Genome-Wide Screen for G6E Interactions on Lipids Figure 2. Lipoprotein subclass particle and key serum lipid concentration correlations with WHR for different genotypes of rs6448771. The height of the bar is the meta-correlation between the lipoprotein particle concentration and waist-to-hip ratio, and the whiskers correspond to standard error of the meta-correlation. The P-values have been taken from the interaction meta-analysis and only P-values,0.01 are shown in the figure. The two cohorts in which the lipid particle concentrations were measured with NMR metabonomics platform were YFS and NFBC1966 with combined number of samples of 6,500. XXL_VLDL: Chylomicrons and extremely large very low-density lipoprotein particles; XL: Very large, L: large, M: Medium, S: Small, XS: Very small; VLDL: very low-density lipoprotein; IDL: intermediate-density lipoprotein; LDL: low-density lipoprotein; HDL: High-density lipoprotein; TG: Triglycerides; TC: Total cholesterol. doi:10.1371/journal.pgen.1002333.g002 these populations are detailed in Table S1A (discovery), S1B (replication) and S1C (further replication). Brief descriptions of the cohorts are provided in the Text S1 section ‘‘Short descriptions of the cohorts’’. Phenotype determination Individuals were excluded from analysis if they were not of European descent or were receiving lipid-lowering medication at the time of sampling. TC, HDL-C, and TG concentrations were measured from serum or plasma extracted from whole blood, typically using standard enzymatic methods. LDL-C was either directly measured or estimated using the Friedewald Equation (LDL-C = TC – HDL-C – 0.456TG for individuals with TG#4.52 mmol/l, samples with TG level higher than 4.52 were discarded in the calculation of LDL-C) [18]. Covariates and epidemiological risk factors were ascertained at the same time that blood was drawn for lipid measurements. BMI was defined as weight in kilograms divided by the square of height in meters. Waist circumference was measured at the mid-point between the lower border of the ribs and the iliac crest; hip circumference was measured at the widest point over the buttocks. Waist-to-hip ratio was defined as the ratio of waist and hip circumferences. Alcohol consumption and smoking habits were determined via interviews and/or questionnaires. Both behaviors were coded as dichotomous (abbreviations: ALC for drinker/ abstainer and SMO for current smoker/current non-smoker) and semi-quantitative traits. Semi-quantitative alcohol usage (ALCq) was based on daily consumption in grams (0: 0 g/day; 1: .0 and #10 g/day; 2: .10 and #20 g/day; 3: .20 and #40 g/day; 4: .40 g/day). Semi-quantitative smoking (SMOq) was assessed based on the number of cigarettes per day (0: 0 cigarettes/day; 1: .0 and #10 cigarettes/day; 2: .10 and #20 cigarettes/day; 3: .20 and #30 cigarettes/day; 4: .30 cigarettes/day). Genotyping and imputations Affymetrix, Illumina or Perlegen arrays were used for genotyping in the discovery cohorts. Each study filtered both individuals and SNPs to ensure robustness for genetic analysis. After quality control, these data were used to impute genotypes for approximately 2.5 million autosomal SNPs based on the LD patterns observed in the HapMap 2 CEU samples. Imputed genotypes were coded as dosages, fractional values between 0 and 2 reflecting the estimated number of copies of a given allele for a given SNP for each individual. Cohort specific details concerning quality control filters, imputation reference sets and imputation software are described in Table S4. In silico replication Replication cohorts utilized genome-wide imputed data, as described above, where available. Details on the genotyping methods implemented in the replication samples are described in Table S4. Serum NMR metabonomics, lipoprotein subclasses Proton NMR spectroscopy was used to measure lipid, lipoprotein subclass and particle concentrations in native serum samples. NMR methods have been previously described in detail [16,19]. Serum concentrations of total triglycerides (TG), total cholesterol (TC) together with LDL-C and HDL-C were determined. In addition, total lipid and particle concentrations in 14 lipoprotein subclasses were measured. The measurements of these subclasses have been validated against high-performance liquid chromatography [20]. The subclasses were as follows: chylomicrons and largest VLDL particles (particle diameters from approx 75 nm upwards), five different VLDL subclasses: very large VLDL (average particle diameter 64.0 nm), large VLDL (53.6 nm), medium-size VLDL (44.5 nm), small VLDL PLoS Genetics | www.plosgenetics.org 6 October 2011 | Volume 7 | Issue 10 | e1002333