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A polymorphism of the gene coding for cholesterol ester transfer protein (CETP) that affects transfer of plasma cholesterol ester and its sensitivity to regulation
Biomed
& Pharmac~orher
1997;s 1:404-40s 8) Elswier, Paris
Correspondance
A polymorphism of the gene coding for cholesterol ester transfer protein (CETP) that affects transfer of plasma cholesterol ester and its sensitivity to regulation A Girard-Globa Laboratoire l’ilnriquniile, L?on, France
de MPtabolismr
et CNRS
drs UPRESA
Lipides, 5014.
Hcipiral UnlvrrsitP
de Leon
1,
Cholesterol is continuously being synthesized by all cells in the organism, yet cannot be degraded. Its only mode of clearance is through conjugation with bile acids followed by bile secretion. This unique mechanism ensures optimal utilization of a molecule which is, at once, indispensable and energetically costly. To transfer from tissues to the liver it must associate with lipoproteins which are the main carriers of hydrophobic molecules. Uptake of excess cholesterol from cells is operated by high density lipoproteins (HDL) which accomodate unesterified cholesterol (UC) on their phospholipid (PL)-rich surface. This UC is esterified and cholesterol esters (CE) move to the hydrophobic core of the particle. HDL can deliver their CE to the liver with limited efficiency. The main pathway for CE delivery to the liver is a transfer from HDL to low density lipoproteins (LDL) followed by endocytosis of LDL via a specific receptor. When this receptor functions optimally, the pathway is highly efficient. When it fails to do so, however, the LDL become oxidized, are taken up by a scavenger receptor located on arterial wall cells and macrophages and become a major factor of cholesterol deposition and atherosclerosis. It is easily understood, therefore, that the higher the ratio of LDL-C to HDL-C, the greater the atherogenic risk. The transfer from HDL to LDL is mediated by a
choSlesterol ester protein (CETP) which operates a bi-directional exchange between a CE molecule in HDL and a TG molecule in LDL. Thus, when LDL uptake is efficient CETP promotes rapid CE clearance, but when it is not, CETP delivers CE to an enlarged pool of potentially atherogenic LDL (for a review see [I]). CETP is a 72 kD highly hydrophobic protein which is coded for by a 2.1 kb messag’e in 16 exons distributed over an unusually large space of 25 kb [2, 31. A frequent biallelic Taql polymorphism (CE:TP/TaqlB) of the gene has been evidenced. the absence of cutting site (B2 allele in 40% of sukjects) being associated with higher concentrations of HDL-C [4]. A number of modulating factors affect CETP activity, among which obesity (increases CETP activity) and alcohol intake (lowers CETP activity) potentially account for opposite effects on HDL-C as well as on risk [S]. CETP activity is low in alcoholics and increases after alcohol withdrawal [6] while moderate alcohol intake induces a reduction of variable importance depending on individuals [7]. This led us to hypothesize that alcohol had a ‘different impact dependent upon the TaqIB-related genetic polymorphism. A recent population study [8] focusing on the con’trol population of the ECTIM (&de custe’vwin de I’infurctus du mywar-de/a case-witness study of myocardial infarctus) project has indeed shown that B2 carriers had lower CETP and higher HDL, but that the effect could only be evidenced in subjects drinking over 25 g of alcohol daily. Accordingly the odds-ratio for myocardial infarction of B2 homozygotes compared to Bl decreased from 1 in non-drinkers to 0.75 in drinkers of 75 g per day or more. The absence of a similar interaction for CETP, despite parallel variation, suggested the presence of two independent functional variables linked t’o the B2 allele. A search for variants on the coding sequence of the CETP gene, however, failed to identify them. We have tested the gene-environment interaction in an intervention study [submitted for publication] focusing on confirmed alcoholics (mean intalke 170 f 88 g per day) undergoing weaning
CETP
by deprivation. A distinct genotypic effect was evidenced under alcohol, with lower CETP activity (P = 0.0002) and higher HDL-C (P = 0.014) in B2 homozygotes, but no effect of allelic makeup on apoA1 which is the major protein of HDL. After withdrawal, CETP increased and HDL-C decreased significantly in allele B2 bearers, only so that the genotypic effect lost significance, while apoAI decreased to the same extent in all. The results are best explained by a co-dominant effect of allele B2 on sensitivity to alcohol ,while the independent variation of apoA1 accounts for the often reported low degree of correlation between CETP and HDL-C. The independent effect of alcohol on apoA1 can have several origins. One of them is the stimulation by alcohol of lipoprotein lipase which, by reducing the triglyceride content of HDL, might diminish their catabolism [9]. An independent stimulation of apoA1 synthesis by alcohol would also account for the low correlation and its improvement by weaning. Its existence has been demonstrated in the rat [IO] and, more recently, in human subjects [Ill. Our interpretation is that alcohol has a dual effect on HDL: 1) an enlargement of the apoA1 pool in all subjects, causing a permissive effect and 2) an enlargement of the HDL-C pool related to a reduction of CETP activity, most effective in the presence of the “sensitive” B2 allele while the persistently high levels of CETP associated with the Bl allele preclude modulation by alcohol, in spite of elevated concentrations of apoA1. More recently, we have demonstrated an impact of this same TaqlB polymorphism on the welldocumented differences in HDL-C concentrations between males and females 151. In a population of 400 non-insulin dependent diabetics we have been able to demonstrate that the presence of the B2 allele in men increases HDL-C concentrations sufficiently so they are not different from those of females. On the contrary, female hormones abolish genotypic differences. As a result the difference between the HDL-C of males and females is significant only in bearers of the Bl allele.
gene coding
405
This interesting system provides a model for environmental and hormonal modulation of gene expression. It shows that nutritional conclusions reached through epidemiological studies, such as the beneficial effects of alcohol on cardiovascular risk, or the sex differences in risk may not apply equally to all individuals, depending on sometimes subtle differences in genetic make-up. Acknowledgements We would like to thank those who participated in our original contribution: 1 Toury, F Lam&e. V Durlach, A Durlach. A Zahouani. References Tall AR. IPlasma lipid transfer protein:, J Lipid Res 1986:27:361-7 Drayna D, Jarnagin AS. McLean J et al. Cloning and sequencing of the human CETP IDNA. Nuturr 1987;327:632-4 Agellon LB, Quinet EM. Gillette GG et al. Organiration elf the human CETP gene. L(iochem I990:29: 1372-h Kondo I, Berg D, Drayna D et al. DNA Polymorphsm of the gene coding for CETP is associated with high density lipoprotein cholesterol and apolipoprotein level. C(in Genetics 19X%35:49-56 L.anger RD. Criqui MH. Reed DM. Lipoproteins and blood pressure as biological pathways for effect of moderate alcohol consumption on coronary heart disease. Cirwlotion 1992;89:910-I5 Hannuksela M, Marcel Y, Kesaniemi et al. Reduction in the concentration and activitv of CETP bv alcohol. J Lipid Rrs
1992:33:737-44
.
Hagiage M, Marti C. Girard-Globa A et al. Effect of a moderate alcohol intake on the lipoproteins of normo trif,lyceridemvz obese subjects compared with normoponderal controls. Wr,tab C/in Exp 1992;41:856-6 I F‘umeron F. Betoulle D, Cambien F et al. Alcohol intake modulates the effect of the CETP gene on plasma HDL and the risk of myocardiul infarction. J Clin I’AI.PS~ 1995:96: 1664-7 I Goldberg IJ, Blaner WS, Vanni TM et al. Role of lipoprotein lipaae in rhe regulation of HDL metabolism. J Chin /rlur.sr 1990:86:463-73 IO Okamoto Y, Fujimori Y. Naknno H et al. .I Lrzh Clin Med 1988; I I 1:482-5 I I De Oliveira c Silva ER, Foster D. MC gee Harper M, Brinton EA. Alcohol raises HDL levels by increasing production of apoAl and apoAlI. Circulation 1996;94(suppl I ‘I: 265
& Pharmac~orher
1997;s 1:404-40s 8) Elswier, Paris
Correspondance
A polymorphism of the gene coding for cholesterol ester transfer protein (CETP) that affects transfer of plasma cholesterol ester and its sensitivity to regulation A Girard-Globa Laboratoire l’ilnriquniile, L?on, France
de MPtabolismr
et CNRS
drs UPRESA
Lipides, 5014.
Hcipiral UnlvrrsitP
de Leon
1,
Cholesterol is continuously being synthesized by all cells in the organism, yet cannot be degraded. Its only mode of clearance is through conjugation with bile acids followed by bile secretion. This unique mechanism ensures optimal utilization of a molecule which is, at once, indispensable and energetically costly. To transfer from tissues to the liver it must associate with lipoproteins which are the main carriers of hydrophobic molecules. Uptake of excess cholesterol from cells is operated by high density lipoproteins (HDL) which accomodate unesterified cholesterol (UC) on their phospholipid (PL)-rich surface. This UC is esterified and cholesterol esters (CE) move to the hydrophobic core of the particle. HDL can deliver their CE to the liver with limited efficiency. The main pathway for CE delivery to the liver is a transfer from HDL to low density lipoproteins (LDL) followed by endocytosis of LDL via a specific receptor. When this receptor functions optimally, the pathway is highly efficient. When it fails to do so, however, the LDL become oxidized, are taken up by a scavenger receptor located on arterial wall cells and macrophages and become a major factor of cholesterol deposition and atherosclerosis. It is easily understood, therefore, that the higher the ratio of LDL-C to HDL-C, the greater the atherogenic risk. The transfer from HDL to LDL is mediated by a
choSlesterol ester protein (CETP) which operates a bi-directional exchange between a CE molecule in HDL and a TG molecule in LDL. Thus, when LDL uptake is efficient CETP promotes rapid CE clearance, but when it is not, CETP delivers CE to an enlarged pool of potentially atherogenic LDL (for a review see [I]). CETP is a 72 kD highly hydrophobic protein which is coded for by a 2.1 kb messag’e in 16 exons distributed over an unusually large space of 25 kb [2, 31. A frequent biallelic Taql polymorphism (CE:TP/TaqlB) of the gene has been evidenced. the absence of cutting site (B2 allele in 40% of sukjects) being associated with higher concentrations of HDL-C [4]. A number of modulating factors affect CETP activity, among which obesity (increases CETP activity) and alcohol intake (lowers CETP activity) potentially account for opposite effects on HDL-C as well as on risk [S]. CETP activity is low in alcoholics and increases after alcohol withdrawal [6] while moderate alcohol intake induces a reduction of variable importance depending on individuals [7]. This led us to hypothesize that alcohol had a ‘different impact dependent upon the TaqIB-related genetic polymorphism. A recent population study [8] focusing on the con’trol population of the ECTIM (&de custe’vwin de I’infurctus du mywar-de/a case-witness study of myocardial infarctus) project has indeed shown that B2 carriers had lower CETP and higher HDL, but that the effect could only be evidenced in subjects drinking over 25 g of alcohol daily. Accordingly the odds-ratio for myocardial infarction of B2 homozygotes compared to Bl decreased from 1 in non-drinkers to 0.75 in drinkers of 75 g per day or more. The absence of a similar interaction for CETP, despite parallel variation, suggested the presence of two independent functional variables linked t’o the B2 allele. A search for variants on the coding sequence of the CETP gene, however, failed to identify them. We have tested the gene-environment interaction in an intervention study [submitted for publication] focusing on confirmed alcoholics (mean intalke 170 f 88 g per day) undergoing weaning
CETP
by deprivation. A distinct genotypic effect was evidenced under alcohol, with lower CETP activity (P = 0.0002) and higher HDL-C (P = 0.014) in B2 homozygotes, but no effect of allelic makeup on apoA1 which is the major protein of HDL. After withdrawal, CETP increased and HDL-C decreased significantly in allele B2 bearers, only so that the genotypic effect lost significance, while apoAI decreased to the same extent in all. The results are best explained by a co-dominant effect of allele B2 on sensitivity to alcohol ,while the independent variation of apoA1 accounts for the often reported low degree of correlation between CETP and HDL-C. The independent effect of alcohol on apoA1 can have several origins. One of them is the stimulation by alcohol of lipoprotein lipase which, by reducing the triglyceride content of HDL, might diminish their catabolism [9]. An independent stimulation of apoA1 synthesis by alcohol would also account for the low correlation and its improvement by weaning. Its existence has been demonstrated in the rat [IO] and, more recently, in human subjects [Ill. Our interpretation is that alcohol has a dual effect on HDL: 1) an enlargement of the apoA1 pool in all subjects, causing a permissive effect and 2) an enlargement of the HDL-C pool related to a reduction of CETP activity, most effective in the presence of the “sensitive” B2 allele while the persistently high levels of CETP associated with the Bl allele preclude modulation by alcohol, in spite of elevated concentrations of apoA1. More recently, we have demonstrated an impact of this same TaqlB polymorphism on the welldocumented differences in HDL-C concentrations between males and females 151. In a population of 400 non-insulin dependent diabetics we have been able to demonstrate that the presence of the B2 allele in men increases HDL-C concentrations sufficiently so they are not different from those of females. On the contrary, female hormones abolish genotypic differences. As a result the difference between the HDL-C of males and females is significant only in bearers of the Bl allele.
gene coding
405
This interesting system provides a model for environmental and hormonal modulation of gene expression. It shows that nutritional conclusions reached through epidemiological studies, such as the beneficial effects of alcohol on cardiovascular risk, or the sex differences in risk may not apply equally to all individuals, depending on sometimes subtle differences in genetic make-up. Acknowledgements We would like to thank those who participated in our original contribution: 1 Toury, F Lam&e. V Durlach, A Durlach. A Zahouani. References Tall AR. IPlasma lipid transfer protein:, J Lipid Res 1986:27:361-7 Drayna D, Jarnagin AS. McLean J et al. Cloning and sequencing of the human CETP IDNA. Nuturr 1987;327:632-4 Agellon LB, Quinet EM. Gillette GG et al. Organiration elf the human CETP gene. L(iochem I990:29: 1372-h Kondo I, Berg D, Drayna D et al. DNA Polymorphsm of the gene coding for CETP is associated with high density lipoprotein cholesterol and apolipoprotein level. C(in Genetics 19X%35:49-56 L.anger RD. Criqui MH. Reed DM. Lipoproteins and blood pressure as biological pathways for effect of moderate alcohol consumption on coronary heart disease. Cirwlotion 1992;89:910-I5 Hannuksela M, Marcel Y, Kesaniemi et al. Reduction in the concentration and activitv of CETP bv alcohol. J Lipid Rrs
1992:33:737-44
.
Hagiage M, Marti C. Girard-Globa A et al. Effect of a moderate alcohol intake on the lipoproteins of normo trif,lyceridemvz obese subjects compared with normoponderal controls. Wr,tab C/in Exp 1992;41:856-6 I F‘umeron F. Betoulle D, Cambien F et al. Alcohol intake modulates the effect of the CETP gene on plasma HDL and the risk of myocardiul infarction. J Clin I’AI.PS~ 1995:96: 1664-7 I Goldberg IJ, Blaner WS, Vanni TM et al. Role of lipoprotein lipaae in rhe regulation of HDL metabolism. J Chin /rlur.sr 1990:86:463-73 IO Okamoto Y, Fujimori Y. Naknno H et al. .I Lrzh Clin Med 1988; I I 1:482-5 I I De Oliveira c Silva ER, Foster D. MC gee Harper M, Brinton EA. Alcohol raises HDL levels by increasing production of apoAl and apoAlI. Circulation 1996;94(suppl I ‘I: 265