Melatonin as an Adjunctive Therapy for Chronic Periodontitis.

Is Dietary Melatonin Supplementation a Viable Adjunctive Therapy for Chronic Periodontitis? A Preliminary Randomized Clinical Trial.

Background: Melatonin is an endogenous indoleamine synthesized mainly by pineal gland and showed anti-inflammatory and antioxidant properties. Moreover, melatonin promotes osteoblastic differentiation and suppresses osteoclastic formation. This randomized clinical trial (RCT) was aimed to assess the additive effect of melatonin supplementation in generalized chronic periodontitis (gCP) patients with insomnia who received scaling and root planing (SRP). Methods: Seventy four gCP patients with primary insomnia participated in this 6-month RCT and randomly distributed between two groups. Melatonin group (MTN+SRP group, n=38) included patients who were subjected to SRP with a 2- month regimen of 10 mg oral melatonin supplementation capsule once daily at bed-time. In the control group (Placebo+SRP group, n=36), SRP was performed for participants provided with matching placebo capsules. The primary treatment outcome included the clinical attachment gain (CAG) after 3 and 6 months of therapy, whereas, the changes in pocket depth ,bleeding on probing (BOP%) and salivary tumor-necrosis factor-α (TNF-α) levels represented the secondary outcomes.

INTRODUCTION Chronic periodontitis is a local chronic inflammation of the tooth supporting apparatus initiated by specific microorganisms including Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola, which are collectively known as the "red complex". It was revealed that periodontal tissue damage begins as inflammatory neutrophil mediated reaction followed by chronic infiltration of monocytes. Previous investigations have reported that most of periodontal tissue destruction is directly caused by the host immune response to the aforementioned specific periodontopathogenic bacteria. Few years ago, host modulatory therapy (HMT) has emerged as a new concept for the treatment of destructive periodontal diseases. The aim of HMT is to decrease the periodontal tissue damage by reducing the destructive arm of the host immune response and boosting its regenerative features. HMT includes systemically or locally delivered therapeutics which are utilized as adjuncts to conventional periodontal treatment. A variety of different drugs have been assessed as host modulatory therapeutics such as non-steroidal anti-inflammatory drugs (NSAIDs), tetracyclines and bisphosphonates. Systemic or topical NSAIDs had significant serious adverse events when used for a long time and after stopping their use, a 'rebound' effect' was observed. Subantimicrobial dose of doxycycline (SDD) revealed beneficial outcomes when combined with non surgical treatment of chronic periodontitis; nevertheless, some adverse reactions associated with tetracyclines particularly photosensitivity were noted. Bisphosphonates were proven to significantly increase alveolar bone density and possess anticollgenase activity , however, their long term use could lead to avascular necrosis of jaw bones especially after tooth extraction. Thus, researchers worldwide are continually searching for new host modulatory agents to be used as adjunctive treatments for periodontal disease. To-date, melatonin (N-acetyl-5-methoxytryptamine) is a well-investigated, endogenous indoleamine which is primarily secreted from the pineal gland and responsible for regulation of sleep/waking cycle. Melatonin possesses a variety of essential properties such as anti-inflammatory, antioxidant, oncostatic and neuroprotective actions. Moreover, melatonin was recently found to promote osteoblastic differentiation and suppress osteoclastic formation through downregulation of the receptor activator of nuclear к-B ligand (RANKL). In lipopolysaccharide-induced experimental periodontitis in rats, locally applied melatonin significantly decreased bone resorption in comparison with rats which received no treatment. Thus, it was proposed that topically applied melatonin can serve as an adjunct to conventional periodontal treatments including scaling and root planing (SRP), as well as, surgical periodontal therapy. Recent studies have revealed that salivary melatonin levels were significantly reduced in subjects with periodontal disease, suggesting that melatonin could serve as a biomarker for periodontal diagnosis and used as a potential therapeutic in different periodontal diseases. Recently, topical application of 1% melatonin gel on the attached gingivae in diabetic individuals with periodontal disease resulted in significantly decreased pocket depth and gingival index as well as lower levels of serum IL-6 and C-reactive protein after therapy. Accordingly, we hypothesized that periodontal treatment with daily dietary supplementation of melatonin adjunctive to SRP in generalized chronic periodontitis (gCP) patients suffering from primary insomnia could improve the periodontal outcomes. Hence, the objective of the present clinical trial is to assess whether the adjunctive therapy of daily melatonin supplementation to SRP compared to placebo with SRP, ameliorates clinical periodontal parameters and reduces salivary TNF-α levels after 3 and 6 months of therapy in patients with gCP and primary insomnia. MATERIALS AND METHODS Study Population: The present study was conducted in accordance with the seventh revision of Helsinky declaration in 2013 and approved by the Institutional Review Board (IRB) of Mansoura University. Individuals were selected during their visits to the Periodontics clinics at the Faculty of Dentistry, Mansoura University, from July, 2016 till September , 2017. Individuals were asked to answer prepared questionnaires about their general medical status and sleep/awakening habits followed by diagnosing the periodontal condition to select eligible participants. Patients were considered eligible for participation if they had Athens Insomnia Scale (AIS) score ≥ 6 (provided that individuals have primary insomnia which not related to any other systemic causes or drug intake according to the International Classification of Sleep Disorders , ICSD-3). Each selected patient should have at least 20 teeth. The enrolled patients were diagnosed to have moderate to severe gCP based on Armitage's classification. Exclusion criteria included diabetes mellitus , smokers, individuals having night work shifts, cancer patients , patients with autoimmune diseases or osteoporosis, users of antibiotics or non-steroidal anti-inflammatory drugs within the last 3 months and patients who were subjected to any periodontal therapy during the last year. Enrolled patients signed informed consents prior to their participation in the clinical trial. Trial design: Seventy four patients diagnosed with gCP and primary insomnia were randomly assigned by a coin toss to either receive 10 mg oral melatonin capsule (Puritan's Pride, Inc., Holbrook, NY, USA) once per day at bedtime for two months in conjunction with SRP (MTN+SRP group, n = 38), or matching placebo capsules for 2 months plus SRP (Placebo+SRP group, n = 36) in this double-blinded parallel randomized controlled trial (RCT). The randomization process was performed in absence of the working investigators. By a third party (pharmacist), sealed glass bottles were packed with 30 capsules of either melatonin or matching placebo and then coded with specific labels unknown to working investigators and patients. Patients in both groups were instructed to have one capsule at bedtime from the given bottle. Compliance of patients were assessed by counting the left capsules in the returned bottle at the end of each month. Adverse effects of the given drugs were monitored on a weekly basis for two months. All patients received meticulous thorough SRP with an ultrasonic scaler and hand curettes in a single visit by experienced periodontist. For all participants, strict oral hygiene instructions were given and 0.12% chlorhexidine mouthwash was prescribed for two weeks following SRP. Periodontal records: Periodontal measures including pocket depth (PD), clinical attachment level (CAL), dichotomous bleeding on probing (BOP) expressed as percentage, gingival (GI) and plaque (PI) indices were recorded at baseline, 3 and 6 months of therapy by a single examiner. To achieve intra-examiner calibration, periodontal mea-surements of 10 patients were performed twice within 2 days prior to RCT conduction. Calibration was approved when measurements of PD and CAL in the two times were within 1 mm variance in more than 90% of all measurements. Saliva Collection: Saliva samples were collected in the morning between 8 to 10 a.m. from all participants after fasting overnight. All participants were asked not to have any drinks except water after arousing. Whole unstimulated saliva samples were harvested by expectoration into 5 mL sterile polypropylene tubes before recording the clinical measurements. All saliva samples were centrifuged to remove debris, and immediately frozen and stored at -80οC until evaluation time. Biochemical Assessment: TNF-α level was determined in saliva by enzyme-linked immunosorbent assay (ELISA). Saliva samples were pipetted into clean microcap tubes and centrifuged at 10,000 × g for 5 minutes. Then, the supernatants were placed in clean microcap tubes and used immediately for ELISA assay. Human- TNF-α ELISA development kit (R&D systems, Abingdon Science Park,Abingdon, UK) was used to quantify this molecule in saliva samples according to the manufacturers' recommendations. The results of salivary TNF-α assay are expressed as concentrations in nanograms per milliliter. Trial outcomes: The primary outcome of the present RCT was the change in clinical attachment gain (CAG) after 3 and 6 months of therapy, whereas, the secondary end-points included the change in pocket depth,BOP% and salivary TNF-α levels after 3 and 6 months. Assessment of untoward drug effects: Systemic signs and symptoms were recorded every week after SRP and initiation of medication intake during the first two months. At the end of RCT, patients indicated for additional periodontal treatments were scheduled as necessary. Statistical Analyses: This RCT was powered to 80% level to detect an average reduction of PD and CAL equal to 0.5 mm variation between groups. The calculated minimum sample size required to achieve 80% power was 30 patients per group. Kolmogrov-Smirnov test of normality was used to explore data normality. Parametric data were presented as mean ± SD except salivary TNF-α concentrations which were expressed as mean± SEM (standard error of the mean). Baseline data were compared by using Student's t-test. One-way analysis of variance (ANOVA) with Holm-Sidak post hoc correction for multiple comparisons were used to determine significant differences at different time points. The level of significance was adjusted at 5%. Statistical analyses were performed by using the Statistical Package for the Social Sciences v. 20, SPSS, Chi-cago, IL.

10 mg melatonin capsule was given to participants in the test group once per day for only 2 months after performing scaling and root planing (SRP) during the whole 6- month period of the study.

Matching placebo capsule was given to the control group once daily for 2 months after receiving scaling and root planing (SRP) during the whole 6- month period of the study.

After performing scaling and root planing (SRP) in the test group, melatonin 10 mg oral capsule was taken once daily at night prior to bedtime for only 2 months during the 6-month study period

After finishing scaling and root planing (SRP) in the control group, placebo capsule was taken once daily at bedtime for two months.

Clinical attachment level was assessed at baseline,3 and 6 months after receiving scaling and root planing.

Bleeding on probing (0 for absent and 1 for present) was evaluated at baseline and after 3 and 6 months of therapy.

Inclusion Criteria: Patients were considered eligible for participation if they had Athens Insomnia Scale (AIS) score ≥ 6. Each selected patient should have at least 20 teeth. The enrolled patients were diagnosed to have moderate to severe gCP based on Armitage's classification. Exclusion Criteria: diabetes mellitus. smokers. individuals having night work shifts. cancer patients. patients with autoimmune diseases or osteoporosis. users of antibiotics or non-steroidal anti-inflammatory drugs within the last 3 months. patients who were subjected to any periodontal therapy during the last year.

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