Abstract
Migraine is a common complex disorder characterized by severe recurrent headache and usually accompanied by nausea and vomiting. Previous studies in our laboratory have utilized three large multigenerational Australian pedigrees affected with migraine to indicate that the disease is genetically heterogeneous, with linkage results implicating genomic susceptibility regions on both chromosomes 19p and Xq. The present study explores the possibility of a correlation between genetic and clinical heterogeneity in these affected pedigrees. Specifically, the clinical characteristics of migraine including subtype, age of onset, frequency, duration, and disease symptoms were compared between the migraine pedigrees, and gender differences were also assessed. Our exploratory analyses revealed no significant differences in any of the clinical characteristics tested between the chromosome 19-linked family and the two X-linked families. Also, we did not detect any differences in male vs. female clinical features for these pedigrees. In conclusion, migraine is considered to be a clinically and genetically heterogeneous disorder; however, our study provided no conclusive evidence that variation in genomic susceptibility region is related to heterogeneity at the clinical level in these migraine-affected pedigrees.
Introduction
Familial typical migraine is a common complex disease characterized by severe recurrent headache. The condition is usually also accompanied by symptoms such as nausea, vomiting, photophobia, and phonophobia. A significant proportion of the population is affected by migraine and the prevalence varies amongst males and females. As indicated by Launer et al., up to 25% of women vs. 8% of men suffer from migraine (1). Migraine shows strong familial aggregation, with first-degree relatives of sufferers having an overall increased risk of being affected compared with the general population (2). The disorder has been classified into two major subtypes; migraine with aura (MA) and migraine without aura (MO), although it is not yet clear whether these subtypes represent genetically distinct entities or are influenced by a common genetic background. Previous studies in our laboratory have provided strong evidence for the location of migraine genes on both chromosome 19p13 (3) and chromosome Xq24-28 (4). Four large multigenerational pedigrees (designated MF1, MF7, MF14, and MF15) were used in the initial linkage studies. These studies provided evidence that migraine is a genetically heterogeneous disorder with MF1 indicating that at least one gene for the disorder is located on chromosome 19p13 (Non-Parametric Linkage Score = 6.64, P = 0.0026) (3). Two of the pedigrees (MF7 and MF14) also provided evidence for the localization of a migraine susceptibility locus on chromosome Xq24-28 (combined LOD = 2.388, P = 0.0005) (4).
Several migraine case-control studies have reported an association between a particular migraine subtype (either MA or MO) and a candidate gene variant(s) (5–7). It is also worthwhile exploring the possibility of a relationship between migraine subtype and different genomic susceptibility regions, as implicated in affected pedigree linkage studies. Similarly, it is possible that different unlinked genetic variants may influence particular migraine characteristics such as frequency or duration of attack. Indeed, migraine case-control studies have already shown associations between polymorphisms in the angiotensin-converting enzyme and serotonin transporter genes and the frequency of migraine attack (8, 9). Furthermore, preliminary twin research has suggested that some migraine symptoms such as nausea and vomiting may also display partial heritability (10). With the identification of an X-linked component of migraine and given the female preponderance, it is also of interest to explore gender differences in linked pedigrees with respect to the clinical features that define the disease. Thus, this study was an exploratory investigation into clinical and gender differences between migraine families linked to chromosome 19 and chromosome X (MF1 and MF7/MF14, respectively).
Methods
Study subjects and variables
All individuals included in this analysis were affected with migraine and participated in the original genetic linkage studies (3, 4). These individuals gave informed consent before participating in the research and were all of Caucasian origin. Prior to the original linkage analyses, all individuals were personally interviewed by a qualified clinical neurologist and diagnosed as being affected with either MA or MO (Dr P. Brimage). All patient information was documented in questionnaires carefully designed and structured to diagnose migraine based on criteria specified by the International Headache Society (IHS) (11). Subsequent to the initial linkage analyses, all pedigree members were re-interviewed for possible changes in affection status by an independent qualified neurologist blinded to the original diagnostic and genetic data (Dr J. MacMillan). Seven individuals originally diagnosed as MO were found to have experienced symptoms associated with MA. These patients, who may represent cases of MA/MO co-occurrence or cases of complete transition from MO to MA, were all classified as being affected with MA for this analysis. In addition, 17 of the unaffected pedigree members have since reported symptoms of migraine. These individuals were also classified as either MA or MO according to the IHS diagnostic criteria (nine MA and eight MO).
The primary variables of interest in the statistical analyses were ‘migraine subtype’, ‘gender’, ‘age of onset’. Also ‘frequency of attack’ and ‘duration of attack’ were considered primary variables, where a patient's average migraine duration was a post-medication estimate. We also explored the possibility that differences in migraine patient symptoms may be influenced by pedigree-specific genetic susceptibility. The secondary variables were patient symptoms chosen for analysis since they are characteristic features of the different phases of disease progression and comprise an integral part of the current migraine diagnostic criteria (11, 12). These migraine symptoms were grouped into those that occurred prior to a headache (aura phase) and symptoms that occurred during a headache (headache phase). Aura symptoms that occurred usually up to 60 min prior to a headache included ‘visual disturbances’, ‘speech problems’ and ‘numbness and tingling’, while symptoms that occurred during the headache phase of a migraine included ‘nausea’, ‘vomiting’, ‘increased sensitivity to sound or light’, ‘increased sensitivity to smell’, ‘vision alteration’, ‘speech problems’ and ‘numbness or tingling’.
The differences in both the primary and secondary variables were compared statistically with respect to the genetically linked affected pedigrees. To allow comparison of clinical characteristics between the implicated susceptibility regions on chromosome 19 and X, clinical data for MF7 and MF14 were combined to represent the ‘X-linked pedigrees’ group. In addition, the differences in these variables were explored between males and females.
Statistical analysis
An exploratory data analysis was performed using clinical information for the three multigenerational pedigrees that previously provided linkage evidence to chromosome 19p13 (MF1) and chromosome Xq24-28 (MF7 and MF14). The count data were compared between the pedigrees using contingency table analysis. The standard χ2 test for association was implemented to compare ‘frequency’ and ‘duration of attacks’ and patient symptoms between the genetically linked pedigrees and between males and females. Fisher's exact test was applied in situations where sparse contingency tables deemed the standard χ2 test as inappropriate. In addition, the Cochran–Mantel–Haenszel statistic was calculated to investigate overall association whilst adjusting for migraine family.
Parametric methods to analyse the age of onset were deemed inappropriate by the Shapiro–Wilk test for normality. Therefore, the non-parametric Kruskal–Wallis test was applied to compare the age of onset of migraine between all migraine families. The difference between males and females with respect to age of onset was compared implementing the Wilcoxon rank sums test. To determine equal distribution of migraine subtypes across pedigrees and gender, the χ2 test goodness-of-fit or Fisher's exact test was applied. All statistical tests were based on exploratory two-sided hypotheses and were performed using SAS® software version 8 for windows. Probabilities calculated for each test are stated as raw P-values and the conventional α-level of 0.05 is used to highlight differences in the data. However, the threshold for claiming statistical significance was adjusted to α= 0.002 using Bonferroni correction based on 30 comparisons.
Results
Analysis of primary patient variables
Three multigenerational families (MF1, MF7, and MF14) were incorporated into the analyses for a total of 62 affected individuals. There were 27 migraineurs belonging to the chromosome 19-linked pedigree (MF1), of which 15 (55.6%) where diagnosed with MA. The X-linked pedigrees (MF7 and MF14) had a combined total of 35 migraineurs, of which 25 (71.4%) were diagnosed with MA (Table 1). Although the X-linked pedigrees had a higher proportion of MA sufferers, there was no statistically significant association between migraine subtype and genomic susceptibility region (as represented by migraine pedigrees showing linkage to chromosome 19 and X, respectively) (χ2 = 1.68 and P = 0.195). Similarly, gender proportions were not different between families showing X-linkage and the chromosome 19-linked pedigree (χ2 = 0.15 and P = 0.697). When looking at the distribution of migraine subtypes across gender for the individual migraine families, MF14 only showed some notable difference. This pedigree had an unequal proportion of females diagnosed with MA compared with males, although this was not statistically significant. It should be noted that, although these migraine subtype frequencies may be subject to some change due to the onset of certain patient symptoms subsequent to this analysis, we do not expect the proportional changes to alter these results significantly.
Primary clinical characteristics of migraine-affected pedigrees∗
The χ2 and Kruskal–Wallis sums tests indicated no significant differences (P > 0.05).
†The pedigrees showing X-linkage (MF1 and MF14) were combined for comparison with the chromosome 19-linked MF1).
The age of onset for all migraine families varied considerably, ranging from 4 to 50 years of age with a median age of 17. The median age of onset for MF1 and the X-linked pedigrees was 18 and 16 years, respectively (Table 1). The Kruskal–Wallis test was implemented to determine if these migraine families differed with respect to this variable. The analysis clearly indicated no significant difference (χ2 = 0.083, P = 0.773).
Fisher's exact test was implemented to determine if the variation in the duration of migraine was associated with genomic susceptibility region. This test produced a P-value of 0.546, which indicates that there was not a significant association between these variables (Table 2). Fisher's exact test was also applied to determine if males and females differ with respect to the duration of migraine for all pedigrees. This analysis produced a P-value of 0.549, indicating no significant difference between the males and females with respect to the duration of migraine in all pedigrees.
∗Fisher's exact test showed no significant difference between families for migraine duration (P > 0.546).
†The pedigrees showing X linkage (MF7 and MF14) are combined for comparison with the chromosome 19 linked, MF1.
‡Fisher's exact test showed no significant difference between male and female groups for duration of migraine (P > 0.549).
Frequency of migraine attacks was categorized into four possible groups: < 1 a month, 1–2 a month, 3–4 a month, or > 4 a month (Table 3). Fisher's exact test produced a P-value of 0.770, which also indicates that the frequency of migraine attack was not significantly associated with genomic susceptibility region. Fisher's exact test was also applied to determine if males and females differ with respect to frequency of migraine attacks for all migraine families. No significant difference between males and females was found (P = 0.687).
∗Fisher's exact test showed no significant difference (P > 0.770).
†The pedigrees showing X linkage (MF7 and MF14) are combined for comparison with the chromosome 19 linked, MF1.
‡Fisher's exact test showed no significant difference between male and female groups for frequency of migraine attacks (P > 0.687).
Analysis of secondary (symptom) variables
Analysis of the patient symptom variables was performed to examine potential differences between genomic susceptibility regions and gender (Tables 4 and 5). Of these variables, a substantial difference in the frequency of numbness and tingling between MF1 and the X-linked pedigrees was observed (P = 0.031), with the data suggesting that members of MF14, specifically, had a higher prevalence of ‘numbness and tingling’ prior to headache (47%) (P = 0.026). Although the counts were low, an over-representation of ‘increased sensitivity to smell’ in the chromosome 19-linked migraine pedigree (MF1) was also apparent (P = 0.034).
Frequencies of symptoms for migraine-affected pedigrees
∗P-values were calculated using χ2 test or Fisher's exact test where appropriate.
Frequencies of symptoms for gender between migraine-affected pedigrees∗
∗P-values were calculated using Cochran–Mantel–Haenszel statistics. If Cochran–Mantel–Haenszel statistic was deemed not appropriate due to insufficient data or non-homogeneous association across strata, Fisher's exact test was applied to pedigrees.
When looking at symptom differences for gender among and between families (Table 5), MF14 again showed evidence of a higher occurrence of visual disturbances prior to a migraine headache in females compared with males (P = 0.013). Further gender difference analysis indicated that males and females differ with respect to vomiting during a migraine headache. Specifically, these data suggest that females have an over-representation of ‘vomiting’ after adjusting for all migraine families (P = 0.020). Despite these notable differences in patient symptoms between pedigree and gender, no result was statistically significant at the adjusted threshold of α= 0.002.
Discussion
This study was carried out to determine if certain clinical characteristics of migraine patients, such as frequency and duration of attack, tend to be different between families showing chromosome 19 linkage compared with those with a chromosome X component. For all pedigrees, comparisons were also conducted to determine if males and females differ with respect to these clinical variables. The association analyses indicated that there were no significant differences in migraine subtype, gender, age of onset, and frequency and duration of attack between our genetically linked migraine families. In addition, there was no indication that males and females differ significantly with respect to duration and frequency of attack in these pedigrees.
The association analyses of specific patient symptoms indicated a notable over-representation in the frequency of sensitivity to smell in MF1. Apparent differences in numbness and tingling prior to migraine headache were also found between the pedigrees, largely due to MF14. In addition, males and females in MF14 were shown to differ with respect to visual disturbances that occurred prior to migraine headache and vomiting that occurred during migraine headache. It should be noted that these differences were not statistically significant at the specified α level and could therefore represent chance occurrences or small sample bias. Nevertheless, given MF14 had an unequal proportion of females diagnosed with MA and visual disturbances are symptoms indicative of MA, the differences found in MF14 may reflect the difference in the proportions of female MA vs. males with MA. It is possible that the MA differences have also contributed to the overall difference in the level of ‘numbness and tingling’ across the pedigrees.
Given the lack of statistical significance for any tested variable, we conclude that, despite some interesting differences, there was no conclusive evidence that clinical characteristics differed between the genetically linked migraine families examined in this analysis. Thus, this study did not provide evidence for a relationship between migraine genetic and clinical heterogeneity. Also, we did not detect any significant differences in clinical characteristics for male vs. female groups in these migraine families.