Correlation Between Tumor Necrosis Factor-α, Interleukin-1 And Interleukin-6 Serum Level And Pain Severity In Chronic Tension Type Headache Patients

CORRELATION BETWEEN TUMOR NECROSIS FACTOR-α, INTERLEUKIN-1 AND INTERLEUKIN-6 SERUM LEVEL
AND PAIN SEVERITY IN CHRONIC TENSION TYPE HEADACHE PATIENTS
Aldy S. Rambe*, Hasan Sjahrir*, Moh. Hasan Machfoed**, Darulkutni Nasution* *Department of Neurology University of Sumatera Utara, School of Medicine, Medan Indonesia **Department of Neurology Airlangga University, School of Medicine, Surabaya Indonesia
ABSTRACT
Aim : Experimental studies suggest that central sensititization plays important role in pathophysiology of chronic pain and chronic tension-type headache (CTTH). Studies on pro-inflammatory cytokines, interleukin (IL)-1β and tumor necrosis factor (TNF)-α serum levels during headache attack showed inconsistent results. This study was aimed to see the serum levels of TNF-α, IL-1, IL-6 in patients with CTTH and its correlation with pain intensity. Methodology : This cross sectional study was conducted at the Adam Malik Hospital and Bukit Barisan II Army Hospital from January 2013 to June 2014 after approved by the Ethics Committee for Health Research University of North Sumatera School of Medicine. The subjects were recruited consecutively from study population and agree to participate in this study. Diagnosis of CTTH was conducted through anamnesis, physical and neurological examination and based on the diagnostic criteria as stated in the ICH X. Subjects underwent physical and neurological examination, including assessment of pain intensity by using the Numeric Rating Scale ( NRS ) for pain. Furthermore, venous blood was taken to measure serum levels of TNF-α , IL-1, and IL-6. Results : The mean of TNF-α serum level is 1.81528 pg/dl (SD ± 0.890686), whereas the mean of IL–1 serum level is 0.20226 pg/dl (SD ± 0.193930), and the mean value of IL-6 serum level is 1.45089 pg/dl (SD ± 1.189502). There was very weak positive correlation (r=0.035) and non significant (p=0.806) between NRS scores and TNF-α serum level. There was non significant (p=0.793) and very weak negative correlation (r =-0037) between NRS

scores and IL-1 serum level. NRS scores have a very weak negative correlation (r=-0.107) and also not statistically significant (p=0.447) with IL-6 serum level. Conclusions : Previous studies showed positive correlation between NRS score and levels of TNF-α, IL-1 and IL-6. The higher NRS score, the higher serum level of TNF-α, IL-1 and IL6. This study shows weak negative correlation between NRS scores and IL-1 and IL-6 serum level. The positive weak correlation only found between NRS scores and TNF-α serum level. All of these correlation were statistically non significant. These findings perhaps because the sample from serum did not describe the biological mechanisms that occur in the brain.
Key words : chronic tension-type headache, cytokines, TNF-α, IL-1, IL-6

INTRODUCTON Tension-type headache (TTH) is the most common form of primary headache.
International Headache Society classification differs three forms of TTH mainly on the basis of headache frequency: (1) infrequent episodic tension-type headache (ETTH, less than 12 headache days per year), (2) frequent ETTH (between 12 and 180 days per year), and (3) chronic tension-type headache (CTTH, at least 180 days per year) 1. Chronic tension-type headache differs from the episodic forms not only in frequency but also with respect to pathophysiology, lack of effect to most treatment strategies, more medication overuse, more disability, and higher personal and socioeconomic costs 2 . Globally, the percentage of the adult population with an active headache disorder is 46% for headache in general, 11% for migraine, 42% for TTH and 3% for chronic daily headache.3
Recent advances in molecular neurobiology have increased our knowledge about mechanisms underlying this chronic headache. Experimental studies suggest that central sensititization plays important role in pathophysiology of chronic pain and CTTH. The discovery of neurotransmitters and neuromodulators, such as : nitric oxide (NO), calcitonin gene-related peptide (CGRP), substance P (SP), neuropeptida Y (NPY) and vasoactive intestinal polypeptide (VIP) that involved in the pain process has provided new insights to our understanding about the biological aspects of chronic headache.4
A number of studies have also measured the levels of cytokines in the blood in conjunction with headache, generally in migraine. Pro-inflammatory cytokines, interleukin (IL)-1β and tumor necrosis factor (TNF)-α were the most studied, where the levels during headache attack was found increasing or normal. Bo et al, studied the level of cytokines in cerebrospinal fluid (CSF) in headache patients and found the elevated levels of IL-1, TGF-b1 (transforming growth factor-b1), and MCP-1 (monocyte chemoattractant protein-1) on ETTH and migraine compared to controls, and there were significant differences in MCP-1 between cervicogenic headache and migraine without aura.5
Kocer found an increasing level of IL-6 in patients with ETTH and CTTH compared to controls. Therefore, they believe that IL-6 is involved in the induction of pain or inflammatory mechanisms in TTH.6 Backonja found an elevated receptor levels of TNF in CSF and blood, also found elevated level of IL-1β in CSF and associated with pain intensity, whereas IL-10 was inversely correlated with pain symptoms.7
Prior studies have found a positive relationship between the number of cytokines with some type of headache. Unfortunately, most measurements of cytokine levels were done in the CSF that is relatively difficult to do routinely in daily practice. This study was aimed to

measure the serum levels of TNF-α, IL-1, IL-6 in patients with chronic TTH and its correlation with pain intensity.
METHODS This was a cross sectional study that measure the levels of TNF α, IL-1 and IL-6 in
serum and its relationship with level of pain intensity in patients with CTTH. Research conducted at the Adam Malik Hospital and Rumkitdam II Bukit Barisan in
the period January 2013 - June 2014 after receiving approval from the Ethics Committee for Health Research School of Medicine in University of North Sumatera. The subjects were recruited consecutively from population and in writing willing to participate in this study. Diagnosis of CTTH was conducted through anamnesis, physical and neurological examination and based on the diagnostic criteria as stated in the ICH X. Subjects underwent physical and neurological examination, including determining the intensity of pain by using the Numeric Rating Scale ( NRS ) for pain. Furthermore, venous blood was obtained to measure serum levels of TNF-α , IL-1, and IL-6.
RESULTS At the beginning, there were 57 subjects met the study criteria. Four of them were

excluded from the study because they had never come for blood examination. Data from 53 subjects who completed the study procedure were analyzed. Characteristics of the subjects can be seen in Table 1.
Fifty three patients with CTTH who participate this study, consist of 8 men (15.1%) and 45 women (84.9%). Most of subjects (86.8%) were married. The majority of the study subjects ( 52.8 % ) came from Batak Toba tribe. The majority of the subjects (45.3%) worked as a civil servant. Twenty six subjects (49.1%) had a high school education background and only 4 subjects (7.5%) who had elementary education. Most subjects had no history of hypertension (81.1 %) , no diabetes mellitus (94.3%), non-smokers (96.2%) and not drug users (100%).

Table 1. Subject Demographics

Variable

Total

N (%)
Sex (%) Male Female
Marital Status (%) Married Not Married
Tribe (%) Batak Toba Mandailing Karo Javaneese Others
Occupation (%) Civil Servant Entrepreneur Housewife Others
Education (%) Primary School Junior High School Senior High School College
Hypertension Yes No
Diabetes Mellitus Yes No
Smoking Yes No
Drug Abuse
Yes

No

53 (100)
8 (15.1) 45 (84.9)
46 (86.8) 7 (13.2)
28 (52.8) 5 (9.4) 5 (9.4)
10 (18.9) 5 (9.4)
24 (45.3) 11 (20.8) 15 (28.3)
3 (5.7)
4 (7.5) 6 (11.3) 26 (49.1) 17 (32.1)
10 (18.9) 43 (81.1)
3 (5.7) 50 (94.3)
2 (3.8) 51 (96.2)
0 (0.0) 53 (100)

Table 2. Mean of age, TNF-α, IL-1 and IL- serum level

Age (years) NRS TNF-α (pg/dl) IL-1 (pg/dl) IL-6 (pg/dl)

Minimum 19 2 0.642 0.057 0.290

Maximum 65 8 4.499 1.307 5.530

Mean 46.04 4.72 1.84221 0.20170 1.51462

SD 10.751 1.725 0.866272 0.186891 1.224567

The mean age of the subjects is 46.06 years old (SD±10.751). NRS score range from 2 to 8 with mean score is 4.72 (SD±1.725). The TNF-α serum levels vary from 0.057 pg/dl to 4.499 pg/dl (mean 1.84221 pg/dl, SD±0.866272). The mean value of IL-1 serum level is 0.20170 pg/dl ((SD±0.186891). The minimum serum level of IL-6 was 0.290 pg/dl and maximum value was 5.530 pg/dl (mean 1.51462 pg/dl, SD±1.224567). (Table 2)
Since the data is normally distributed, we used the Pearson test to see the association between the NRS scores and of TNF-α , IL-1 and IL-6 serum level. We found that there were very weak positive correlation (r=0.035) and non significant (p=0.806) relationship between NRS scores and TNF-α serum level. There were also non significant (p=0.793) and very weak negative correlation (r =-0037) between NRS scores and IL-1 level. NRS scores have a very weak negative correlation (r=-0.107) and also not statistically significant (p=0.447) with IL-6 level. (Table 3)

Table 3. Correlation between NRS Scores with TNF-α, IL-1 and IL-6 Serum Level

TNF alfa

IL-1

IL-6

NRS R

0,035

- 0,037

- 0,107

Score P

0,806

0,793

0,447

N 53 53 53

Pearson Correlation Test

DISCUSSIONS Total subjects was 53, 84.9% ot them were female. These results were supported by
previous studies which TTH was more common in female than male.8,9 Study on chronic daily headache (CDH), which mostly consists of CTTH patients (70 %), found that ratio

between female : male = 5.6 : 1.8.8 One of TTH dissertation research shows that ratio between men and female = 83% : 17%.10 Although some studies show different result, it can be concluded that the prevalence of TTH is higher in female than male.8-10 Meta-analysis of some population-based studies show that prevalence between female : male = 1.3 : 1.11
Many factors influence higher prevalence of TTH in female than male : (1) differences in the perception of symptoms and sensation of pain; (2) behavior differences, (3) differences in personality traits and psychological; and (4) the effect of hormonal was higher in female than in male.11 An experimental pain studies in healthy people using a pressure algometer, concluded that female have a lower pain threshold than male.9 Female with TTH have lower tolerance for pain than male. Low tolerance is correlated with the frequency of headache.12 Hormonal factors also contributed to the occurrence of headache.13 Socioeconomic status plays an important role in coping strategy in TTH, and it differs by gender. Studies conducted by Chu concluded that female are more susceptible to socio-economic impact than male.14
In this study, the highest rate of tribe was Batak Toba. There were not many research study about relationship between pain and races (ethnic). Study by Plesh about prevalence of races/ethnic and gender in relation to general pain concluded that pattern of pain is vary widely between racial/ethnic and gender. Every pain has its own characteristics.15 A previous study found decreased in prevalence, incidence and persistence of pain in young black female than young white female.16 If only the frequency of these inter-ethnic can be analyzed, most likely the result would be not statistically significant. Unlike the black and white ethnicity that differ genetically and phenotypically, these tribes relatively has the same phenotype.
In this study, we found low frequency of hypertension (18.9%), diabetes mellitus (5.7%) and smoking (3.8%). Previous studies showed that these three risk factors play less important roles in TTH compare with as stroke risk factor. Most hypertension is rarely accompanied by other symptoms. Most people with high blood pressure complain about headache, especially on the back part of the head. Hypertensive patients will complain about headache usually when their blood pressure is high.17 Headache with severe hypertension, including hypertensive headache and not classified as primary headaches like TTH.
This test results found a non significant (p=0.806) and very weak positive correlation (r=0.035) between the NRS scores and TNF-α serum level. There were non significant (p=0.793) and very weak negative correlation ( r =-0037) between NRS scores and IL-1 level. NRS scores had a very weak negative correlation (r=-0.107) and not statistically significant with IL-6 level (p=0.447). Clinical significance of this fact showed that : (1) there

were weak negative correlation between NRS score with levels of TNF-α, IL-1 and IL-6; and (2) there was non significant correlation between NRS score with TNF-α, IL-1 and IL-6 levels.
The results are consistent with study by Rozen who found an increased of CSF TNF-α levels in patients with new daily persistent headache (NDPH) and migraine, but not in serum. Increased levels of CSF TNF-α found in almost patients with NDPH in Rozen’s study, indicating the role of TNF-α in the pathogenesis of this condition.18 Tanure et al compare the levels of TNF-α, TNF-α receptor 1 soluble (sTNFR1), TNF-α receptor 2 soluble (sTNFR2), and BDNF during attacks of migraine and headache-free period. They found non significant differences in the levels of TNF-α, sTNFR1 and sTNFR2 during migraine attacks and headache-free period.19
These results differ from study by Bo et al, who examined the cytokine levels in CSF patients with headache and found elevated levels of IL-1, TGF-b1 (transforming growth factor-b1), and MCP-1 (monocyte chemoattractant protein-1) the ETTH and migraine compared to controls, and there were significant differences in MCP-1 between cervicogenic headache and migraine without aura.5 Study by Backonja, 2008, also found an increased in of TNF receptors level in blood and CSF, as well as an increased in IL-1β level in CSF and associated with pain intensity, while IL-10 was inversely related to the symptoms of pain. The imbalance between pro- and anti- cytokine inflammation appears to be a relevant picture that contribute to persistence of chronic pain.7 Other study demonstrated that IL-1 and IL-6 lead to sensitization of trigeminal nociceptors and plays an important role in the pathogenesis of migraine by lowering the threshold of stimulation of other inflammatory stimuli.20 Based on these result, allegedly increased levels of IL-1 and IL-6 will facilitate an increase in the expression of proteins that generate signals in nerve cells and glial cells in ganglion and Trigeminal Nucleus Caudal, which will ultimately contribute to the peripheral and central sensitization, and plays an important role in the pathology of migraine.20 Kocer et al, also found an increased of IL-6 level in patients with ETTH and CTTH compared with controls. Therefore, they believed that IL-6 is involved in induction of pain or inflammatory mechanisms in TTH.6 It has been known that there was increased levels of IL-6 during attack in patient with migraine. Study by Yan et al, showed that IL-6 reinforce dural afferent nerve excitability resulting in sensitization that contribute to migraine pathogenesis. 21
Theoretically, and from several previous studies, there were positive correlation between NRS score and levels of TNF-α, IL-1 and IL-6. The higher NRS score pain intensity, the level of TNF-α IL-1 and IL-6 are also higher. In other words, these results were not in

accordance with the existing theory. In addition of this negative correlation, there was also non significant correlation between NRS score and levels of TNF-α, IL-1 and IL-6. This occurs because the sample from serum did not describe the biological mechanisms that occur in the brain.
CONCLUSIONS From all subjects, the mean value of TNF-α serum level is 1.81528 pg/dl (SD ±
0.890686), whereas the mean of IL–1 serum level is 0.20226 pg/dl (SD ± 0.193930), and the mean value of IL-6 serum level is 1.45089 pg/dl (SD ± 1.189502).
We found a very weak positive and non significant correlation between NRS scores and TNF-α level (r=0.035; p=0.806), and we also found non significant and very weak negative relationship between NRS scores and IL-1 (r=-0.037; p=0.793). NRS scores had a very weak negative relationship and also not statistically significant (r=-0.107; p=0.447).
Acknowledgement : Authors acknowledge the immense help receive from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors/editors/publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed.
Source of funding : All study expenses are covered by the authors themselves.
Conflict of interest : All authors have no conflict of interest in this study. Authors have no afiliation with the laboratory where the blood samples were analyzed.

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