC, Bir Biro Gy, Hotzel D (eds): (eds): Nutrition and Cardiovascular Cardiovascular Risks. Risks. Somogyi Bibl Nutr Dieta. Basel, Basel, Karger, 1992, 1992, No. Bib! No 49, 49, pp pp 66-78
High-Density Lipoprotein Particles, and Arteriosclerosis Arteriosclerosis Nutrition and J. J. C. C. Fruchart Institut Pasteur de Lille, France
High-density lipoproteins clinical High-density lipoproteins (HDL) (HDL) have have been the focus of many clinical epidemiological investigations elevated concentration in and epidemiological investigations because because their their elevated of coronary artery disease [41]. plasma is associated with a reduced incidence of HDL consist consist of of aa collection collection of of particles differing in size, density Human HDL apolipoprotein content content[[I]. and apolipoprotein 1 ].On On the the basis basis of of their their hydrated density, two = 1.063-1.125 1.063-1.125g/m1) glml) and subfractions have have been identified as HDL2 (d = main subfractions HDL3 (d == 1.125-1.21 1.125-1.21 g/ml) glml) [22]; [22]; each of these ΗDL3 these subfractions subfractions can be be further into discrete discrete subclasses subclasses by different techniques [12, [12, 43]. fractionated into metabolism is is not not yet yet well well known and and no no specific specific function(s) The HDL metabolism have been assigned to HDL subfractions defined by physical ultracentrifugal flotation criteria. It seems that HDL particles are produced both in the liver and intestine intestine [57] [57] and and continuously continuouslyconvert convertin inthe theplasma plasma[[1,43]. and 1, 43]. Moreover, of triglyceride (TG)-rich surface material derived from the catabolism surface material derived from the catabolism of triglyceride (TG)-rich contribute to to their their synthesis synthesis [26, [26, 50]. 50]. Several studies lipoproteins appears to contribute reverse cholesterol cholesterol transport, a chain of have shown that HDL can promote reverse remove excess excess cholesterol cholesterol from metabolic reactions that remove from peripheral tissues transport cholesterol cholesterol directly or indirectly, with other lipoproteins to the and transport is generally generally accepted process involves different steps. It is liver [32, 41]. This process that interactions interactions between between ΗDL3 HDL3 and HDL HDL binding binding sites sites facilitate facilitate egress egress of cholesterol from cells cells to to the the plasma lipoproteins [6, 45] 45] and particularly to a pre-~ migrating HDL [[16]. 16].This This incorporation incorporation of free free cholesterol cholesterol into small pre-β acceptor lipoproteins, lipoproteins, facilitating facilitating its its esterification esterification by by lecithin:cholesterol lecithin:cholesterol acceptor cholesteryl ester protein (CETP), (CETP), acyltransferase (LCAT) acyltransferase (LCAT)and and its its cholesteryl ester transfer protein transfer to to very-low-density very-loW-density lipoproteins lipoproteins (VLDL) (VLDL) and exchange for mediates transfer triglycerides [28, 36].
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selected immunosorption techniques designed to Recent studies, using selected isolate subpopulations subpopulations of isolate of lipoproteins, lipoproteins, have have shown shown that that HDL HDL cannot be (apo) A-I A-II. regarded as homogeneous particles with apolipoproteins (αρο) Α-I and Α-II. of particles particles within within HDL HDL is is heterogeneous heterogeneous not only only with with The distribution of respect to their hydrated density but also in relation to their apolipoprotein 17, 40]. 40]. It is now recognized recognized that [I, 17, composition [1, that HDL contain at least two apo A-I-containing lipoprotein particles which might have different types of αρο [8, 20] 20] and and clinical clinical significance significance [47]. [47]. One species metabolic function function [8, species contains contains A-I and andA-II A-II (LpA-I:A-II) (LpA-I:A-II) while in the protein components componentsboth bothapo apoA-I as main protein such lipoprotein lipoprotein one αρο apo A-II A-II is absent absent (LpA-I). (LpA-I). The definition of such The definition other one particles by their apolipoprotein content has been proposed to more particles by their apolipoprotein content has been proposed to be more appropriate for for studying studying the chemical complexity and metabolic functions of appropriate system. the plasma lipoprotein system.
LpA-I and LpA-I:A-II are plasma by by LpA-I and LpA-I:A-II are currently currently purified purified from from total total plasma do sequential immunoaffinity immunoaffinity chromatography [17, 38, 44]. Some authors chromatography [17, 38, 44]. Some authors find any any difference difference in lipid composition composition between between LpA-I LpA-I and and LpA-I:A-II LpA-I:A-II not find [[17], 17],but but others others have have claimed claimed that that the percentage of triglyceride triglyceride and and the ratio ester/total cholesterol cholesterol are are lower lower in in L.ρΑ-I LpA-I than in in LpA-I:A-II LpA-I:A-II [44]. cholesteryl ester/total tobe behigher higherin inLpA-I LpA-I than than in inLpA-I:A-II LpA-I:A-II [2]. [2]. lipid/protein ratio ratioappears appearsto The lipid/protein apo A-I to αρο apo A-II in LpA-I:A-II LpA-I:A-II is is 1.5. 1.5. Small Small quantities quantities The molar ratio of αρο of apolipoproteins A-IV; fractions [2, [2, 19]. 19]. of A-Iv; C's, D, E are found in both fractions Of considerable considerable significance significance isis the finding that proteins stimulating the finding proteins stimulating (LCAT, CETP) or other reverse cholesterol transport (LCAT, other apolipoproteins such as apo J are mainly mainly present present in in LpA-I LpA-I and andnot notininLpA-I:A-II LpA-I:A-II [[19, 19, 20, 23]. αρο apo A-I A-I containing particles metaboOur understanding with respect to αρο However, it has been shown recently lism is limited. However, recently that both particles are LpA-I are are produced produced by by the the intestine intestineonly only[ [14]. synthesized by the liver but LpA-I 14]. between the two two subpopulations is is not not well well The metabolic interrelationship between established but seems that LpA-I are faster rate than established but itit seems that LpA-I are catabolized catabolized at at aa faster LpA-I:A-II [48]. is whether whether LpA-I LpA-I and and LpA-I:A-II LpA-I:A-II LpA-I:A-II [48]. One One of the key questions is physiological roles. have different physiological order to gain some insights into the mechanisms mechanisms by by which cholesterol In order movement takes takes place place in peripheral peripheral cells, cells, cultured cultured adipose adipose cells cells were used as movement a model. model. Indeed, the fact that adipose adipose tissue tissue has has the the ability ability to to accumulate, accumulate,
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store store and, and, when when needed, needed, mobilize mobilize aalarge large pool poolofofunesterified unesterified cholesterol cholesterol means that that these these cells cells meet the the requirement requirement for for the the study studyof ofreverse reverse cholescholesmeans terol transport transport [7]. [7]. Cholesterol Cholesterol efflux efflux mediated mediated by by A-I-containing A-I-containing particles particles terol has been been studied studied in in mouse mouse adipose adipose cells cells after aftercholesterol cholesterol preloading preloading with with has LDL [1 [11]. cholesterol efflux efflux LDL 1 ].Long-term Long-termexposure exposure to to LpA-I LpA-I particles particles promotes promotes cholesterol ofLpA-I:A-II. LpA-I:A-II. whereas no no efflux efflux was observed in the the presence presence of whereas These results, behave as as a distinct results, which which emphasize emphasize that LpA-I:A-II LpA-I:A-II behave distinct [21,37,54]. metabolic entity entity have have been been confirmed confirmed in other other studies studies [21, metabolic 37, 54]. ligands which which recognize recognize the cell cell surface surface HDL-binding HDL-binding sites sites have have The ligands apo A-I, A-I, αρο apo A-IV and αρο apo A-II [9, 51]. 51]. It was was proposed identified as aρo been identified that αρο apo A-I A-I and αρο apo A-IV A-IV play play the role role of of agonist agonist and and αρο apo A-II A-II that of of that of cholesterol cholesterol efflux efflux [[10]. [51] have suggested suggested that antagonist of 10]. Slotte Slotte et et al. al. [51] of ΗDL3 HDL3 to human human fibroblasts fibroblasts or bovine bovine endothelial endothelial cells cells induce a addition of protein kinase kinase C (PKC)-dependent translocation translocation of ofcholesterol from intracelprotein We have have recently recently demonstrated that membranes to the the cell cell surface. surface. We lular membranes efflux from adipose cells cells is is coupled coupled to to diacylglycerol diacylglycerol (DAG) (DAG) cholesterol efflux cholesterol from adipose αρο production and and protein protein kinase kinase C C activation activation [54]. [54]. The fact that binding of ofapo production A-I1DMPC complexes complexes produces of αρο apo A-II/ A-III A-I/DMPC produces diacylglycerol, diacylglycerol,but butnot not that that of complexes, strongly role of of αρο apo Α-II A-II as as that of of an an DMPC complexes, strongly supports supports the the role antagonist to produce produce cholesterol cholesterol efflux. efflux. In agreement with this interpretaIn agreement with this tion, has recently recently been been shown shown that that LpA-I:A-II LpA-I:A-II can can inhibit inhibit the the LpAILpAIn, itit has tíο ofcholesterol-preloaded cholesterol-preloadedadipose adiposecells cells[ [11]. promoted cholesterol cholesterolefflux efflux of promoted 11 ]. methods for direct direct quantification quantification of αρο apo A-I-containing A number of methods plasma have have been been developed: developed: immunoprecipitation immunoprecipitation[[18], particles in human plasma 18], two-phase electroimmunoassay electroimmunoassay [5], [5], enzyme-linked enzyme-linked differential-antibody differential-antibody two-phase immunosorbent assay assay [38]. These three methods are well well adapted immunosorbent adapted in research of accuracy. accuracy. The laboratories but are are time-consuming time-consuming and suffer suffer from lack of recent development of of a differential differential electro-immunoassay allows allows the direct measurement of LpA-I. LpA-I. By By using using a large large excess excess of of anti anti A-II, A-II, LpA-I:A-II LpA-I:A-II measurement first peak and LpA-I LpA-I migrates further as a second particles are retained in a first peak [46]. [46]. This This new new system systemcan canprovide provide specific specificand andreproducible reproducible determinadeterminaLpA-I in plasma. tions of LpA-I in human plasma.
Both A-I-containing particles particles were were detected αρο A-I-containing Both lipoprotein lipoprotein forms forms of apo mainly relative proportion proportion of of LpA-I LpA-I was was higher higher in HDL2 mainly in in HDL but the relative than HDL3. ΗDL3.
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Concentrations of LpA-I LpA-I and LpA-I:A-II LpA-I:A-II were plasma Concentrations were determined determined in plasma samples from normolipemic men and women. The average concentrations concentrations of LpA-I were 10% higher LpA-I were approximately approximately 10% higher in in women women than than in men. LpA-I but not LpA-I:A-II LpA-I:A-II are angiographiLpA-I are decreased decreased in normolipemic, angiographically documented CAD patients when compared to a group of asymptomatic asymptomatic subjects and a group group of ofpatients patients with with arteriographically arteriographically normal normal coronary coronary subjects arteries [47]. However, However, in a recent similar study where where the the patient group had higher triglyceride triglyceride levels levels than than controls, controls, we we have have found found that that both LpA-I and LpA-I:A-II are similar degree degree (one (one can can LpA-I:A-II arereduced reducedinin CAD CAD patients patients to to a similar ofLpA-I:A-II LpA-I:A-II particles particles in in hypertriglyceridemic hypertriglyceridemic the decrease decrease of postulate that the of such such particles during patients may be related to the the decreased decreased formation of lipolysis) [30]. A case case control study of apo A-I-containing particles has been performed populations at at contrasting contrastingrisk risk for for CAD CAD (ECTIM (ECTIM Study) Study) [15]. [15]. Male in three populations patients with with a myocardial myocardial infarction infarction and controls controls were were recruited recruited in two two patients Irish Center Center (Belfast). (Belfast). French centers (Strasbourg, Toulouse) and a Northern Irish restandardized mortality mortality rates rates in in Belfast, Belfast, Strasbourg, Strasbourg, Toulouse Toulouse are, reThe standardized spectively, 348, 78/100,000 for population. LpA-I LpA-I and spectively, 348, 102 102 and and 78/100,000 for the tested population. LpA-I:A-II levels but the the level level of ofLpA-I LpA-I:A-II levels are are lower lower in in patients patients than in controls but LpA-I differs significantly, on a statistical basis, between three populations: LpA-I differs significantly, on a basis, LpA-I was much lower in Belfast Belfast than than in the the French centers in controls and in cases. multivariateanalysis analysis suggests suggests that the the LpA-I/HDL LpA-I1HDL cholescholesIn this study, the multivariate is very very significant. significant. terol ratio ratio is of LpA-I LpA-I (but not not Recently, itit has also been been observed observed that level of Recently, has also that the level LpA-I:A-II) in LpA-I:A-II) in children children whose whose parents parents suffer sufferaa premature premature form form of CAD is control group with no no familial familial history history of of CAD CAD [3]. [3]. lower than in a control
Effect of of Alcohol Intake Particles Intake on HDL Particles Among dietary factors influence on plasma HDL Among factors studied for a possible influence levels, alcohol most intriguing intriguing because because it has has been been levels, alcohol intake intake isis one one of the most 27]. This be positively positively associated with HDL concentrations [4, 27]. reported to be some investigators investigators to suggest suggest that moderate moderate alcohol alcohol consumption consumption has led some may be be protective protective against against coronary coronary artery artery disease. disease. Epidemiological Epidemiological and may have been clinical data have provided some support for this idea, but there have many inconsistencies inconsistencies among demonstrate the many among studies studies that that have have sought sought to to demonstrate this context context [[10, beneficial effect effect of of alcohol alcohol consumption consumption in this 10, 27, 42].
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1. Characteristics of study groups Table 1. groups (mean (mean values values ± SD)
Group 1I (n = 68)
2 (n = 65)
3 (n = 69)
4 (n = 69)
55(n=63) (n = 63)
Age, years Age,
10.1 47.6 ± 10.1
47.6± 10.2 47.6 ± 10.2
47.7 ± 9.7
47.9 ± 9.7
47.9 ± 9.8
Alcohol consumption, Alcohol g/week
130 ± 40
59* 289 ± 59*
492 ± 60*
53* 676 ± 53*
189* 993 ± 189*
kg Body weight, kg
10.9 74.2 ± 10.9
77.2 ± 11.5 11.5
78.3 ± 12.3 12.3 78.3
1.8 1.8
13.6 75.6 ± 13.6
173 ± 6.1 6.1 173
172 ± 5.6
Height, cm
171.4 ± 6.3
172.4 ± 6.3
Erythrocyte corpuscular Jlm3 11 volume, µm3
92.9 ± 3.5
93.5 ± 3.9
94.7 ± 4.9**
14.9* 96.7 ± 14.9*
11.7** 96.3 ± 11.7**
y-Glutamyl transpeptidase, IU IV
24.1 ± 20.1 20.1 24.1
29.2 ± 22.9
48.5 ± 55.7 48.5
67.8 ± 77.6* 77.6*
114* 79.5 ± 114*
173.4±6 173.4 ±6
Although there is no no doubt doubt about the increase of plasma HDL concentrations on alcohol intake, the subfraction of of HDL HDL responsible responsible for this biologibiological observation has not yet been formally identified. Indeed, studies in which HDL has been been fractionated fractionated based on hydrated hydrated density density have have been been inconinconHDL clusive. Contradictory reputedly `anti'anticlusive. Contradictory data have been published about the reputedly atherogenic' HDL, i.e. the less dense HDL2 subfraction [13, 24, 25, 33-35, atherogenic' HDL, i.e. less dense HDL2 subfraction [13, 24, 25, 56]. 39, 49, 53, 55, 56]. Because of because of the potential Because of the the heterogeneity heterogeneity within within HDL HDL and because clinical and and metabolic metabolic importance importance of of newly newly defined defined apo apo AI-containing AI-containing clinical particles, we we decided to investigate investigate whether plasma concentralipoprotein particles, of LpA-I and LpA-I:A-II LpA-I:A-II were tions of were influenced influenced by by alcohol alcohol consumption. consumption. 25 to to 63 63 years) years) who who 334 men men (aged (aged from 25 Our study was carried out in 334 were undergoing were undergoing aa routine routine health health examination examination in in the the Health Center of the Lille, France. They completed a form, covering the past 6 Pasteur Institute, Lille, eating habits habits and cigarette cigarette and months, that allowed allowed the assessment assessment of eating months, alcohol consumption. with a alcohol consumption. Alcohol Alcohol intake intake was was recorded recorded by by an interview with specially trained physician. specially ofalcohol alcohol per per week), week), the men From the alcohol consumption consumptiondata data(grams (gramsof five groups were divided into five were groups with with either 63 or 69 subjects in each. of the subjects in each The main clinical and biochemical characteristics of 1. group are summarized in table 1.
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5: *p < 0.001, **p < 0.1. Significance of differences Significance differences among among group group 11 and and groups groups 2 through 5:
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study group (mean values SD) values ± SD) Table 2. Lipoprotein and apoprotein concentrations in study Group 2
3
4
3
5
LpA-I, mg/dl
51.9±26.6 51.9 ± 26.6
43.7±23.1 43.7 ± 23.1
43.9 ± 25.3
35.8 ± 18.3
19.2 37.2 ± 19.2
LpA-I:A-II, mg/dl LpA-Ι:Α-ΙΙ,
15.8 80.9 ± 15.8
85.0± 15.9 85.0 ± 15.9
86.8 ± Ι16.3 86.8 6.3
19.5 96.2 ± 19.5
10.3 ± 22.0
HDL cholesterol, mmol/I (mg/dl) mmol/1
1.41 1.41 ± 0.33 (54 ± 12)
1.46 ± 0.39 15) (56 ± 15)
1.54 ± 0.42 (59±16) (59 ± 16)
1.61 ± 0.52** 1.61 (62 ± 20)
1.94 ± 0.82* (75±31) (75 ± 31)
136.8 ± 32.4
128.8 ± 27.8
130.8±29.8 130.8 ± 29.8
10.2 39.9 ± 10.2
10.9 40.2 ± 10.9
10.1 42.2 ± 10.1
A-I, Apolipoprotein Α-Ι, mg/dI A-II, Apoliprotein Α-ΙΙ, mg/dI
26.1 132.0 ± 26.1 11.1 ** 44.3 ± 11.1**
140.7 ± 29.4
14.1* 48.5 ± 14.1*
The mean mean ±±SD SDage age of ofthe the subjects subjects in in each each group group was was 48 48 ± ± 10 years (table 1). All All groups groups were were comparable comparable for height, height, weight, weight, systolic systolic and diastolic diastolic 1). blood pressure, pressure, levels plasma glucose, glucose, plasma plasma blood levels of of plasma plasma uric uric acid acid and plasma of the erythrocyte corpuscular volume and aminotransferases, only in terms of y-glutamyl transpeptidase 1). of y-glutamyl transpeptidase activity (table 1). physical activity, activity, cigarette cigarette consumption, The mean physical consumption, and and eating habits were the same in the groups groups based were the same based on a questionnaire administered by the physician in charge of the subjects. Only HDL cholesterol cholesterol and api apo A-II A-II levels levels seemed seemed to to be besignificantly significantly Only :::::;200 200 g/week) glweek) was compared different when when group group 1 (alcohol (alcohol consumption, consumption, < 800 with either either group group 44 (alcohol (alcohol consumption consumption ranged ranged between between 601 with 601 and 800 glweek) or (alcohol consumption consumption > 800 800 g/week) glweek) (table mean g/week) or 5 (alcohol (table 2). 2). The mean increases of HDL, cholesterol and αρο apo A-II levels levels from increases from groups groups 1 through 5 were 37.6 and and 21.6%, 21.6%, respectively, respectively, while while αρο apo A-I A-I levels levels were were unaltered. unaltered. were correlation study study confirmed confirmed this this strong strong relationship relationship between between Results of a correlation Results 3). alcohol consumption cholesterol and αρο apo A-II A-II levels levels (table 3). alcohol consumption and and HDL cholesterol However, no correlation was found with apo A-I levels or with any of the However, no was αρο A-I levels or with any other parameters measured in this study. LpA-I:A-II levels The LpA-I:A-II levels were were increased increased from from groups 1 through 5, while the LpA-I levels LpA-I levels decreased decreased significantly significantly by by 28.3% 28.3% (fig. (fig. I). 1).The The correlation correlation study
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I: *p < 0.001, 0.00 I, **p < Significance of Significance of differences differencesamong among groups groups 22 through through 5 and group 1: < 0.1.
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analysis of ofy-glutamyl y-glutamyl transpeptidase transpeptidase activity, lipoproTable 3. Spearman's correlational analysis of alcohol intake with reported reported values values of teins, and apoproteins with Alcohol intake Alcohol
pp
y-Glutamyl transpeptidase
+0.30
High-Density Lipoprotein Particles, and Arteriosclerosis Arteriosclerosis Nutrition and J. J. C. C. Fruchart Institut Pasteur de Lille, France
High-density lipoproteins clinical High-density lipoproteins (HDL) (HDL) have have been the focus of many clinical epidemiological investigations elevated concentration in and epidemiological investigations because because their their elevated of coronary artery disease [41]. plasma is associated with a reduced incidence of HDL consist consist of of aa collection collection of of particles differing in size, density Human HDL apolipoprotein content content[[I]. and apolipoprotein 1 ].On On the the basis basis of of their their hydrated density, two = 1.063-1.125 1.063-1.125g/m1) glml) and subfractions have have been identified as HDL2 (d = main subfractions HDL3 (d == 1.125-1.21 1.125-1.21 g/ml) glml) [22]; [22]; each of these ΗDL3 these subfractions subfractions can be be further into discrete discrete subclasses subclasses by different techniques [12, [12, 43]. fractionated into metabolism is is not not yet yet well well known and and no no specific specific function(s) The HDL metabolism have been assigned to HDL subfractions defined by physical ultracentrifugal flotation criteria. It seems that HDL particles are produced both in the liver and intestine intestine [57] [57] and and continuously continuouslyconvert convertin inthe theplasma plasma[[1,43]. and 1, 43]. Moreover, of triglyceride (TG)-rich surface material derived from the catabolism surface material derived from the catabolism of triglyceride (TG)-rich contribute to to their their synthesis synthesis [26, [26, 50]. 50]. Several studies lipoproteins appears to contribute reverse cholesterol cholesterol transport, a chain of have shown that HDL can promote reverse remove excess excess cholesterol cholesterol from metabolic reactions that remove from peripheral tissues transport cholesterol cholesterol directly or indirectly, with other lipoproteins to the and transport is generally generally accepted process involves different steps. It is liver [32, 41]. This process that interactions interactions between between ΗDL3 HDL3 and HDL HDL binding binding sites sites facilitate facilitate egress egress of cholesterol from cells cells to to the the plasma lipoproteins [6, 45] 45] and particularly to a pre-~ migrating HDL [[16]. 16].This This incorporation incorporation of free free cholesterol cholesterol into small pre-β acceptor lipoproteins, lipoproteins, facilitating facilitating its its esterification esterification by by lecithin:cholesterol lecithin:cholesterol acceptor cholesteryl ester protein (CETP), (CETP), acyltransferase (LCAT) acyltransferase (LCAT)and and its its cholesteryl ester transfer protein transfer to to very-low-density very-loW-density lipoproteins lipoproteins (VLDL) (VLDL) and exchange for mediates transfer triglycerides [28, 36].
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selected immunosorption techniques designed to Recent studies, using selected isolate subpopulations subpopulations of isolate of lipoproteins, lipoproteins, have have shown shown that that HDL HDL cannot be (apo) A-I A-II. regarded as homogeneous particles with apolipoproteins (αρο) Α-I and Α-II. of particles particles within within HDL HDL is is heterogeneous heterogeneous not only only with with The distribution of respect to their hydrated density but also in relation to their apolipoprotein 17, 40]. 40]. It is now recognized recognized that [I, 17, composition [1, that HDL contain at least two apo A-I-containing lipoprotein particles which might have different types of αρο [8, 20] 20] and and clinical clinical significance significance [47]. [47]. One species metabolic function function [8, species contains contains A-I and andA-II A-II (LpA-I:A-II) (LpA-I:A-II) while in the protein components componentsboth bothapo apoA-I as main protein such lipoprotein lipoprotein one αρο apo A-II A-II is absent absent (LpA-I). (LpA-I). The definition of such The definition other one particles by their apolipoprotein content has been proposed to more particles by their apolipoprotein content has been proposed to be more appropriate for for studying studying the chemical complexity and metabolic functions of appropriate system. the plasma lipoprotein system.
LpA-I and LpA-I:A-II are plasma by by LpA-I and LpA-I:A-II are currently currently purified purified from from total total plasma do sequential immunoaffinity immunoaffinity chromatography [17, 38, 44]. Some authors chromatography [17, 38, 44]. Some authors find any any difference difference in lipid composition composition between between LpA-I LpA-I and and LpA-I:A-II LpA-I:A-II not find [[17], 17],but but others others have have claimed claimed that that the percentage of triglyceride triglyceride and and the ratio ester/total cholesterol cholesterol are are lower lower in in L.ρΑ-I LpA-I than in in LpA-I:A-II LpA-I:A-II [44]. cholesteryl ester/total tobe behigher higherin inLpA-I LpA-I than than in inLpA-I:A-II LpA-I:A-II [2]. [2]. lipid/protein ratio ratioappears appearsto The lipid/protein apo A-I to αρο apo A-II in LpA-I:A-II LpA-I:A-II is is 1.5. 1.5. Small Small quantities quantities The molar ratio of αρο of apolipoproteins A-IV; fractions [2, [2, 19]. 19]. of A-Iv; C's, D, E are found in both fractions Of considerable considerable significance significance isis the finding that proteins stimulating the finding proteins stimulating (LCAT, CETP) or other reverse cholesterol transport (LCAT, other apolipoproteins such as apo J are mainly mainly present present in in LpA-I LpA-I and andnot notininLpA-I:A-II LpA-I:A-II [[19, 19, 20, 23]. αρο apo A-I A-I containing particles metaboOur understanding with respect to αρο However, it has been shown recently lism is limited. However, recently that both particles are LpA-I are are produced produced by by the the intestine intestineonly only[ [14]. synthesized by the liver but LpA-I 14]. between the two two subpopulations is is not not well well The metabolic interrelationship between established but seems that LpA-I are faster rate than established but itit seems that LpA-I are catabolized catabolized at at aa faster LpA-I:A-II [48]. is whether whether LpA-I LpA-I and and LpA-I:A-II LpA-I:A-II LpA-I:A-II [48]. One One of the key questions is physiological roles. have different physiological order to gain some insights into the mechanisms mechanisms by by which cholesterol In order movement takes takes place place in peripheral peripheral cells, cells, cultured cultured adipose adipose cells cells were used as movement a model. model. Indeed, the fact that adipose adipose tissue tissue has has the the ability ability to to accumulate, accumulate,
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store store and, and, when when needed, needed, mobilize mobilize aalarge large pool poolofofunesterified unesterified cholesterol cholesterol means that that these these cells cells meet the the requirement requirement for for the the study studyof ofreverse reverse cholescholesmeans terol transport transport [7]. [7]. Cholesterol Cholesterol efflux efflux mediated mediated by by A-I-containing A-I-containing particles particles terol has been been studied studied in in mouse mouse adipose adipose cells cells after aftercholesterol cholesterol preloading preloading with with has LDL [1 [11]. cholesterol efflux efflux LDL 1 ].Long-term Long-termexposure exposure to to LpA-I LpA-I particles particles promotes promotes cholesterol ofLpA-I:A-II. LpA-I:A-II. whereas no no efflux efflux was observed in the the presence presence of whereas These results, behave as as a distinct results, which which emphasize emphasize that LpA-I:A-II LpA-I:A-II behave distinct [21,37,54]. metabolic entity entity have have been been confirmed confirmed in other other studies studies [21, metabolic 37, 54]. ligands which which recognize recognize the cell cell surface surface HDL-binding HDL-binding sites sites have have The ligands apo A-I, A-I, αρο apo A-IV and αρο apo A-II [9, 51]. 51]. It was was proposed identified as aρo been identified that αρο apo A-I A-I and αρο apo A-IV A-IV play play the role role of of agonist agonist and and αρο apo A-II A-II that of of that of cholesterol cholesterol efflux efflux [[10]. [51] have suggested suggested that antagonist of 10]. Slotte Slotte et et al. al. [51] of ΗDL3 HDL3 to human human fibroblasts fibroblasts or bovine bovine endothelial endothelial cells cells induce a addition of protein kinase kinase C (PKC)-dependent translocation translocation of ofcholesterol from intracelprotein We have have recently recently demonstrated that membranes to the the cell cell surface. surface. We lular membranes efflux from adipose cells cells is is coupled coupled to to diacylglycerol diacylglycerol (DAG) (DAG) cholesterol efflux cholesterol from adipose αρο production and and protein protein kinase kinase C C activation activation [54]. [54]. The fact that binding of ofapo production A-I1DMPC complexes complexes produces of αρο apo A-II/ A-III A-I/DMPC produces diacylglycerol, diacylglycerol,but butnot not that that of complexes, strongly role of of αρο apo Α-II A-II as as that of of an an DMPC complexes, strongly supports supports the the role antagonist to produce produce cholesterol cholesterol efflux. efflux. In agreement with this interpretaIn agreement with this tion, has recently recently been been shown shown that that LpA-I:A-II LpA-I:A-II can can inhibit inhibit the the LpAILpAIn, itit has tíο ofcholesterol-preloaded cholesterol-preloadedadipose adiposecells cells[ [11]. promoted cholesterol cholesterolefflux efflux of promoted 11 ]. methods for direct direct quantification quantification of αρο apo A-I-containing A number of methods plasma have have been been developed: developed: immunoprecipitation immunoprecipitation[[18], particles in human plasma 18], two-phase electroimmunoassay electroimmunoassay [5], [5], enzyme-linked enzyme-linked differential-antibody differential-antibody two-phase immunosorbent assay assay [38]. These three methods are well well adapted immunosorbent adapted in research of accuracy. accuracy. The laboratories but are are time-consuming time-consuming and suffer suffer from lack of recent development of of a differential differential electro-immunoassay allows allows the direct measurement of LpA-I. LpA-I. By By using using a large large excess excess of of anti anti A-II, A-II, LpA-I:A-II LpA-I:A-II measurement first peak and LpA-I LpA-I migrates further as a second particles are retained in a first peak [46]. [46]. This This new new system systemcan canprovide provide specific specificand andreproducible reproducible determinadeterminaLpA-I in plasma. tions of LpA-I in human plasma.
Both A-I-containing particles particles were were detected αρο A-I-containing Both lipoprotein lipoprotein forms forms of apo mainly relative proportion proportion of of LpA-I LpA-I was was higher higher in HDL2 mainly in in HDL but the relative than HDL3. ΗDL3.
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Concentrations of LpA-I LpA-I and LpA-I:A-II LpA-I:A-II were plasma Concentrations were determined determined in plasma samples from normolipemic men and women. The average concentrations concentrations of LpA-I were 10% higher LpA-I were approximately approximately 10% higher in in women women than than in men. LpA-I but not LpA-I:A-II LpA-I:A-II are angiographiLpA-I are decreased decreased in normolipemic, angiographically documented CAD patients when compared to a group of asymptomatic asymptomatic subjects and a group group of ofpatients patients with with arteriographically arteriographically normal normal coronary coronary subjects arteries [47]. However, However, in a recent similar study where where the the patient group had higher triglyceride triglyceride levels levels than than controls, controls, we we have have found found that that both LpA-I and LpA-I:A-II are similar degree degree (one (one can can LpA-I:A-II arereduced reducedinin CAD CAD patients patients to to a similar ofLpA-I:A-II LpA-I:A-II particles particles in in hypertriglyceridemic hypertriglyceridemic the decrease decrease of postulate that the of such such particles during patients may be related to the the decreased decreased formation of lipolysis) [30]. A case case control study of apo A-I-containing particles has been performed populations at at contrasting contrastingrisk risk for for CAD CAD (ECTIM (ECTIM Study) Study) [15]. [15]. Male in three populations patients with with a myocardial myocardial infarction infarction and controls controls were were recruited recruited in two two patients Irish Center Center (Belfast). (Belfast). French centers (Strasbourg, Toulouse) and a Northern Irish restandardized mortality mortality rates rates in in Belfast, Belfast, Strasbourg, Strasbourg, Toulouse Toulouse are, reThe standardized spectively, 348, 78/100,000 for population. LpA-I LpA-I and spectively, 348, 102 102 and and 78/100,000 for the tested population. LpA-I:A-II levels but the the level level of ofLpA-I LpA-I:A-II levels are are lower lower in in patients patients than in controls but LpA-I differs significantly, on a statistical basis, between three populations: LpA-I differs significantly, on a basis, LpA-I was much lower in Belfast Belfast than than in the the French centers in controls and in cases. multivariateanalysis analysis suggests suggests that the the LpA-I/HDL LpA-I1HDL cholescholesIn this study, the multivariate is very very significant. significant. terol ratio ratio is of LpA-I LpA-I (but not not Recently, itit has also been been observed observed that level of Recently, has also that the level LpA-I:A-II) in LpA-I:A-II) in children children whose whose parents parents suffer sufferaa premature premature form form of CAD is control group with no no familial familial history history of of CAD CAD [3]. [3]. lower than in a control
Effect of of Alcohol Intake Particles Intake on HDL Particles Among dietary factors influence on plasma HDL Among factors studied for a possible influence levels, alcohol most intriguing intriguing because because it has has been been levels, alcohol intake intake isis one one of the most 27]. This be positively positively associated with HDL concentrations [4, 27]. reported to be some investigators investigators to suggest suggest that moderate moderate alcohol alcohol consumption consumption has led some may be be protective protective against against coronary coronary artery artery disease. disease. Epidemiological Epidemiological and may have been clinical data have provided some support for this idea, but there have many inconsistencies inconsistencies among demonstrate the many among studies studies that that have have sought sought to to demonstrate this context context [[10, beneficial effect effect of of alcohol alcohol consumption consumption in this 10, 27, 42].
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1. Characteristics of study groups Table 1. groups (mean (mean values values ± SD)
Group 1I (n = 68)
2 (n = 65)
3 (n = 69)
4 (n = 69)
55(n=63) (n = 63)
Age, years Age,
10.1 47.6 ± 10.1
47.6± 10.2 47.6 ± 10.2
47.7 ± 9.7
47.9 ± 9.7
47.9 ± 9.8
Alcohol consumption, Alcohol g/week
130 ± 40
59* 289 ± 59*
492 ± 60*
53* 676 ± 53*
189* 993 ± 189*
kg Body weight, kg
10.9 74.2 ± 10.9
77.2 ± 11.5 11.5
78.3 ± 12.3 12.3 78.3
1.8 1.8
13.6 75.6 ± 13.6
173 ± 6.1 6.1 173
172 ± 5.6
Height, cm
171.4 ± 6.3
172.4 ± 6.3
Erythrocyte corpuscular Jlm3 11 volume, µm3
92.9 ± 3.5
93.5 ± 3.9
94.7 ± 4.9**
14.9* 96.7 ± 14.9*
11.7** 96.3 ± 11.7**
y-Glutamyl transpeptidase, IU IV
24.1 ± 20.1 20.1 24.1
29.2 ± 22.9
48.5 ± 55.7 48.5
67.8 ± 77.6* 77.6*
114* 79.5 ± 114*
173.4±6 173.4 ±6
Although there is no no doubt doubt about the increase of plasma HDL concentrations on alcohol intake, the subfraction of of HDL HDL responsible responsible for this biologibiological observation has not yet been formally identified. Indeed, studies in which HDL has been been fractionated fractionated based on hydrated hydrated density density have have been been inconinconHDL clusive. Contradictory reputedly `anti'anticlusive. Contradictory data have been published about the reputedly atherogenic' HDL, i.e. the less dense HDL2 subfraction [13, 24, 25, 33-35, atherogenic' HDL, i.e. less dense HDL2 subfraction [13, 24, 25, 56]. 39, 49, 53, 55, 56]. Because of because of the potential Because of the the heterogeneity heterogeneity within within HDL HDL and because clinical and and metabolic metabolic importance importance of of newly newly defined defined apo apo AI-containing AI-containing clinical particles, we we decided to investigate investigate whether plasma concentralipoprotein particles, of LpA-I and LpA-I:A-II LpA-I:A-II were tions of were influenced influenced by by alcohol alcohol consumption. consumption. 25 to to 63 63 years) years) who who 334 men men (aged (aged from 25 Our study was carried out in 334 were undergoing were undergoing aa routine routine health health examination examination in in the the Health Center of the Lille, France. They completed a form, covering the past 6 Pasteur Institute, Lille, eating habits habits and cigarette cigarette and months, that allowed allowed the assessment assessment of eating months, alcohol consumption. with a alcohol consumption. Alcohol Alcohol intake intake was was recorded recorded by by an interview with specially trained physician. specially ofalcohol alcohol per per week), week), the men From the alcohol consumption consumptiondata data(grams (gramsof five groups were divided into five were groups with with either 63 or 69 subjects in each. of the subjects in each The main clinical and biochemical characteristics of 1. group are summarized in table 1.
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5: *p < 0.001, **p < 0.1. Significance of differences Significance differences among among group group 11 and and groups groups 2 through 5:
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study group (mean values SD) values ± SD) Table 2. Lipoprotein and apoprotein concentrations in study Group 2
3
4
3
5
LpA-I, mg/dl
51.9±26.6 51.9 ± 26.6
43.7±23.1 43.7 ± 23.1
43.9 ± 25.3
35.8 ± 18.3
19.2 37.2 ± 19.2
LpA-I:A-II, mg/dl LpA-Ι:Α-ΙΙ,
15.8 80.9 ± 15.8
85.0± 15.9 85.0 ± 15.9
86.8 ± Ι16.3 86.8 6.3
19.5 96.2 ± 19.5
10.3 ± 22.0
HDL cholesterol, mmol/I (mg/dl) mmol/1
1.41 1.41 ± 0.33 (54 ± 12)
1.46 ± 0.39 15) (56 ± 15)
1.54 ± 0.42 (59±16) (59 ± 16)
1.61 ± 0.52** 1.61 (62 ± 20)
1.94 ± 0.82* (75±31) (75 ± 31)
136.8 ± 32.4
128.8 ± 27.8
130.8±29.8 130.8 ± 29.8
10.2 39.9 ± 10.2
10.9 40.2 ± 10.9
10.1 42.2 ± 10.1
A-I, Apolipoprotein Α-Ι, mg/dI A-II, Apoliprotein Α-ΙΙ, mg/dI
26.1 132.0 ± 26.1 11.1 ** 44.3 ± 11.1**
140.7 ± 29.4
14.1* 48.5 ± 14.1*
The mean mean ±±SD SDage age of ofthe the subjects subjects in in each each group group was was 48 48 ± ± 10 years (table 1). All All groups groups were were comparable comparable for height, height, weight, weight, systolic systolic and diastolic diastolic 1). blood pressure, pressure, levels plasma glucose, glucose, plasma plasma blood levels of of plasma plasma uric uric acid acid and plasma of the erythrocyte corpuscular volume and aminotransferases, only in terms of y-glutamyl transpeptidase 1). of y-glutamyl transpeptidase activity (table 1). physical activity, activity, cigarette cigarette consumption, The mean physical consumption, and and eating habits were the same in the groups groups based were the same based on a questionnaire administered by the physician in charge of the subjects. Only HDL cholesterol cholesterol and api apo A-II A-II levels levels seemed seemed to to be besignificantly significantly Only :::::;200 200 g/week) glweek) was compared different when when group group 1 (alcohol (alcohol consumption, consumption, < 800 with either either group group 44 (alcohol (alcohol consumption consumption ranged ranged between between 601 with 601 and 800 glweek) or (alcohol consumption consumption > 800 800 g/week) glweek) (table mean g/week) or 5 (alcohol (table 2). 2). The mean increases of HDL, cholesterol and αρο apo A-II levels levels from increases from groups groups 1 through 5 were 37.6 and and 21.6%, 21.6%, respectively, respectively, while while αρο apo A-I A-I levels levels were were unaltered. unaltered. were correlation study study confirmed confirmed this this strong strong relationship relationship between between Results of a correlation Results 3). alcohol consumption cholesterol and αρο apo A-II A-II levels levels (table 3). alcohol consumption and and HDL cholesterol However, no correlation was found with apo A-I levels or with any of the However, no was αρο A-I levels or with any other parameters measured in this study. LpA-I:A-II levels The LpA-I:A-II levels were were increased increased from from groups 1 through 5, while the LpA-I levels LpA-I levels decreased decreased significantly significantly by by 28.3% 28.3% (fig. (fig. I). 1).The The correlation correlation study
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I: *p < 0.001, 0.00 I, **p < Significance of Significance of differences differencesamong among groups groups 22 through through 5 and group 1: < 0.1.
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analysis of ofy-glutamyl y-glutamyl transpeptidase transpeptidase activity, lipoproTable 3. Spearman's correlational analysis of alcohol intake with reported reported values values of teins, and apoproteins with Alcohol intake Alcohol
pp
y-Glutamyl transpeptidase
+0.30
