Salivary Cortisol Levels and Burning Symptoms in Patients with Burning Mouth Syndrome before and after Low Level Laser Therapy: a Double Blind Controlled Randomized Clinical Trial

Abstract

Objectives

Burning mouth syndrome (BMS) is a disorder which is described as burning sensation of the oral mucosa without pathological changes. Most of the patients have some underlying stressful conditions. Stress induces an increase in secretion of different endocrine glands resulting in higher levels of glucocorticoids. One of the options for treatment of BMS includes low level laser therapy (LLLT). The aim of this research was to determine salivary cortisol levels and intensity of burning symptoms in BMS patients before and after LLLT.

Materials and Methods

Twenty-three participants were allocated by randomization in two groups: 12 patients in the study group and 11 patients in the placebo group. Cortisol levels in all patients were analyzed from the sample of saliva collected without stimulation. In both groups, the LLLT was performed once a day for ten consecutive days (excluding weekend) with Ga-Al-As light-emitting diode type of laser, with a wavelength of 685nm. In the control group, LLLT was done with inactive laser probe which was only emitting audio signal. The intensity of burning symptoms was measured by a visual analogue scale (VAS). The VAS and unstimulated saliva were measured at baseline and on the last day of the LLLT. A quantitative analysis of saliva was performed using competitive commercial ELISA-kit.

Results

VAS scores and salivary cortisol levels were significantly lower in both groups after LLLT.

Conclusions

LLLT can be useful in patients with BMS for reducing burning symptoms and salivary cortisol level. Future studies on a larger number of patients should clarify whether the positive results are an outcome of laser effectiveness or of placebo effect.

Introduction

Burning mouth syndrome (BMS) is a disorder which is described as burning sensation of oral mucosa without pathological changes (1). The etiology of BMS is unclear, although disorders of the central (2) or peripheral nervous system are possible causes. The most frequently affected sites are the anterior two thirds of the tongue although the BMS can affect any other intraoral site (3). Peri-menopausal and post – menopausal women are the most affected patients. It is hard to estimate the real epidemiology of BMS because of non-universal diagnostic criteria and, also, because of a low awareness about BMS among dentists and general health practitioners. Some studies from the literature have shown some nonspecific health problems in BMS patients, such as irritable bowel syndrome, headaches, temporomandibular joint pain, dermatological, musculoskeletal and psychiatric disorders (4) as well as higher level of catastrophizing is some patients (5). The majority of the patients have some underlying stressful conditions. Also, the burning symptoms itself could increase the level of stress in patients with BMS.

Stress induces an increase in secretion of different endocrine glands resulting in higher levels of glucocorticoids (6). Cortisol is a glucocorticoid which is secreted from the adrenal cortex. It is involved in the metabolism of carbohydrates, proteins and fats. In addition, it has a role in water metabolism, affects the sensitivity of the nervous system and affects the human stress response (3). Results from the literature point out that cortisol levels in saliva are increased in BMS patients as indicator of stress conditions (3, 7). However, Nosratzehi et al. (8) and de Souza et al. (9) have shown that differences between salivary cortisol levels among BMS patients and control were not significant.

The estimation of cortisol in saliva is a non-invasive procedure as opposed to blood collection which induces stress in some patients leading to an increase in cortisol levels (10). Due to an advance in analytical methods, the levels of cortisol and cortisone in saliva can be measured with high reliability. The reference interval is determined by the manufacturer of the specific kit.

A recent study (11) has shown that women with temporomandibular disorders and salivary cortisol above 10 ng/ml reported higher pain scores. This result points out the involvement of stress in other painful conditions in the orofacial region.

The treatment of BMS is difficult and represents a serious clinical problem. Various treatment possibilities have been suggested but with limited success. According to a recent systematic review, a certain intervention for patient with BMS has not been proposed (12).

Low level laser therapy (LLLT) as a treatment option includes the use of light to a biologic system in order to promote tissue regeneration, reduce inflammation and relieve pain (13). The results regarding LLLT in BMS patients are controversial and require further investigation.

The aim of this research was to determine the salivary cortisol levels and intensity of burning symptoms in BMS patients before and after LLLT. To our knowledge, this is the first study evaluating the effect of LLLT on salivary cortisol levels in BMS patients.

Materials and methods

This was a double-blinded randomized clinical trial approved by the Ethics Committee of the School of Dental Medicine, University of Zagreb, Croatia. The written consent form was signed by each patient consistent with the Declaration of Helsinki. There were 12 patients in the study group and 11 patients in the control group. Sample size was calculated according to previously published data (14) by power analysis with significance level α=0.05 and power β=0.8. Participants were randomly allocated into two groups by random number generator.

Inclusion criteria were patients with burning sensations for at least three months before the treatment and with normal appearance of the oral mucosa. Burning sensations were present on only one intraoral site – the tongue, lip or hard palate. The patients with BMS came to our Department for the first time and they had not had any previous treatments for BMS Excluding criteria were patients with diabetes, serum iron and vitamin B deficiency, previous head and neck radiotherapy, patients with autoimmune diseases and those taking antidepressants, anxiolytics, anticonvulsants and hormonal therapy which could affect cortisol level.

Primary outcome measure was a patient's subjective complaint of burning sensation detected by a visual analogue scale (VAS) grading 0 to 10, with level 10 representing the strongest level of symptoms.

The secondary outcome measure was the salivary cortisol level in unstimulated saliva. Saliva was collected between 9AM and 10AM by spitting into grading tubes which were immediately frozen. Teeth brushing and eating or drinking was not allowed for 60 minutes before saliva sampling.

VAS scale and unstimulated saliva were measured at baseline and last day of the LLLT.

On the assay day, saliva samples were defrosted and concentrations of cortisol were determined by manufacturer's instructions for Salimetrics commercial ELISA-kit (Salimetrics, LCC, State College, PA, USA). The range of manufacturer’s reference values was from 0.007-0.115 µg/dL.

In the study group, the LLLT was done daily for ten consecutive days (excluding weekend) (15) with Ga-Al-As light-emitting diode type of laser by a third person – nurse from our clinical Department in what way randomization was enabled. Laser wavelength was 685nm and treatment was performed on three reported burning sites. LLLT parameters are shown in Table 1

Table 1. Relevant technical data for LLLT (BTL2000 Medical Technologies, s.r.o., Czech Republic).

Parameter (unit)	value
Wavelenght (nm)	685
Dose (J/cm2)	2.00
Power (mW)	30
Power density (W/cm2)	0.003
Number of treatments	10
Single treatment duration (sec)	381
Treated surface area (cm2)	3
Distance (cm)	0.5
Frequency (Hz)	5.20
Cumulative dose (J/cm2)	60

The treatment protocol was identical for the control group, but LLLT was done with inactive laser probe which was only emitting the audio signal.

Statistical analysis

Statistical analysis was performed by MedCalc statistical software, version 18.10.2. (Ostend, Belgium). Differences between groups regarding age and sex were tested by the Mann-Whitney and Fischer-exact test respectively. The Wilcoxon signed-rank test for related samples was used to compare the differences between VAS scores and salivary cortisol level before and after LLLT. The level of significance was set at 0.05 (p<0.05).

Results

A total of 23 patients, 20 women and three men (one in the study group and two in the control group) participated in the study. Median age of the patients in the study group was 61 (47 - 70) years and in the control group 62 (50 - 69) years. Age and sex distribution did not differ between groups (Table 2). All participants have completed the study without any reported side effects of the therapy as evidenced by their subjective responses and control examination of the oral mucosa. The results have shown that VAS scores were significantly lower in both groups after LLLT (study group: median 5.5 (4-9) to median 4 (3-7); control group: median 5 (0-8) to median 3 (1.5-6.5)) (Table 3).

Table 2. Caracteristics of the study and control group.

	Study group	Control group	P
Male (N)	1	2	>0.05*
Female (N)	11	9
Total (N)	12	11
Age Median (range)	61 (47-70)	62 (50-69)	>0.05**

*Fisher-exact test

**Mann-Whitney test

Table 3. VAS scores and salivary cortisol levels before and after LLLT.

	VAS scores before LLLT (median (range))	VAS scores after LLLT (median (range))	P*	Salivary cortisol level before LLLT (median (range)µg/dL)	Salivary cortisol level after LLLT (median (range)µg/dL)	P*
Laser probe turned on	5.5 (4-9)	4 (3-7)	<0.05	0.377 (0.198-1.333)	0.305 (0.122-0.831)	<0.05
Laser probe turned off	5 (0-8)	3 (1.5-6.5)	<0.05	0.313 (0.137-0.85)	0.222 (0.122-0.498)	<0.05

* Wilcoxon signed rank test

Our results have shown that all patients had elevated salivary cortisol levels prior to the LLLT (> 0.115 µg/dL). Decreased salivary cortisol levels were detected in both groups after LLLT (study group: median 0.337 (0.198-1.333) µg/dL to median 0.305 (0.122-0.831) µg/dL; control group: median 0.313 (0.137-0.85) µg/dL to median 0.222 (0.122-0.498) µg/dL (Table 3).

Discussion

The results of scientific studies published so far regarding the treatment of BMS patients with LLLT are not consistent. Additionally, previous studies have shown controversial results regarding the salivary cortisol levels in BMS patients. Amenábar et al. (3) and Kim et al. (7) have shown elevated cortisol levels in BMS patients, which is in disagreement with Nosratzehi et al. (8) and de Souza et al. (9).

In our study, we have reported decreased salivary cortisol levels in BMS patients after the LLLT when the laser diode was switched on and when the laser diode was switched off. Bearing in mind the fact that the cortisol level increases in stress situations (6), we may assume that patients were less stressed after the therapy. The patients also had a smaller number of burning symptoms after the therapy, which was confirmed by decreased VAS scores that were significantly lower after the LLLT in both groups. Similar results were obtained by Sikora et al (15) who also reported decreased VAS scores in BMS patients regardless of the laser probe being on or off.

A recent study of Spanemberg et al. (14) has shown that the effect of LLLT was satisfactory in a small number of BMS patients and that LLLT could be alternative to treatment with psychoactive drugs. Their study included 12 patients in the study group and 9 patients in the control group. This study differs from ours in the treatment protocol – 2 weeks sessions for 4 weeks (14) contrary to 5 weeks sessions for 2 weeks (ours) and laser wavelength – infrared (14) contrary to red (ours). A group of authors in 2015 (16) have reported a reduction of burning symptoms in BMS patients treated with three different laser treatment protocols including infrared and red diode and different weekly application. These data point out that different modality of LLLT could be effective in treatment of patients with BMS but also a need for defining a standard protocol for low level laser therapy in BMS patients.

Valenzuela and Lopez-Jornet (17) reported that LLLT reduces symptoms slightly in BMS patients which they obtained by use of OHIP-14, VAS and Hospital anxiety and depression scale. Their results have shown that OHIP-14 and VAS scores were significantly lower after two weeks. However, no significant reduction was found from 2 to 4 weeks therapy.

Al-Maweri et al. (18) made a systemic meta-analysis of the published literature on BMS and LLLT and included results of five randomized clinical trials (16, 19-22), three uncontrolled clinical trials (23-25) and two case series (26, 27). Sugaya et al. (19) have found that LLLT has the same effect as placebo in BMS patients. Pezelj-Ribaric et al. (22) did not find any significant reductions of VAS scores after LLLT. Although the abovementioned studies have shown negative results, Al-Maweri et al. (18) concluded that LLLT could be effective for reducing pain in BMS patients. They pointed out that well designed double-blinded randomized clinical trials are necessary to evaluate the role of LLLT in patients with BMS.

The results of previous studies have shown an association between pain intensity, salivary cortisol levels and psychological stress (28, 29). Also, the patients with chronic pain show greater improvement when they get treatment, whether or not it is a placebo, than without any therapy at all (30). Every possible treatment option is associated with beneficial cognitive effects such as stress reduction (31, 32). Although Vukoja et al. (33) suggest that positive effect of LLLT is probably the result of a placebo, previous data (30-32) justifies the use of LLLT despite possible placebo effects.

The advantages of our study are that, to our knowledge, this is the first study measuring salivary cortisol levels in patients with BMS before and after LLLT. The limitations of the study refer to cortisol measurement that may be affected by wake up-time and it cannot be influenced by the researcher (34).

Conclusion

Despite the limitations of the study, our results have shown that LLLT can be useful for reducing burning symptoms and the salivary cortisol level, thus reducing stress. However, further research on a larger number of BMS patients is needed to clarify whether the positive results are attributed to LLLT effectiveness or to placebo effect.