Eva Boes

Stefan Coassin

Barbara Kollerits

Iris M Heid

MD Florian Kronenberg
email: florian.kronenberg@i-med.ac.at

Genetic-epidemiological evidence on genes associated with HDL cholesterol levels: A systematic in-depth review

Keywords: HDL cholesterol, genetic association, candidate genes, systematic review, genome-wide association study

High-density lipoprotein (HDL) particles exhibit multiple antiatherogenic effects. They are key players in the reverse cholesterol transport which shuttles cholesterol from peripheral cells (e.g. macrophages) to the liver or other tissues. This complex process is thought to represent the basis for the antiatherogenic properties of HDL particles. The amount of cholesterol transported in HDL particles is measured as HDL cholesterol (HDLC) and is inversely correlated with the risk for coronary artery disease: an increase of 1 mg/dL of HDLC levels is associated with a 2% and 3% decrease of the risk for coronary artery disease in men and women, respectively. Genetically determined conditions with high HDLC levels (e.g. familial hyperalphalipoproteinemia) often coexist with longevity, and higher HDLC levels were found among healthy elderly individuals.

HDLC levels are under considerable genetic control with heritability estimates of up to 80%. The identification and characterization of genetic variants associated with HDLC concentrations can provide new insights into the background of longevity. This review provides an extended overview on the current genetic-epidemiological evidence from association studies on genes involved in HDLC metabolism. It provides a path through the jungle of association studies which are sometimes confusing due to the varying and sometimes erroneous names of genetic variants, positions and directions of associations.

Furthermore, it reviews the recent findings from genome-wide association studies which have identified new genes influencing HDLC levels.

The yet identified genes together explain only a small amount of less than 10% of the HDLC variance, which leaves an enormous room for further yet to be identified genetic variants. This might be accomplished by large population-based genome-wide metaanalyses and by deep-sequencing approaches on the identified genes. The resulting findings will probably result in a re-drawing and extension of the involved metabolic pathways of HDLC metabolism. 'A' allele decreases risk for CVD: OR=0.95 (95%CI 1.93-2.63) An increase of HDLC of 2.28 mg/dL decreases the for OR=0.92 (95%CI 0.91-0.93) A B

Introduction 1.HDL metabolism

High-density lipoproteins (HDL) are a heterogeneous group of particles composed of a core of cholesteryl ester and triglycerides surrounded by an amphipathic layer of free cholesterol, phospholipids and apolipoproteins (apo) (Klos et al., 2007).

HDL particles exhibit multiple antiatherogenic effects (Kontush et al., 2006). They shuttle cholesterol from peripheral cells (e.g. macrophages) to the liver or other tissues in need of large amounts of cholesterol [START_REF] Bruce | Plasma lipid transfer proteins, high-density lipoproteins, and reverse cholesterol transport[END_REF], an important step that relieves the peripheral cells from cholesterol burden (Figure 1). The concept for this complex process was first porposed by Glomset et al. 40 years ago [START_REF] Glomset | The plasma lecithins:cholesterol acyltransferase reaction[END_REF]) and is called reverse cholesterol transport. It is thought to represent the basis for the antiatherogenic properties of HDL (Kontush et al., 2006;[START_REF] Von Eckardstein | High Density Lipoproteins and Arteriosclerosis: Role of Cholesterol Efflux and Reverse Cholesterol Transport[END_REF]. Furthermore, HDL has antioxidative properties due to associated antioxidative enzymes and expresses antiinflammatory activity in various pathways. Figure 1 illustrates the major routes of lipoprotein metabolism to allow a better understanding of the HDL metabolism [START_REF] Kataoka | Apolipoprotein E polymorphism in American Indians and its relation to plasma lipoproteins and 98[END_REF]. HDL precursor particles are secreted as disc-shaped structures by the liver and intestine and can absorb free cholesterol from cell membranes, a process mediated by ABCA1, apoA-I and apoA-IV. While ABCA1 presents the best understood active cholesterol efflux mechanism, the HDL-mediated removal of free cholesterol from the peripheral cells is also aided by other active transporters such as ABCG1 as well as by passive diffusion which can be enhanced by plasma membrane receptors such as SCARB1 [START_REF] Cavelier | Lipid efflux by the ATP-binding cassette transporters ABCA1 and ABCG1[END_REF]. ApoA-I is the major apolipoprotein of HDL and activates the enzyme lecithin:cholesteryl acyltransferase (LCAT), which esterifies the accepted free cholesterol to allow more efficient packaging of the cholesterol for transport. By acquisition of additional apolipoproteins, cholesteryl esters, and triglycerides, HDL 3 particles are transformed into spherical HDL 2 particles [START_REF] Dieplinger | The in vitro formation of HDL2 during the action of LCAT: the role of triglyceride-rich lipoproteins[END_REF]. Reverse cholesterol transport can take three different routes. First, large HDL particles with multiple copies of apoE can be taken up by the liver via the LDL receptor [START_REF] Bruce | Plasma lipid transfer proteins, high-density lipoproteins, and reverse cholesterol transport[END_REF]. Second, the accumulated cholesteryl esters from HDL can be selectively taken up by the liver mediated by SR-B1 [START_REF] Acton | Identification of scavenger receptor SR-BI as a high density lipoprotein receptor[END_REF]. This receptor is expressed primarily in liver and nonplacental steroidogenic tissues. Third, cholesteryl esters are transferred by the cholesteryl ester transfer protein (CETP) from HDL to triglyceride-rich lipoproteins [START_REF] Bruce | Plasma lipid transfer proteins, high-density lipoproteins, and reverse cholesterol transport[END_REF]. Serum HDL cholesterol levels are influenced by the complexity of these reverse cholesterol transport processes. Disturbances in the concentrations of apoproteins, function of enzymes, transport proteins, receptors, other lipoproteins, and their clearance from plasma can have a major impact on the antiatherogenic properties in HDL.

HDLC and epidemiological evidence for antiatherogenic properties

High-density lipoprotein (HDL) particles exhibit multiple antiatherogenic effects (Kontush et al., 2006) and HDL cholesterol (HDLC) concentrations show a strong inverse correlation with the risk of coronary artery disease (CAD) [START_REF] Lewington | Blood cholesterol and vascular mortality by age, sex, and blood pressure: a meta-analysis of individual data from 61 prospective studies with 55,000 vascular deaths[END_REF]. Epidemiological studies highlighted the antiatherogenic function of HDLC and showed that an increase of 1 mg/dL of HDLC levels is associated with a 2% and 3% decrease of the risk for CAD in men and women, respectively [START_REF] Wilson | High-density lipoprotein, low-density lipoprotein and coronary artery disease[END_REF][START_REF] Linsel-Nitschke | HDL as a target in the treatment of atherosclerotic cardiovascular disease[END_REF]. A recent meta-analysis [START_REF] Lewington | Blood cholesterol and vascular mortality by age, sex, and blood pressure: a meta-analysis of individual data from 61 prospective studies with 55,000 vascular deaths[END_REF] including prospective observational studies with HDLC measurements available in 150,000 individuals demonstrated a strong negative association with ischemic heart disease mortality in every age group, with no evidence of a threshold beyond which higher HDL cholesterol was no longer associated with lower mortality. On average, 13 mg/dL higher HDL cholesterol was associated with about a third lower ischemic heart disease mortality. Within every age group, the strength of this association was comparable for men and women. Moreover, clinical trials have established that increasing HDLC levels by drugs could reduce CAD risk [START_REF] Gotto | Low high-density lipoprotein cholesterol as a risk factor in coronary heart disease: a working group report[END_REF][START_REF] Manninen | Lipid alterations and decline in the incidence of coronary heart disease in the Helsinki Heart Study[END_REF][START_REF] Rubins | Gemfibrozil for the secondary prevention of coronary heart disease in men with low levels of high-density lipoprotein cholesterol. Veterans Affairs High-Density Lipoprotein Cholesterol Intervention Trial Study Group[END_REF]Kronenberg 2004), thus bringing longer life expectancy. This hypothesis is compatible with observations that familial hyperalphalipoproteinemia often coexists with longevity [START_REF] Patsch | Lipoproteins in familial hyperalphalipoproteinemia[END_REF], and that higher HDLC levels are found among healthy elderly aged 85-89 years as compared to those in middle-aged subjects [START_REF] Nikkila | High-density lipoprotein cholesterol and longevity[END_REF].

Accordingly, HDL and molecules involved in HDL metabolism seem to be attractive candidates for longevity-promoting factors [START_REF] Arai | Aging and HDL metabolism in elderly people more than 100 years old[END_REF].

Since HDLC levels are under considerable genetic control with heritability estimates of up to 80% (Kronenberg et al., 2002;[START_REF] Perusse | Familial resemblance of plasma lipids, lipoproteins and postheparin lipoprotein and hepatic lipases in the HERITAGE Family Study[END_REF][START_REF] Wang | Genetics of variation in HDL cholesterol in humans and mice[END_REF][START_REF] Goode | Heritability of longitudinal measures of body mass index and lipid and lipoprotein levels in aging twins[END_REF], the identification and characterization of genetic variants associated with HDLC concentrations can provide useful information related to genotype-phenotype relationships and give new insights in the background of longevity. This article reviews the current genetic-epidemiological evidence from association studies on genes involved in HDL metabolism. Genetic association studies establish estimates for a difference of mean HDLC levels among subjects with a certain genetic variant compared to the others and thus a measure of association between the genetic background and HDLC outcome in human beings. This overview should provide a path through the jungle of association studies often reporting different names and positions for the same genetic variant under study. Furthermore, the associations are sometimes reported in the direction of the "risk allele", that is the genetic variant associated with increased CAD risk and thus lower HDLC levels, or in the direction of the "minor allele", that is the less frequently appearing variant in the population, which may even differ between study populations. Furthermore, it reviews the recent findings from genome-wide association studies, which have identified new genes influencing HDLC levels and provides a first taste on the future. It is not intended to provide an overview on functional studies.

Methods on literature search

The literature search for genes previously reported for association with HDLC was performed in PubMed using search terms such as "(meta-analysis OR associat* OR epidemiolog*) AND (polymorphism OR genetic OR mutation) AND HDL AND human" with an Entrez Date in PubMed until April 2008 (EDAT: the date the citation was added to PubMed).

Results were complemented by knowledge of the lipid-experienced investigators of this study.

Eligible studies for data extraction were meta-analyses, population-based studies, studies in the general population, healthy control populations, hospital-based controls, control populations selected for not having type 2 diabetes mellitus (T2DM), and case-control studies for cardiovascular disease (CVD) or T2DM. Studies which were included in a metaanalysis were only indirectly considered by showing the results of this meta-analysis in the tables instead of each single study. We did not consider studies with special patient populations other than cardiovascular disease or T2DM (e.g. patients with chronic kidney disease).

Exclusion criteria for our search were studies in a non-English language where only the abstract was available in English and small studies with less than 500 individuals.

Studies which investigated a large number of genetic variants were only considered to provide statistically significant association, if the associations were strong enough to hold for a significance level corrected for the multiple comparisons. This was usually not sufficiently accounted for in the reported associations. Moreover, we omitted studies on polymorphisms with a too low minor allele frequency (MAF) yielding less than about 30 study participants with the minor allele considering the number of individuals investigated.

If the results of a particular polymorphism of a certain study population were considered in more than one publication, only one of these publications was referenced in our tables.

A gene was considered an HDLC candidate gene if at least two eligible studies reported a statistically significant association of genetic variation within or related to the particular gene with HDLC levels. An exception from this rule was made for LCAT for which strong functional evidence was available combined with one large and a few small association studies.

Rarely investigated genetic variations that showed a significant association with HDLC levels in at least one large study (n>1000) are listed in an additional table (Table 11). and the Supplementary Table show the results of all eligible candidate gene association studies which were accessible in PubMed until the Entrez Date April 2008. Even though we draw our attention to the fact not to miss the most important studies, we are aware that it might not have been possible to have complete lists of all studies which deal with candidate genes for HDL cholesterol.

It should be noted that identifying the precise rs numbers of some polymorphisms was very tedious if not impossible as rs numbers were often lacking or mistakenly reported and in some of these cases we were only able to derive rs numbers by combining literature reports with databases such as HapMap or NCBI. In some cases the effect size of the investigated SNP on HDL cholesterol was only displayed in a figure and allowed only an approximate estimate of the numerical value.

For the single nucleotide polymorphisms (SNPs) of the candidate genes reported in the Tables 1-9 we imported data from HapMap and constructed an r 2 -plot using HaploView 4.1 to illustrate the correlation of the SNPs reported for each gene. For APOA1, APOC3 and PON1 no r 2 -plot was constructed since only one or none of the tabulated SNPs was available in HapMap. Furthermore, rs2303790 of CETP and rs2066718 of ABCA1 are not displayed in the r 2 -plots since the MAF of these SNPs was 0.0% for the imported HapMap data.

A further chapter reviews the evidence recently derived from genome-wide association (GWA) studies. SNPs derived from these studies are not listed in but are provided in an extra table (Table 12).

Results from candidate gene association studies

Cholesteryl ester transfer protein (CETP)

The CETP gene is located on chromosome 16 and consists of 16 exons and 15 introns and is a member of the lipopolysaccharide binding protein gene family (Yamashita et al., 2001). The gene locus is highly polymorphic with several common polymorphisms as well as rare mutations [START_REF] Thompson | High-density genotyping and functional SNP localization in the CETP gene[END_REF].

CETP is a key plasma protein that influences circulating levels of HDLC by mediating the transfer of esterified cholesterol from HDL to apoB-containing particles in exchanges for triglycerides [START_REF] Tall | Plasma cholesteryl ester transfer protein[END_REF]) (Figure 1). The effect of this transfer is an increase in atherogenic apoB-containing particles and a decrease of HDL [START_REF] Carlquist | Inconsistencies in the genetic prediction of HDL cholesterol versus atherosclerosis[END_REF]. The lipid profile typically seen in subjects with CETP deficiency includes elevated HDL cholesterol levels with LDL cholesterol levels generally in the normal range [START_REF] Miller | Genetics of HDL regulation in humans[END_REF]. Complete loss of CETP activity due to mutations in the CETP gene can result in up to five times the normal HDLC levels, heterozygous deficiency of CETP result in milder (10-30%) increases in HDLC levels (Klos et al., 2007).

A great number of studies investigated the association of CETP polymorphisms with HDLC concentrations and the evidence for an association is very clear (Table 1). Although these SNPs are in strong linkage disequilibrium they are not necessarily strongly correlated [START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF][START_REF] Thompson | High-density genotyping and functional SNP localization in the CETP gene[END_REF][START_REF] Mccaskie | Cholesteryl ester transfer protein gene haplotypes, plasma high-density lipoprotein levels and the risk of coronary heart disease[END_REF]. SNPs reported in Table 1 that were genotyped in HapMap were for the most part only weakly correlated with each other (Figure 2). Some of the investigated SNPs showed pronounced additive influences on HDLC levels with changes of 2-3 mg/dL per copy of the minor allele. Meta-analyses including more than 10,000 individuals are available for rs708272 (Taq1B) and rs5882 (Ile405Val)

showing a clear association of these SNPs with HDLC levels [START_REF] Boekholdt | Natural genetic variation as a tool in understanding the role of CETP in lipid levels and disease[END_REF][START_REF] Boekholdt | Cholesteryl ester transfer protein TaqIB variant, high-density lipoprotein cholesterol levels, cardiovascular risk, and efficacy of pravastatin treatment: individual patient metaanalysis of 13,677 subjects[END_REF]. Recently, a dense genotyping approach in more than 2000 individuals provided a very clear overview on the genetic variability within this gene and its associations with HDLC levels [START_REF] Thompson | High-density genotyping and functional SNP localization in the CETP gene[END_REF]. During the review process of this manuscript, a further meta-analysis on CETP was published [START_REF] Thompson | Association of cholesteryl ester transfer protein genotypes with CETP mass and activity, lipid levels, and coronary risk[END_REF], which is not included in Table 1 since the Entrez Date in PubMed was after our deadline of April 2008. The results of this meta-analysis which included 26 to 72 studies with 39,581 to 68,134 participants for the three common SNPs rs708272 (Taq1B), rs5882 (I405V), and rs1800775 (-629C>A) are in line with the results of the other studies listed in Table 1 and underline the significant influence of the CETP gene on HDLC levels.

Lipoprotein lipase (LPL)

Lipoprotein lipase is a critical enzyme involved in lipolysis of triglyceride-rich lipoproteins, chylomicrons and VLDL [START_REF] Miller | Genetic determinants of low high-density lipoprotein cholesterol[END_REF]. LPL does not act directly on HDL, but its action on triglyceride-rich lipoproteins has an important indirect effect on HDL metabolism [START_REF] Lewis | New insights into the regulation of HDL metabolism and reverse cholesterol transport[END_REF] (Figure 1). Familial LPL deficiency is a rare autosomal recessive disorder characterized by absence of LPL activity and a massive accumulation of chylomicrons in plasma. Homozygous deficiency is associated with severe hypertriglyceridemia and marked reductions in high-and low-density cholesterol levels (Klos et al., 2007).

Our literature search identified nine polymorphisms investigated for an association with HDL cholesterol (Table 2 andSupplementary Table). Most of these SNPs showed a significant association with HDLC levels (Table 2), whereas no study found a significant association for the common polymorphism rs285, which is only weakly correlated with the other SNPs (Supplementary Table andFigure 2). Five of the seven SNPs available in HapMap are located in a block of strong linkage disequilibrium [START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF] and especially rs320, rs326, rs13702 and rs10105606 show high correlations (Figure 2). A metaanalysis is available for four common polymorphisms which demonstrated significant associations with HDLC [START_REF] Wittrup | Lipoprotein lipase mutations, plasma lipids and lipoproteins, and risk of ischemic heart disease : A meta-analysis[END_REF]: the minor allele of rs328 (S447X respectively S474X according to NCBI) in >4000 individuals was associated with increased HDLC levels, whereas the minor alleles of rs268 (N291S) in almost 15,000 individuals, Gly188Glu (no rs number available) in >10,000 individuals and rs1801177 (D9N) in >5000 individuals were associated with lower HDLC levels.

Hepatic lipase (LIPC, HL, HTGL)

The human hepatic lipase gene, LIPC, is located on chromosome 15q21. It comprises 9 exons and 8 introns, and spans a length of more than 30 kb. It encodes a protein of 449 amino acids with a signal peptide of 23 amino acids [START_REF] Cai | Structure of the human hepatic triglyceride lipase gene[END_REF][START_REF] Ameis | Isolation and characterization of the human hepatic lipase gene[END_REF].

Hepatic lipase is a glycoprotein of approximately 65 kDa and is synthesized in hepatocytes and then secreted and bound to hepatocytes and hepatic endothelial surfaces. Hepatic lipase specifically catalyzes the hydrolysis of triglycerides, diglycerides and phospholipids in native lipoproteins [START_REF] Miller | Genetics of HDL regulation in humans[END_REF]. It appears to be involved in the selective uptake of cholesterol ester from HDL [START_REF] Lambert | Effects of apolipoprotein A-I genetic variations on plasma apolipoprotein, serum lipoprotein and glucose levels[END_REF] (Figure 1). To date, few patients with true hepatic lipase deficiency have been identified. All individuals with complete deficiency have had elevated plasma cholesterol and triglyceride concentrations. Furthermore, deficiency is characterized by abnormally triglyceride-rich HDL and LDL particles as well as high levels of HDLC [START_REF] Cohen | Hepatic lipase: new insights from genetic and metabolic studies[END_REF][START_REF] Holleboom | The value of HDL genetics[END_REF].

Seven SNPs in LIPC were reported to be associated with HDLC levels, two of them showing a very consistent association: rs1800588 and rs2070895 (Table 3). On six of the seven SNPs data are available in HapMap, and the r 2 -plot indicates a strong correlation between three of these SNPs which are all located in the promoter region of LIPC (Figure 2).

A meta-analysis including more than 24,000 individuals is available for rs1800588 (also described as C-514T or C-480T) showing that one or two copies of the minor allele increase HDLC levels by 1.5 and 3.5 mg/dL, respectively [START_REF] Isaacs | The -514 C->T hepatic lipase promoter region polymorphism and plasma lipids: a meta-analysis[END_REF].

Endothelial lipase (EL, LIPG)

Endothelial lipase (encoded by the LIPG gene) was discovered in 1999 independently by two different laboratories [START_REF] Jaye | A novel endothelial-derived lipase that modulates HDL metabolism[END_REF][START_REF] Hirata | Cloning of a unique lipase from endothelial cells extends the lipase gene family[END_REF] and is a member of the triglyceride lipase gene family. This enzyme derived its name because of its expression by endothelial cells [START_REF] Jaye | Endothelial lipase and HDL metabolism[END_REF]. Since overexpression of LIPG in mice resulted in marked reduction of HDLC levels [START_REF] Jaye | A novel endothelial-derived lipase that modulates HDL metabolism[END_REF], this has led to the concept that LIPG may play an important role in HDL metabolism. It promotes the turnover of HDL components and it increases the catabolism of apolipoprotein A-I (Figure 1). It was proposed that the endothelial lipase together with the hepatic lipase converts large HDL particles to smaller particles by its phospholipase activity [START_REF] Jaye | Endothelial lipase and HDL metabolism[END_REF]. Evidence is increasing that endothelial lipase might play a role in the etiology of the lipoprotein profile characteristic of the metabolic syndrome and an increased activity of this enzyme is linked to the underlying proinflammatory state in the metabolic syndrome [START_REF] Lamarche | Endothelial lipase and the metabolic syndrome[END_REF].

In our literature search we found five SNPs of LIPG which were significantly associated with HDL levels (Table 4). The evidence for an association is less strong as for CETP, LPL and LIPC and the studies available are much smaller than for these three genes. Four of these SNPs were available in HapMap, three of them are strongly correlated with r 2 ranging from 0.31-0.96 (Figure 2).

Lecithin:cholesteryl acyltransferase (LCAT)

LCAT is associated with HDL and esterifies cholesterol in plasma (Figure 1). ApoA-I serves as a cofactor for this reaction [START_REF] Miller | Genetic determinants of low high-density lipoprotein cholesterol[END_REF]Klos et al., 2007). Important knowledge on the function of this gene was derived from patients with LCAT deficiency which results in free cholesterol and phosphatidylcholine deposition in membranes followed by corneal opacification (fish-eye disease), anemia, and renal failure [START_REF] Miller | Genetic determinants of low high-density lipoprotein cholesterol[END_REF].

Homozygous LCAT deficiency is an underlying cause of two conditions: familial LCAT deficiency and fish-eye disease. While the former is associated with the complete loss of LCAT activity, the latter is associated with a change in the substrate specificity of LCAT that becomes inactive toward HDL cholesterol, while retaining its activity toward LDL cholesterol.

Both conditions are characterized by severe hypoalphalipoproteinemia, but only familial LCAT deficiency is strongly associated with premature CAD [START_REF] Sviridov | Genetic factors affecting HDL levels, structure, metabolism and function[END_REF].

Even though LCAT is due to its physiological function an important HDL candidate gene, only few large studies investigated the possible role of this gene on HDLC levels. Five polymorphisms of LCAT were investigated for an association with HDLC levels, but the findings are not entirely consistent (Table 5). The two SNPs available in HapMap (rs4986970 and rs2292318) are not correlated at all with each other (Figure 2) which could explain the presence of association with HDLC for one SNP (rs2292318) and the lack of association for the other SNP (rs4986970). An influence of LCAT on HDLC levels is supported by genomewide association studies (see below). There is an urgent need for large studies investigating the influence of common and rare genetic variations of this candidate gene on HDLC levels.

Scavenger receptor class B type 1 (SCARB1, SR-B1)

The SCARB1 gene has been localized to chromosome 12 spanning a region of 75 kb containing 13 exons. The gene encodes a receptor protein of approximately 80 kDa, whose weight can vary based on its extent of glycosylation. SCARB1 is highly expressed in liver and steroidogenic tissue (adrenal, ovaries, and testes) [START_REF] Cao | Structure and localization of the human gene encoding SR-BI/CLA-1. Evidence for transcriptional control by steroidogenic factor 1[END_REF]. It was isolated and characterized as a functional hepatic receptor for HDL [START_REF] Acton | Identification of scavenger receptor SR-BI as a high density lipoprotein receptor[END_REF]. This receptor participates in the selective uptake of cholesterol ester [START_REF] Miller | Genetics of HDL regulation in humans[END_REF], and binds a number of ligands with high affinity, including native HDL [START_REF] Cao | Structure and localization of the human gene encoding SR-BI/CLA-1. Evidence for transcriptional control by steroidogenic factor 1[END_REF]. The selective uptake involves the transfer of cholesterol from the HDL particle and the release of the lipidpoor HDL particle into the plasma [START_REF] Acton | Identification of scavenger receptor SR-BI as a high density lipoprotein receptor[END_REF] (Figure 1). It should be mentioned that control levels of plasma HDLC and non-HDL cholesterol as well as the propensity for atherosclerosis [START_REF] Miller | Genetics of HDL regulation in humans[END_REF], but it has to be determined whether this receptor has an equally important function in humans.

Despite the obvious functional evidence for an influence of SCARB1 on HDLC levels, the genetic-epidemiological evidence is relatively weak. Our literature search identified five polymorphic sites studied with HDLC levels with inconsistent results (Table 6). The two variants found in HapMap (rs5888 and rs5891) show no correlation with each other (Figure 2). Large genetic epidemiological studies on this gene are required before a final conclusion can be drawn.

ATP binding cassette transporter A1 (ABCA1)

The ATP binding cassette group of transporter proteins is involved in the transfer of various substances across plasma membranes, including ions, peptides, vitamins, and hormones. The ABCA1 transporter is expressed in liver, macrophages and steroidogenic tissues and plays a pivotal role in the initial phase of reverse cholesterol transport by mediating cholesterol and phospholipid efflux from macrophages to HDL [START_REF] Miller | Genetics of HDL regulation in humans[END_REF][START_REF] Cavelier | Lipid efflux by the ATP-binding cassette transporters ABCA1 and ABCG1[END_REF] (Figure 1). Far more than 1000 polymorphic sites within this gene are known. We appreciate the most important knowledge on ABCA1 and its influence on HDL metabolism to a rare monogenic recessive disorder, called Tangier disease in which patients carry homozygous mutations in ABCA1 [START_REF] Bodzioch | The gene encoding ATP-binding cassette transporter 1 is mutated in Tangier disease[END_REF][START_REF] Rust | Tangier disease is caused by mutations in the gene encoding ATP-binding cassette transporter 1[END_REF][START_REF] Brooks-Wilson | Mutations in ABC1 in Tangier disease and familial high-density lipoprotein deficiency[END_REF]. This causes a deposition of cholesteryl esters in the reticuloendothelial organs which results in orange-colored tonsils. Cholesterol is entrapped in macrophages due to the heavily disturbed cholesterol efflux. This results in lipid-depleted HDL particles rapidly catabolized and HDL deficiency.

Since the ABCA1 transporter plays an important physiological role in HDL metabolism, many studies attempted to show an association of this candidate gene with HDLC levels.

Table 7 and the Supplementary Table list 19 SNPs of ABCA1 that were investigated in the literature. However, most studies did not reveal large effects of these polymorphisms on HDLC levels although many included a large number of individuals. An explanation for this failing might be that a lot of rare (population frequency below 1%) and very rare (also called "private") mutations in ABCA1 with strong effects for each, could sum to an important influence on HDL metabolism [START_REF] Cohen | Multiple rare alleles contribute to low plasma levels of HDL cholesterol[END_REF]. These rare mutations are hardly to be pinpointed by association studies due to the small number of mutation carriers even in large studies. A typical example was recently shown by the Copenhagen City Heart Study (Frikke-Schmidt et al., 2008b) which could demonstrate in more than 9000 participants a pronounced effect for four rare mutations in ABCA1 on HDL cholesterol levels (a reduction of 17 mg/dL for heterozygotes vs noncarriers, p<0.001).

The r 2 -plot of the 9 available SNPs in HapMap shows that most SNPs are only weakly correlated which suggests that the missing associations of the single SNPs with HDL levels are independent of each other (Figure 2).

Apolipoprotein A-I (APOA1)

APOA1 is located in the APOA1/C3/C4 gene cluster on human chromosome 11q23 which harbors also APOA5.

As the major protein component of the HDL particle, apoA-I is an important ligand for HDL binding to cellular receptors, including scavenger receptor class B type 1 and ABCA1 [START_REF] Rigotti | Scavenger receptor BI--a cell surface receptor for high density lipoprotein[END_REF][START_REF] Remaley | Apolipoprotein specificity for lipid efflux by the human ABCAI transporter[END_REF]. Therefore, apoA-I serves as a cofactor for cholesterol esterification and is an important component of the reverse cholesterol transport [START_REF] Miller | Genetics of HDL regulation in humans[END_REF] (Figure 1). The clinical importance of apoA-I is underscored by a premature CVD in families with APOA1 deficiency and by a marked atherosclerosis in knockout mouse models [START_REF] Miller | Genetics of HDL regulation in humans[END_REF]. Patients carrying two functionally-relevant mutations in APOA1 being either homozygous or compound heterozygous, have extremely rare states of complete HDLC deficiency [START_REF] Funke | Genetic determinants of high density lipoprotein levels[END_REF]) and present with two different clinical hallmarks, namely xanthomas or corneal opacities [START_REF] Von Eckardstein | Differential diagnosis of familial high density lipoprotein deficiency syndromes[END_REF].

In our literature search we found three SNPs (rs670, rs5069 and rs5070) which were investigated for an association with HDLC levels. Mostly, large studies found a significant association with HDLC levels, but further large studies are required (Table 8 andSupplementary Table).

Apolipoprotein C-III (APOC3)

Apolipoprotein C-III is a component of triglyceride-rich lipoproteins and HDL particle and is transferred to HDL during the hydrolysis of triglyceride-rich lipoproteins [START_REF] Miller | Genetic determinants of low high-density lipoprotein cholesterol[END_REF].

The major physiological role of ApoC-III appears to be an inhibiting effect on LPL (Figure 1).

For the APOC3 gene, data on the influence of genetic variability on HDLC levels are heterogeneous. If indeed present, the influence seems to be weak (Table 8).

Apolipoprotein A-V (APOA5)

APOA5 is located proximal to the APOA1/C3/C4 gene cluster and was recently described by two groups [START_REF] Pennacchio | An apolipoprotein influencing triglycerides in humans and mice revealed by comparative sequencing[END_REF][START_REF] Van Der Vliet | Apolipoprotein A-V: A novel apolipoprotein associated with an early phase of liver regeneration[END_REF]. The human APOA5 gene consists of four exons and codes a 369 amino acid protein, expressed only in the liver [START_REF] Hubacek | Apolipoprotein A5 and triglyceridemia. Focus on the effects of the common variants[END_REF]. ApoA-V is predominantly located on TG-rich particles, chylomicrons and VLDL but also on HDL, and there is evidence that ApoA-V serves as an activator of LPL [START_REF] Hubacek | Apolipoprotein A5 and triglyceridemia. Focus on the effects of the common variants[END_REF]) (Figure 1). Among the apolipoproteins, APOA5 is the gene with the most pronounced associations with HDLC levels. Many polymorphisms of APOA5 were investigated in the literature, and most of them show an association with decreased levels of HDLC (Table 8). Only four SNPs are available in HapMap which are only weakly associated except rs662799 and rs651821 with a strong correlation (Figure 2).

Ghrelin (GHRL)

Ghrelin was only recently described by Kojima et al. (Kojima et al., 1999). Ghrelin is an endogenous peptide, its active form is composed of 28 aminoacids and circulates in plasma in a concentration of ~800-1000 pg/mL. Ghrelin is an orexigenic peptide predominantly produced by the stomach. Accumulating evidence suggests that ghrelin is a key regulator of body weight [START_REF] Cummings | Roles for ghrelin in the regulation of appetite and body weight[END_REF].

Two studies found a significant inverse association of four polymorphisms with HDLC levels whereas another study found no association of five SNPs with HDLC (Table 9). The effects of the SNPs on HDLC levels seem to be independent from each other since most of the SNPs available in HapMap are not highly correlated (Figure 2).

LDL-receptor (LDLR)

The LDL-receptor is a cell surface receptor that mediates the uptake of LDL particles from the circulation via receptor-mediated endocytosis [START_REF] Hobbs | The LDL receptor locus in familial hypercholesterolemia: mutational analysis of a membrane protein[END_REF]. Since this receptor plays an important role in plasma lipoprotein homeostasis [START_REF] Muallem | Quantitative effects of common genetic variations in the 3' UTR of the human LDL-receptor gene and their associations with plasma lipid levels in the Atherosclerosis Risk in Communities study[END_REF] (Figure 1), genetic variations in the LDLR gene that alter its expression could contribute to inter-individual differences in lipoprotein levels. Some of the five literature-reported polymorphisms in the LDLR gene show an association with HDLC levels, which was not confirmed by other SNPs (Table 9). For the four SNPs available in HapMap, rs688 and rs5925 showed strong correlations (r 2 =0.96) (Figure 2).

Paraoxonase 1 (PON1)

Paraoxonase 1 is a member of the paraoxonase family that includes three isoenzymes; their genes are clustered on chromosome 7q21. 3-22.1 (Primo-Parmo et al., 1996). PON1, a HDL-associated enzyme, is partly responsible for the antioxidation properties of HDL by protecting low-density lipoproteins against oxidation [START_REF] Mackness | Paraoxonase prevents accumulation of lipoperoxides in low-density lipoprotein[END_REF].

The PON1 gene has two widely investigated polymorphisms, the Gln192Arg and the Met55Leu, but the results are not entirely convincing (Table 9). A strong linkage disequilibrium exists between the two polymorphisms [START_REF] Arca | PON1 L55M polymorphism is not a predictor of coronary atherosclerosis either alone or in combination with Q192R polymorphism in an Italian population[END_REF].

Apolipoprotein E (APOE)

ApoE is a part of chylomicrons, VLDL, IDL and of some subspecies of HDL [START_REF] Sviridov | Genetic factors affecting HDL levels, structure, metabolism and function[END_REF] (Figure 1). The role of APOE polymorphism in determining VLDL-LDL levels is well known, but its independent influence on HDLC concentration is still controversial [START_REF] Sviridov | Genetic factors affecting HDL levels, structure, metabolism and function[END_REF].

A connection between the APOE polymorphism and HDLC levels was found in many but not all studies (Table 10). Since APOE is the best investigated of all known polymorphisms with far more than 50 publications that investigated an association with HDLC levels, we only referenced large population-based studies with more than 1000 individuals per study. Almost all of these studies found the E2 allele with increased and the E4 allele with decreased HDL levels compared to the E3 allele.

Results on rarely investigated genes

Table 11 provides the results on genes rarely investigated which showed a significant association with HDLC levels in at least one large study (n>1000). It might be a worthwhile attempt to replicate the shown associations with HDLC in further large studies.

Negative results on often investigated genes

Many studies investigated the association of the APOA4 gene and HDLC levels. The most frequently investigated polymorphisms are the T347S (rs675) and Q360H (rs5110).

Especially the large studies with sample sizes above 1000 did not show a significant association of these polymorphisms with HDLC levels (data not shown).

The same holds true for APOB: several studies investigated the association of this gene with HDLC levels. Large studies, however, showed non-significant results (data not shown).

A recent genome-wide association study with almost 18,000 individuals from 16 populationbased cohorts found apoB to be associated with HDLC levels (p=4.4x10 -8 ) [START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF].

Common versus rare variants

Common variants are those that appear in the general population with a frequency >1% or even >5% of subjects. These variants often show, individually, rather small associations.

Kathiresan et al. illustrated, however, that a combining of the information on several of these low-impact common variants can contribute to some extent to the prediction of HDLC levels (Kathiresan et al., 2008a). They studied nine SNPs at nine loci in 5287 subjects, created a genotype score on the basis of the number of unfavorable alleles and found decreasing HDLC levels with increasing genotype scores. Another study [START_REF] Zietz | Kandidatengene des Diabetes mellitus Typ 2: Gibt es einen Gen-Dosiseffekt für Risikofaktoren sowie mikro-und makrovaskuläre Folgeerkrankungen?[END_REF]) investigated five polymorphisms in different genes and found a significant association between the number of rarely occurring genotype combinations and HDLC levels, whereas each single polymorphism was not associated with HDLC levels.

Rare variants are those that appear with <1% in the general population. Cohen and colleagues [START_REF] Cohen | Multiple rare alleles contribute to low plasma levels of HDL cholesterol[END_REF]) investigated whether rare DNA sequence variants collectively contribute to variation in HDLC levels by sequencing ABCA1, APOA1, and LCAT in individuals from a population-based study. Nonsynonymous sequence variants were eight times more common (16% vs. 2%) in individuals with low HDLC (<5th percentile) than in those with high HDLC levels (>95th percentile). Biochemical studies indicated that most sequence variants in the low HDLC group were functionally important. The authors concluded that rare alleles with major phenotypic effects contribute significantly to low plasma HDLC levels in the general population. This seminal work started a new era with a focus on rare variants, and it is a consequent step to search these variants after a gene has been pinpointed by common variants to contribute to a certain phenotype.

Other examples for genes with rare variants having an important influence on HDLC levels are the angiopoietin-like 4 (ANGPTL4) gene (variant E40K, MAF=2%) which was associated with significantly higher levels of HDLC (p=4.0x10 -7 ) in 8726 individuals of the ARIC Study [START_REF] Romeo | Population-based resequencing of ANGPTL4 uncovers variations that reduce triglycerides and increase HDL[END_REF] or rare variants in the alcohol dehydrogenase 2 (ADH2) gene (MAF=2.8%) [START_REF] Whitfield | ADH genotype does not modify the effects of alcohol on high-density lipoprotein[END_REF] or in the cytochrome P450, family 3, subfamily A, polypeptide 4 (CYP3A4) gene (MAF=0.5%) [START_REF] Yamada | Prediction of genetic risk for dyslipidemia[END_REF] which showed significant associations in 901 and 3787 individuals, respectively. These studies highlight several important aspects for quantitative traits: 1) multiple common alleles influence trait variation, with each allele conferring a modest effect; 2) although each SNP exerts a modest effect, a combination of SNPs in aggregate can have a substantial influence on HDL levels; and 3) many rare alleles of candidate genes with a strong effect of each allele might have a pronounced influence on the trait under investigation in the studied population. Considering that all common alleles together explain less than 5-10% of HDLC levels in the general population and that HDLC levels are under considerable genetic control with heritability estimates of up to 80% (Kronenberg et al., 2002;[START_REF] Perusse | Familial resemblance of plasma lipids, lipoproteins and postheparin lipoprotein and hepatic lipases in the HERITAGE Family Study[END_REF], leaves a wide space for rare variants as well as for genetic regulatory

mechanisms not yet well understood.

New insights from genome-wide association studies

Advantages of genome-wide association studies

The recent introduction of microarray technology for genotyping allows the genotyping of several hundreds of thousands genetic variants in a single person in one step. This enables genome-wide association (GWA) studies by genotyping a large number of individuals with phenotypes of interest at reasonable costs. Compared to a hypothesis-driven candidate gene approach as described in the chapters above the hypothesis-free GWA studies can identify new susceptibility genes without making any a priori biological assumptions. They permit to identify genes involved in pathways which until now were unknown to be involved in a certain phenotype. GWA studies are therefore a new and very powerful tool to identify genetic contributors to phenotypes and have revolutionized gene hunting (for review see [START_REF] Coassin | An Optimized Procedure for the Design and Evaluation of Ecotilling Assays[END_REF][START_REF] Mccarthy | Genome-wide association studies for complex traits: consensus, uncertainty and challenges[END_REF]. There was never a time before in which in such a short time an unbelievable high number of genes for complex diseases and phenotypes has been identified. This holds true for complex diseases such as T2DM [START_REF] Frayling | Genome-wide association studies provide new insights into type 2 diabetes aetiology[END_REF], cancer as well as continuous variables such as lipids.

The introduction of GWA studies allowed to leave the path of hypothesis-driven search for genes influencing lipid levels, which was based on a limited a priori knowledge on biological and physiological processes involved in lipid metabolism. GWA studies allow to widen the horizon and to detect new genes involved in lipid metabolism. The identification of genes, however, is strongly dependent on the power of GWA studies which is usually quite low considering the fact that 500,000 SNPs and more are investigated at once, which creates a pronounced multiple testing problem with up to 25,000 false positive findings if the conventional significance threshold of p<0.05 is used. A decrease of this threshold to a genome-wide significance level (e.g. p<10 -7 ) and an increase in the number of samples studied increases the power and allows to bring the "real" genes out of the jungle of false positive associations. Therefore, the step from the single study to the meta-analysis was never before as short as in times of GWA studies. The researchers in this field realized immediately that they are more successful when combining their studies, which allows to identify even gene variants modifying the disease risk by only 10% or explaining less than 0.5% of the variance of continuous parameters.

Findings on HDLC by genome-wide association studies

Until now eight GWA studies on HDLC levels are available [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]Kathiresan et al., 2008b;Kooner et al., 2008;[START_REF] Wallace | Genome-wide association study identifies genes for biomarkers of cardiovascular disease: serum urate and dyslipidemia[END_REF][START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF]Kathiresan et al., 2007;[START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF][START_REF] Cashman | Genetic loci associated with plasma concentration of low-density lipoprotein cholesterol, highdensity lipoprotein cholesterol, triglycerides, apolipoprotein A1, and apolipoprotein B among 6382 white women in genome-wide analysis with replication[END_REF]. Results from these studies are summarized in Table 12. On the one hand, they have confirmed the following genes which were already known from earlier functional as well as association studies: CETP, LPL, LIPC, LIPG, ABCA1, LCAT, and the APOA1C3A4A5 gene cluster. On the other hand, these studies have identified new candidate genes influencing HDLC levels which need further functional characterization.

The most pronounced association of these newly identified genes was observed for GALNT2 [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]Kathiresan et al., 2008b). This gene would not have been identified in an hypothesis-driven approach since it does not have a known function immediately connected to lipid metabolism. GALNT2 encodes a widely expressed glycosyltransferase that could potentially modify a lipoprotein or receptor. This gene was also found to be associated with triglyceride concentrations [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]Kathiresan et al., 2008b). Another newly identified gene region was located near MVK and MMAB. These neighboring genes are regulated by SREBP2. MVK encodes mevalonate kinase, which catalyzes an early step in cholesterol biosynthesis, and MMAB encodes a protein that participates in a metabolic pathway that degrades cholesterol [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]. GRIN3A was identified in the initial GWA with a genome-wide significant p value of 2.5x10 -8 , but was not confirmed in the combined analysis with the replication samples [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF].

CLPTM1 was found in two independent GWA studies with a combined p value of 5.79x10 -6 .

However, this SNP was not found by other GWA studies [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF][START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF][START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF][START_REF] Cashman | Genetic loci associated with plasma concentration of low-density lipoprotein cholesterol, highdensity lipoprotein cholesterol, triglycerides, apolipoprotein A1, and apolipoprotein B among 6382 white women in genome-wide analysis with replication[END_REF].

Future GWA studies under way with even larger study samples and meta-analyses in up to 50,000 to 100,000 study participants will elucidate whether the above mentioned new genes or previously known candidate genes mentioned in chapter 3 and 4 but not yet identified in the hitherto published GWA studies will be confirmed. It can be taken for granted that new candidates will be identified with even smaller effect sizes than for the already identified genes.

Except three GWA studies [START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF][START_REF] Cashman | Genetic loci associated with plasma concentration of low-density lipoprotein cholesterol, highdensity lipoprotein cholesterol, triglycerides, apolipoprotein A1, and apolipoprotein B among 6382 white women in genome-wide analysis with replication[END_REF][START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF], previous studies were mainly based on study participants ascertained for T2DM or the metabolic syndrome [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]Kathiresan et al., 2008b;[START_REF] Wallace | Genome-wide association study identifies genes for biomarkers of cardiovascular disease: serum urate and dyslipidemia[END_REF]Kooner et al., 2008;Kathiresan et al., 2007), which could have obscured the results. Therefore, meta-analyses of population-based studies might have an advantage in identifying new candidates for HDLC levels that are not yet distorted by disease processes or medication. Such a meta-analysis of 16 European population-based cohorts has been published very recently [START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF]. For HDL cholesterol, eight regions showed genome-wide significant results. The results of the SNPs with the lowest p value for each of these regions are displayed in Table 12. Five of these regions (LPL, ABCA1, LIPC, CETP, and LIPG) have been identified in earlier GWA studies, APOB was for the first time identified in a GWA study for HDL cholesterol, and two regions (CTCF-PRMT8 and MADD-FOLH1) represent entirely novel regions. The MADD-FOLH1 locus of chromosome 11 represents a gene desert close to the centromere with no known gene on the 550 kb flanking region, and the two genes MADD and FOLH1 flanking the locus have not been implicated in lipid metabolism. The CTCF-PRMT8 gene encodes a transcriptional regulator, potentially involved in hormone-dependent gene silencing [START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF]. Further studies are needed to examine the potential role of these or neighbouring genes in HDL metabolism.

It is of interest that some loci (LCAT, APOA1C3A4A5, GALNT2, MVK/MMAB, CLPTM1 and GRIN3A) identified in earlier GWA studies of mostly diseased populations could not be confirmed in the meta-analysis of 16 European population-based cohorts. It can not be excluded that they were false positives in earlier studies considering that some were of borderline significance in these studies. Another explanation could be that genetic variation in these genes might play a more pronounced role in populations with metabolic diseaserelevant disturbances compared to the general population.

Conclusion from GWA studies on intergenic regions

One of the most important observation of our and other GWA studies is the fact that strong association signals are observed for SNPs in regions hitherto regarded as "intergenic". For example, we observed for HDLC very strong signals up to 70kb downstream of LIPG and LPL, and 10kb upstream of CETP [START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF]. This is particularly remarkable as the usual current candidate gene studies focus on SNPs within the gene ±5kb. While our downstream LPL SNPs showed some correlation with previously literaturereported SNPs within the gene, our LIPG signals were completely independent from SNPs within the gene or from any SNP reported previously in candidate gene studies. The identified CETP SNP 10kb upstream (rs8999419) was independent from all of the numerous SNPs reported in candidate gene studies and was even located in a recombination hotspot which strongly supports this region as an independent locus. These findings are in line with ideas that several regions within and outside of a gene can have independent effects on the gene function and that even regions far away from the gene harboring transcription factor binding sites, miRNAs, other enhancer elements or yet unknown regulatory elements can remotely control gene function. A similar example is reported for chromosomal region 9p21, which was shown to be associated with myocardial infarction [START_REF] Samani | Genomewide Association Analysis of Coronary Artery Disease[END_REF] and T2DM [START_REF] Scott | A genome-wide association study of type 2 diabetes in Finns detects multiple susceptibility variants[END_REF][START_REF] Saxena | Genome-wide association analysis identifies loci for type 2 diabetes and triglyceride levels[END_REF]. SNPs with the most significant signals were located more than 100kb upstream of the cyclindependent kinase inhibitors CDKN2A and CDKN2B, which supports either long-range effects on one of these genes or the influence of a gene not yet annotated. This supports the relevance of intergenic regions and calls for future functional studies to address this issue.

As we discussed recently [START_REF] Heid | A genome-wide association analysis of HDL cholesterol in the population-based KORA Study sheds new light on intergenic regions[END_REF], the relevance of genetic variation far outside of the gene might have far reaching consequences for candidate gene association studies in general. It is conceivable that plausible candidate genes in the past have been dropped prematurely when no association between intragenic variation and the investigated phenotypes was observed. If these regions are not considered and the variation located there is not in strong linkage disequilibrium with intragenic variation, an association would be missed. A higher-weighting for intragenic regions in previous candidate gene studies results in a biased search of phenotype-influencing gene regions and should probably be avoided.

In line with these observations, the GWA study approach thus evolves as "the better candidate gene study" as it enables a much more comprehensive analysis of all known candidate genes with ad libitum extension beyond gene boundaries and comparability across studies. Previously, candidate gene studies were each investigating different SNPs in different studies with different HDLC measurements and often different analysis models and in most of the studies with a strong focus on the promoter and intragenic regions of these genes.

Heterogeneity of low-HDL syndromes

HDL cholesterol levels have a strong genetic determination, with heritability estimates ranging in most cases between 40% and 60% [START_REF] Wang | Genetics of variation in HDL cholesterol in humans and mice[END_REF][START_REF] Goode | Heritability of longitudinal measures of body mass index and lipid and lipoprotein levels in aging twins[END_REF]Kronenberg et al., 2002). As it is the case with many intermediate cardiovascular risk phenotypes, low HDLC can be either monogenic or purely environmental or, in most cases is multifactorial / polygenic in origin [START_REF] Von Eckardstein | Differential diagnosis of familial high density lipoprotein deficiency syndromes[END_REF]. The most common inherited form of low HDLC is familial hypoalphalipoproteinemia, which is defined as an HDLC level below the 10th percentile, without secondary cause, and associated with a family history of low HDLC levels. The genetic cause is not fully characterized: some of the cases are due to mutations in HDL structural genes (mostly in one of the three genes APOA1, LCAT or ABCA1), whereas in other cases there appears to be accelerated catabolism of HDL and its apolipoproteins without a specific genetic mutation identified (Klos et al., 2007). In contrast to the rare occurrence of isolated low HDL in hypoalphalipoproteinemia, very low plasma levels of HDLC are also found in patients with genetically disturbed metabolic pathways which are indirectly linked to HDL metabolism. For example, many patients with lipid storage diseases like Gaucher´s disease, Nieman-Pick disease, diabetes mellitus or hypertriglyceridemia present with low HDL cholesterol (Von Eckardstein 2006). In the context of high triglycerides, low HDL cholesterol is not only an early symptom, but also provides a very sensitive marker of impaired glucose tolerance and increased lipolysis [START_REF] Rohrer | High density lipoproteins in the intersection of diabetes mellitus, inflammation and cardiovascular disease[END_REF]. These examples reflect the very heterogeneous nature of HDLC which makes HDL genetics very complex.

Relationship between HDLC and triglyceride genes

Many genes control lipolysis of plasma triglycerides, a process that also affects HDLC levels through the delivery of apolipoproteins and phospholipids to HDL [START_REF] Holleboom | The value of HDL genetics[END_REF]. There are several genes influencing both HDLC and triglyceride levels which have been identified by candidate gene and hypothesis-free GWA study approaches. For example associations between common SNPs at the APOA5 locus and triglyceride concentrations as well as HDLC levels are significant and relatively consistent across studies and populations [START_REF] Lai | The apolipoprotein A5 locus is a strong determinant of plasma triglyceride concentrations across ethnic groups in Singapore[END_REF]. Certain variants of LPL are associated with elevated plasma triglyceride and decreased HDLC levels [START_REF] Busch | Variation of candidate genes in triglyceride metabolism[END_REF]. The variation of the recognition site for SstI within the 3´-untranslated region of APOC3 has consistently shown an association with both plasma triglyceride and HDLC levels, whereas common variants of some other promising candidate genes, such as LIPC, were not as consistent [START_REF] Busch | Variation of candidate genes in triglyceride metabolism[END_REF].

Recent GWA analyses confirmed on the one hand the association of several loci with prior evidence of an association with triglyceride and HDLC levels (LPL, APOA1C3A4A5, and partly LIPC) [START_REF] Saxena | Genome-wide association analysis identifies loci for type 2 diabetes and triglyceride levels[END_REF][START_REF] Wallace | Genome-wide association study identifies genes for biomarkers of cardiovascular disease: serum urate and dyslipidemia[END_REF]Kooner et al., 2008;[START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]Kathiresan et al., 2008b;[START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF][START_REF] Cashman | Genetic loci associated with plasma concentration of low-density lipoprotein cholesterol, highdensity lipoprotein cholesterol, triglycerides, apolipoprotein A1, and apolipoprotein B among 6382 white women in genome-wide analysis with replication[END_REF], and identified on the other hand new loci (GALNT2, APOB) with an influence on both HDL and triglyceride levels [START_REF] Willer | Newly identified loci that influence lipid concentrations and risk of coronary artery disease[END_REF]Kathiresan et al., 2008b;[START_REF] Aulchenko | Genome-wide association study in 16 European population cohorts: Major loci influencing lipid levels and coronary heart disease risk[END_REF].

HDLC genes and atherosclerosis

The epidemiological evidence for an inverse association of HDLC concentrations with the risk of coronary artery disease is very strong [START_REF] Lewington | Blood cholesterol and vascular mortality by age, sex, and blood pressure: a meta-analysis of individual data from 61 prospective studies with 55,000 vascular deaths[END_REF], but the causality of this relationship is hard to prove. Since HDL levels are strongly determined by genetic variants (besides environmental factors), a causal association between HDLC and CAD can be demonstrated by showing an association of these genetic variants with risk of CAD. The idea behind is the concept of Mendelian Randomization [START_REF] Katan | Apolipoprotein E isoforms, serum cholesterol, and cancer[END_REF][START_REF] Davey | Mendelian randomization' : can genetic epidemiology contribute to understanding environmental determinants of disease?[END_REF][START_REF] Kataoka | Apolipoprotein E polymorphism in American Indians and its relation to plasma lipoproteins and 98[END_REF] which is based on the fact that it is randomly determined at the time of conception which of the two alleles from the father as well as from the mother will be transmitted to the child. Since the transmitted alleles are of lifelong persistence, these alleles determine to a certain amount also whether a person is exposed e.g. to low HDLC levels and therefore to the CVD risk associated with low HDLC levels. Therefore, the association between the polymorphism and CVD is less influenced by reverse causation or confounding.

Reverse causation would mean that CVD influences the polymorphism, which can practically be excluded. Confounding would mean that e.g. a lifestyle factor such as smoking is associated with the disease (which is often the case) as well as with the polymorphism (which is less probable). Therefore this method is well appropriate to underline a causal relationship which is hardly possible with conventional epidemiological observation studies.

The idea of Mendelian Randomization was originally proposed by Katan in the mid eighties [START_REF] Katan | Apolipoprotein E isoforms, serum cholesterol, and cancer[END_REF]) and was more and more used during the last five years [START_REF] Davey | Mendelian randomization' : can genetic epidemiology contribute to understanding environmental determinants of disease?[END_REF]. To our knowledge, it was the first time applied in 1992 when Sandholzer and colleagues clearly showed that the apolipoprotein(a) gene locus determines the risk for CVD by its strong influence on lipoprotein(a) concentrations [START_REF] Sandholzer | Apo(a) isoforms predict risk for coronary heart disease: a study in six populations[END_REF]. This seminal work demonstrated that high lipoprotein(a) concentrations are a primary, genetically determined risk factor for CVD, which was discussed controversially at that time since some researchers had believed that high lipoprotein(a) concentrations are a consequence rather than a cause of CVD. The causal relationship between lipoprotein(a) and atherosclerosis by using the Mendelian Randomization approach has been confirmed in the meanwhile by several studies (Kronenberg et al., 1999b;Kronenberg et al., 1999a;[START_REF] Wild | A prospective case-control study of lipoprotein(a) levels and apo(a) size and risk of coronary heart disease in Stanford Five-City Project participants[END_REF][START_REF] Kamstrup | Apolipoprotein(a) pentanucleotide repeat polymorphism, lipoprotein(a) levels and risk of ischemic heart disease[END_REF]Koch et al., 1997).

Figure 3A shows some considerations concerning the sample size for Mendelian Randomization projects. If one assumes that a particular polymorphism explains an unusual high 30% of an intermediate phenotype and that this intermediate phenotype explains about 10% of the clinical endpoint, it is expected that the polymorphism itself explains about 3% of the clinical endpoint (0.30 x 0.10 = 0.03). Single polymorphisms of HDLC genes, however, explain at best 3% of the HDLC levels. This would mean that such a polymorphism would explain 0.3% of the clinical endpoint (0.03 x 0.10 = 0.003). These considerations warn us that very large case-control studies are required to demonstrate an association between a polymorphism and a clinical endpoint. This might be the reason why although many association studies found an association between various polymorphisms and HDLC levels, most of them were markedly underpowered to consistently find an association of these polymorphisms with clinical endpoints.

A recent meta-analysis of three polymorphisms within the CETP gene (Taq1B, I405V and -629C>A) included 92 studies with up to 113,833 healthy participants, and 46 studies on 27,196 coronary cases and 55,338 controls [START_REF] Thompson | Association of cholesteryl ester transfer protein genotypes with CETP mass and activity, lipid levels, and coronary risk[END_REF]. The minor allele of each of the three polymorphisms was associated with a moderate inhibition of CETP activity and, in line with that, modestly higher HDLC levels and, most importantly, with a weakly inverse association with coronary risk. The odds ratios (OR) for coronary disease were compatible with the expected reductions in risk for equivalent increases in HDLC concentrations demonstrated in prospective studies [START_REF] Lewington | Blood cholesterol and vascular mortality by age, sex, and blood pressure: a meta-analysis of individual data from 61 prospective studies with 55,000 vascular deaths[END_REF]. Figure 3B illustrates these associations for the Taq1B polymorphism. These results clearly support a causal association of low HDLC levels with CVD. It also demonstrates that large studies are required to detect an association of this kind with certainty: the meta-analysis on the Taq1B polymorphism included 38 studies with a total of 19,035 cases and 32,368 controls. Since the effects of the investigated polymorphisms on HDLC levels were in the usual range expected in association studies, the results and the required sample size will be rather the rule than the exemption.

Even if the Mendelian Randomization approach worked fine e.g. for CETP, the situation is less clear for ABCA1. A large study of almost 50,000 individuals from Copenhagen showed that heterozygosity for rare loss-of-function mutations in ABCA1 were associated with substantial, lifelong lowering of plasma levels of HDLC (-17 mg/dL). These variants, however, were not associated with an increased risk of ischemic heart disease (Frikke-Schmidt et al., 2008b). It was speculated that this might be explained by the observation that the lower HDLC levels in these loss-of-function mutations are not accompanied by high triglyceride concentrations and that low HDLC levels per se are not atherogenic. A dysfunctionality of the HDL lipoprotein not necessarily reflected in the HDLC levels could be more important than HDLC levels themself. This is in line with another study in the Danish population showing that common non-synonymous genetic variation in ABCA1 predicts risk of ischemic heart disease independent of HDLC concentrations (Frikke-Schmidt et al., 2008a).

Conclusions

Several decades of research on the genetic contribution to HDL metabolism have identified a large number of genes contributing to the antiatherogenic HDLC concentrations.

The variants detected in these genes together explain only a small amount of less than 10% of the HDLC variance, but the heritability of HDLC is estimated to be up to 80%. This provides an enormous room for further yet to be identified genetic variants. This might be accomplished by genome-wide association studies. Since the low-hanging fruits might already be detected by earlier candidate gene studies and the first genome-wide association studies, future studies in large population-based genome-wide meta-analyses will identify new genes with even smaller effects on HDLC levels. These findings will probably result in a redrawing or extension of the involved metabolic pathways of HDLC metabolism. On the other hand, in-depth analysis of particular genes by deep-sequencing approaches or similar techniques [START_REF] Coassin | An Optimized Procedure for the Design and Evaluation of Ecotilling Assays[END_REF] will extend the previously made observation that a large number of rare alleles considerably contributes to a complex phenotype. polymorphism within the CETP gene and its association with HDLC levels as well as coronary disease [START_REF] Thompson | Association of cholesteryl ester transfer protein genotypes with CETP mass and activity, lipid levels, and coronary risk[END_REF] (for details see chapter 10 of the text).

diabetes. The Strong Heart Study. Arteriosclerosis, Thrombosis and Vascular Biology 16, 918-925. 79. Kathiresan, S., Manning, A.K., Demissie, S., D' Agostino, R.B., Surti, A., Guiducci, C. et al., 2007. A genome-wide association study for blood lipid phenotypes in the Framingham Heart Study. BMC.Med.Genet. 8 Suppl 1, S17. 80. Kathiresan, S., Melander, O., Anevski, D., Guiducci, C., Burtt, N.P., Roos, C. et al., 2008a. Polymorphisms associated with cholesterol and risk of cardiovascular events. N.Engl.J Med. 358, 1240-1249. 81. Kathiresan, S., Melander, O., Guiducci, C., Surti, A., Burtt, N.P., Rieder, M.J. et al., 2008b. Six new loci associated with blood low-density lipoprotein cholesterol, high-density lipoprotein cholesterol or triglycerides in humans. Nat.Genet. 40, 189-197. 82. Katsuya, T., Baba, S., Ishikawa, K., Mannami, T., Fu, Y., Inamoto, N. et al., 2002. Epsilon 4 allele of apolipoprotein E gene associates with lower blood pressure in young Japanese subjects: the Suita Study. J Hypertens. 20, 2017-2021. 83. Klerkx, A.H., Tanck, M.W., Kastelein, J.J., Molhuizen, H.O., Jukema, J.W., Zwinderman, A.H. et al., 2003. Haplotype analysis of the CETP gene: not TaqIB, but the closely linked -629C-->A polymorphism and a novel promoter variant are independently associated with CETP concentration. Hum. Mol.Genet. 12, 111-123. 84. Klos, K.L., Kullo, I.J., 2007. Genetic determinants of HDL: monogenic disorders andcontributions to variation. Curr.Opin.Cardiol. 22, 344-351. 85. Klos, K.L., Sing, C.F., Boerwinkle, E., Hamon, S.C., Rea, T.J., Clark, A. et al., 2006. Consistent effects of genes involved in reverse cholesterol transport on plasma lipid and apolipoprotein levels in CARDIA participants. Arteriosclerosis, Thrombosis and Vascular Biology 26, 1828-1836. 86. Ko, Y.L., Hsu, L.A., Hsu, K.H., Ko, Y.H., Lee, Y.S., 2004. The interactive effects of hepatic lipase gene promoter polymorphisms with sex and obesity on high-density-lipoprotein cholesterol levels in Taiwanese-Chinese. Atherosclerosis 172, 135-142. 87. Koch, M., Kutkuhn, B., Trenkwalder, E., Bach, D., Grabensee, B., Dieplinger, H. et al., 1997. Apolipoprotein B, fibrinogen, HDL cholesterol and apolipoprotein(a) phenotypes predict coronary artery disease in hemodialysis patients. Journal of the American Society of Nephrology 8, 1889-1898. 88. Kojima, M., Hosoda, H., Date, Y., Nakazato, M., Matsuo, H., Kangawa, K., 1999. Ghrelin is a growth-hormone-releasing acylated peptide from stomach. Nature 402, 656-660. 89. Komurcu-Bayrak, E., Onat, A., Poda, M., Humphries, S.E., Acharya, J., Hergenc, G. et al., 2007. The S447X variant of lipoprotein lipase gene is associated with metabolic syndrome and lipid levels among Turks. Clinica Chimica Acta 383, 110-115. 90. Kontush, A., Chapman, M.J., 2006. Functionally defective high-density lipoprotein: a new therapeutic target at the crossroads of dyslipidemia, inflammation, and atherosclerosis. Pharmacol.Rev. 58, 342-374. 91. Kooner, J.S., Chambers, J.C., guilar-Salinas, C.A., Hinds, D.A., Hyde, C.L., Warnes, G.R. et al., 2008. Genome-wide scan identifies variation in MLXIPL associated with plasma triglycerides. Nat.Genet. 40, 149-151. 92. Kronenberg, F., 2004. Epidemiology, pathophysiology and therapeutic implications of lipoprotein(a) in kidney disease. Expert.Rev.Cardiovasc.Ther. 2, 729-743. 93. Kronenberg, F., 2008. Genome-wide association studies in aging-related processes such as diabetes mellitus, atherosclerosis and cancer. Exp.Gerontol. 43, 39-43. 94. Kronenberg, F., Coon, H., Ellison, R.C., Borecki, I.B., Arnett, D.K., Province, M.A. et al., 2002. Segregation analysis of HDL cholesterol in the NHLBI Family Heart Study and in Utah pedigrees. Eur.J. Hum.Genet. 10, 367-374. 95. Kronenberg, F., Heid, I.M., 2007. Genetik intermediärer Phänotypen. Medizinische Genetik 19, 304-308. 96. Kronenberg, F., Kronenberg, M.F., Kiechl, S., Trenkwalder, E., Santer, P., Oberhollenzer, F. et al., 1999a. Role of lipoprotein(a) and apolipoprotein(a) phenotype in atherogenesis: prospective results from the Bruneck Study. Circulation 100, 1154-1160. 97. Kronenberg, F., Neyer, U., Lhotta, K., Trenkwalder, E., Auinger, M., Pribasnig, A. et al., 1999b. The low molecular weight apo(a) phenotype is an independent predictor for coronary artery disease in hemodialysis patients: a prospective follow-up. Journal of the American Society of Nephrology 10, 1027-1036. Abbreviations and explanations:

• Chrom. position: means chromosomal position according to NCBI Build 36 as included in Ensembl Release 49.

• Allele: means order of reference allele > risk allele; del, deletion; ins, insertion.

• MAF, minor allele frequency; Car., carriers; if MAF is not available, % Car. is provided.

• Study type: M, meta-analysis; P, population-based; GP, general population; Co, controls; CVD, cardiovascular disease; MI, myocardial infarction; NGT, normal glucose tolerance subjects; DM, diabetes mellitus; HoCo, hospital-based controls; HoSta, hospital staff; HBP, hypertensive patients; HeEx, health examination; Non-DM Co, nondiabetic control subjects; CorAng, coronary angiography; hyperCH, hypercholesterolemic patients. • Ethnicity: W, Whites; B, Blacks; A, Asians; W-Bra, Caucasian-Brazilians; B-Bra, African-Brazilians; AA, African Americans; NHW, non-Hispanic Whites; H, Hispanics; UK, United Kingdom; Asian-Ind., Asian-Indians. • HDLC, HDL cholesterol; M, males; F, females; sel., selected; n.s., not significant; n.a., not available. For abbreviations and explanations, see Table 1. For abbreviations and explanations, see Table 1. For abbreviations and explanations, see Table 1. SNPs rs1433099 and rs2738466 of LDLR never showed an association in any study but data on these studies are provided in the Supplementary Table .   For abbreviations and explanations, see Table 1. GRIN3A is shown here because it was significant in the initial scan (p=2.5x10 -8 ).

b z-sc: The ENGAGE consortium provided the effect size on the z-scale.

For abbreviations and explanations, see Table 1.

Boes et al. Boes et al. Figure 3 CETP TaqIB (rs708272)

Figure 1 .Figure 2 .

 12 Figure 1. Illustration of the major normal lipoprotein metabolic pathways. Refer to the text for detailed explanation. Blue arrows refer to points of action of the respective genes in blue. Red arrows refer to activating or inhibiting actions of certain apolipoproteins. Abbreviations: ABCA1, ATP binding cassette transporter 1; apo, apolipoprotein; CE, cholesterol ester; CETP, cholesteryl ester transfer protein; FFA, free fatty acid; HDL, highdensity lipoprotein; IDL, intermediate-density lipoprotein; LCAT, lecithin:cholesteryl acyltransferase; LDL, low-density lipoprotein; LDLR, low-density lipoprotein-receptor; LIPC, hepatic lipase; LIPG, endothelial lipase; Lp(a), lipoprotein(a); LPL, lipoprotein lipase; SR-B1, scavenger receptor B1; TG, triglycerides; VLDL, very low-density lipoprotein. (Figure adaptedfrom Kwan BCH, Kronenberg F, Beddhu S, Cheung AK: Lipoprotein metabolism and lipid management in chronic kidney disease. J. Am.Soc. Nephrol. 18:1246-1261[START_REF] Frayling | Genome-wide association studies provide new insights into type 2 diabetes aetiology[END_REF]

Figure 3 .

 3 Figure 3. Mendelian Randomization Approach. Panel A shows some considerations concerning the sample size for Mendelian Randomization projects (for explanation see chapter 10 of the text). Panel B shows the results of a recent meta-analysis of the Taq1B

c

  rs number not fully clarified.

a

  Eleven base pair deletion ranging from position -140 to -150. -140 to -150del mutation identified in the transcription factor binding region of the proximal promoter of the SCARB1 gene.

  o g e n o u s P a t h w a y E o g e n o u s P a t h w a y R e v e r s e C h o l e s t e r o l T r a n s p o r t

Table 1 :

 1 Genetic variants within the CETP gene coding for the cholesteryl ester transfer protein and their association with HDLC concentrations.

				Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	rs12149545 G-2708A	55550662	G>A	30	(McCaskie et al., 2007)	GP, CVD	W	2683 GP, 556 CVD	+1.9 per copy	<0.001
					G>A	31	(Klerkx et al., 2003)	CVD	W	709	+1.5 / +3.5 (1/2 copies)	0.0016
	rs3764261		55550825	G>T	28	(Pare et al., 2007)	CVD, Co	W	1442	n.a.	9x10 -6
	rs4783961 G-971A	55552395	G>A	49	(Klerkx et al., 2003)	CVD	W	709	+1.2 / +1.9 (1/2 copies)	0.09
	rs1800775 C-629 A	55552737	C>A	48	(Borggreve et al., 2005)	P	W	7083	+2.7 / +5.4 (1/2 copies)	<0.001
					C>A	51	(Bernstein et al., 2003)	P	W	847 M, 873 F	+4.2 for homozygotes	a	<0.002	a
					C>A	49	(Kathiresan et al., 2008a)	GP	W	5287	+3 / +5 (1/2 copies)	2x10 -29
					C>A	42-58	(Tai et al., 2003b)	GP	A	4050	+2.2 / +3.4 (1/2 copies)	a	3.28x10 -9 a
					C>A	48	(McCaskie et al., 2007)	GP, CVD	W	2683 GP, 556 CVD	+2.7 per copy	<0.001
					C>A	40	(Blankenberg et al., 2004)	CVD	W	CVD: 1214	CVD: +2.0 / +3.5 (1/2 copies)	0.02
								Co		Co: 574	Co: +3.3 / +6.1 (1/2 copies)	0.005
					C>A	44	(Klerkx et al., 2003)	CVD	W	709	+0.8 / +3.9 (1/2 copies)	<0.0001
					C>A	50	(Eiriksdottir et al., 2001)	MI, Co	W	309 MI, 757 Co	+1.9 / +6.1 (1/2 copies)	a	<0.0001	a
					C>A	48	(Freeman et al., 2003)	CVD, Co	W	498 CVD, 1107 Co	+2.9 / +4.4 (1/2 copies) a	<0.001 a
	rs708272	b	Taq1B, TaqIB	55553789	B1>B2 40-45	(Boekholdt et al., 2005)	M	-	13,677	+1.2 / +3.8 (1/2 copies)	<0.0001
			G+279A		B1>B2 n.a.	(Boekholdt et al., 2003)	M	-	>10,000	+4.6 for 2 copies	<0.00001
			(G=B1, A=B2)		B1>B2 42	(Borggreve et al., 2005)	P	W	7083	+2.7 / +5.0 (1/2 copies)	<0.001
					B1>B2 44	(Ordovas et al., 2000)	P	W	2916	+2.5 / +4.7 (1/2 copies)	a	<0.001	a
					n.a.	43W/26B (Klos et al., 2006)	P	W/B	2056 W, 1943 B	n.a. / n.a.	<0.01 / <0.02
					G>A	W: 44	(Nettleton et al., 2007)	P	W	W: 8764	W: +2.3 / +5.8 (1/2 copies)	<0.001
						B: 27			B	B: 2695	B: +3.8 / +9.8 (1/2 copies)	<0.001
					B1>B2 41	(Sandhofer et al., 2008)	P	W	1503	+2 / +5 (1/2 copies)	<0.001
					B1>B2 33-43	(Tai et al., 2003b)	GP	A	4207	+2.5 / +4.4 (1/2 copies)	a	1.25x10 -10 a
					B1>B2 40	(Tsujita et al., 2007)	GP	A	1729	M: +1.2 / +3.5 (1/2 copies)	0.096
											F: +1.9 / +6.2 (1/2 copies)	<0.001
					B1>B2 42	(McCaskie et al., 2007)	GP, CVD	W	2683 GP, 556 CVD	+2.7 per copy	<0.001
					B1>B2 42	(Whiting et al., 2005)	CVD, Co	W	2392 CVD, 827 Co	+1.7 / +3.6 (1/2 copies)	<0.001
					B1>B2 40	(Carlquist et al., 2003)	CVD	W	1464	+2.1 / + 3.0 (1/2 copies)	0.003

a pooled estimates and p values. b Studies of this SNP with sample sizes <1000 are provided in the Supplementary Table.

Table 2 :

 2 Genetic variants within the LPL gene coding for the lipoprotein lipase and their association with HDLC concentrations. a SNPs rs285 and rs10105606 never showed an association in any study but data on these studies are provided in the Supplementary Table.Studies with sample sizes <1000 are provided in the Supplementary Table.Position 474 instead of 447 according to NCBI database. For abbreviations and explanations, see Table1.

					Chrom.
	rs number Alias		position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	rs1801177 D9N; Asp9Asn 19849988	G>A	4% Car. (Wittrup et al., 1999)	M	-	5067	-3.1 for heterozygotes	0.002
	no rs #	Gly188Glu	19856022	Gly>Glu 1% Car. (Wittrup et al., 1999)	M	-	10,434	-9.7 for heterozygotes	<0.001
	rs268		N291S,	19857809	A>G	5% Car. (Wittrup et al., 1999)	M	-	14,912	-4.6 for heterozygotes	<0.001
			Asn291Ser
	rs320		HindIII; Int8	19863357	H+>H-30	(Senti et al., 2001)	P	W	520	+5.5 in H-H-vs. H+H+	0.025
						H2>H1 26	(Holmer et al., 2000)	P	W	1361	M: +3.5 for heterozygotes	0.0018
												F 25-44yrs: no association	n.s.
												F 45-64yrs: +4.2 for heterozy.	0.0212
						H+>H-32	(Corella et al., 2002)	GP	W	906	+1.9 for carriers	0.003
						T>G	17	(Ukkola et al., 2001)	GP	W	568	no association	n.s.
						H+>H-42% Car. (Radha et al., 2006)	NGT, DM	A	NGT: 550	NGT: +3.0 in carriers	<0.05
											DM: 465	DM: +1.0 in carriers	<0.05
						+>-	27-31	(Ahn et al., 1993)	DM, Co	NHW, H 615 NHW, 579 H	higher values in carriers	0.005
	rs326				19863719	n.a.	44	(Klos et al., 2006)	P	B	1943	M / F	0.013 / 0.004
	rs328	b	S447X	c	19864004	S>X	20% Car. (Wittrup et al., 1999)	M	-	4388	+1.5 for heterozygotes	<0.001
			1595CG		C>G	W: 10	(Nettleton et al., 2007)	P	W	W: 8968	W: +2.8 / +4.0 (1/2 copies)	<0.001
							B: 7			B	B: 2677	B: +3.1 / +12.6 (1/2 copies)	<0.001
						S>X	11	(Lee et al., 2004)	P	A	4058	+3.1 for carriers	<0.001
						S>X	11	(Komurcu-Bayrak et al., 2007) P	Turks	1586	M: no association	n.s.
												F: +4.0 for carriers	<0.001
						S>X	29	(Pallaud et al., 2001)	P	W	1552	no association	n.s.
						n.a.	n.a.	(Costanza et al., 2005)	P	W	1543	+ 2.7 for carriers	0.0017
						C>G	9	(Kathiresan et al., 2008a)	GP	W	5287	+3 / +5 (1/2 copies)	3x10 -12
						S>X	10	(Talmud et al., 2002b)	GP	W	2773	no association	n.s.
	rs13702			19868772	n.a.	50	(Klos et al., 2006)	P	B	1943	M / F	0.009 / 0.006

a b c

Table 3 :

 3 Genetic variants within the LIPC gene coding for the hepatic lipase and their association with HDLC concentrations.Studies with sample sizes <1000 are provided in the Supplementary Table.It is also possible that the known SNP 3b-280 is meant (rs12909325; chrom. position 56620995).

					Chrom.
	rs number Alias		position		Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	rs1077834 T-710C		56510771		T>C	22	(Andersen et al., 2003)	P	W	9121	+3-4% per copy	<0.001
	rs1800588	b C-514T	a	56510967		C>T	25	(Isaacs et al., 2004)	M	Various	>24,000	+1.5 / +3.5 (1/2 copies)	<0.001
			Pos. -480T	a		C>T	W: 21	(Nettleton et al., 2007)	P	W	W: 8897	W: +2.2 / +3.8 (1/2 copies)	<0.001
								B: 53			B	B: 2909	B: +1.6 / +4.0 (1/2 copies)	<0.001
							C>T	21	(Isaacs et al., 2007)	P	W	6239	+1.3 / +4.3 (1/2 copies)	d	<0.001	d
							C>T	38	(Tai et al., 2003a)	P	A	2170	+2.3 / +2.7 (1/2 copies)	0.001
							C>T	21	(Kathiresan et al., 2008a)	GP	W	5287	+1 / +4 (1/2 copies)	4x10 -10
							C>T	25	(Talmud et al., 2002b)	GP	W	2773	+1.5 per copy	0.04
							C>T	24	(Whiting et al., 2005)	CVD, Co	W	3319 CVD, 1385 Co	+1.0 / +3.8 (1/2 copies)	0.001
							C>T	51	(Yamada et al., 2007)	HoCo, P	A	5207	+2.5 per copy	<0.001
							C>T	21-24	(McCaskie et al., 2006)	GP, CVD	W	6412	+2.0-2.5 per copy	<0.001
	rs2070895 G-250A		56511231		G>A	22	(Andersen et al., 2003)	P	W	9121	+3-4% per copy	<0.001
			1b-280,HL1-280		n.a.	n.a.	(Costanza et al., 2005)	P	W	1543	+ 1.5 for carriers	0.020
							G>A	32	(de Andrade et al., 2004)	P	W	514	M: assoc., F: no assoc.	0.001 / n.s.
							G>A	23 22	(Grarup et al., 2008)	P P	W W	5585 8407	+3.9 / +3.9 (1/2 copies) +3.9 / +3.9 (1/2 copies)	-10 8x10 2x10 -7
							G>A	51	(Yamada et al., 2007)	HoCo, P	A	5213	+2.7 per copy	<0.001
							G>A	39	(Ko et al., 2004)	HeEx	A	716	+2.1 for carriers	0.026
	rs12594375		56519760		G>A	37	(Iijima et al., 2008)	GP	A	2970	n.a.	0.00003
	rs8023503			56527880		C>T	38	(Iijima et al., 2008)	GP	A	2970	n.a.	0.0001
								33		GP	A	1633	+2.6 / +3.4 (1/2 copies)	0.0007
	no rs #		3b-279,HL3-279 56620996	c	n.a.	n.a.	(Costanza et al., 2005)	P	W	1543	+ 1.8 for carriers	0.024
	rs3829462 +1075C		56640371		A>C	5	(Fang et al., 2002)	HoSta	A	823	+8.0 for heterozygotes	<0.05
	a	C-514T and -480T are the same polymorphisms (depending on the nucleotide taking as the transcription start site).
	d	pooled estimates and p values.
	For abbreviations and explanations, see

b c

Table 1 .

 1

Table 4 :

 4 Genetic variants within the LIPG gene coding for the endothelial lipase and their association with HDLC concentrations.

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL	P value
	rs3813082 -384A>C	45342041 A>C	12	(Hutter et al., 2006)	Co		A	541	+1.3 / +10.2 (1/ 2 copies)	0.021
				A>C	12	(Yamakawa-Kobayashi et al.,	Kids		A	340	+0.7 / +9.8 (1/ 2 copies)	0.0086
						2003)
	rs2000813 584 C/T	45347862 T>I	32	(Paradis et al., 2003)	GP		W	495	M: +1.2 / +2.7 (1/2 copies)	0.82
		T111I									F: -0.4 / +1.9 (1/2 copies)	0.09
				C>T	24	(Hutter et al., 2006)	Co		A	541	+0.5 / +6.1 (1/ 2 copies)	0.048
				C>T	18 / 30	(Tang et al., 2008)	CVD, Co		A	265 CVD, 265 Co	+3.7 in carries	a	<0.02	a
				C>T	24	(Yamakawa-Kobayashi et al.,	Kids		A	340	-0.6 / +2.6 (1/ 2 copies)	n.s.
						2003)
				C>T	29	(Ma et al., 2003)	CVD		90% W	372	+1.6 / +6.0 (1/ 2 copies)	0.035
	rs2276269 C+42T/ln5	45356000 C>T	44	(Mank-Seymour et al., 2004)	HDL sel. b	W	594	decreases HDLC	0.007
	rs6507931 T+2864C/ln8	45367006 T>C	42	(Mank-Seymour et al., 2004)	HDL sel.	b	W	594	decreases HDLC	0.004
	rs3744841 2237G>A	45371372 G>A	36	(Yamakawa-Kobayashi et al.,	Kids		A	340	-4.0 / -4.3 (1/ 2 copies)	0.011
						2003)
	a	pooled estimates and p values.
		Selected according to HDLC levels: high HDLC group with values >60 mg/dL (n=355) and low HDLC group with values <35 mg/dL (n=239).

b

Table 5 :

 5 Genetic variants within the LCAT gene coding for the lecithin:cholesteryl acyltransferase and their association with HDLC concentrations.

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	no rs #	Gly230Arg	66531921	Gly>Arg 5% Car.	a (Miettinen et al., 1998)	see	a	W	156 low HDLC,	variant only in low HDLC
										160 high HDLC	group
	rs4986970 Ser208Thr	66533821	S>T	3	(Talmud et al., 2002b)	GP		W	2773	no association	n.s.
		Ser208Thr		S>T	6% Car. (Boekholdt et al., 2006)	CVD	W	546	no association	n.s.
	rs5922	608C/T	66533918	C>T	n.a.	(Zhang et al., 2003)	CVD, Co	A	CVD: 203	increase in HDL	0.015
										Co: 209	no association	n.s.
				C>T	14% Car. (Zhu et al., 2006)	Stroke, Co A	150 stroke, 122 Co	lower HDLC in heterozyg.	<0.05
	no rs #	P143L	66534134	C>T	6% Car. (Zhang et al., 2004)	CVD, Co	A	190 CVD, 209 Co	association with low HDLC	<0.01
		+511C>T
	rs2292318	66543207	G>A	12	(Pare et al., 2007)	CVD, Co	W	1442	increases HDLC	2x10 -5
	a	5% carriers in 156 persons with low HDL (8-27 mg/dL) vs. 0% carriers in 160 persons with high HDL (73-139 mg/dL) selected from a large population
		study.

Table 6 :

 6 Genetic variants within the SCARB1 (SR-B1) gene coding for the scavenger receptor class B type 1 and their association with HDLC concentrations.

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	rs5888	A350A	123850701 C>T	48	(Osgood et al., 2003)	P	W	M: 1177	M: +1.2 / +2.3 (1/2 copies)	n.s.
									F: 1239	F: 0.0 / +1.2 (1/2 copies)	n.s.
		(exon8 C-T)		C>T	44	(Morabia et al., 2004)	P	W	M: 865	M: +1.9 / 2.7 (1/2 copies)	0.0062
									F: 891	F: +0.4 / +0.8 (1/2 copies)	n.s.
		C1050T		n.a.	n.a.	(Costanza et al., 2005)	P	W	1543	-5.1 for carriers	n.s.
				C>T	41	(Roberts et al., 2007)	Families	Amish	M: 428	M: no association	n.s.
									F: 491	F: +4 / +3 (1/ 2 copies)	0.04
				C>T	49	(Boekholdt et al., 2006)	CVD	W	546	+2.3 / +1.9 (1/2 copies)	0.03
	rs5891		123865495 G>A	3	(Roberts et al., 2007)	Families	Amish	M: 428	M: no association	n.s.
									F: 491	F: +9 / +10 (1/ 2 copies)	<0.001
	rs4238001 Ex.1(G>A)	123914216 G>A	12	(Osgood et al., 2003)	P	W	2465	+0.4 for carriers	n.s.
	no rs #	11-bp del a	123914499-	ins>del a 4% Car. (Hsu et al., 2003)	HeEx	A	690	+6.0 for deletion carriers	0.035
			509
	no rs #	C-142T	123914515 C>T	14	(Hsu et al., 2003)	HeEx	A	664	+1 / -1 (1/ 2 copies)	n.s.

Table 7 :

 7 Genetic variants within the ABCA1 gene coding for the ATP binding cassette transporter A1 and their association with HDLC concentrations. a

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	rs2246298 C(-297)T	106730554 C>T	25	(Shioji et al., 2004b)	GP	A	1625	n.a.	0.0455
	rs1800976 G(-273)C	106730530 G>C	40	(Shioji et al., 2004b)	GP	A	GP: 1626	+1.9 / +2.7 (1/2 copies)	0.026
							HBP	A	HBP: 735	/ +5.0 (1/2 copies)	0.026
	rs1800977 C69T; C-14T	106730271 C>T	36	(Porchay et al., 2006)	GP	W	5040	no association	n.s.
				C>T	38	(Hodoglugil et al., 2005)	GP (no DM) Turks	2332	M: +0.7 / +1.9 (1/2 copies)	<0.02
										F: -0.3 / 0.0 (1/2 copies)	n.s.
	rs1800978 G378C	106705799 G>C	13	(Porchay et al., 2006)	GP	W	5040	-1.2 / -2.7 (1/2 copies)	0.03
	rs3890182		106687476 G>A	13	(Kathiresan et al., 2008a)	GP	W	5287	-1 / -3 (1/2 copies)	0.003
	rs2275542 IMS-JST071749 106684901 n.a.	n.a.	(Shioji et al., 2004b)	GP	A	<1880	n.a.	0.006
	rs2515602		106639989 n.a.	27	(Klos et al., 2006)	P	B	1943	M / F	0.034 / <0.001
	rs2853578 G596A	106633129 G>A	28	(Whiting et al., 2005)	CVD, Co	W	2468 CVD, 834 Co	-0.2 / +2.8 (1/2 copies)	0.02
	rs2066718 V771M	106629076 G>A	3	(Frikke-Schmidt et al., 2004)	P	W	9123	M: trend to higher levels	0.12
		2310G>A								F: higher levels in carriers	0.02
		G2706A		V>M	5	(Hodoglugil et al., 2005)	GP (no DM) Turks	2458	M: +2.0 for heterozygotes	<0.01
										F: +1.9 for heterozygotes	n.s.
				V>M	3	(Clee et al., 2001)	CVD	W	635	no association	n.s.
	rs2066715 V825I	106627854 G>A	6	(Frikke-Schmidt et al., 2004)	P	W	9123	M: no association	n.s.
		2472G>A								F: +3.1 in carriers	0.0005
		G2868A		V>I	6	(Hodoglugil et al., 2005)	GP (no DM) Turks	2105	no association	n.s.
				V>I	8	(Clee et al., 2001)	CVD	W	682	no association	n.s.
	rs4149313 I883M	106626574 A>G	12	(Frikke-Schmidt et al., 2004)	P	W	9123	M: no association	n.s.
										F: increase in hetercygotes	0.05
		2648A>G		I>M	22	(Hodoglugil et al., 2005)	GP (no DM) Turks	2532	no association	n.s.
	rs33918808 E1172D	106619453 G>C	3	(Frikke-Schmidt et al., 2004)	P	W	9123	M: no association	n.s.
										F: higher levels in carriers	0.05
		3515G>C		E>D	5	(Hodoglugil et al., 2005)	GP (no DM) Turks	2238	no association	n.s.
				E>D	3	(Clee et al., 2001)	CVD	W	644	no association	n.s.
	no rs #	32b.+30, ABC32 106606881 n.a.	n.a.	(Costanza et al., 2005)	P	W	1543	-2.2 for carriers	0.0040

Table 8 :

 8 Genetic variants within the genes APOA1, APOC3 and APOA5 coding for apolipoprotein A-I, C-III and A-V, respectively and their associations with HDLC concentrations. a

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	APOA1
	rs5070	T84C (1280)	116213230 T>C	W:33/B:47 (Brown et al., 2006)	P	Multi	3831	no association	n.s.
		(HaeIII)		T>C	23	(Shioji et al., 2004a)	GP	A	1637	+1.9 / +5.4 (1/2 copies)	0.0005
	rs5069	MspI RFLP	116213464 T>C	W: 3	(Brown et al., 2006)	P	W, B	3831	W: M / F	0.0001 / 0.027
					B: 31					B: M / F	n.s. / 0.022
		C-151T		+>-	3	(Kamboh et al., 1996)	GP	NHW	M: 248	M: -3.0 in carriers	n.s.
									F: 280	F: +7.5 in carriers	0.009
		+83bp; +83 C/T		+>-	40	(Kamboh et al., 1999)	GP	A	M: 490	M: -2.3 / -0.9 (1/2 copies)	n.s.
									F: 291	F: +0.1 / -0.9 (1/2 copies)	n.s.
		1046 (C->T)		M1>M2 44	(Zaman et al., 1997)	Co	A	1328	-1.2 / -1.3 (1/2 copies)	n.s.
				+>-	5	(Larson et al., 2002)	Co	W	730	+2.0 for carriers	n.s.
	APOC3
	rs2854117 -482C>T (9154) 116205352 G>A	W: 73	(Brown et al., 2006)	P	Multi	3831	W: M / F	n.s. / n.s.
					B: 33					B: M / F	n.s. / 0.014
				C>T	27	(Pallaud et al., 2001)	P	W	772 M, 780 F	no association	n.s.
				T>C	27	(Qi et al., 2007)	GP (DM)	W	875	+0.6 / +0.1 (1/2 copies)	n.s.
	rs2854116 C-455T, 9127	116205379 A>G	W: 62	(Brown et al., 2006)	P	Multi	3831	W: M / F	n.s. / n.s.
					B: 30					B: M / F	n.s. / 0.045
				T>C	41	(Lahiry et al., 2007)	P	Inuits	1308	-3.1 / -5.4 (1/2 copies)	<0.05
				T>C	38	(Qi et al., 2007)	GP (DM)	W	846	+0.2 / -1.7 (1/2 copies)	n.s.
	rs618354	c PvuII	116205987 1>2	49	(Kamboh et al., 1999)	GP	A	M: 478 F: 291	M: -0.9 / -1.3 (1/2 copies) F: +0.1 / -4.2 (1/2 copies)	n.s. 0.029
	rs5128	+3175	116208850 C>G	9	(Pallaud et al., 2001)	P	W	772 M, 780 F	no association	n.s.
		Sst1 RFLP		S1>S2 9	(Russo et al., 2001)	P	W	M: 1219	M: -1.8 in carriers	0.04
									F: 1266	F: +0.4 in carriers	n.s.
		3´-UTR/Sac I		->+	9	(Hegele et al., 1995)	P	Hutteries 713	-5.0 for heterozygotes	0.0014
		3238C>G		S1>S2 7	(Corella et al., 2002)	GP	W	906	+1.9 for carriers	0.079
				S1>S2 35	(Arai et al., 2005)	GP	A	852	-0.8 / -1.5 (1/2 copies)	0.41

Table 9 :

 9 Genetic variants within the genes GHRL, LDLR and PON1 coding for ghrelin, LDL-receptor and paraoxonase 1, respectively and their associations with HDLC concentrations. a

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	GHRL
	rs3755777 -1500C>G	10308364	C>G	49	(Choi et al., 2006)	Non-DM Co A	641	-2 for carriers	0.0020
	rs26311	-1062G>C	10307926	A>G	12	(Martin et al., 2008)	GP	Canadian 1464	no association	n.s.
				G>C	37	(Choi et al., 2006)	Non-DM Co A	640	-4 for carriers	<0.0001
	rs26312	-994C>T	10307858	C>T	39	(Choi et al., 2006)	Non-DM Co A	639	-3 for carriers	0.0004
	rs26802		10307365	T>G	30	(Martin et al., 2008)	GP	Canadian 1464	no association	n.s.
	rs696217	+408C>A	10306457	C>A	18	(Choi et al., 2006)	Non-DM Co A	636	-2 for carriers	0.0042
		Leu72Met		Leu>Met 7	(Hubacek et al., 2007)	P	W	1189 M, 1364 F	-2.3 for carriers	0.01
	rs2075356		10303809	T>C	11	(Martin et al., 2008)	GP	Canadian 1464	no association	n.s.
	rs4684677		10303453	T>A	6	(Martin et al., 2008)	GP	Canadian 1464	no association	0.09
	rs35684		10301686	A>G	27	(Martin et al., 2008)	GP	Canadian 1464	no association	n.s.
	LDLR
	rs2228671 Exon 2	11071912	n.a.	n.a.	(Costanza et al., 2005)	P	W	1543	+3.8 for carriers	0.0056
	rs688 =	1866C>T	11088602	C>T	12	(Yamada et al., 2008)	HoCo	A	2417	+1.5 / +8.5 (1/2 copies)	0.0155
	rs57911429	Asn591Asn
		Exon 12/HincII		->+	39	(Hegele et al., 1995)	P	Hutteries 713	-2.3 / -4.3 (1/2 copies)	0.047
	rs5925 =	2052T>C	11091881	T>C	17	(Yamada et al., 2008)	HoCo	A	2417	+1.2 / +5.4 (1/2 copies)	0.043
	rs57369606	Val653Val
	PON1
	rs662 =	Q192R	94775382	A>G	30	(Srinivasan et al., 2004)	P	W	W: 1232	W: +0.1 / +2.3 (1/2 copies)	0.041
	rs60480675	Gln192Arg			67			B	B: 554	B: -5.4 / -6.7 (1/2 copies)	0.008
				Q>R	31	(Manresa et al., 2006)	P	W	1364	-0.6 for carriers	n.s.
				Q>R	31	(Senti et al., 2002)	P	W	M: 724	M: 0.0 / -0.6 (1/2 copies)	n.s.
									F: 656	F: +0.6 / -1.6 (1/2 copies)	n.s.
				Q>R	29	(Hegele et al., 1995)	P	Hutteries 738	-3.1 / -3.1 (1/2 copies)	0.001
				Q>R	32	(van Aalst-Cohen et al., 2005) hyperCH	W	1002	0.0 / -0.8 (1/2 copies)	n.s.
				Q>R	36	(Rios et al., 2007)	CVD, Co	W-Bra	M: 261	M: +1.5 / +2.7 (1/2 copies)	0.035

Table 10 :

 10 Association of the APOE polymorphism coding for apolipoprotein E with HDLC concentrations. a Due to the high number of studies investigating the APOE polymorphism, only large (n>1000) population-based studies are provided in this Table.

	Study name	Sample size	Influence of E2 on HDL	Influence of E4 on HDL	P value b
	EPIC Norfolk Study (Wu et al., 2007)	22,915			<0.001
	ARIC Study (Volcik et al., 2006)	9304 Whites			<0.05
		3187 African Americans			<0.05
	Copenhagen City Heart Study (Frikke-Schmidt et al.,	5025 women			<0.001
	2000)	4035 men			n.s. / 0.001
	Rotterdam Study (Slooter et al., 2001)	5401
	Strong Heart Study (Kataoka et al., 1996)	1296 nondiabetic women			0.007
		1040 nondiabetic men			n.s.
		1322 diabetic women			0.040
		752 diabetic men			n.s.
	Suita Study (Katsuya et al., 2002)	3997	n.d.		0.003
	Framingham Offspring Study (Wilson et al., 1994)	916 women	OR for HDL<35 = 0.80	OR for HDL<35 = 1.15	n.s.
		1034 men	OR for HDL<35 = 1.36	OR for HDL<35 = 1.37	0.11 / 0.06
	Cardiovascular Risk in Young Finns Study (Gronroos et	1736			0.04
	al., 2007)
	(Costanza et al., 2005)	1543		n.d.	0.00078
	Bogalusa Heart Study (Srinivasan et al., 1999)	1477			<0.05 / n.s.

a b P value from ANOVA comparing the major genotype groups.

n.s., not significant; n.d., not done.

Table 11 :

 11 Genetic variants in genes rarely investigated, which showed significant association with HDLC levels in at least one large study (n>1000).

			Chrom.
	rs number Alias	position	Allele	MAF	Study	Study type Ethnicity Sample size	Change in HDLC in mg/dL P value
	ALDH2 = aldehyde dehydrogenase 2
	rs671	Glu487Lys	110726149 Glu>Lys 30	(Takagi et al., 2001)	P	A	M: 1919	M: -2.3 / -8.5 (1/2 copies)	<0.0001
					28				F: 2138	F: -1.9 / -1.9 (1/2 copies)	0.0254
		ALDH2 *1-*2		1>2	31	(Nakamura et al., 2002)	GP	A	M: 826	M: -1.8 / -3.5 (1/2 copies)	n.s.
					28				F: 1295	F: -1.2 / -0.2 (1/2 copies)	n.s.
	CD36 = CD36 antigen
	rs10499859 a		80096746	A>G	33	(Love-Gregory et al., 2008)	HyperGen	b AA	2020	+2.5 per copy	2.8x10 -4
	DGAT1 = acyl CoA:diacylglycerol acyltransferase 1
	rs7003945 C79T	145521273 T>C	43	(Ludwig et al., 2002)	P	W	1322	-1.8 / -1.4 (1/2 copies)	0.019
	EDN1 = endothelin-1
	rs5370		12404241	G>T	21	(Pare et al., 2007)	CVD, Co	W	1442	decreases HDLC	1x10 -5
	MCC = mast cell chymase
	rs1800875 G3255A	24049178	G>A	20	(Fukuda et al., 2002)	P	A	1046	+2.5 / +1.8 (1/2 copies)	0.038
	NR1H3 = liver X receptor alpha
	rs11039155 -6G>A	47237338	G>A	11	(Legry et al., 2008)	P	W	2290	+2.1 for carriers	0.007

PPAR-= peroxisome proliferator activated receptor alpha

	rs1800206 L162V	44992938	L>V	4	(Manresa et al., 2006)	P	W	1204	-4.0 for carriers	<0.05

PPAR-2 = peroxisome proliferator activated receptor gamma 2

			45413088	rs7240405	17 (Heid et al., 2008)		1643	-2.77 mg/dL	3.19x10 -4	8213	4.67x10 -10
	ABCA1	9	106696891 rs3905000	14 (Aulchenko et al., 2008)		17,913	-4.37 (z-sc) b	8.6x10 -13
			106687041 rs4149268	36 (Willer et al., 2008)		8656		3.3x10 -7	8656+11,327	1.2x10 -10
			106687476 rs3890182	13 (Kathiresan et al., 2008b)		2758	-2.21 mg/dL	3x10 -5	2758+18,554	3x10 -10
			106687476 rs3890182	12 (Wallace et al., 2008)		1636		3.8x10 -2	1636+2631	2.09x10 -6
			106660656 rs9282541	0-9 (Kooner et al., 2008)				10,536	4.8x10 -8
	LCAT	16 66582496	rs255052		17 (Willer et al., 2008)		8656		1.5x10 -6	8656+4534	1.2x10 -7
	APOA1C3A4A5 11 116233840 rs12225230	18 (Cashman et al., 2008)		6382	1.5 mg/dL	5.3x10 -5
			116124283 rs28927680	7	(Kathiresan et al., 2008b)		2758	-0.78 mg/dL	0.31	2758+18,554	2x10 -5
	APOB	2	21059688	rs6754295	25 (Aulchenko et al., 2008)		17,915	2.63 (z-sc) b	4.4x10 -8
	CTCF-PRMT8	16 66459571	rs2271293	13 (Aulchenko et al., 2008)		17,910	4.99 (z-sc)	b	8.3x10 -16
	MADD-FOLH1 11 48475469	rs7395662	39 (Aulchenko et al., 2008)		17,917	2.82 (z-sc) b	6.0x10 -11
	GALNT2	1	228361539 rs2144300	40 (Willer et al., 2008)		8656		6.6x10 -7	8656+11,406	2.6x10 -14
			228362314 rs4846914	40 (Kathiresan et al., 2008b)		2758	-1.30 mg/dL	3x10 -4	2758+18,554	2x10 -13
	MVK/MMAB	12 108379551 rs2338104	45 (Willer et al., 2008)		8656		1.9x10 -6	8656+11,399	3.4x10 -8
	CLPTM1	19 50169221	rs16979595	16 (Wallace et al., 2008)		1636		6.11x10 -3	1636+2631	5.79x10 -6
	GRIN3A	9	103402758 rs1323432	12 (Willer et al., 2008)		8656	1.93 mg/dL	2.5x10 -8 a	8656+8176	7.7x10 -4 a
	a
	rs1805192 Pro12Ala		12396238	Pro>Ala 5	(Tai et al., 2004)	P	A	4038	+1.6 for carriers	0.028
	rs3856806 C1431T		12450557	C>T	23	(Tai et al., 2004)	P	A	4038	no association	n.s.
	PRKAA2 = AMP-kinase 2 subunit
	rs2796498 1 -2		56916574	1>2	46	(Spencer-Jones et al., 2006)	GP	W	2275	-1.5 / -2.3 (1/2 copies)	0.01
	ZNF202 = Zinc Finger Protein 202
	rs3183878 c.*2T>G Exon10 123101913 T>G	39	(Stene et al., 2006)	P	W	F: 5039	F: no association	n.s.
										M: 4064	M: -1.9 for homozygous Car.	0.05

'A' allele increases HDLC by 2.28 mg/dL (95%CI 1.93-2.63)

Acknowledgments

Our own research discussed in this review was funded by grants from the "Genomics of Lipid-associated Disorders -GOLD" of the "Austrian Genome Research Programme GEN-AU", the Austrian National Bank (Project 12531) and the Austrian Heart Fund to F. Kronenberg and by the German National Genome Research Net, the Munich Center of Health Sciences (MC Health) as part of LMUinnovativ, Germany, and the NIH-subcontract from the Children's Hospital, Boston, USA, under the prime grant 1 R01 DK075787-01A1, CFDA 93.848 supporting the work of I.M. Heid.

The HyperGen (Hypertension Genetic Epidemiology Network) Study recruited hypertensive siblings diagnosed before age 60 from sibships with two or more affected persons, normotensive offsprings and random controls. Details on the study are provided in reference [START_REF] Williams | NHLBI family blood pressure program. Methodology and recruitment in the HyperGEN network[END_REF].

For abbreviations and explanations, see Table 1.