Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection

Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection Afrah Abdulsahib Abbas, Fawaz Aswad This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6227107/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Migraine is a highly prevalent and disabling disorder. Because stress appears to be a prominent trigger of this condition. the aim of this study was to evaluate the therapeutic effects of botulinum toxin injection in migraine and tension headache patients and to estimate salivary cortisol, salivary alpha amylase in these groups of patients before and after treatment with botulinum toxin injection. A total of 45 subjects were divided into two groups 25 patients with chronic migraine and 20 patients with chronic tension headaches. The salivary biomarker (salivary cortisol, salivary alpha amylase) was detected by enzyme-linked immunosorbent assay (ELISA) kits. The result showed the increased statistically significant (p < 0.05) in salivary cortisol, and decreased statistically significant (p < 0.05) in salivary alpha amylase after intervention with botulinum toxin in both study groups (chronic migraine and chronic tension headaches). Salivary amylase decreased significantly after therapy in both groups (migraine and tension headache). Salivary cortisol increased significantly after therapy. In addition, Nausea and vomiting responded best to therapy, followed by orofacial discomfort. Migraine Headache Tension Headache cortisol salivary alpha amylase Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction Headaches are a frequent symptom of central nervous system convergence [ 1 ]. Sensitization and activation of trigeminal ganglion components may contribute to headache and facial pain problems [ 1 ]. Migraine is the most researched headache condition. Migraine and other headache problems may be more prevalent in orofacial pain (OFP) patients [ 2 ]. Chronic orofacial pain may be neuropathic, neurovascular or musculoskeletal, musculoskeletal pain involves tempromandibular joint and masticatory muscles [ 3 ]. TTH is the most frequent main headache, with a lifetime frequency of 52% compared to 18% for migraine. Chronic headache affects 1.7–4% of adults worldwide. Headache diseases impact individuals of all ages, ethnicities, economic levels, and regions worldwide. Child headaches are infrequent but grow with age. Adolescence, especially in childbearing women, increases headaches and migraines [ 4 ] . The neurotoxic protein botulinum toxin (BTX) is generated by the bacteria Clostridium botulinum and related species. BTX inhibits the release of the neurotransmitter acetylcholine from axon terminals at the neuromuscular junction. This effect paralyses the muscles [ 5 ]. Botulinum toxin type A (BTX-A) is utilised in TTH therapy. Wiliam Binder discovered its therapeutic benefit in alleviating headaches [ 6 ]. He reported a reduction in migraine recurrence after a cosmetic injection of BTX-A to diminish forehead creases. Saliva has long been thought to be a mirror of the body since it includes the serum elements which are evaluated in standard blood tests for disease and health monitoring, saliva as a diagnostic fluid provides a lot of advantages over serum for disease diagnosis [ 7 ]. Salivary alpha-amylase (sAA) has evolved as a viable and reliable marker of ANS activity in stress studies during the last 15 years. Alpha-amylase is a salivary enzyme that aids in carbohydrate and starch digestion. It can be measured quickly and non-invasively through saliva collection, and as such, it is frequently used as a proxy measure of sympathetic arousal in the acute stress literature, particularly in studies looking to reduce participant load or when paradigms require participants to collect samples at home [ 8 ]. The most abundant protein in saliva is alpha amylase(sAA), accounting for approximately 20% of salivary proteins [ 9 ]. Cortisol is the primary glucocorticoid hormone released by the adrenal cortex. Cortisol is a steroid hormone based on its chemical structure. Cortisol influences carbohydrate, lipid, and protein metabolism. Cortisol is involved in the stress response and is required for an organism to operate properly. Corticosteroids are a class of hormones secreted by the adrenal gland cortex. Corticosteroids are classified into three types based on their primary physiological influence: glucocorticoids (cortisol and corticosterone), mineral corticoids (aldosterone), and androgen hormones (dehydro-epi androsterone and androstenedione). Cortisol is tested in blood and urine, and more recently, in instances when a blood sample is difficult to obtain, cortisol has been detected in saliva [ 10 ]. Material and Methods Field research Total subjects composed of forty-five participants were recruited and concentrated for this study. They were carefully informed about the aim of the investigation and they were free to accept or refuse to participate. A longitudinal study was done in Al-Imam Alsadiq General Hospital in Hilla City and the laboratory work was done in the lab of the hospital. Approval was obtained from Ministry of Health for patient's Subjects: The participants were examined from the period of June - 2022 to October-2022.The study protocol was approved by oral medicine department, college of Dentistry, Baghdad University. The sample was obtained from 45 subjects who were divided into two groups: Group 1: Twenty-five patients with chronic migraine. Group 2 : Twenty patients with chronic tension headaches. Inclusions criteria Good general health, age≥18years. Participants had also to be free off ever/or cold Exclusion criteria Fibromyalgia, rheumatoid arthritis, whiplash-associated disorder, neurological disorders, pregnancy or lactation, high blood pressure, tobacco usage , or muscle relaxants , a known BTX allergy; and oral complaint , such as oral dryness or mucosal lesions, were excluded from further involvement in the study. Saliva collection The whole sample of saliva was taken from patients in the morning from 9 a.m to 10 a.m patient come fasting and then asked to collect sample of Saliva after an individual was asked to rinse his/her mouth thoroughly with distal water to insure the removal of any possible debris or contaminating materials. The first mouth-full of saliva was discarded to allow clearance of water then they were asked to collect all the saliva until the recommended amount of saliva was obtained pre and post botox injection by one month were done 10 millilitres of unstimulated whole saliva was collected ..The collected saliva was centrifuged at 4000 r.p.m (rotation per minute) for 10 minutes and then centrifuged clear supernatant saliva kept frozen and stored at -20 °C until the whole saliva samples were collected , then SAMY1 and COR determination done using Human salivary alpha-amylase ELISA kit and Human cortisol ELISA kit. BTX injections In this study, onabotulinumtoxinA (OBTA) (nabota) was used for prophylaxis of chronic migraine and chronic tension headache, and this type of botulinum toxin was FDA-approved in February 2019.One handful of MU of onabotulinumtoxin A (nabota) is diluted with 2.5 ml of saline, yielding a concentration of 5 MU per 12.5 ml.Each intramuscular injection site is injected with 5 MU of onabotulinumtoxinA.The injection paradigm for chronic migraine consists of 31 fixed sites in the following muscles: mm. frontalis 20 MU (four sites), mm. corrugatores 10 MU (two sites), m. procerus 5 MU (1 site), mm. occipitalis 30 MU (six sites), mm. temporalis 40 MU (eight sites), mm. trapezii 30 MU (in six sites), cervical paraspinal muscle Group 20 MU (four sites) In these fixed sites, a total dose of 155 MU of onabotulinumtoxin A is applied. The injection paradigm for chronic tension headache consists of 10 fixed sites in the following muscles: mm. frontalis 30 MU (six sites),, mm. corrugatores 10 MU (two sites),, mm. procerus 5 MU (1 site),, and in the middle of the forehead 5 MU (1 site).. In these fixed sites, a total dose of 50 MU of onabotulinumtoxin A is applied. Constant site, constant dose injection scheme applied in PREEMPT studies [ 11 ]. Statistical Analysis Statistical analyses were done using IBMSPSS version 27 computer software (IBM Statistical Package for Social Sciences) in association with Microsoft Excel. The Wilcoxon Signed Ranks Test determined the statistical significance of the average (median) change in a quantitative outcome following intervention. A P value below 0.05 indicated statistical significance. All significance tests were bilateral. Kruskal-Wallis Test compared medians of non-normally distributed variables between more than two groups. ANOVA analysed the statistical significance of the mean difference of a continuous response variable that is known or presumed to be regularly distributed (like age) between more than two groups. Ethical approval The Ethics committee, College of Dentistry, University of Baghdad, gave ethical approval for this study . The study was conducted in accordance with the ethical principles that have their origin in Declaration of Helsinki . it was carried out with patients , verbal and analytic approval before the sample was taken , the study protocol and the subject information and consent form were reviewed and approved by local ethics committee to the document number ( Ref # 459, date : January 19, 2022 ) to get this approval . Results Migraine headaches group After obtaining informed permission, 25 people with a confirmed Migraine diagnosis were enrolled in the study. According to Table 1, the bulk of the Migraine group (88%) were females, and 40% were 30 years old, with just 20% being 40 years old. Table 1 : Frequency distribution of migraine headache study group by age and sex. N % Gender Female 22 88.0 Male 3 12.0 Total 25 100.0 Age (years)-categories <30 10 40.0 (30-39) 10 40.0 40+ 5 20.0 Total 25 100.0 Range = 20-25 Mean = 32.6 SD = 8.1 The result in table 2 showed, among the migraine group 60% complained of associated TMJ problems and about three quarters (76%) complained of paresthesia, 80% with orofacial pain, Everybody (100%) of the recruited study group reported nausea or vomiting. Table 2: The relative frequency of selected clinical features at baseline (before intervention) among migraine headache study group. (Total N=25) N % Orofacial pain 20 80.0 Severe pain 25 100.0 Nausea or vomiting 25 100.0 Level of salivary Amylase among migraine headache The intervention was also associated with a statistically significant reduction of 58.6 (ng/l) for salivary amylase and modifications (reduction) in salivary amylase from a baseline median of 130.23 (ng/l) to a post-intervention median of 63.74 (ng/l). As demonstrated in tables 3 and Figure 1. Table 3: The magnitude and statistical significance of changes after intervention in salivary Amylase concentration among migraine headache study group. Amylase Before intervention After intervention Changes after intervention Range (86.18 to 758.11) (31.44 to 103.46) (-4.4 to -696.8) Median 130.23 63.74 -58.6 Interquartile range (106.01 to 254.78) (60.71 to 74.4) (-43.6 to -171) N 25 25 25 P (Wilcoxon-signed ranks test) <0.001 Level of salivaryCortisol among migraine headache The intervention was also associated with a statistically significant increase of 6.4 (ng/ml) in salivary cortisol, as shown in Table 4. Figure 2 depicts the alterations in salivary Cortisol from a baseline median of 8.52 (ng/ml) to a post-intervention median of 13.84 (ng/ml). Table 4: The magnitude and statistical significance of changes after intervention in salivary cortisol concentration among migraine headache study group. Cortisol Before intervention After intervention Changes after intervention Range (0.1 to 10.82) (10.13 to 17.91) (17.5 to 1.2) Median 8.52 13.84 6.4 Interquartile range (4.59 to 9.54) (12.46 to 15.6) (8.6 to 4.3) N 25 25 25 P (Wilcoxon-signed ranks test) <0.001 Tension headache group Twenty individuals with a confirmed diagnosis of tension headache were recruited after completing an informed consent form. Seventy percent of the tension headache group consisted of females, of which 45 percent were <30 years old and only 15 percent were ≥40 years old. As shown in Table 5. Table 5: Frequency distribution of tension headache study group by age and sex. N % Gender Female 14 70.0 Male 6 30.0 Total 20 100.0 Age group (years) <30 9 45.0 (30-39) 8 40.0 40+ 3 15.0 Total 20 100.0 Range = 20-48 Mean = 31.6 SD = 7.3 As shown in table 30, among the tension headache group, only 10% claimed that the pain is aggravated by routine physical activity, 35% reported orofacial pain and another 35% nausea or vomiting, 40% complained of associated TMJ problems. Table 6: The relative frequency of selected clinical features at baseline (before intervention) among tension headache study group. Pretreatment clinical features (N=20) N % Orofacial pain 7 35.0 Nausea or vomiting 7 35.0 Associated with TMJ 8 40.0 Level of salivary Amylase among tension headache The average reduction in salivary amylase of 83.7after intervention was statistically significant, with changes ranging between a minimum decrease of 2.56 to -a maximum decrease of 335.7, table 7. Figure 3, shows the reduction (changes) in salivary amylase from a median of 161.235at baseline to a median of 74.36after intervention. Table 7: The magnitude and statistical significance of changes after intervention in salivary Amylase concentration among tension headache study group. Amylase Before intervention After intervention Changes after intervention Range (74.71 to 384.26) (48.45 to 105.21) (-2.56 to -335.7) Median 161.235 74.36 -83.7 Interquartile range (108.85 to 253.925) (65.44 to 83.56) (-43.585 to -186.39) N 20 20 20 P (Wilcoxon-signed ranks test) <0.001 Level of salivary Cortisol among tension headache The average increase in salivary cortisol of 7.605 after intervention was statistically significant, with changes ranging between a minimum increase of 14.92to a maximum increase of 1.43, table 8. Figure 4, shows the increase (changes) in salivary cortisol from a median of 3.37 at baseline to a median of 12.13 after intervention. Table 8 : The magnitude and statistical significance of changes after intervention in salivary cortisol concentration among tension headache study group. Cortisol Before intervention After intervention Changes after intervention Range (2.16 to 12.43) (7.01 to 18.59) (14.92 to 1.43) Median 3.37 12.13 7.605 Interquartile range (3.075 to 6.235) (10.455 to 15.505) (9.69 to 6.22) N 20 20 20 P (Wilcoxon-signed ranks test) <0.001 Discussion Headaches are the most ancient kind of human suffering. Despite the fact that headaches are a prevalent complaint in medical practice, little is known regarding their occurrence and epidemiology in various groups. In recent years, there has been a greater emphasis on studies in this area, which have attempted to shed some light on a few of the important aspects of migraine and tension headaches and understand its impact on people and society [12] . At motor neurons, botulinum toxin type A inhibits exocytosis of acetylcholine from presynaptic nerve endings through proteolytic cleavage of a specific target protein, SNAP-25 (synaptosomal protein with a molecular weight of 25 kDa), that is essential for vesicle fusion with neuronal membranes leading to decreased muscle contractions [13].The Migraine group was 88% female and 12% male in this research. 40% of migraine sufferers were under 30, whereas 20% were over 40. The tiny sample size makes these findings unreliable [14]. Demographics matched Lipton et al. [15] and Schwenker [16]. In their research, women outnumbered males in migraine (80% vs. 20%), and migraine headaches generally have a throbbing feel, are moderate to severe, and affect one side of the head more than the other. Most patients report photophobia. Sociodemographic, genetic, and environmental factors may explain this. However, numerous earlier research supported Ramage-Morin et al. [17] finding that migraine prevalence was greater among 30–40-year-olds, with women being more vulnerable. Stovner et al. [18] found a 14.4% global age-standardized prevalence estimate for migraine in 2016 (18.9% for women and 9.8% for men . Botulinum toxins reduced attack/months by 15 on average. Botulinum toxins reduce peripheral sensitization of nociceptive fibres, reducing central sensitization [19]. BoNTA( Botulinum toxin A) suppresses peripheral sensitization, which indirectly reduces central sensitization [20]. BoNTA prevented central sensitization in migraine sufferers in this research.The study by Jakubowski et al. [21] showed that BoNTA did not prevent allodynia during migraine in both responders (those who reported significant drops in attack frequency, attack duration, and headache intensity) and nonresponders. However, allodynia predicts BoNTA response. Our research findings vary from Jakubowski et al. [21] owing to patient population variances. Rockett et al. [22] found that the headache reduced over hours, as expected. The latest research also found that "critical" sleep often relieved headaches. Inhibiting these neurotransmitters may avoid neurogenic inflammation and peripheral sensitization. This reduces peripheral-to-central nervous system pain signals. Thus, BoNT/A indirectly prevents migraine and other pain-related central sensitization. In this research, salivary amylase decreased by 58.6 (ng/l). Salivary amylase decreased from 130.23 (ng/l) at baseline to 63.74 following intervention. Botulinum toxin may reduce salivary gland activity (submandibular and parotid glands). This may reduce saliva output and salivary amylase levels [23]. This result is consistent with Guntinas-Lichius [24], who found that botulinium toxin A blocks SNAP-25 (synaptomsome-associated protein of molecular weight 25 KD), which regulates acetylcholine release into the synaptic space [23,24].The intervention in the finding of present study increased salivary Cortisol by 6.4 ng/ml. Salivary Cortisol increased from 8.52 (ng/ml) at baseline to 13.84 (ng/ml) after intervention. Before intervention, cortisol levels may rise, triggering migraines or reacting to chronic migraines , Cortisol is a steroid hormone . It is made in the cortex of the adrenal glands and then released into the blood in response to stress[25]. Simeoni et al. [26] observed that chronic migraine patients treated with BoNT-A had lower salivary cortisol levels. BoNT-A reduced migraine-induced cortisol. The therapy reduced significantly [26]. This contradicts the research. The present study has shown that BoNT-A treatment was associated with the migraine-induced increase of cortisol. Migraines are often associated with distress, which is highlighted as a trigger factor of headaches but may also appear as a consequence of migraine attacks , Salivary cortisol levels increase under a variety of physical and psychological challenges. Salivary and plasma cortisol levels always correlate with each other following stress, confirming that the two pathways are the same salivary cortisol increases with psychological stress and correlates serum cortisol[27]. Cortisol is generated by two adrenal glands on top of each kidney. Stress releases cortisol. Overproduction or underproduction of cortisol might harm health. Cortisol, a "flight or fight" stress hormone, fights inflammation, opens pupils, and pumps blood to the brain and heart. It catabolizes and inhibits the immune system [26] . Stress raises cortisol levels and relaxing lowers them. Unfortunately, migraine attacks and prolonged discomfort may cause stress. Stress may perpetuate migraines and headaches. Cortisol variation may cause an attack [26,28]. Tension headache group The result showed the majority of the tension headache were females (70%), 45% were <30 years old, and only 15% were ≥40 years. The previous study suggest that tension-type headache is a more complex phenomenon influenced by several psychosocial factors. The female preponderance may be explained by clinical factors related to female hormones [29].Tension headache is very closely associated with psychiatric conditions, primarily affective and anxiety disorders [30].Lyngberg et al. [33] found that in their study, the annual incidence of TTH was 14.2 per 1000 person-years for frequent tension-type headaches (female-to-male ratio: 3:1), decreasing with age [31,32,33]. According to finding in the current study, among the tension headache group, only 10% claimed that the pain is aggravated by routine physical activity, 35% reported orofacial pain and another 35% nausea or vomiting, 40% complained of associated TMJ problems, and 65% reported more than one episode. According to Chowdhury, [34] who indicated that nausea and vomiting are excluded from tension headaches, or they may occur in a mild form. This study supports the current findings however, about 20% patients of TTH can complain mild-to-moderate anorexia (as per ICHD I criteria) which needs to be distinguished from nausea [25]. Tension headache is often reported to start at some time during the day and to increase slowly. Then the headache remains throughout the day and is often unaltered during widely varying activities although some people may have an aggravation by late evening . Similarly, Olesen et al. [35] stated 18% may have pulsatile headache, 10% unilateral pain, 28% aggravation on routine physical activity, 18% anorexia, 4% nausea, and 11% photophobia. In addition to Chowdhury, [34], these authors indicated the absence of unilaterality, absence of pulsatile and throbbing character, lack of aggravation by physical activity, lack of nausea and vomiting, and lack of photophobia.The present study showed a reduction significantly in salivary amylase levels in tension headache patients after treatment with the botulinum toxin. The reason for this may be the effect of BTXA on the submandibular glands as the application of BTXA within the glands leads to glandular atrophy and reduces salivary secretion, which leads to a decrease in the level of salivary amylase [36] .Proper doses of BTXA reduce muscle tension without weakening the muscle, which creates a good situation for restoring its function [31] .Actually, amylase has been known as a stress biomarker. Since psychological stress and painful stimuli could activate SAM system, SAA may work as an easy index for assessing SAM activity and an indirect marker for the objective assessment of pain [37].According to Bugdayci et al. [38]the of levels SAA reduced during the attack period in moderate and severe pain, which is inconsistent with the present study. On the other hand, in a study on patients with chronic pain, a significant positive correlation was reported between SAA levels and pain scale [39]. In another study, the level of SAA was correlated with pain severity in patients with symptomatic irreversible pulpitis [39]. According to the present study, the average salivary cortisol increased significantly after the intervention among tension headache study group. Actually, it is difficult to explain this result due to the lack of sufficient sources, as the current study is the first to our knowledge. However, there may be a mechanism of BTXA that stimulates an increase in cortisol concentration. More studies should be done to verify the mechanism and effectiveness of treatment and the course of adrenal gland disease in patients with tension headaches. However, cortisol is a well-established stress hormone, which is released in response to a kaleidoscope of stimuli (e.g., fear, emotions, pain) by the adrenal glands. The leading functions of this hormone include the regulation of glucose homeostasis, the suppression of the immune system, along with the activation of a number of anti-stress and anti-inflammatory pathways [40]. Conclusion In the migraine group, the quantitative measures of salivary amylase showed a significant reduction in response to treatment. While salivary cortisol showed a significant increase in response to treatment. In addition to Nausea and vomiting were the most responsive parameters to treatment, followed by orofacial pain. In the tension headache group, the quantitative measures of salivary amylase showed a significant reduction in response to treatment. But salivary cortisol showed a significant increase in response to treatment. Having orofacial pain and associated TMJ complaints was the most responsive parameter to treatment. The last respondent characteristic was nausea and vomiting. Declarations Consent to participate declaration I, the undersigned, consent to participate in the study titled "Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection." I understand the study's purpose, procedures, potential risks, and my right to withdraw at any time without affecting my care. Participant Name: ____________________ Signature: __________________________ Date: Ethics Approval declaration The Ethics committee, College of Dentistry, University of Baghdad, gave ethical approval for this study. The study was conducted in accordance with the ethical principles that have their origin in Declaration of Helsinki . it was carried out with patients , verbal and analytic approval before the sample was taken , the study protocol and the subject information and consent form were reviewed and approved by local ethics committee to the document number ( Ref # 459, date : January 19, 2022 ) to get this approval . Funding Declaration in the manuscript : No funding Clinical trial number: not applicable. Funding Declaration in the manuscript: No funding Author Contribution Afrah Abdulsahib Abbas contributed to the conceptualization of the study, data collection, and analysis of results. She was responsible for writing and revising the manuscript, ensuring that all aspects of the research were accurately represented.Fawaz Aswad assisted in the design of the study and provided critical insights during data interpretation. 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Clinical Characteristics of Headache among a Sample of Iraqi Medical and Dental Students. Medico-Legal Update, 20(1). Lyngberg, A. C., Rasmussen, B. K., Jørgensen, T., & Jensen, R. (2005). Prognosis of migraine and tension-type headache: a population-based follow-up study. Neurology, 65(4), 580–585. Chowdhury, D. (2012). Tension type headache. Annals of Indian Academy of Neurology, 15(Suppl 1), S83. Olesen, J., Bousser, M. G., Diener, H. C., Dodick, D., First, M., Goadsby, P. J.,… Steiner, T. J. (2006). New appendix criteria open for a broader concept of chronic migraine. Cephalalgia, 26(6), 742–746. Ellies, M. A., Laskawi, R., Götz, W., Arglebe, C., &Tormählen, G. (1999). Immunohistochemical and morphometric investigations of the influence of botulinum toxin on the submandibular gland of the rat. European archives of oto-rhino-laryngology, 256, 148–152. Tandon, M., Singh, A., Saluja, V., Dhankhar, M., Pandey, C. K., & Jain, P. (2016). Validation of a new “objective pain score” vs.“numeric rating scale” for the evaluation of acute pain: a comparative study. Anesthesiology and pain medicine, 6(1). Bugdayci, G., Yildiz, S., Altunrende, B., Yildiz, N., &Alkoy, S. (2010). Salivary alpha amylase activity in migraine patients. Autonomic Neuroscience, 155(1–2), 121–124. Ahmadi-Motamayel, F., Goodarzi, M. T., Jamshidi, Z., Mahdavinezhad, A., &Rafieian, N. (2016). Evaluation of salivary and serum alpha amylase level in dental caries of adolescence. Brazilian Dental Science, 19(2), 40–46. Fukuda, S., & Morimoto, K. (2001). Lifestyle, stress and cortisol response: Review II: Lifestyle. Environmental health and preventive medicine, 6, 15–21. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6227107","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":442076993,"identity":"9f0e4444-99d4-46c3-b826-af30130926dd","order_by":0,"name":"Afrah Abdulsahib Abbas","email":"data:image/png;base64,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","orcid":"","institution":"University of Baghdad","correspondingAuthor":true,"prefix":"","firstName":"Afrah","middleName":"Abdulsahib","lastName":"Abbas","suffix":""},{"id":442076994,"identity":"2ded156d-0bdb-4f81-8fed-01bfe8423fb8","order_by":1,"name":"Fawaz Aswad","email":"","orcid":"","institution":"University of Baghdad","correspondingAuthor":false,"prefix":"","firstName":"Fawaz","middleName":"","lastName":"Aswad","suffix":""}],"badges":[],"createdAt":"2025-03-14 14:08:29","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6227107/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6227107/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":80811842,"identity":"8259da4b-4c73-4099-9cba-255130867727","added_by":"auto","created_at":"2025-04-17 10:29:48","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":282242,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eDot diagram showing the changes after intervention in salivary amylase concentration among migraine headache study group\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Picture1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6227107/v1/cbc6dac75f08c160a46548a4.jpg"},{"id":80811847,"identity":"d92443e6-ca65-4734-8142-ed5ca4a59096","added_by":"auto","created_at":"2025-04-17 10:29:48","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":70043,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eDot diagram showing the changes after intervention in salivary cortisol concentration among migraine headache study group\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Picture2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6227107/v1/5606662fe893a462e4bc9afb.jpg"},{"id":80811843,"identity":"2579586b-dd34-4139-8c52-0c013cc74aca","added_by":"auto","created_at":"2025-04-17 10:29:48","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":70099,"visible":true,"origin":"","legend":"\u003cp\u003eDot diagram showing the changes after intervention in salivary amylase concentration among tension headache study group.\u003c/p\u003e","description":"","filename":"Picture3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6227107/v1/c1a012bfb2d023ecf4358e8c.jpg"},{"id":80811845,"identity":"547e1e62-171e-40d0-8b5d-1a401ff5e974","added_by":"auto","created_at":"2025-04-17 10:29:48","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":66897,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eDot diagram showing the changes after intervention in salivary cortisol concentration among tension headache study group\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Picture4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6227107/v1/9fe17ab980a81357c30fc0e2.jpg"},{"id":80811850,"identity":"260ef755-8deb-439b-882a-a6c5af228557","added_by":"auto","created_at":"2025-04-17 10:29:48","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":97031,"visible":true,"origin":"","legend":"\u003cp\u003eUnnumbered image in the \u003cstrong\u003eMaterial and Methods \u003c/strong\u003esection.\u003c/p\u003e","description":"","filename":"Picture5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6227107/v1/9c52c9e969969a01e5a2d06e.jpg"},{"id":96362901,"identity":"e0df444a-b964-49de-9c38-7e3355d9010a","added_by":"auto","created_at":"2025-11-20 10:02:39","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2103719,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6227107/v1/668d9e0f-889c-49b8-aff0-add8e82221c1.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection","fulltext":[{"header":"Introduction","content":"\u003cp\u003eHeadaches are a frequent symptom of central nervous system convergence [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Sensitization and activation of trigeminal ganglion components may contribute to headache and facial pain problems [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Migraine is the most researched headache condition. Migraine and other headache problems may be more prevalent in orofacial pain (OFP) patients [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Chronic orofacial pain may be neuropathic, neurovascular or musculoskeletal, musculoskeletal pain involves tempromandibular joint and masticatory muscles [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eTTH is the most frequent main headache, with a lifetime frequency of 52% compared to 18% for migraine. Chronic headache affects 1.7\u0026ndash;4% of adults worldwide. Headache diseases impact individuals of all ages, ethnicities, economic levels, and regions worldwide. Child headaches are infrequent but grow with age. Adolescence, especially in childbearing women, increases headaches and migraines [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e] .\u003c/p\u003e \u003cp\u003eThe neurotoxic protein botulinum toxin (BTX) is generated by the bacteria Clostridium botulinum and related species. BTX inhibits the release of the neurotransmitter acetylcholine from axon terminals at the neuromuscular junction. This effect paralyses the muscles [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Botulinum toxin type A (BTX-A) is utilised in TTH therapy.\u003c/p\u003e \u003cp\u003eWiliam Binder discovered its therapeutic benefit in alleviating headaches [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. He reported a reduction in migraine recurrence after a cosmetic injection of BTX-A to diminish forehead creases.\u003c/p\u003e \u003cp\u003eSaliva has long been thought to be a mirror of the body since it includes the serum elements which are evaluated in standard blood tests for disease and health monitoring, saliva as a diagnostic fluid provides a lot of advantages over serum for disease diagnosis [ 7 ]. Salivary alpha-amylase (sAA) has evolved as a viable and reliable marker of ANS activity in stress studies during the last 15 years. Alpha-amylase is a salivary enzyme that aids in carbohydrate and starch digestion. It can be measured quickly and non-invasively through saliva collection, and as such, it is frequently used as a proxy measure of sympathetic arousal in the acute stress literature, particularly in studies looking to reduce participant load or when paradigms require participants to collect samples at home [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The most abundant protein in saliva is alpha amylase(sAA), accounting for approximately 20% of salivary proteins [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eCortisol is the primary glucocorticoid hormone released by the adrenal cortex. Cortisol is a steroid hormone based on its chemical structure. Cortisol influences carbohydrate, lipid, and protein metabolism. Cortisol is involved in the stress response and is required for an organism to operate properly. Corticosteroids are a class of hormones secreted by the adrenal gland cortex. Corticosteroids are classified into three types based on their primary physiological influence: glucocorticoids (cortisol and corticosterone), mineral corticoids (aldosterone), and androgen hormones (dehydro-epi androsterone and androstenedione). Cortisol is tested in blood and urine, and more recently, in instances when a blood sample is difficult to obtain, cortisol has been detected in saliva [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e].\u003c/p\u003e"},{"header":"Material and Methods","content":"\u003cp\u003e\u003cstrong\u003eField research\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTotal subjects composed of forty-five participants were recruited and concentrated for this study. They were carefully informed about the aim of the investigation and they were free to accept or refuse to participate. A longitudinal study was done in Al-Imam Alsadiq General Hospital in Hilla City and the laboratory work was done in the lab of the hospital. Approval was obtained from Ministry of Health for patient\u0026apos;s\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSubjects:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe participants were examined from the period of June - 2022 to October-2022.The study protocol was approved by oral medicine department, college of Dentistry, Baghdad University.\u003c/p\u003e\n\u003cp\u003eThe sample was obtained from 45 subjects who were divided into two groups:\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGroup 1:\u003c/strong\u003e Twenty-five patients with chronic migraine.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGroup 2\u003c/strong\u003e: Twenty patients with chronic tension headaches.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eInclusions criteria\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eGood general health, age\u0026ge;18years. Participants had also to be free off ever/or cold\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExclusion criteria\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFibromyalgia, rheumatoid arthritis, whiplash-associated disorder, neurological disorders, pregnancy or lactation, high blood pressure, tobacco usage , or muscle relaxants , a known BTX allergy; and oral complaint , such as oral dryness or mucosal lesions, were excluded from further involvement in the study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSaliva collection\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe whole sample of saliva was taken from patients in the morning from 9 a.m to 10 a.m \u0026nbsp;patient come fasting and then asked to collect sample of \u0026nbsp; Saliva after an individual was asked to rinse his/her mouth thoroughly with distal water to insure the removal of any possible debris or contaminating materials. The first mouth-full of saliva was discarded to allow clearance of water then they were asked to collect all the saliva until the recommended amount of saliva was obtained pre and post botox injection by one month were done 10 millilitres of unstimulated whole saliva was collected ..The collected saliva was centrifuged at 4000 r.p.m (rotation per minute) for 10 minutes and then centrifuged clear supernatant saliva kept frozen and stored at -20 \u0026deg;C until the whole saliva samples were collected , then SAMY1 and COR determination done using Human salivary alpha-amylase ELISA kit and Human cortisol ELISA kit.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eBTX injections\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn this study, onabotulinumtoxinA (OBTA) (nabota) was used for prophylaxis of chronic migraine and chronic tension headache, and this type of botulinum toxin was FDA-approved in February 2019.One handful of MU of onabotulinumtoxin A (nabota) is diluted with 2.5 ml of saline, yielding a concentration of 5 MU per 12.5 ml.Each intramuscular injection site is injected with 5 MU of onabotulinumtoxinA.The injection paradigm for chronic migraine consists of 31 fixed sites in the following muscles: mm. frontalis 20 MU (four sites), mm. corrugatores 10 MU (two sites), m. procerus 5 MU (1 site), mm. occipitalis 30 MU (six sites), mm. temporalis 40 MU (eight sites), mm. trapezii 30 MU (in six sites), cervical paraspinal muscle Group 20 MU (four sites) In these fixed sites, a total dose of 155 MU of onabotulinumtoxin A is applied. The injection paradigm for chronic tension headache consists of 10 fixed sites in the following muscles: mm. frontalis 30 MU (six sites),, mm. corrugatores 10 MU (two sites),, mm. procerus 5 MU (1 site),, and in the middle of the forehead 5 MU (1 site).. In these fixed sites, a total dose of 50 MU of onabotulinumtoxin A is applied.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConstant site, constant dose injection scheme applied in PREEMPT studies [ 11 ].\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStatistical Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eStatistical analyses were done using IBMSPSS version 27 computer software (IBM Statistical Package for Social Sciences) in association with Microsoft Excel.\u003c/p\u003e\n\u003cp\u003eThe Wilcoxon Signed Ranks Test determined the statistical significance of the average (median) change in a quantitative outcome following intervention. A P value below 0.05 indicated statistical significance. All significance tests were bilateral. Kruskal-Wallis Test compared medians of non-normally distributed variables between more than two groups. ANOVA analysed the statistical significance of the mean difference of a continuous response variable that is known or presumed to be regularly distributed (like age) between more than two groups.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;Ethical approval\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe Ethics committee, College of Dentistry, University of Baghdad, gave ethical approval for this study . The study was conducted in accordance with the ethical principles that have their origin in Declaration of Helsinki . \u0026nbsp;it was carried out with patients\u003csup\u003e,\u0026nbsp;\u003c/sup\u003everbal and analytic approval before the sample was taken , the study protocol and the subject information and consent form were reviewed and approved by local ethics committee to the document number ( Ref # 459, date : January 19, 2022 ) to get this approval .\u0026nbsp;\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003eMigraine headaches group\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAfter obtaining informed permission, 25 people with a confirmed Migraine diagnosis were enrolled in the study. According to Table 1, the bulk of the Migraine group (88%) were females, and 40% were 30 years old, with just 20% being 40 years old.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e1\u003c/strong\u003e\u003cstrong\u003e: Frequency distribution of migraine headache study group by age and sex.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"532\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eN\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e%\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003eGender\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003eFemale\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e88.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003eMale\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e12.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003eTotal\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e100.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003eAge (years)-categories\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003e\u0026lt;30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e40.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003e(30-39)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e40.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003e40+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e20.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4.13534%;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 63.9098%;\"\u003e\n \u003cp\u003eTotal\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 15.9774%;\"\u003e\n \u003cp\u003e100.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eRange = 20-25\u003c/p\u003e\n\u003cp\u003eMean = 32.6\u003c/p\u003e\n\u003cp\u003eSD = 8.1\u003c/p\u003e\n\u003cp\u003eThe result in table 2 showed, among the migraine group 60% complained of associated TMJ problems and about three quarters (76%) complained of paresthesia, 80% with orofacial pain, Everybody (100%) of the recruited study group reported nausea or vomiting.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2: The relative frequency of selected clinical features at baseline (before intervention) among migraine headache study group.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"510\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 66.6667%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e(Total N=25)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eN\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e\u003cstrong\u003e%\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 66.6667%;\"\u003e\n \u003cp\u003eOrofacial pain\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e80.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 66.6667%;\"\u003e\n \u003cp\u003eSevere pain\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e100.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 66.6667%;\"\u003e\n \u003cp\u003eNausea or vomiting\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 16.6667%;\"\u003e\n \u003cp\u003e100.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eLevel of salivary Amylase among migraine headache\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe intervention was also associated with a statistically significant reduction of 58.6 (ng/l) for salivary amylase and modifications (reduction) in salivary amylase from a baseline median of 130.23 (ng/l) to a post-intervention median of 63.74 (ng/l). As demonstrated in tables 3 and Figure 1.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3: The magnitude and statistical significance of changes after intervention in salivary Amylase concentration among migraine headache study group.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"600\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 416px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAmylase\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBefore intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAfter intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eChanges after intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eRange\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(86.18 to 758.11)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(31.44 to 103.46)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(-4.4 to -696.8)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eMedian\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e130.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e63.74\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e-58.6\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eInterquartile range\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(106.01 to 254.78)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(60.71 to 74.4)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(-43.6 to -171)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eP (Wilcoxon-signed ranks test)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eLevel of salivaryCortisol among migraine headache\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe intervention was also associated with a statistically significant increase of 6.4 (ng/ml) in salivary cortisol, as shown in Table 4. Figure 2 depicts the alterations in salivary Cortisol from a baseline median of 8.52 (ng/ml) to a post-intervention median of 13.84 (ng/ml).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 4: The magnitude and statistical significance of changes after intervention in salivary cortisol concentration among migraine headache study group.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"600\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 416px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eCortisol\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBefore intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAfter intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eChanges after intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eRange\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(0.1 to 10.82)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(10.13 to 17.91)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(17.5 to 1.2)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eMedian\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e8.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e13.84\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e6.4\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eInterquartile range\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(4.59 to 9.54)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(12.46 to 15.6)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(8.6 to 4.3)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eP (Wilcoxon-signed ranks test)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eTension headache group\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTwenty individuals with a confirmed diagnosis of tension headache were recruited after completing an informed consent form. Seventy percent of the tension headache group consisted of females, of which 45 percent were \u0026lt;30 years old and only 15 percent were \u0026ge;40 years old. As shown in Table 5.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 5: Frequency distribution of tension headache study group by age and sex.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"599\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eN\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e%\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eGender\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eFemale\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e70.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eMale\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e30.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eTotal\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e100.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eAge group (years)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003e\u0026lt;30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e45.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003e(30-39)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e40.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003e40+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e15.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eTotal\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e100.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eRange = 20-48\u003c/p\u003e\n\u003cp\u003eMean = 31.6\u003c/p\u003e\n\u003cp\u003eSD = 7.3\u003c/p\u003e\n\u003cp\u003eAs shown in table 30, among the tension headache group, only 10% claimed that the pain is aggravated by routine physical activity, 35% reported orofacial pain and another 35% nausea or vomiting, 40% complained of associated TMJ problems.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 6: The relative frequency of selected clinical features at baseline (before intervention) among tension headache study group.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"599\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003e\u003cstrong\u003ePretreatment clinical features (N=20)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eN\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e%\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eOrofacial pain\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e35.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eNausea or vomiting\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e35.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 362px;\"\u003e\n \u003cp\u003eAssociated with TMJ\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 118px;\"\u003e\n \u003cp\u003e40.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eLevel of salivary Amylase among tension headache\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe average reduction in salivary amylase of 83.7after intervention was statistically significant, with changes ranging between a minimum decrease of 2.56 to -a maximum decrease of 335.7, table 7. Figure 3, shows the reduction (changes) in salivary amylase from a median of 161.235at baseline to a median of 74.36after intervention.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 7: The magnitude and statistical significance of changes after intervention in salivary Amylase concentration among tension headache study group.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"600\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 416px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAmylase\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBefore intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAfter intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eChanges after intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eRange\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(74.71 to 384.26)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(48.45 to 105.21)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(-2.56 to -335.7)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eMedian\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e161.235\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e74.36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e-83.7\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eInterquartile range\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(108.85 to 253.925)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(65.44 to 83.56)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(-43.585 to -186.39)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eP (Wilcoxon-signed ranks test)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eLevel of salivary Cortisol among tension headache\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe average increase in salivary cortisol of 7.605 after intervention was statistically significant, with changes ranging between a minimum increase of 14.92to a maximum increase of 1.43, table 8. Figure 4, shows the increase (changes) in salivary cortisol from a median of 3.37 at baseline to a median of 12.13 after intervention.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 8\u003c/strong\u003e\u003cstrong\u003e: The magnitude and statistical significance of changes after intervention in salivary cortisol concentration among tension headache study group.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"600\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 416px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eCortisol\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBefore intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAfter intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eChanges after intervention\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eRange\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(2.16 to 12.43)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(7.01 to 18.59)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(14.92 to 1.43)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eMedian\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e3.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e12.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e7.605\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eInterquartile range\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(3.075 to 6.235)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(10.455 to 15.505)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e(9.69 to 6.22)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 184px;\"\u003e\n \u003cp\u003eP (Wilcoxon-signed ranks test)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 139px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Discussion","content":"\u003cp\u003eHeadaches are the most ancient kind of human suffering. Despite the fact that headaches are a prevalent complaint in medical practice, little is known regarding their occurrence and epidemiology in various groups. In recent years, there has been a greater emphasis on studies in this area, which have attempted to shed some light on a few of the important aspects of migraine and tension headaches and understand its impact on people and society [12] . At motor neurons, botulinum toxin type A inhibits exocytosis of acetylcholine from presynaptic nerve endings through proteolytic cleavage of a specific target protein, SNAP-25 (synaptosomal protein with a molecular weight of 25 kDa), that is essential for vesicle fusion with neuronal membranes leading to decreased muscle contractions [13].The Migraine group was 88% female and 12% male in this research. 40% of migraine sufferers were under 30, whereas 20% were over 40. The tiny sample size makes these findings unreliable [14]. Demographics matched Lipton et al. [15] and Schwenker [16]. In their research, women outnumbered males in migraine (80% vs. 20%), and migraine headaches generally have a throbbing feel, are moderate to severe, and affect one side of the head more than the other. Most patients report photophobia. Sociodemographic, genetic, and environmental factors may explain this. However, numerous earlier research supported Ramage-Morin et al. [17] finding that migraine prevalence was greater among 30\u0026ndash;40-year-olds, with women being more vulnerable. Stovner et al. [18] found a 14.4% global age-standardized prevalence estimate for migraine in 2016 (18.9% for women and 9.8% for men . Botulinum toxins reduced attack/months by 15 on average. Botulinum toxins reduce peripheral sensitization of nociceptive fibres, reducing central sensitization [19]. BoNTA( Botulinum toxin A) \u0026nbsp;suppresses peripheral sensitization, which indirectly reduces central sensitization [20]. BoNTA prevented central sensitization in migraine sufferers in this research.The study by Jakubowski et al. [21] showed that BoNTA did not prevent allodynia during migraine in both responders (those who reported significant drops in attack frequency, attack duration, and headache intensity) and nonresponders. However, allodynia predicts BoNTA response. Our research findings vary from Jakubowski et al. [21] owing to patient population variances. Rockett et al. [22] found that the headache reduced over hours, as expected. The latest research also found that \u0026quot;critical\u0026quot; sleep often relieved headaches. Inhibiting these neurotransmitters may avoid neurogenic inflammation and peripheral sensitization. This reduces peripheral-to-central nervous system pain signals. Thus, BoNT/A indirectly prevents migraine and other pain-related central sensitization.\u003c/p\u003e\n\u003cp\u003eIn this research, salivary amylase decreased by 58.6 (ng/l). Salivary amylase decreased from 130.23 (ng/l) at baseline to 63.74 following intervention. Botulinum toxin may reduce salivary gland activity (submandibular and parotid glands). This may reduce saliva output and salivary amylase levels [23]. This result is consistent with Guntinas-Lichius [24], who found that botulinium toxin A blocks SNAP-25 (synaptomsome-associated protein of molecular weight 25 KD), which regulates acetylcholine release into the synaptic space [23,24].The intervention in the finding of present study increased salivary Cortisol by 6.4 ng/ml. \u0026nbsp;Salivary Cortisol increased from 8.52 (ng/ml) at baseline to 13.84 (ng/ml) after intervention. Before intervention, cortisol levels may rise, triggering migraines or reacting to chronic migraines ,\u0026nbsp;Cortisol is a steroid hormone . It is made in the cortex of the adrenal glands and then released into the blood in response to stress[25].\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSimeoni et al. [26] observed that chronic migraine patients treated with BoNT-A had lower salivary cortisol levels. \u0026nbsp;BoNT-A reduced migraine-induced cortisol. The therapy reduced significantly [26]. This contradicts the research.\u003c/p\u003e\n\u003cp\u003eThe present study has shown that BoNT-A treatment was associated with the migraine-induced increase of cortisol. Migraines are often associated with distress, which is highlighted as a trigger factor of headaches but may also appear as a consequence of migraine attacks ,\u0026nbsp;Salivary cortisol levels increase under a variety of physical and psychological challenges. Salivary and plasma cortisol levels always correlate with each other following stress, confirming that the two pathways are the same salivary cortisol increases with psychological stress and correlates serum cortisol[27].\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eCortisol is generated by two adrenal glands on top of each kidney. Stress releases cortisol. Overproduction or underproduction of cortisol might harm health. Cortisol, a \u0026quot;flight or fight\u0026quot; stress hormone, fights inflammation, opens pupils, and pumps blood to the brain and heart. It catabolizes and inhibits the immune system [26] . Stress raises cortisol levels and relaxing lowers them. Unfortunately, migraine attacks and prolonged discomfort may cause stress. Stress may perpetuate migraines and headaches. Cortisol variation may cause an attack [26,28].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTension headache group\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe result showed the majority of the tension headache were females (70%), 45% were \u0026lt;30 years old, and only 15% were \u0026ge;40 years. The previous study suggest that tension-type headache is a more complex phenomenon influenced by several psychosocial factors. The female preponderance may be explained by clinical factors related to female hormones [29].Tension headache is very closely associated with psychiatric conditions, primarily affective and anxiety disorders [30].Lyngberg et al. [33] found that in their study, the annual incidence of TTH was 14.2 per 1000 person-years for frequent tension-type headaches (female-to-male ratio: 3:1), decreasing with age [31,32,33].\u003c/p\u003e\n\u003cp\u003eAccording to finding in the current study, among the tension headache group, only 10% claimed that the pain is aggravated by routine physical activity, 35% reported orofacial pain and another 35% nausea or vomiting, 40% complained of associated TMJ problems, and 65% reported more than one episode. According to Chowdhury, [34] who indicated that nausea and vomiting are excluded from tension headaches, or they may occur in a mild form. This study supports the current findings however, about 20% patients of TTH can complain mild-to-moderate anorexia (as per ICHD I criteria) which needs to be distinguished from nausea [25]. Tension headache is often reported to start at some time during the day and to increase slowly. Then the headache remains throughout the day and is often unaltered during widely varying activities although some people may have an aggravation by late evening . Similarly, Olesen et al. [35] stated 18% may have pulsatile headache, 10% unilateral pain, 28% aggravation on routine physical activity, 18% anorexia, 4% nausea, and 11% photophobia.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn addition to Chowdhury, [34], these authors indicated the absence of unilaterality, absence of pulsatile and throbbing character, lack of aggravation by physical activity, lack of nausea and vomiting, and lack of photophobia.The present study showed a reduction significantly in salivary amylase levels in tension headache patients after treatment with the botulinum toxin. The reason for this may be the effect of BTXA on the submandibular glands as the application of BTXA within the glands leads to glandular atrophy and reduces salivary secretion, which leads to a decrease in the level of salivary amylase [36] .Proper doses of BTXA reduce muscle tension without weakening the muscle, which creates a good situation for restoring its function\u0026nbsp;[31]\u0026nbsp;.Actually, amylase has been known as a stress biomarker. Since psychological stress and painful stimuli could activate SAM system, SAA may work as an easy index for assessing SAM activity and an indirect marker for the objective assessment of pain [37].According to Bugdayci et al. [38]the of levels SAA reduced during the attack period in moderate and severe pain, which is inconsistent with the present study. On the other hand, in a study on patients with chronic pain, a significant positive correlation was reported between SAA levels and pain scale [39]. In another study, the level of SAA was correlated with pain severity in patients with symptomatic irreversible pulpitis [39].\u003c/p\u003e\n\u003cp\u003eAccording to the present study, the average salivary cortisol increased significantly after the intervention among tension headache study group. Actually, it is difficult to explain this result due to the lack of sufficient sources, as the current study is the first to our knowledge. However, there may be a mechanism of BTXA that stimulates an increase in cortisol concentration. More studies should be done to verify the mechanism and effectiveness of treatment and the course of adrenal gland disease in patients with tension headaches. However, cortisol is a well-established stress hormone, which is released in response to a kaleidoscope of stimuli (e.g., fear, emotions, pain) by the adrenal glands. The leading functions of this hormone include the regulation of glucose homeostasis, the suppression of the immune system, along with the activation of a number of anti-stress and anti-inflammatory pathways [40].\u0026nbsp;\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIn the migraine group, the quantitative measures of salivary amylase showed a significant reduction in response to treatment. While salivary cortisol showed a significant increase in response to treatment. In addition to Nausea and vomiting were the most responsive parameters to treatment, followed by orofacial pain. In the tension headache group, the quantitative measures of salivary amylase showed a significant reduction in response to treatment. But salivary cortisol showed a significant increase in response to treatment. Having orofacial pain and associated TMJ complaints was the most responsive parameter to treatment. The last respondent characteristic was nausea and vomiting.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eConsent to participate declaration\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eI, the undersigned, consent to participate in the study titled \u003cstrong\u003e\u0026quot;Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection.\u0026quot;\u003c/strong\u003e I understand the study\u0026apos;s purpose, procedures, potential risks, and my right to withdraw at any time without affecting my care.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eParticipant Name:\u003c/strong\u003e ____________________\u003cbr\u003e\u003cstrong\u003eSignature:\u003c/strong\u003e __________________________\u003cbr\u003e\u003cstrong\u003eDate:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics Approval declaration\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe Ethics committee, College of Dentistry, University of Baghdad, gave ethical approval for this study. The study was conducted in accordance with the ethical principles that have their origin in Declaration of Helsinki . \u0026nbsp;it was carried out with patients\u003csup\u003e,\u0026nbsp;\u003c/sup\u003everbal and analytic approval before the sample was taken , the study protocol and the subject information and consent form were reviewed and approved by local ethics committee to the document number ( Ref # 459, date : January 19, 2022 ) to get this approval .\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding Declaration in the manuscript\u003c/strong\u003e\u003cstrong\u003e\u003cspan dir=\"RTL\"\u003e:\u003c/span\u003e\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;No funding\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical trial number: not applicable.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eDeclaration in the manuscript: No funding\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAfrah Abdulsahib Abbas contributed to the conceptualization of the study, data collection, and analysis of results. She was responsible for writing and revising the manuscript, ensuring that all aspects of the research were accurately represented.Fawaz Aswad assisted in the design of the study and provided critical insights during data interpretation. He played a key role in writing and reviewing the manuscript, focusing on the clinical implications of the findings.Both authors have approved the final version of the manuscript and agree to be accountable for all aspects of the work.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eCady, R., Schreiber, C., Farmer, K., \u0026amp;Sheftell, F. (2002). Primary headaches: a convergence hypothesis. Headache: The Journal of Head and Face Pain, 42(3), 204\u0026ndash;216.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKumar, K. H., \u0026amp; Elavarasi, P. (2016). Definition of pain and classification of pain disorders. 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Headache: The Journal of Head and Face Pain,50(8), 1306\u0026ndash;1312.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAli, M. R., \u0026amp; Abdulkareem, E. H. (2020). Efficiency of BTX-A in the Alleviation of Hemifacial Pain. Journal of International Dental and Medical Research, 13(1), 321\u0026ndash;326.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMohammed Ali, N. S., \u0026amp; Basim, R. (2020). Clinical Characteristics of Headache among a Sample of Iraqi Medical and Dental Students. Medico-Legal Update, 20(1).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLyngberg, A. C., Rasmussen, B. K., J\u0026oslash;rgensen, T., \u0026amp; Jensen, R. (2005). Prognosis of migraine and tension-type headache: a population-based follow-up study. Neurology, 65(4), 580\u0026ndash;585.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChowdhury, D. (2012). Tension type headache. Annals of Indian Academy of Neurology, 15(Suppl 1), S83.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOlesen, J., Bousser, M. G., Diener, H. C., Dodick, D., First, M., Goadsby, P. J.,\u0026hellip; Steiner, T. J. (2006). New appendix criteria open for a broader concept of chronic migraine. Cephalalgia, 26(6), 742\u0026ndash;746.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eEllies, M. A., Laskawi, R., G\u0026ouml;tz, W., Arglebe, C., \u0026amp;Torm\u0026auml;hlen, G. (1999). Immunohistochemical and morphometric investigations of the influence of botulinum toxin on the submandibular gland of the rat. European archives of oto-rhino-laryngology, 256, 148\u0026ndash;152.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTandon, M., Singh, A., Saluja, V., Dhankhar, M., Pandey, C. K., \u0026amp; Jain, P. (2016). Validation of a new \u0026ldquo;objective pain score\u0026rdquo; vs.\u0026ldquo;numeric rating scale\u0026rdquo; for the evaluation of acute pain: a comparative study. Anesthesiology and pain medicine, 6(1).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBugdayci, G., Yildiz, S., Altunrende, B., Yildiz, N., \u0026amp;Alkoy, S. (2010). Salivary alpha amylase activity in migraine patients. Autonomic Neuroscience, 155(1\u0026ndash;2), 121\u0026ndash;124.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAhmadi-Motamayel, F., Goodarzi, M. T., Jamshidi, Z., Mahdavinezhad, A., \u0026amp;Rafieian, N. (2016). Evaluation of salivary and serum alpha amylase level in dental caries of adolescence. Brazilian Dental Science, 19(2), 40\u0026ndash;46.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFukuda, S., \u0026amp; Morimoto, K. (2001). Lifestyle, stress and cortisol response: Review II: Lifestyle. Environmental health and preventive medicine, 6, 15\u0026ndash;21.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Migraine Headache, Tension Headache, cortisol, salivary alpha amylase","lastPublishedDoi":"10.21203/rs.3.rs-6227107/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6227107/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eMigraine is a highly prevalent and disabling disorder. Because stress appears to be a prominent trigger of this condition. the aim of this study was to evaluate the therapeutic effects of botulinum toxin injection in migraine and tension headache patients and to estimate salivary cortisol, salivary alpha amylase in these groups of patients before and after treatment with botulinum toxin injection.\u003c/p\u003e \u003cp\u003eA total of 45 subjects were divided into two groups 25 patients with chronic migraine and 20 patients with chronic tension headaches. The salivary biomarker (salivary cortisol, salivary alpha amylase) was detected by enzyme-linked immunosorbent assay (ELISA) kits.\u003c/p\u003e \u003cp\u003eThe result showed the increased statistically significant (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) in salivary cortisol, and decreased statistically significant (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) in salivary alpha amylase after intervention with botulinum toxin in both study groups (chronic migraine and chronic tension headaches).\u003c/p\u003e \u003cp\u003eSalivary amylase decreased significantly after therapy in both groups (migraine and tension headache). Salivary cortisol increased significantly after therapy. In addition, Nausea and vomiting responded best to therapy, followed by orofacial discomfort.\u003c/p\u003e","manuscriptTitle":"Evaluation of Stress Biomarker in Migraine and Tension Headache Patients Before and After Botulinum Toxin Injection","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-17 10:29:43","doi":"10.21203/rs.3.rs-6227107/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"caa95502-b74d-4c6b-be8f-c0d221db9fe5","owner":[],"postedDate":"April 17th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-11-13T17:23:33+00:00","versionOfRecord":[],"versionCreatedAt":"2025-04-17 10:29:43","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6227107","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6227107","identity":"rs-6227107","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}