Dr. Takao Takeshima, Associate Professor of Neurology, Department of Neurology, Institute of Neurological Sciences, Faculty of Medicine, Tottori University, 36-1 Nishicho, Yonago, 683-8504, Japan.
Background and Objective.— Cortical spreading depression and neurogenic inflammation have been hypothesized to be key steps in the development of migraine headache. Recent studies have highlighted matrix metalloproteinase-9 (MMP-9) in cortical spreading depression, neurogenic inflammation, and cerebral ischemia. To seek their possible association, we investigated plasma MMP-9 levels in migraineurs during headache-free periods.
Methods.— Plasma MMP-9 levels in 84 migraine subjects and 61 controls were determined by enzyme-linked immunosorbent assay. In addition, 23 patients with tension type headache were included in the study as comparative subjects.
Results.— The MMP-9 levels in migraineurs (42.5 ± 4.6 ng/mL, mean ± SE) were significantly higher than those in controls (25.4 ± 2.7 ng/mL, P < .005). Those levels in tension type headache subjects (24.6 ± 4.8 ng/mL) did not differ from those in controls. There was no significant difference between subjects having migraine with aura and those without aura. The MMP-9 levels did not correlate with age, duration of illness, frequency of migraine attack, duration of headache attack, or medication for headache. Mean plasma MMP-9 levels were the highest in subjects from whom blood samples were taken 2-4 days after their latest attack.
Conclusions.— The degradation of extracellular matrix showing the increase of MMP-9 in migrainurs may be associated with an abnormality in their blood vessel permeability. MPP-9 plays some role in migraine pathophysiology. Further studies of MMPs are necessary to elucidate their role.
Migraine headache, a highly prevalent and disabling disease, is characterized by the peripheral and central sensitization of pain perceptive systems. Although the pathophysiology is not fully under stood, cortical spreading depression (CSD) and neurogenic inflammation have been hypothesized to be major steps in the development of migraine headache.1,2 Recent studies suggests possible association of migraine and ischemic brain lesions, especially posterior lobes and cerebellum.3 The matrix metalloproteinases (MMPs) are a family of enzymes with more than 20 members identified to date that are all extracellular endopeptidases requiring Zn2+. The major targets are the proteins of the extracellular matrix (ECM).4 The role for MMPs has also been suggested in the pathogenesis of both acute and chronic neurological disorders such as stroke, Alzheimers's disease, HIV-associated dementia, and multiple sclerosis.5-7
Matrix metalloproteinase-9 (MMP-9 or gelatinase B) is a relevant member of MMPs and has a broad range of specific substrates such as gelatine, collagens, and fibronectin.6 Significant pathological roles of MMP-9 have been reported in cerebral ischemia and neural inflammation.8-10
Cortical spreading depression is a propagating wave of neuronal and glial depolarization in cerebral cortex and has been implicated in disorders of neurovascular regulation such as stroke, head trauma, and migraine. Gursoy-Ozdemir et al11 demonsrated CSD increased the cortical tissue MMP-9 level, in animal models and altered the levels of inflammatory cytokines. We have reported alteration of plasma transforming growth factor-β, a relevant cytokine, in interictal migrainures.12 Alterations of plasma interleukins have also been reported in migrainures.13
Changes of plasma MMPs in patients have been reported in some neurological disorders such as ischemic stroke14 and multiple sclerosis.15 However, there has been no study concerning plasma MMPs in migraine subjects. To explore the possible association of the MMPs in migraine, we measured plasma MMP-9 in migraineurs during the headache-free periods.
SUBJECTS AND METHODS
Eighty-four volunteer migraineurs, who visited a headache clinic at Tottori University Hospital, participated in this study. All subjects were given a general physical and neurological examination. The diagnosis of headache type was established according to the diagnostic criteria of the International Classification of Headache Disorders II.16 Twenty-one subjects suffered from migraine with aura and 63 from migraine without aura. All headache sufferers were generally normal except for their headaches. We recruited 61 healthy subjects without headache as controls. They comprised 58 volunteers recruited from hospital workers, 2 students of the University, and 1 family member of a patient, who were generally normal and received no medication. In addition, 23 patients with tension type headache were included in the study as comparative subjects. Mean age and male to female ratio of the subjects are shown in Table 1. Clinical characteristics and the current medication of migraine subjects are summarized in Table 2. All participants gave their informed consent following their full understanding of the nature and aim of the study. We obtained venous blood samples from each participant during the headache-free period, at least 2 days after the last headache attack. Sample blood was cooled immediately in EDTA-containing tubes. We obtained plasma by centrifugal separation. The plasma samples were frozen and stored at −30°C until assay. MMP-9 levels in plasma were determined by enzyme-linked immunosorbent assay (MMP-9, human, Biotrak ELISA System; Amersham Bioscience, Piscataway, NJ, USA). Statistical analyses were performed using SPSS software version 11.01-J (Tokyo, Japan). The data were evaluated using the non-parametric Kruskal-Wallis test followed by Mann–Whitney U-test. The Spearman coefficient was used to evaluate correlations between variables.
Table 1.—. Mean Age and Sex Ratio of the Subjects
Male : female
Mean ± SD.
32.5 ± 11.1
33.6 ± 13.0
Migraine with aura
27.0 ± 8.2
Migraine without aura
36.0 ± 13.7
51.3 ± 18.2
Table 2.—. Headache Characteristics and Medication of Migraine Subjects
Migraine with aura (n = 22)
Migraine without aura (n = 62)
Total (n = 84)
Mean ± SD.
NSAID = non-steroidal anti-inflammatory drug.
Age of onset (years)
17.1 ± 8.7
23.0 ± 8.6
21.5 ± 9.0
Duration of illness (years)
9.2 ± 8.6
12.9 ± 10.7
12.0 ± 10.3
Frequency of headache (per month)
3.4 ± 2.8
4.5 ± 4.1
4.2 ± 3.8
Duration of headache (hours)
17.8 ± 11.3
23.6 ± 15.9
22.1 ± 15.0
Medication for headache (%)
No acute medication
The plasma MMP-9 levels in controls, migraine, and tension type headache subjects are presented in Table 3. The MMP-9 levels in migraineurs were significantly higher than those in controls. Those levels in tension type headache subjects did not differ from those in controls. There was no significant difference between MMP-9 levels in migraine subjects with aura and those having migraine without aura. The MMP-9 levels did not correlate with age, duration of illness, frequency of migraine attack, duration of headache attack, or medication for headache. Mean plasma MMP-9 levels were the highest in subjects from whom blood samples were taken 2-4 days after their latest attack (Figure).
Table 3.—. Plasma MMP-9 Levels
P < .005 compared with controls,
P < .05 compared with controls.
Mean ± SE; Kruskal-Wallis test, P < .01; Mann-Whitney U-test.
This study demonstrated that plasma MMP-9 levels were increased in migraineurs during the headache-free periods. Although there might be some possibility that high MMP-9 is a preparative state for a migraine attack or that migraine attacks cause an increase of MMP-9, the elevation of plasma MMP-9 levels seemed to be a consequence of migraine attack, because they were associated with the period from the latest headache attack in our findings.
Matrix metalloproteinases have been shown to contribute to: (1) the breakdown of the blood–brain barrier (BBB); (2) cytokine production and the propagation of an inflammatory response including demyelination; (3) tumor invasion, metastasis, and angiogenesis; and (4) inappropriate degradation of the ECM leading to an alteration of structural integrity in various diseases.6 Since neurogenic inflammation is a key feature of migraine, increased MMP-9 levels in migraineurs might play an important role in its pathophysiology. Acute attack of multiple sclerosis, which is a demyelinating disorder associated with neurogenic inflammation, causes an elevation of plasma MMP-9 levels.15,17 Stimulation with lipopolysaccharide is known to mimic the neurogenic inflammation in experimental animal models.18 Lipopolysaccharide can enhance MMP-9 mediated by activation for inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β).
Cortical spreading depression is a neural phenomenon associated with the migraine aura, such as fortification spectra. CSD in rat models increases the production of TNF-α and IL-1β in the brain tissure.19 Gursoy-Ozdemir et al demonstrated that CSD upregulates MMP-9 in the brain, and then alters the permeability of BBB in murine models.11 Mean plasma MMP-9 level in migraine with aura was significantly higher than in the controls. It was unexpectedly lower than in migraine without aura, although the difference was not statistically significant. Plasma MMP-9 levels in migraine with aura were not higher than those in migraine without aura in our clinical study. There are some opinions that CSD might even occur in the brains of patients with migraine without aura. Premonitory symptoms such as change of mood or feeling of hunger – the so called clinically silent aura – often appear before migraine attack, when a change of neuronal activity occurs in areas that do not present any neuronal symptoms. If we hypothesize that silent CSD occurs in migraine without aura, this facilitates an understanding of the high MMP-9 levels seen in subjects with migraine without aura in this study.
Previous reports have shown that plasma MMP-9 is increased in patients with ischemic stroke,9 and that MMP-9 promoted injury of BBB.8 In an animal model of brain ischemia, activation of MMP-9 appeared in endothelial cells 48 hours after reperfusion following the release of TNF-α and IL-1β. Activated MMP-9 causes the disruption of BBB.20 Some recent reports have claimed that migraine is a risk factor for cerebral ischemia. It has been reported that subjects with migraine show a higher prevalence of white matter lesions in their brain as demonstrated by MRI.21 Elevated plasma MMP-9 levels in migraine may be related to proteolytic degradation of the BBB. Increased MMP-9 in migraineurs may support the hypothesis that migraine may share a pathophysiology with ischemic stroke.
The origin of MMP-9 in migraineurs is unclear. In ischemic stroke, endothelial cells in cerebral vessels seem to produce MMP-9.20 Astrocytes and microglia are demonstrated to be associated with the induction of MMP-9 in a culture model of inflammatory stimuration.10 From the point of view of the relationship between migraine and neurogenic inflammation or CSD, it may be hypothesized that increased plasma MMP-9 in migraineurs could be derived from the brain tissue, but systemic ECM. However, this view remains speculative.
As far as we are aware, this is the first report investigating increased plasma MMP-9 concentration in migraine subjects. We found that plasma MMP-9 levels in migraineurs were increased during headache-free periods. The increased level of MMP-9 in migraineurs may be associated with the characteristics of their blood vessels. Our results are an important contribution to the understanding of migraine headache. Further studies of MMPs in migraine are necessary to fully elucidate their role.
Acknowledgment: This study was partly supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science, and Culture of Japan (K.I. and T.T.).