Introduction and the Literature review
1.3 The Literature Review
1.3.8 Single Nucleotide Polymorphisms and the Metabolic syndrome
1.3.8.1 LIPOPROTEIN LIPASE
Circulating lipids are carried in lipoproteins, which consist of esterified and unesterified cholesterol, triglycerides, phospholipids and protein (Preiss-Landi et al., 2002).
Lipoprotein lipase (LPL) is the major rate-limiting enzyme that catalyses the hydrolysis of triglyceride from circulating triglyceride-rich lipoproteins, very low density proteins and chylomicrons (Wang, 2009). During this process, surface-free cholesterol and phospholipids are transferred to HDL particles, increasing the concentration of HDL
48 (Preiss-Landi et al., 2002). The products from these catalytic reactions, like fatty acids, are taken up by the tissues and processed differentially; hence LPL acts as a bridging protein which mediates the cellular binding and up-take of lipoproteins (Stein, 2003).
Lipoprotein lipase is found throughout the body, with the highest activity and mRNA levels found in the heart, skeletal muscle, adipose tissue, and to a lesser degree, in the lungs and brain (Zechner, 1997). It is expressed in the adipocytes, macrophages, smooth muscle cells, and most importantly, on the luminal surface of the capillary endothelium (Zechner, 1997). Since LPL plays a pivotal role in overall lipoprotein metabolism, it is involved in forward cholesterol transport and contributes to the maturation of high- density lipoprotein (HDL) precursors, which are themselves involved in reverse cholesterol transport (Pillarisetti, 2003). Lipoprotein lipase also controls the delivery of triglyceride-derived free-fatty acids to muscle, adipose tissue, and vascular wall macrophages, where lipid uptake influences peripheral insulin sensitivity, central obesity, and foam cell formation (Mead, 2002).
Most of the identified SNPs with functional effects have been reported to lead to loss of enzymatic function, and hence, higher triglyceride and lowered HDL levels (Kathiresan et al., 2008). A reduction in LPL activity could result in adverse metabolic consequences and contribute to the clinical phenotype (Zhang et al., 1996), which is supported by the strong association that has been reported between reduced LPL activity, major cardiovascular risk factors (Mead, 2002) and MS (Aguilera et al., 2008). Therefore, diminished LPL activity is suggested to be one mechanism that results in impaired clearance of circulating
49 lipoproteins, with subsequent hypertriglyceridemia (Wang et al., 2009). An increase in LPL activity, however, is associated with a favourable lipid profile, namely, a lower triglyceride and higher HDL levels (Preiss-Landi et al., 2002). Hayden et al. (1991) reported that lipid abnormalities have genetic determinants, with other researchers postulating that the metabolic parameters that determine serum lipid and lipoprotein levels are modulated by multiple gene-gene and gene-environmental interactions (Corella et al., 2002; Lee et al., 2004).
1.3.8.1.1 The Lipoprotein lipase gene
The gene that codes for LPL is located on chromosome 8p22, spans close to 35 kb, and contains 10 exons (Oka et al., 1990). Most of the over 100 identified LPL gene mutations are rare, with 20% occurring in the non-coding regions (Murthy et al., 1996). Three common exonic variants, namely, D9N, N291S and S447X have been identified (Van Bockxmeer, 2001).
Abnormalities of LPL function are associated with adverse pathophysiological conditions like hyperlipidaemia. Prospective case-control studies have shown a strong and consistent association of LPL polymorphisms with raised triglyceride levels and CAD (Sagoo et al., 2008). A recent cross-sectional and longitudinal study of 2045 African Americans and 2116 Europeans was undertaken by Tang et al. (2010) to determine the association of eight LPL polymorphisms with the lipid profile. Apart from the population-
50 related heterogeneity that was observed over the 20 year follow-up, they reported that lipid variations were influenced by the effect of ageing on the LPL polymorphisms. LPL variants have also been associated with obesity in South Indian Asians (Radha et al., 2007).
Since raised triglyceride levels and lowered HDL levels make up two of the defining features of MS the study of the LPL gene was considered important in this project.
Furthermore, since there are clearly mechanistically opposing effects of LPL polymorphisms on lipid profile, the S447X polymorphism (which has been found to have a favourable effect on lipid profile), and the N291S polymorphism (which has been reported as having an adverse association with HDL and triglyceride levels) were studied.
1.3.8.1.2 Polymorphisms in the LPL gene
The S447X polymorphism that is located on exon 9 is reported as being one of the most common polymorphisms, with a frequency of 5.6% to 21.1 % (Sagoo et al., 2008;
Humphries et al., 1998). This SNP is characterized by a C-G transversion, where the X- allele encodes a prematurely truncated LPL protein, Serine, converting the 447 codon prematurely to a termination codon (Rip et al., 2006). In contrast to other LPL SNPs, the S447X variant is an exception within the coding region, and has been linked to increased lipolytic activity (Ross et al., 2005). Hokanson (1999) suggests that the functional properties of this truncated protein results in an enhanced bridging function, which leads
51 to increased clearance of triglyceride-rich lipoproteins. In a case-control study of subjects with combined hyperlipidaemia, Wung et al. (2006) showed that individuals with this mutation had significantly lower triglyceride, low-density lipoprotein cholesterol and TG/HDL-C ratio, which are established risk factors for CVD (Hokanson et al., 1996) in contrast to individuals without the SNP. This was also shown in a recent meta-analysis (Kathiresan et al., 2008), and has been supported in a large cohort of 1577 Chinese Canadian subjects (McGladdery, 2001), and in other twin and candidate gene association studies (Huang et al., 2006; Groop, 2001).
There have been subsequent studies which show that carriers of the X447 allele have a higher degree of protection against developing MS (Jensen et al., 2009). For example, in the Turkish Adult Risk Factor (TARF) study, Komurcu-Bayrak et al. (2007) evaluated the relationship of the S447X variant with serum lipid levels and MS. They found that individuals with this variant had higher levels of HDL and lower fasting glucose when compared to those with the wild-type. Furthermore, in a study of two hundred Egyptian patients with acute myocardial infarction, S447X was found to be associated with a favourable lipid profile. Conversely, this profile was reversed with the addition of DM (Tarek et al., 2011) as a risk factor. Another study examining the association between the S447X variant, hypertension-induced LV growth and CAD risk, showed that the hypertensive carriers of the variant were at an increased risk of LVH and risk for CAD. This was in contrast to the protective trend observed in normotensive subjects with the variant (Talmud et al., 2007).
52 Since considerable variation has been reported between study populations (Kathiresan et al., 2008) in the association of the S447X polymorphism with plasma lipids, its association with risk of CHD, in addition to genetic factors (Corella et al., 2002) and environmental exposures like smoking (Lee et al., 2004), we investigated the association between S447X with MS and its components. Furthermore, the influence of this SNP on the lipid profile or on the MS has not been studied in a community-based cohort of randomised men and women. This is particularly pertinent to the Phoenix community, in South Africa, whose inhabitants, being of predominantly Asian Indian origin, are known to have a high propensity for adverse lipid profiles and DM (Seedat et al., 1990).
In contrast to the cardio-protection conferred by the S447X mutation, the N291S polymorphism has been associated with low lipoprotein lipase enzymatic activity, and hence, an unfavourable lipid profile (Zhanget al., 1996; Hayden et al., 1991), since identified in 1994. This may be due to the location of the N291S SNP in a heparin-binding cluster, which is thought to affect the interaction of LPL with the cell wall glycosaminoglycan, as well as the result of the substitutions of the amino acid in the N- terminal domain of LPL which is responsible for catalytic function (Mead et al., 2002).
A large meta-analysis comprising 17 630 subjects (Hokanson et al., 1996) showed that this mutation was associated with a marginal risk for CVD, with stronger associations reported in certain populations. Another meta-analysis of 13 studies by Wittrup et al.
(1999) reported that this mutation was associated with a significant increase in triglyceride levels and low HDL, with Souverein et al. (2005) reporting a significant
53 association between this mutation and triglyceride levels in a study of 512 males with CAD. This was also shown in EARS study (Humphries et al., 1998) where younger subjects with N291S mutation were more likely to have elevated TG and decreased HDL concentrations, potentiated by moderate obesity. The N291S SNP has also been associated with increased atherosclerotic risk, as reported by the Framingham Offspring study (Kastelein, et al., 1999). A meta-analysis by Yaomin et al. (2006) found that this mutation conferred additional risk for a dyslipidaemia profile and was associated with DM and CAD, with López-Ruiz et al. (2009) and Sagoo et al. (2008) reporting that N291S could identify subjects with a high CV risk and was associated with adverse lipid profiles.
Therefore, in view of the high propensity of Asian Indians for dyslipidaemia and DM, the study of the associations between this polymorphism, DM and dyslipidaemia in this sample was determined to ascertain if additional CV risk for MS was conferred by the presence of the N291S polymorphism.