Concentrated Growth Factor Versus Platelets-rich Fibrin in Revascularization of Immature Necrotic Mandibular First Permanent Molar.
Phase: NA
Conditions: Revascularization of Immature Young Permenant Teeth
Sponsor: Minia University

 
 
 
 
 
CONCENTRATED GROWTH FACTOR VERSUS 
PLATELETS-RICH FIBRIN IN REVASCULARIZATION 
OF IMMATURE NECROTIC MANDIBULAR FIRST 
PERMANENT MOLAR 
Parallel Randomized Controlled Trial  
 
Protocol 
Submitted to 
Faculty of Dentistry, Minia University in Partial Fulfillment of 
the Requirements of the Doctor Degree in Dental Science 
(Pediatric Dentistry)  
By 
Bothaina Ahmed Ibrahim Ahmed 
Assistant lecturer in Pediatric and Community Department, 
South Valley University Faculty of Dentistry 
 
Faculty of Dentistry 
Minia University 
2025 

 
 
 
 
 
 
Supervisors 
 
                 Assistant Prof. Ahmad Abdel Hamid Abdel Kader Elheeny 
Assistant professor of Pediatric and Community Dentistry 
Faculty of Dentistry, Minia University 
 
Dr. Kareem Mohamed Mahmod Eissawy 
Lecturer of Pediatric and Community Dentistry 
Faculty of Dentistry, Minia University 
 
 

 
Introduction 
The contemporary non-surgical endodontic management of immature teeth has 
shown favorable outcome rates of 95% in teeth diagnosed with irreversible pulpitis1 
and 85% in necrotic cases.2 
There is a paradigm shift in the treatment protocol of such teeth from conventional 
root canal treatment to regenerative endodontic treatment (RET) 3-4. Hargreaves et 
al., in 2008 reported that regenerative endodontic procedures are possible by 
application of the principles of tissue engineering that require the spatial orientation 
of stem cells, signaling molecules, and the scaffold. Such three elements of tissue 
engineering as seed cells, scaffold materials and growth factors have an essential 
impact on the efficacy of RET.5 
A suitable scaffold can provide a suitable location for seed cells and biological 
conditions conducive to cell metabolism, and regulate their differentiation and 
proliferation. The main scaffolds for regenerative endodontic therapy are autologous 
blood clots, autologous platelet concentrate, biomaterial scaffolds, and so on.6-8 
The traditional method in RET is to stimulate apical bleeding, fill the root canal with 
blood, and use blood clots (BC) as scaffolds for pulp revascularization. Blood clots 
as a scaffold in RET had a good practicability and success rate [6]. However, it was 
found that clinically there were quite a few cases with no or insufficient blood clots 
which were disable to fill the root canal or support the pressure from the crown 
sealing material, leading to treatment failure. Therefore, it is necessary to find 
scaffolds to replace blood clots. Endogenous materials replacing blood clots as 
scaffolds include platelet-rich plasma (PRP), platelet-rich fibrin (PRF), and 
concentrated growth factor (CGF).8 

The use of platelets for regenerative medicine has increased in recent years. Platelets, 
which contain growth factors, play major roles in cell migration, proliferation, 
differentiation and angiogenesis and are associated with the tissue regeneration 
process. Autologous platelet concentrates (APCs) are produced by the centrifugation 
of venous blood at different speeds and the use or non-use of thrombin and 
anticoagulant. As a result of these processing protocols, a fibrin clot is formed that 
contains platelets and leukocytes. 9 
The main generations of APCs are platelet-rich plasma (PRP), platelet-rich fibrin 
(PRF) and concentrated growth factor (CGF). The efficacy of platelet concentrates 
in promoting wound healing and tissue regeneration has been at the center of 
scientific interest over the past few decades.10 
Platelets include growth factors (GFs) such as basic fibroblast growth factor (bFGF), 
vascular endothelial growth factor (VEGF), insulin-like growth factor-1 (IGF-1), 
transforming growth factor-1 (TGF-1) and platelet-derived growth factor-BB 
(PDGF-BB). There are significant differences in the amounts of GFs produced using 
the three different APC techniques (CGF, PRF and PRP). PRF and CGF produce 
significantly more GFs during the procedure as compared to PRP.11 
  CGF may play an role in vascular maintenance and angiogenesis due to its 
inclusion of CD34-positive cells 12 Besides CGF with a stronger regeneration ability 
contains more growth factors and fibrin matrix than PRF on account of the 
preparation of CGF adopts differential centrifugation with higher separation 
efficiency.12-13 
  Platelet concentrate products have good biological characteristics and are easy to 
produce. In recent years, they have been widely used in regenerative treatment of 

oral medicine, such as periodontal tissue regeneration and orthodontic tooth 
movement .14-15 
  Biodentine is a recently released tricalcium silicate-based material developed as a 
permanent dentin substitute to replace the damaged dentin.16 
  Biodentine is a two component material. The powder is mainly composed of 
Tricalcium silicates. It also contains Di Calcium Silicate as a second core material 
and Calcium Carbonate and Oxide as filler. The powder contains Zirconium oxide 
as a radio-opacifier. The liquid contains Calcium Chloride as a setting accelerator 
and a water reducing agent. The presence of a setting accelerator allows the material 
setting in 12 min and the presence of a water reducing agent avoids the formation of 
cracks within the material.17 
 Biodentine is a highly biocompatible material. This biocompatibility of the material 
was investigated through its direct application to human pulp cells simulating the 
direct pulp condition and indirectly through a dentin slice to simulate its indirect 
pulp capping in vivo. Under both conditions Biodentine was not found to affect 
target cell viability under in vivo application conditions.18Additionally, when 
Biodentine was applied onto human pulp cells to investigate its effects on their 
specific functions by studying expression of odontoblast specific functions such as 
expression of Nestin (a human odontoblast specific marker) and Dentin Sialoprotein, 
Bio dentine was not found to inhibit the expression of these proteins but rather 
induce their expression and the cells mineralization capacity.18-19 Further 
investigations demonstrated the absence of toxicity of Biodentine to human MG63 
human osteoblast cells with the MTT assay with properties comparable to that of 
MTA. 

  When Compared to MTA, Biodentine is easier to handle, stronger mechanically 
and has a shorter setting time.20It can be used as a temporary enamel substitute up to 
6 months and in different applications as a permanent dentin substitute without any 
surface treatment. Additionally, while discoloration with MTA21 and its derivatives 
have been reported in regenerative endodontics and seem to be mainly due to the 
presence of Bismuth oxide as a radio-opacifier, no discoloration of tooth crown has 
been reported after 48 months with Biodentine which does not contain Bismuth 
oxide but Zirconium oxide as a radio-opacifier.22-23 
 
 

Aim of the study 
 
The aim of this study is to compare and evaluate clinically and radiographically the 
effect of CGF and PRF with the application of biodentine on regeneration of necrotic 
immature lower first permanent molar and assessment of the clinical success rate 
over 18months. 
 
 

Materials and Methods 
This study will be based on the ethics committee of Minia University- 
Faculty of Dentistry. 
Ethical regulation:  
All patients and their caregivers will be informed in an easy detailed manner 
about the treatment procedures, expected outcome of the current treatment, 
possibility of any complications and assuring them to the presence of plan B in case 
of any complication. Verbal consent will be obtained from all children who will 
accept participation in the study, Parents will sign an informed consent laid down by 
the ethical committee of the Faculty of Dentistry / Minia University.  
Study design: 
 Parallel Randomized Control Trial.  
Two groups will be done:  
Group (1): Include children who will be treated with CGF and biodentine (control 
group). 
 Group (2): Include children who will be treated with PRF and biodentine (study 
group). 
Randomization and Concealment:  
The included patients will be randomized into one of the study groups. 
Randomization will be performed using a computer-generated block randomization 
system (Microsoft Office Excel, 2007). 

Concealment: A plan of management will be sealed in closed envelopes and 
numbered according to the randomization tables. Packing, sealing, and numbering 
will all be performed by a neutral health care provider other than the investigator.  
Sample size justification: 
The study will be with two parallel arms (1:1 allocation ratio). The sample size per 
group will be calculated according to the following equation for binary outcomes; 
N = (Zα/2 + Zβ) 2 ×p1(1 – p1) + p2(1 – p2)/δ 2 so the minimal sample size for an 
equal size clinical trial is 27 teeth in each group. We will recruit 30 teeth in each 
group for possible attrition. 
Study sample: 
 6-8 years old will be selected with no sex predilection for the study from 
outpatients of the Pediatric and Community Dentistry Department, Minia 
University, according to the following eligibility criteria: 
Inclusion Criteria: 
1. Children with immature necrotic lower fitrst permanent molar. 
2. No history of repeated pathosis or cellulitis. 
3. Patients accept the treatment modality. 
4. Patient-free from systemic, allergic or genetic problems. 
Exclusion Criteria:  
1. Tooth mobility. 
2. Teeth with non-restorable crowns. 
3. Children with facial cellulitis 
4. Children with draining fistula 

5. Children with a history of recurrent preapical  pathosis 
6. Children who will not comply to recall program. 
Clinical steps: 
 First Appointment: 
 1- Child communication and psychological management. 
2- Local anesthesia. 
3-Rubber dam application 
4- Disinfection of the tooth and the working field. 
5- Access cavity preparation and remove the necrotic tissue in the pulp chamber 
6- Disinfection of the root canal is accomplished by gently irrigating the root canal 
with a minimum of 20 mL 1.5% NaOCl for 5 minutes. It will be dispensed through 
a syringe and a 20-gauge needle 24-25. NaOCl is a potent antimicrobial agent and 
effectively dissolves necrotic and organic tissue. 26 Its solvent potential is dependent 
on its concentration and the frequency of fluid exchange. 27-29  
When irrigating with NaOCl, the needle should be introduced into the root canal to 
a point 2 mm short of the apical foramen28-29, and the NaOCl is slowly expressed 
from the syringe to prevent its introduction into the periapical tissues. .30 Irrigation 
is made with ultrasonic passive activation.  
7- Initial NaOCl irrigation is followed by irrigation with 20 mL sterile saline for 5 
minutes, followed by drying with paper point. 
8. The root canal will be filled with calcium hydroxide (UltraCal, Ultradent, Paris, 
France) as an intracanal medication using a hand plugger (Medesy srl, Maniago) 
9. A sterile cotton pellet will placed over the root canal medication then the access 
cavity will be sealed with a glass ionomer temporary restoration (RIVA self-cure, 
SDI Ltd, Paris, France) will be placed to seal the access cavity. 

10. Three weeks later, the tooth condition will be assessed. If the signs of infection 
still persist, the irrigation protocol and intracanal medication will be repeated. 
  Second Appointment (After 3 Weeks):  
1. Before proceeding to the next phase, there should be no adverse clinical 
signs and symptoms. 
2. Local anesthesia without a vasoconstrictor will be used. 
3. Rubber dam will be applied. 
4. The temporary filling and medicament will be removed by gently irrigating 
the root canal with a minimum of 20 mL of 1.5% NaOCl, 0.9% saline (5 
mL), and 17% EDTA (20 mL) (Cerkamed, Dental Products, Stalowa Wola, 
Poland). Then, an ultrasonic noncutting tool will be passively activated for 
one minute. Finally, the root canal will be dried with sterile, absorbent tips. 
5. Introduction of a sterile #20 K-file into the apical tissues 2 mm past the 
apical foramen to initiate bleeding into the root canal. .28, 30 Bleeding should 
be controlled so that it does not extend beyond a point approximately 3 mm 
apical to the CEJ. This is done by applying intracanal pressure with a sterile 
saline-soaked cotton pellet until a clot is formed. The estimated mean time 
for the establishment of a stable blood clot is 15 minutes 28-30 
 Preparation of PRF and CGF Clots: 
  A 10-mL blood sample will be drawn through venipuncture of the antecubital 
vein using a 10-mL sterile hypodermic syringe (Luer-Slip type, El Dawlia ico nos. 
19 and 67). The venous blood sample will be collected into sterile, specialized 
plain-type single-use centrifuge tubes (Hebei Xinle Sci&Tech Co., Ltd, 
Shijiazhuang, China) without anticoagulant additives.  
The centrifugation process will be performed using the Benchtop Low-Speed 
Centrifuge ET-12M device. For PRF preparation, the tubes were centrifuged at 
3000 revolutions per minute (rpm) for 10 minutes.(31) . 

For CGF preparation, the centrifugation process will follow a specific sequence: 
acceleration for 30 seconds, 2 minutes at 2700 rpm, 4 minutes at 2400 rpm, 4 
minutes at 2700 rpm, 3 minutes at 3000 rpm, and finally, deceleration for 36 
seconds19. After separation of the blood components, the topmost plasma-poor 
layer (straw-colored layer) will be removed using a sterile syringe. The fibrin-
gelatinous yellow substance will be carefully retrieved with an HTS 171c66.25 curved stainless steel 
college tweezer, then will be separated from the basal red blood cell fraction by a sterile scissor and 
dragged across a sterile 4 x 4 gauze pad to remove the excess(31) 
6. The prepared CGF fractions will be inserted into the root canal space using 
a small hand plugger (Dentsply Maillefer, Ballaigues, Switzerland) up to 3 
mm below the cementoemamel junction for group I teeth, Then  
Biodentine (Septodont) will be prepared according to the manufacturer’s 
instructions and   will be packed against a collagen restorable matrix 
(PARASORB Cone, Resorba Medical GmbH or Collacone, botiss 
biomaterials GmbH, Germany) using a gold-plated instrument (Nordent 
Dura Lite 1020, Victoria, Australia) 
7. For group II the prepared PRF fractions will be the root canal space using a 
small hand plugger (Dentsply Maillefer, Ballaigues, Switzerland) up to 3 
mm below the cementoemamel junction for group I teeth, Then  
Biodentine (Septodont) will be prepared according to the manufacturer’s 
instructions and   will be packed against a collagen resorbable matrix 
(PARASORB Cone, Resorba Medical GmbH or Collacone, botiss 
biomaterials GmbH, Germany) using a gold-plated instrument (Nordent 
Dura Lite 1020, Victoria, Australia). 
 

8. The access will be sealed with a layer of reinforced glass ionomer base 
(RIVA self-cure, SDI Limited). Finally, a resin composite (Filtek z250 
universal restorative, ESPE, St. Paul, MN) coronal restoration will be 
inserted 
Follow up:  
All patients will be recalled at 3, 6, 12 and 18 for clinical and digital radiographic 
evaluation. 
Clinical assessment: 
1. Absence of signs and symptoms of pain, mobility, sinus tract or periapical 
lesion. 
2. Normal responses to percussion, palpation, and normal pocket probing depths. 
 Radiographical assessment: 
1-Absence of periapical pathosis and furcation involvement.  
2- Elongation of the roots on follow-up periods. 
3- Normal PDL space width 
4-Normal bone trabeculation. 
Statistical methods 
 The collected data will be coded, tabulated, and statistically analyzed using IBM 
SPSS statistics (Statistical Package for Social Sciences) software version 22.0, 
IBM Corp., Chicago, USA, 2013. Quantitative normally distributed data will be 
described as mean± SD (standard deviation) after testing for normality using 
Shapiro-Wilk test, then compared using independent t-test if normally distributed. 
 

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