Clinical and Biochemical Evaluation of the Use of Locally Delivered Metformin as an Adjunctive Therapy to Non Surgical Periodontal Treatment of Periodontitis Patients (Randomized Controlled Study)
Phase: NA
Conditions: Periodontitis
Sponsor: Ain Shams University

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Clinical and Biochemical Evaluation of the Use of 
Locally Delivered Metformin as an adjunctive 
therapy to Non Surgical Periodontal Treatment of 
Periodontitis Patients  
(Randomized Controlled Study) 
Protocol Submitted to Department of Oral Medicine, 
Periodontology and Oral Diagnosis, Faculty of Dentistry,          
Ain Shams University In Partial Fulfillment of the Requirements 
for Masters' Degree in Periodontology 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Faculty of Dentistry 
Ain Shams University 
2019 

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Review of literature 
      Periodontitis is a chronic multifactorial inflammatory disease associated 
with dysbiotic plaque biofilms and characterized by progressive destruction 
of the tooth-supporting apparatus. Its primary features include the loss of 
periodontal tissue support, manifested through clinical attachment loss 
(CAL) and radiographically assessed alveolar bone loss, presence of 
periodontal pocketing (that are a hallmark of the disease and can eventually 
lead to tooth loss) and gingival bleeding (Papapanou et al., 2018).  
 
      The classification of periodontal diseases and conditions in 1999 
included gingival diseases including (dental plaque-induced gingival 
diseases and non-plaque-induced ginigival lesions), chronic periodonitits, 
aggressive periodonitits, periodonitits as a manifestation of systemic 
diseases, necrotizing periodontal diseases, abscess of the periodontium, 
periodontitis with endodontic lesions and developmental or acquired 
deformities (Armitage, 1999). 
 
      The new classification of periodontal and peri-implant diseases and 
conditions in 2017 classified these diseases into four main categories: First 
category concerned with periodontal health & gingival diseases either dental 
biofilm induced gingivitis and non-dental biofilm induced gingivitis. Second 
category concerned with periodonitits which can be classified into 
necrotizing periodontitis, periodontitis as a  manifestation  of  systemic  
disease  and  periodontitis  which  involve  the forms of the disease 
previously recognized as “chronic” or “aggressive”. Third category 
concerned with other conditions affecting the periodontium as systemic 
diseases, traumatic occlusion, mucogingival deformities, tooth related 
factors, periodontal abscesses and endodontic-periodontal diseases. Fourth 
category concerned with peri-implant diseases and conditions as peri-
implant health, peri-implant mucositis, peri-implantitis and peri-implant soft 
& hard tissue deficiencies.  
 
      Periodontitis is described in four stages which depend on clinical 
attachment loss (CAL) ranging from Stage 1: Initial periodontitis (CAL 1-2 
mm), Stage 2: Moderate periodontitis (CAL 3-4 mm), Stage 3: Severe 
periodontitis with potential for additional tooth loss and Stage 4: Severe 
periodontitis with potential for loss of dentition (CAL ≥5 mm). Grading 
focuses on assessing risk factors as smoking, systemic considerations as 
diabetes, and outcomes of non-surgical periodontal therapy. Grade A: 
Slow rate of progression (no CAL loss over 5 years), Grade B: Moderate 
rate of progression (CAL loss <2 mm over 5 years) and Grade C: Rapid rate 
of progression (≥2 mm over 5 years) (Caton et al., 2018). 

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      The pathogenesis of periodontal diseases is mediated by the 
inflammatory response to bacteria in the dental biofilm. The immune 
response to infection is regulated by cytokine and chemokine signals. 
Cytokines and chemokines (chemotactic cytokines) are the messages 
between cells. Cytokines are low-molecular-weight proteins involved in the 
initiation and further stages of inflammation, in which they regulate the 
amplitude and the duration of the response. The genetic regulation leading to 
the secretion of proinflammatory cytokines from a variety of cells is 
generally dependent on the activation of nuclear factor kappa-B 
transcription. Cytokines are produced by resident cells, such as epithelial 
cells and fibroblasts, and by phagocytes (neutrophils and macrophages) in 
the acute and early chronic phases of inflammation, and by immune cells 
(lymphocytes) in established and advanced lesions. After recognition and 
presentation of microbes to the appropriate cells, cytokines of the innate 
response, including tumor necrosis factor alpha, interleukin-1beta and 
interleukin-6, are the first to appear in the periodontal disease pathogenesis 
pathways. It is suggested that individuals who produce high levels of these 
mediators in response to the dysbiosis will experience more and severe 
tissue loss (Hanada &Yoshimura, 2002). 
 
      As an adjunctive measurement to aid proper diagnosis, evaluation of 
levels of various biomarkers is one possible tool. A biomarker is an indicator 
of a biological situation, which can help to distinguish between health and 
disease (Li & Wang, 2014). In periodontology, one of the main challenges is 
to identify diagnostic and prognostic biomarkers for determination of disease 
activity and to diﬀerentiate between health and disease (Buduneli & Kinane, 
2011).  
 
      Sources of biomarkers for periodontal disease include saliva, serum and 
gingival crevicular ﬂuid (Ghallab, 2017). GCF is an inflammatory exudate 
that can be collected at the gingival margin or within the gingival crevice 
which contains microorganisms, their products, and host derived substances 
(cytokines, enzymes, immunoglobulins, tissue degradation products, 
immune system cells) (Bostanci, & Belibasakis, 2018).       
 
      Many studies examined bone resorption biomarkers such as RANKL and 
OPG in GCF or in health and disease. RANKL is a TNF family member 
protein, in health, its principal sources are osteoblasts, in disease, it is 
synthesized mostly from T and B lymphocytes (Kawai et al., 2006). 
Receptor activator of nuclear factor kappa-beta (RANK) is the receptor of 
RANKL, located on pre-osteoclasts. RANK-RANKL interaction activates 
transcription factors of osteoclastogenesis (Teitelbaum & Ross, 2003). OPG 
is a soluble glycoprotein, the decoy receptor of RANKL, is synthesized from 
many diﬀerent types of cells, Formation of RANKL-OPG complex inhibits  

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RANK-RANKL interaction via blocking their binding sites (Baud'huin et 
al., 2013). Elevated concentrations of RANKL in GCF of periodontitis when 
compared to healthy areas were demonstrated by (Gürlek et al., 2017). 
 
      Lowering bacterial load and inhibiting progression of inflammation are 
the primary treatment goals that are achieved by mechanical debridement 
such as scaling and root planing (SRP), a gold standard treatment for 
periodontitis (Socransky & Haffajee, 2000). However, SRP shows physical 
limitations in certain inaccessible areas such as deep periodontal pockets, 
furcations and interproximal areas of misaligned teeth, which does not allow 
complete reduction of anaerobic infection and recurrence (Cobb, 2005). 
 
      Considering these difficulties, various adjunctive treatments have been 
investigated to supplement Scaling and Root Planing as local delivery drugs. 
These include systemic and localized delivery of antibiotics (Clarithromycin 
gel, minocycline microspheres), chlorohexidine, bisphosphonates, statins & 
photodynamic therapy to reduce bacterial counts and improve clinical 
periodontal parameters such as probing depth reduction, gaining clinical 
attachment level and bone defect (BD) fill in periodontitis (Zandbergen et 
al., 2013), (Akram et al., 2016), (Abduljabbar et al., 2017). 
 
      Metformin a biguanide used in treatment of diabetes mellitus type II as 
an oral hypoglycemic drug, has shown stimulating effect on osteoblastic 
lineages leading to osteoblast differentiation and bone formation. MF 
increased type-I collagen production in both osteoblast-like cell lines and 
stimulated alkaline phosphate activity in MC3T3E1 osteoblasts (Cortizo et 
al., 2005). Since MF benefits on fracture reduction and takes positive action 
on osteoblasts, it appears possible that this agent may have protective effects 
on bone loss. Recent in vivo and in vitro studies suggest that metformin 
reduces receptor activator for nuclear factor-kappa B ligand (RANKL) and 
stimulates osteoprotegerin (OPG) expression in osteoblasts, further inhibits 
osteoclast differentiation and prevents bone loss in ovariectomized rats (Mai 
et al., 2011).  In another study on ligature-induced periodontitis, the MF 
treatment of rats induced a significant reduction in alveolar bone loss 
compared to vehicle-treated rats (Bak et al., 2010).  
 
      Randomized clinical trials regarding the use of Locally delivered 
metformin in periodonitits, The 1% MF gel used along with Scaling and 
Root Planing was found to provide improvement in clinical and 
radiographical parameters confirmed by Studies done by (Rao et al., 2013) 
& (Pradeep et al., 2017) in the form of significant increase in the PD 
reduction, CAL gain, and improved IBD depth reduction in vertical bone 
defects. So In this study we will evaluate The 1% MF gel along with SRP 
was found to provide improvement in clinical and radiological parameters 

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confirmed by Studies done by (Rao et al., 2013) & (Pradeep et al., 2017) in 
the form of significant increase in the PD reduction, CAL gain, and 
improved IBD depth reduction in vertical bone defects. So In this study we 
will evaluate the effect of the locally delivered metformin in the form of 
clinical and biochemical assessment. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Aim of Study 
 
Primary outcome :  
      To evaluate the clinical parameters before & after the application of 
locally delivered metfromin as an adjunctive therapy to non surgical 
periodontal treatment of Stage III periodontitis.   
Secondary outcome : 
      To measure the RANKL and OPG levels in GCF before & after 
application of metformin as an adjunctive therapy to non surgical 
periodontal treatment of Stage III periodontitis.   
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Subjects and Methods 
Study design and Sample size calculation : 
          In this randomized controlled study, total sample size of 16 patients (8 
patient/group) were selected for this study. The sample size was calculated 
to yield a 95% power in the results of this study based on the results of a 
previous study (Pradeep et.al., 2017) in which the authors found a significant 
difference in IBD depth between test and placebo groups (effect size=.2.08 
mm, alpha (2 tailed)=.05). The number will be increased to 20 in each group 
to compensate for dropouts. 
Patient Selection : 
         Forty patients, having periodontitis, will participate in this randomized 
clinical study. The subjects will be recruited consecutively from the 
outpatient clinic of Oral Medicine, Periodontology, oral Diagnosis and 
Radiology Department, Faculty of Dentistry, Ain Shams University The 
purpose of the study will be explained to all patients and an informed 
consent will be signed before the conduction of the study. The proposal will 
be presented to the faculty of Dentistry Ain Shams University Research 
Ethics committee and have to be approved before starting the research 
 
Inclusion Criteria will be : 
1. Patients with Stage III & IV Periodontitis. 
2. Age range (30 - 50) years. 
3. Good compliance with the plaque control instructions following initial 
therapy. 
4. Availability for follow up and maintenance program. 
Exclusion Criteria will be : 
1. Smokers  
2. Systemic diseases which could influence the outcome of the therapy                
(According to Cornell Medical Index-Health Questionnaire). 
3. Pregnant and lactating females. 
4. Vulnerable groups of patients’ e.g (prisoners, handicapped patients and 
decisionally impaired individuals). 
 
 
 

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For all patients who are suitable for the study the following clinical 
evaluation parameters will be measured : 
a. Plaque index (PI) (Silness & Löe, 1964) 
b. Gingival index (GI) (Löe & Silness, 1963) 
c. Probing depth (PD) (Caton, 1980). 
d. Clinical attachment level (CAL) (Ramfjord, 1967).  
e. Standardized Periapical Radiograph  
• All evaluations will be done pre operatively (base line) and 6 months post 
operatively. 
• Full mouth assessment will be done and then the deepest areas will be 
evaluated to receive the local drug. 
 
Grouping Criteria : 
     The study consists of two groups, each containing 20 patients. Patients 
will be randomly selected using computer generated randomization 
(www.randomizer.org) and this will be performed by another individual 
other than the investigator.  
Group I (20 patients) : (Treatment Group) 
 Supra and sub gingival scaling and root planning will be performed for 
patients having Stage III Periodonitits. Followed by local application of 
metformin gel. Oral Hygiene measures will be instructed following 
treatment and maintenance visits will be given to them. 
Group II (20 patients) : (Control Group) 
 Supra and sub gingival scaling and root planning will be performed to Stage 
III Periodonitits patients. Followed by local application of placebo gel. Oral 
Hygiene measures will be instructed following treatment and maintenance 
visits will be given to them. 
 
Metfromin Preparation : 
MF gel will be prepared, briefly all the required ingredients of the formulation will 
be weighed accurately. Dry gellan gum powder will be dispersed in distilled water 
maintained at 95°C. The dispersion will be stirred at 95°C for 20 min using a 
magnetic stirrer to facilitate hydration of gellan gum. The required amount of 
mannitol will be added to the gellan gum solution with continuous stirring and the 
temperature will be maintained above 80°C. Weighed amount of MF will be added 
with stirring. Then sucralose, citric acid, and preservatives (methylparaben, 
propylparaben) will be added with stirring. Finally, required amount of sodium 
citrate will be dissolved in 10 ml of distilled water and will be added to the 
mixture. The mixture will be allowed to cool to room temperature to form gel. 
(Mohapatra et al., 2008) 

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Steps of the Research : 
 
1. After enrollment, a detailed case history will be recorded. 
2. Initial therapy by full mouth supra and subgingival scaling and root 
planing using ultrasonic scaler and hand instruments will be done on 
all patients. 
3. The patients will be given instructions for self-performed plaque 
control measures with soft dental brush and interdental cleaning using 
dental floss or interdental brush. 
4. Baseline sampling and the local drug delivery in both study and 
control groups will be performed next day of scaling to avoid 
contamination by blood. Samples from the gingival crevicular fluid 
will be collected from all selected sites using standardized sterile 
periopaper inserted into the deepest part of each periodontal pocket 
and left in situ for 30 seconds. 
5. The area will be completely dried using oil free air syringe, and then 
the site will be isolated with cotton rolls to prevent contamination 
from saliva. The local drug delivery gel will be placed in the 
periodontal pockets using a dedicated syringe until the gel flowed out 
from the gingival margin. 
 
Outcomes Assessment : 
 
a) Clinical Assessment 
Clinical Assessment will be done using William’s graduated probe at 
baseline and six months (180 days) later. 
 
b) Biochemical Assessment  
Biochemical assessment will be done at baseline, and one month after local 
drug delivery. RANKL & OPG concentrations will be determined using a 
commercial human RANKL & OPG ELISA Kits.  Measurements will be 
performed according to the manufacturer’s instructions. 
 
Statistical Analysis : 
- All data will be collected, tabulated and statistically analyzed by 
appropriate statistical program. 
 

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