Associations between modifiable exposures and disease seen in observational epidemiology are sometimes confounded and thus misleading, despite our best efforts to improve the design and analysis of studies. Mendelian randomization-the random assortment of genes from parents to offspring that occurs during gamete formation and conception-provides one method for assessing the causal nature of some environmental exposures. The association between a disease and a polymorphism that mimics the biological link between a proposed exposure and disease is not generally susceptible to the reverse causation or confounding that may distort interpretations of conventional observational studies. Several examples where the phenotypic effects of polymorphisms are well documented provide encouraging evidence of the explanatory power of Mendelian randomization and are described. The limitations of the approach include confounding by polymorphisms in linkage disequilibrium with the polymorphism under study, that polymorphisms may have several phenotypic effects associated with disease, the lack of suitable polymorphisms for studying modifiable exposures of interest, and canalization-the buffering of the effects of genetic variation during development. Nevertheless, Mendelian randomization provides new opportunities to test causality and demonstrates how investment in the human genome project may contribute to understanding and preventing the adverse effects on human health of modifiable exposures.

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‘Mendelian randomization’: can genetic epidemiology contribute to understanding environmental determinants of disease?

This two-part article provides an overview of the global burden of atherothrombotic cardiovascular disease. Part I initially discusses the epidemiologic transition which has resulted in a decrease in deaths in childhood due to infections, with a concomitant increase in cardiovascular and other chronic diseases; and then provides estimates of the burden of cardiovascular (CV) diseases with specific focus on the developing countries. Next, we summarize key information on risk factors for cardiovascular disease (CVD) and indicate that their importance may have been underestimated. Then, we describe overarching factors influencing variations in CVD by ethnicity and region and the influence of urbanization. Part II of this article describes the burden of CV disease by specific region or ethnic group, the risk factors of importance, and possible strategies for prevention.

Global Burden of Cardiovascular Diseases Part I: General Considerations, the Epidemiologic Transition, Risk Factors, and Impact of Urbanization

Levels of low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides and total cholesterol are heritable, modifiable risk factors for coronary artery disease. To identify new loci and refine known loci influencing these lipids, we examined 188,577 individuals using genome-wide and custom genotyping arrays. We identify and annotate 157 loci associated with lipid levels at P < 5 × 10(-8), including 62 loci not previously associated with lipid levels in humans. Using dense genotyping in individuals of European, East Asian, South Asian and African ancestry, we narrow association signals in 12 loci. We find that loci associated with blood lipid levels are often associated with cardiovascular and metabolic traits, including coronary artery disease, type 2 diabetes, blood pressure, waist-hip ratio and body mass index. Our results demonstrate the value of using genetic data from individuals of diverse ancestry and provide insights into the biological mechanisms regulating blood lipids to guide future genetic, biological and therapeutic research.

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Discovery and refinement of loci associated with lipid levels

Observational epidemiological studies suffer from many potential biases, from confounding and from reverse causation, and this limits their ability to robustly identify causal associations. Several high-profile situations exist in which randomized controlled trials of precisely the same intervention that has been examined in observational studies have produced markedly different findings. In other observational sciences, the use of instrumental variable (IV) approaches has been one approach to strengthening causal inferences in non-experimental situations. The use of germline genetic variants that proxy for environmentally modifiable exposures as instruments for these exposures is one form of IV analysis that can be implemented within observational epidemiological studies. The method has been referred to as 'Mendelian randomization', and can be considered as analogous to randomized controlled trials. This paper outlines Mendelian randomization, draws parallels with IV methods, provides examples of implementation of the approach and discusses limitations of the approach and some methods for dealing with these.

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Mendelian randomization: using genes as instruments for making causal inferences in epidemiology.

Medical genetics was revolutionized during the 1980s by the application of genetic mapping to locate the genes responsible for simple Mendelian diseases. Most diseases and traits, however, do not follow simple inheritance patterns. Geneticists have thus begun taking up the even greater challenge of the genetic dissection of complex traits. Four major approaches have been developed: linkage analysis, allele-sharing methods, association studies, and polygenic analysis of experimental crosses. This article synthesizes the current state of the genetic dissection of complex traits—describing the methods, limitations, and recent applications to biological problems.

Genetic Dissection of Complex Traits

https://www.thelancet.com/pdfs/journals/lancet/PIIS0140-6736(00)82009-7.pdf

Large-scale test of hypothesised associations between the angiotensin-converting-enzyme insertion/deletion polymorphism and myocardial infarction in about 5000 cases and 6000 controls. International Studies of Infarct Survival (ISIS) Collaborators.

Background In previous studies, unadjusted comparisons of mortality and major morbidity after acute myocardial infarction have generally indicated that women have a poorer outcome than men. Much larger studies are needed, with more complete adjustment for coexisting conditions, to determine whether this difference is explained by the older age of the women studied or by the presence of other unfavorable prognostic factors, or both. Methods As part of the Third International Study of Infarct Survival (ISIS-3), information was collected on deaths during days 0 to 35 and on major clinical events during hospitalization up to day 35 for 9600 women and 26,480 men with suspected acute myocardial infarction who were considered to have a clear indication for fibrinolytic therapy. We compared the outcome among women and men, first without adjustment, then with adjustment for age, and finally with adjustment for other recorded base-line characteristics by means of multiple logistic regression. Results The unadjusted...

A comparison of the early outcome of acute myocardial infarction in women and men. The Third International Study of Infarct Survival Collaborative Group.

A Comparison of the Early Outcome of Acute Myocardial Infarction in Women and Men

BACKGROUND: Blood concentrations of fibrinogen have been associated with coronary heart disease risk in epidemiological studies, but it is uncertain whether this association is causal or reflects residual confounding by other risk factors. We investigated the relationship between the single nucleotide polymorphism at position -148 in the beta-fibrinogen gene promoter (beta - 148C/T), blood fibrinogen levels, and risk of myocardial infarction (MI) in sufficiently large numbers of coronary disease cases to reliably address this question. METHODS: Genotyping and measurement of blood fibrinogen concentration were carried out in 4,685 cases of confirmed MI and 3,460 controls with no history of coronary disease. A meta-analysis of ISIS and 19 other studies of beta-fibrinogen genotypes involving a total of 12,220 coronary disease cases and 18,716 controls was conducted. RESULTS: Among the ISIS controls, mean plasma fibrinogen concentrations with the C/C, C/T and T/T genotypes were 3.34 (SE 0.015), 3.48 (0.022), and 3.60 (0.064) g/l, respectively, corresponding to an increase of 0.14 (0.024) g/l per T allele (trend P < 0.0001). In the case-control comparison, 0.14 g/l higher usual plasma fibrinogen concentration was associated with an age-adjusted and sex-adjusted risk ratio for MI of 1.17 [95% confidence interval (95% CI) 1.14-1.19; P < 0.0001]. But, after further adjustment for smoking, body mass index, and plasma apolipoprotein B/A(1) ratio, this risk ratio fell to 1.03 (95% CI 1.00-1.05; P = 0.05). Moreover, fibrinogen genotype was not significantly associated with MI incidence: risk ratio of 1.06 (95% CI 0.96-1.16) per higher-fibrinogen allele in ISIS alone and of 1.00 (95% CI 0.95-1.04) per allele in the meta-analysis. CONCLUSIONS: Genotypes that produce lifelong differences in fibrinogen concentrations do not materially influence coronary disease incidence. As these genotype-dependent differences in fibrinogen were allocated randomly at conception (Mendelian randomization), this association is not likely to be confounded by other factors. Consequently, these genetic results provide strong evidence that long-term differences in fibrinogen concentrations are not a major determinant of coronary disease risk.

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Fibrinogen and coronary heart disease: test of causality by 'Mendelian randomization'

Results In whole blood stored at room temperature for up to 7 days, concentrations of albumin, apolipoproteins A1 and B (apoA1 and apoB), cholesterol, high density lipoprotein (HDL), total protein, and triglycerides changed by less than 4%, and low density lipoprotein (LDL) by less than 7%. Whilst alanine transaminase (ALT), creatine kinase (CK), creatinine, and γ-glutamyl transferase (GGT) concentrations changed substantially at room temperature, there was less than 4% change during chilled storage up to 7 days. By contrast, aspartate transaminase (AST) concentrations increased markedly under both conditions. Conclusions A wide range of important analytes, including lipids, change by only a few per cent in whole blood during storage at room temperature for several days. Mailed transport of whole blood samples may, therefore, be a simple and cost-effective option for large-scale epidemiological studies.

Alison Palmer

Papers

Large-scale test of hypothesised associations between the angiotensin-converting-enzyme insertion/deletion polymorphism and myocardial infarction in about 5000 cases and 6000 controls. International Studies of Infarct Survival (ISIS) Collaborators.

A comparison of the early outcome of acute myocardial infarction in women and men. The Third International Study of Infarct Survival Collaborative Group.

A Comparison of the Early Outcome of Acute Myocardial Infarction in Women and Men

Fibrinogen and coronary heart disease: test of causality by 'Mendelian randomization'

Stability of plasma analytes after delayed separation of whole blood: implications for epidemiological studies