Low serum 25-hydroxyvitamin D is associated with higher risk of frequent headache in middle-aged and older men

Vitamin D has been suggested to have a role in various neurovascular diseases, but the data regarding headache is inconclusive. Our aim was to investigate the associations between serum 25-hydroxyvitamin D [25(OH)D], a marker for vitamin D status, and risk of frequent headache. The study population consisted of 2601 men from the population-based Kuopio Ischaemic Heart Disease Risk Factor Study (KIHD) from eastern Finland, aged 42–60 years in 1984–1989. The cross-sectional associations with prevalence of self-reported frequent headache (defined as weekly or daily headaches) were estimated with multivariable-adjusted odds ratios. The average serum 25(OH) concentration was 43.4 nmol/L (SD 18.9, min-max 7.8–136.1 nmol/L). A total of 250 men (9.6%) reported frequent headache. The average serum 25(OH)D concentration among those with frequent headache was 38.3 nmol/L (SD 18.8) and 43.9 nmol/L (SD 18.9) among those without frequent headache, after adjustment for age and year and month of blood draw (P for difference <0.001). After multivariable adjustments, those in the lowest vs. the highest serum 25(OH)D quartile had 113% (95% CI 42, 218%; P for trend <0.001) higher odds for frequent headache. In conclusion, low serum 25(OH)D concentration was associated with markedly higher risk of frequent headache in men.

In three case-reports, supplementation with vitamin D and calcium reduced migraine attacks [12][13][14] . However, results from larger, randomized, placebo-controlled vitamin D supplementation trials have been mixed 15,16 . Recently, combined therapy with simvastatin and vitamin D 3 reduced the number of days with migraine 17 . This finding supported the previous cross-sectional observation from the same study group that statin use was associated with lower odds for severe headache or migraine only among those with higher serum 25(OH)D concentrations 18 .
Overall, the study findings are inconsistent and many studies are small in size. Therefore, the purpose of our study was to investigate the associations between serum 25(OH)D and risk of frequent headache among 2601 middle-aged and older men from a population-based cohort study from eastern Finland.

Methods
Study Population. The Kuopio Ischaemic Heart Disease Risk Factor Study (KIHD) was designed to investigate risk factors for cardiovascular disease, atherosclerosis, and related outcomes in a population-based, randomly selected sample of men from Eastern Finland 19 . The baseline examinations were carried out in 1984-1989 for a total of 2682 men who were 42, 48, 54 or 60 years old (83% of those eligible). The baseline characteristics of the entire study population have been described previously 19 . The KIHD study protocol was approved by the Research Ethics Committee of the University of Kuopio. All subjects gave written informed consent for participation. All methods were performed in accordance with approved guidelines and regulations. Subjects with missing data on serum 25(OH)D concentration (n = 53) or headache (n = 28) were excluded, leaving 2601 men for the analyses.

Measurements.
Fasting venous blood samples were collected between 8AM and 10AM at the baseline examinations. Subjects were instructed to abstain from ingesting alcohol for three days and from smoking and eating for 12 hours prior to giving the sample. Detailed descriptions of the assessment of medical history and medications 20 , smoking 20 , alcohol consumption 20 , blood pressure 20 , and physical activity 21 at baseline have been published. Education was assessed in years by using self-administered questionnaire. Annual income was obtained from a self-administered questionnaire. Hypertension diagnosis was defined as systolic/diastolic blood pressure > 140/90 mmHg or use of hypertension medication. Body mass index (BMI) was computed as the ratio of weight in kilograms to the square of height in meters. Serum 25(OH)D 3 was measured after hexane extraction with high-performance liquid chromatography using coulometric electrode array detection, as previously described 22 . Diagnostic Criteria for Frequent Headache. Headache diagnosis was based on the response to the question in the study questionnaire: "Have you had headache during the previous 12 months?" The choices were "not at all", "less than once per month", "monthly", "weekly" and "daily". In the current study, those with weekly or daily headache were classified as having frequent headache.
Statistical Analysis. The univariate relationships between serum 25(OH)D and characteristics of the study population were assessed by means and linear regression (for continuous variables) or χ 2 -tests (for categorical variables). The mean serum 25(OH)D concentration in the subjects with frequent headache and in the rest of the study population were analyzed using analysis of covariance (ANCOVA), with adjustments for potential confounders. Multivariate-adjusted logistic regression was used to estimate odds ratios (OR) in quartiles of serum 25(OH)D, with the highest category as the reference. The model 1 included age (years), examination year and examination month. The Model 2 included the variables in the Model 1 plus body mass index (kg/m 2 ), systolic and diastolic blood pressure (mmHg), smoking (never smoker, previous smoker, current smoker < 20 cigarettes/d and current smoker ≥ 20 cigarettes/d), education years, income (euros), leisure-time physical activity (kcal/d), self-reported health (extremely good, quite good, average, quite poor, extremely poor), use of analgesics (drugs for headache, back pain, joint pain or other pain, opioids, or acetylsalicylic acid), and alcohol intake (g/d). All quantitative variables were entered in the models as continuous variables. Further adjustments for disease history (coronary heart disease, stroke, cancer, type 2 diabetes), number of concurrent medications used, or use of cardiovascular disease drugs that could potentially be associated with headache (beta-blockers, calcium channel blockers, angiotensin-converting-enzyme inhibitors) did not change the associations (< 5% change in estimates). Cohort mean was used to replace missing values in covariates (< 2.3%). Tests of linear trend were conducted by assigning the median values for each category of exposure variable and treating those as a single continuous variable. All P-values were 2-tailed (α = 0.05). Data were analysed using SPSS 21.0 for Windows (Armonk, NY: IBM Corp.). Data availability. The KIHD data is not publicly available.

Results
Population Characteristics. The characteristics of the study population according to the serum 25(OH) D quartile are shown in the Table 1. Those with a lower serum 25(OH)D concentration were more likely to be younger, have lower leisure-time physical activity, lower income and lower self-reported health. They were also more likely to be smokers.
In the whole study population, the average serum 25(OH)D concentration was 43.4 nmol/L (SD 18.9, min-max 7.8-136.1 nmol/L). Among the subjects, 67.9% had the serum 25(OH)D concentration < 50 nmol/L and 6.9% had concentration ≥ 75 nmol/L. The average serum 25(OH)D concentration among those with frequent headache was 38.3 nmol/L (SD 18.8) and 43.9 nmol/L (SD 18.9) among those without frequent headache, after adjustment for age and year and month of blood draw (p for difference < 0.001).
Scientific RepoRts | 7:39697 | DOI: 10.1038/srep39697 The proportion of subjects with frequent headache was lower among those men whose blood sample was drawn during the high UVB exposure months, i.e. between June and September (45 out of 626 men, 7.2%) than among the other men (205 out of 1975 men, 10.4%) (p for difference = 0.018).

Associations of serum 25(OH)D with risk of frequent headache.
Among the 2601 men, 250 men (9.6%) reported frequent headache. After adjustment for age and year and month of blood draw, those in the lowest serum 25(OH)D quartile had 116% higher odds (OR 2.16; 95% CI 1.49-3.13) for frequent headache (Model 1, Table 2). Further adjustments for potential confounders did not appreciably change the associations (Model 2, Table 2). However, although the p-value for trend across quartiles was statistically significant, the increased odds for frequent headache was present only in the lowest serum 25(OH)D quartile.

Discussion
In this cross-sectional population-based cohort study among middle-aged men from eastern Finland, low serum 25(OH)D concentration was associated with a higher risk of frequent headache.
The role of vitamin D in headaches remains unclear, mainly due to predominantly small studies with inconsistent findings [8][9][10][11][12][13][14][15][16][17][18] and the lack of large randomized supplementation trials that would show benefit after improvement of vitamin D status with vitamin D supplementation 23 . Our study, being one of the largest studies that have investigated the issue, supports the view that vitamin D may be beneficial in headache prevention. Besides the size of the study, the strengths of our study include its population-based recruitment and the extensive examinations for potential confounding factors.
One interesting issue is that how vitamin D can reduce the occurrence of headaches? The classical view suggests that the main signalling effect of vitamin D is mediated by its active metabolite 1,25(OH) 2 Table 2. Odds ratios (95% confidence intervals) of self-reported frequent headache in 2601 men according to serum 25-hydroxyvitamin D concentration. Model 1 is adjusted for age, examination year and month of blood draw. Model 2 is adjusted for Model 1 plus body mass index (kg/m 2 ), systolic and diastolic blood pressure (mmHg), smoking (never smoker, previous smoker, current smoker < 20 cigarettes/d and current smoker ≥ 20 cigarettes/d), education years, income (euros), leisure-time physical activity (kcal/d), self-reported health (extremely good, quite good, average, quite poor, extremely poor), use of analgesics (drugs for headache, back pain, joint pain or other pain, opioids, or acetylsalicylic acid), and alcohol intake (g/d).
regulation of transcription 24 . The regulation of transcription could include expression of 'protective genes' , such as the anti-inflammatory cytokine IL-10 25 or suppression of pro-inflammatory or pro-nociceptive genes, such as IL-1beta, TNF-alpha and others 23 . Vitamin D receptor (VDR) and the vitamin D metabolizing enzymes are expressed in nociceptive sensory neurons 26 , consistent with pro-nociceptive signalling in deficiency of vitamin D 27 . Vitamin D deficiency has also been associated with chronic tension-type headache, perhaps by causing musculoskeletal pain 28,29 .
Recently, activation of various transcriptional factors was linked also with fast non-genomic effects within the nociceptive system 30 . This is likely applicable also to signalling initiated by the vitamin D 25 . Thus, a rapid inhibitory effect of 1,25(OH) 2 D on ATP-gated P2X7 in human immune cells was reported 31 . Notably, P2X7 receptors expressed by almost all types of immune cells, including brain-residing microglial cells, are triggering the pro-inflammatory and pro-nociceptive cascades leading to chronification of pain 32 .
A major limitation of the current study is the cross-sectional design, so we could not assess temporal relationship or the direction of the association. For example, those who suffer from frequent headaches may be less likely to spend time outdoors and would thus be less exposed to the UVB light. However, this may not be that relevant in Finland and in other regions where UVB exposure is strong enough for vitamin D formation in the skin only during few months of a year. Also, despite extensive adjustment for potential confounders, we cannot fully exclude the possibility of residual confounding. Another limitation is the small proportion of subjects with higher serum 25(OH)D concentrations. The risk of headache may have been stronger, if the contrast in the serum 25(OH)D concentrations were larger. Our study also included only middle-aged men, so the findings may not be generalizable to other age-groups or to women. Finally, the outcome definitions were based on self-report, and we did not have information on the type of headache.
In conclusion, the results of this study in a Finnish population suggests an association between the serum 25(OH)D and risk of frequent headache. Large randomized vitamin D supplementation trials are needed to elucidate the role of vitamin D supplementation as a prophylaxis or treatment for headache.