The results of this GWAS do not indicate that any of the 339,596 SNPs are associated with CVD events among migraineurs at the genome-wide level. After lowering the significance threshold to 5×10−6 there was some suggestion that five SNPs at different loci might be implicated. Two of the SNPs suggested an association with ischemic stroke (rs7698623, rs4975709), one with major CVD (rs2143678), and one with CVD death (rs1406961) among women with migraine with aura. In addition, rs1047964 appeared to be associated with CVD death among women with any migraine. There is no indication that migraine aura status modifies the association between previously described markers for MI, ischemic stroke, and silent brain infarcts with any of the CVD events. Although the WGHS population is among the largest with information about both migraine and incident CVD, our results need to be treated with caution, since the number of CVD events was only modest and the MAFs for three of the SNPs was low.
Migraine, in particular migraine with aura, has been associated with CVD. The evidence was summarized in a recent meta-analysis indicating a two-fold increased risk for ischemic stroke, which only appears for migraine with aura 
. The evidence is less clear for other CVD events; however, individual studies suggest that this pattern may also extend to MI, CVD death, and hemorrhagic stroke 
. The association between migraine and ischemic vascular events was independent of many CVD risk factors in many studies 
. Furthermore, the increased risk of ischemic stroke is more apparent among migraineurs without CVD risk factors 
. In addition functional 
, cellular 
, and genetic data 
indicate shared biological vulnerability between vascular disorders and migraine.
Investigating the genetic liability for CVD among migraineurs has only recently begun. Two analyses reported on the influence of single gene variants on the migraine-CVD association and showed a modulatory effect of the MTHFR
polymorphisms among women. The two-fold increased risk of CVD among migraineurs with aura was further raised among those migraineurs with aura also carrying the MTHFR
677TT genotype. This was only seen for ischemic stroke and not for MI. With regard to the ACE
D/I polymorphism, only migraineurs with aura carrying the DD/DI genotype seemed to be at increased risk for ischemic stroke and MI, but not carriers of the II genotype.
The approach of our study is novel in two different ways. First, we compared migraineurs with CVD events to migraineurs without CVD events in contrast to previous studies comparing migraineurs to non-migraineurs. This design eliminates a potential confounding effect of migraine with gene variants on CVD events and specifically addresses the question which gene variants are associated with CVD events among migraineurs. Second, our study investigates the effect of genetic markers among migraineurs on their risk for CVD events from a genome-wide perspective.
None of the investigated SNPs was significantly associated with CVD events among migraineurs at a genome-wide level. However, our analyses provide some indication of an association of five genetic variants with CVD events at a suggestive significance threshold. In addition, our results—similar to previous non-genetic studies—support the concept that the risk for CVD among migraineurs is not strongly confounded by classical CVD risk factors. First, results from the multivariable-adjusted models are very similar to age-adjusted models (), although CVD risk factors are more prevalent among migraineurs with CVD events than among those without CVD events (). This is likely due to a co-linearity between age and classical CVD risk factors. Second, none of the five implicated genetic variants has previously been reported to be associated with CVD risk factors or CVD events.
The potential function for the five variants with regard to an increased CVD risk among migraineurs is unclear. A summary of the known functional characteristics of these variants is presented in . However, it is important to note, that four of the SNPs were identified for the association between migraine with aura and CVD events and two of those specifically for the association with ischemic stroke. This agrees with data from observational studies suggesting a particularly strong link between migraine with aura and ischemic stroke 
Functional characteristics of genetic variants implicated in the association with CVD among migraineurs (with p<5×10−6).
Our study has several strengths, including CVD events being verified by an endpoints committee of physicians, detailed information on many potential CVD risk factors, and genetic information from 339,596 SNPs covering the whole genome. In addition, the homogenous nature of the cohort, consisting only of white women aged ≥45 years, may reduce confounding. For example, while there is no reason to assume that migraine pathophysiology is different between women and men, the phenotypic expression among migraineurs may differ by gender and age. This is suggested, for example, by data showing that the migraine-ischemic stroke association is greater among younger women than older women 
, which may depend on a changing pattern in cardiovascular risk profile 
However, several limitations should be considered. First, and most importantly, data to directly replicate our results are not available to verify our results or perform a meta-analysis. However, we believe that this should not hold back research, since our results may stimulate new research ideas and accelerate ongoing research. Second, we cannot exclude the possibility of spurious associations. The number of women with migraine experiencing a CVD event was limited (n
164) and the odds ratios for the five SNPs are rather large with effect sizes between 3 and 12 compared to usually much smaller odds ratios found in association studies for complex and heterogeneous disorder. For rs7698623, rs1047964, and rs1406961 this may also in part be due to the low MAFs (all <10%; ). Hence, our data should be interpreted with caution. Third, migraine and aura status were self-reported and were not classified according to strict IHS criteria 
. Although previous reports from the WHS have shown good agreement of our classification with modified IHS criteria from 1988 
, and despite excellent agreement between self-reported migraine and migraine classification based on IHS criteria from 2004 in the WHS 
, non-differential misclassification is possible, which may in part explain some of our null findings. However, the prevalence of migraine (18.3%) and migraine aura (39.5% of migraineurs) is similar to those reported in other population-based studies 
. Finally, we only considered an additive genetic model of transmission. However, the vast majority of gene-phenotype associations follow an additive model, which would also have sufficient power to capture dominant modes of transmission. While recessive modes of transmission may be missed, we consider this a minor disadvantage because they are rare and mostly relevant for monogenic disorders.
Our results demand additional epidemiological studies and basic science research and should be considered hypothesis generating at present. First, large well-defined cohorts are needed with standardized information on migraine and aura status, gender, ethnicity, CVD risk factors, and other medical as well as genetic information, to verify the potential role of the gene variants for the CVD risk among migraineurs. In addition, further insight into the biological function of these genes may advance the understanding of the migraine-CVD link.