Revascularization of immature permanent teeth via blood clot or PRF Platelet rich fibrin: A review

Introduction

The regenerative endodontic treatment approach is a biological procedure that is designed to replace an impaired tooth structure, including dentin, root structure, and the pulp dentin complex. Seven possible techniques allow regenerative endodontics. These are postnatal stem cell therapy, root canal revascularization, pulp implant, scaffold implant, injectable scaffolds, three-dimensional cell printing, and gene therapy.¹ Root canal revascularization is one of the approaches to regenerative endodontics with the main goal of converting nonvital teeth into vital teeth. It is a novel approach for the management of immature necrotic permanent teeth.² Historically, apexification was induced using calcium hydroxide in the tooth before placing guttapercha.³ Though it was successful, there are several disadvantages,⁴ such asthe long time required for the formation of the calcified barrier (3–24 months),⁵ multiple appointments, and the effect of long-term calcium hydroxide on the mechanical properties of dentin.⁶

Mineral trioxide aggregate (MTA) is used as an alternative to calcium hydroxide apexification. It has several advantages over calcium hydroxide apexification.^7,8 These include a reduction in treatment time and fewer patient visits.⁹ But apexification treatments will not foster further root development and hence immature teeth remain susceptible to cervical root fracture because there is the formation of an apical barrier without thickening of root dentin.

Recently, a new approach has been introduced to treat immature teeth with pulp necrosis.¹⁰ Intracanal revascularization and continued root development can be achieved through procedures that preserve mesenchymal stem cells and dental pulp stem cells.¹¹ The revascularization of the teeth is based on the fact that the cells of the apical papilla can remain vital even after pulp necrosis which can proliferate into canal space.¹² An appropriate scaffold is required for the stem cell proliferation and differentiation to position the growth factors and stem cells in the canal space. Bloodclot,¹³ collagen,¹⁴ platelet-rich plasma (PRP),¹⁵and platelet-rich fibrin (PRF)¹⁶ can be used as scaffolding material in revascularization procedures. There are many studies on revascularization using blood clots and a few studies on PRF. This comprehensive review discusses the revascularization process by using blood clots and PRF and also histological studies.

Root canal revascularization via blood clot

Revascularization can be achieved by inducing bleeding into the canal space which results in the genesis of a blood clot. The blood clot acts as a matrix that traps the cells which initiate the formation of new tissue when the canal space is adequately disinfected. Disinfection can be achieved by intracanal irrigants with a combination of antibiotics.¹ Calcium hydroxide and triple antibiotic paste are used for disinfecting the canals. Triple antibiotic seems to be more appropriate as calcium hydroxide resulted in problems like weakening of dentinal walls along with tissue necrosis.²

Operative protocol for revascularization

It is a two-step procedure wherein the second step takes place after two to three weeks if the tooth is asymptomatic and shows reducing apical lesion.

First step

• Local anesthesia

• Rubber dam isolation of the tooth to be treated.

• Disinfection with 10% povidone-iodine (iso-Betadine) of the tooth before access opening

• Opening of the pulp chamber till canal entrance (pulpotomy)

• Irrigation with 20 mL sodium hypochlorite (1.25%– 5.25%) and then with physiological serum and finally with 2% chlorhexidine

• Instrumentation should be avoided in root canal • Drying the canal with paper cones

• Placement of triple antibiotic paste into the canal space

• Placing a cotton pellet at the orifice of the root canal

• Sealing the access cavity with a temporary restorative filling

Second step (Done two or three weeks later if the tooth is asymptomatic and/or absence of fistula)

• Local anesthesia without vasoconstrictor

• Rubber dam isolation of the tooth

• Disinfection of the tooth with 10% povidone-iodine (iso-Betadine) before opening it

• Opening the tooth to have access to the root canal

• Removal of the triple antibiotic paste or calcium hydroxide using irrigation with sodium hypochlorite (1.25%–5.25%) followed by physiological serum and finally with chlorhexidine (2%)

• Induction of bleeding at the apex. The blood level should be at the cement-enamel junction.

• Placement of MTA on the clot formed to achieve a hermetic seal

• Placement of a wet cotton ball on MTA.

• Finally, sealing the cavity with a temporary filling.²

There are many case series and case reports on revascularization of the pulp canal space of necrotized immature teeth. Table 1 shows the conventional method of revascularization by inducing bleeding in the canal.

History

In 1961 Nyguard–Ostby and Hijortdal³⁸ evaluated the role of blood clots in healing. They reported that in the root canal, the blood clot was formed by the granulation tissue growing from the periapical area and not from the blood cells originally contained in the clot. Even though these were encouraging findings, the regeneration of a pulp-dentin complex was not confirmed through histologic analysis. Thus, the regenerative procedures were ignored. Iwaya et al.,¹⁷ reported the first successful revascularization of a tooth with apical periodontitis and open apex. Since then revascularization has become a preferred treatment choice over apexification. Banchs and Trope¹⁸ reported that the tooth responded to vitality testing with cold after 2 years of follow-up. Chue and Huang¹⁹ concluded that despite peri radicular abscess,apexogenesis could occur in a permanent tooth with a wide open apex if pulp tissue survived. Cotti et al.,²⁰ reported that apical development and continuous increase in the dentinal wall thickness can occur with the revascularization technique. Shah et al.,¹³ in their pilot study observed that there are several advantages of revascularization. It requires a shorter treatment duration, progressive root development, is cost-effective, and strengthens the root. But long-term clinical results are not yet available. Calcification of the entire canal, compromised esthetics, and difficulty in future endodontic procedures if necessitated are some of the unwanted outcomes. Kim et al.,²⁶ suggested potential esthetic problems should be taken into account when minocycline is used as a canal medicament. Renolds et al.,²⁵ suggested sealing the dentinal tubules of the chamber ignorer forestall the crown discoloration as a consequence of triple antibiotic paste. Petrino et al.,²⁷ suggested the use of an esthetic without a vasoconstrictor. Norsat et al.,²⁸ reported the use of Calcium- enriched mixture (CEM) cement over the blood clot. It should be placed inside the canals to provide a good seal and favorable outcomes.Cehrelib et al., ²⁹ concluded that the tooth gave a positive cold response in cases where uninstrumentation was done. Mahamoud Torabinejad et al.,¹⁵ concluded that PRP is potentially an ideal scaffold for revascularization.

Revascularization using PRF

In the conventional method, bleeding is induced into the canal space for revascularization which involves irritation of the periapical tissues mechanically.²⁰The resultant blood clot acts as a scaffold that facilitates the ingrowth of new tissues into the empty canal space. However, this procedure can be unpleasant for the patients as it mechanically irritates the periapical tissues and is traumatic, mainly in young children.²⁷ Moreover, it is extremely difficult to induce and maintain a revascularization procedure. Petrinoet al²⁷ and Ding et al²³ in their case series concluded that unfavorable treatment outcomescan be expected of the failure of induction of bleeding into the canal space. In addition, desired and optimal level of placement of MTA over blood clots is technically problematic.²⁷ All these factors resulted in the development of biologic approaches in revascularization, like PRF. Choukrounet al developed PRF which is a second-generation platelet concentrate in France³⁹ and has been used by different dental specialties like periodontology, oral and maxillofacial surgery and implantology. Using PRF as a scaffold material in the regenerative endodontic procedure is a novel approach adopted by contemporary nonsurgical endodontics. Lance et al recommended the use of platelet concentrates over blood clots as the former has augmented the concentration of growth factors and increased the cell proliferation.⁴⁰

Preparation of PRF

Preparation of PRF is simple and involves the patient’s blood in specialized glass test tubes without anticoagulants which are centrifuged for 12 minutes at 2400 rpm. This results in the separation of blood samples into three layers: a red cell base at the bottom with acellular plasma on the topmost layer, and a clot of PRF in the middle. The resultant PRF clot is then pressed in between gauze pieces to attain a firm membrane and is placed in the root canal space.¹⁶

New protocol

Stephanie and Melanie Namour suggested a new protocol for revascularization after reviewing and analyzing the literature about the choice of material. They suggested isolatingthe tooth with a rubber dam and usingpovidone-iodine (10%) as a disinfectant to reduce the oral bacterial concentration. Instrumentation of root canals is still not advised as it increases dentinal wall fragility, harm stem cells in apical papillae, and damage other cells and growth factors. For irrigation, a combination of ethylenediaminetetraacetic acid (EDTA) and 6% sodium hypochlorite is recommended. EDTA allows irrigants to penetrate better into the crevices and tubules of the canal space. Triple antibiotic paste is a biocompatible root canal medicament. In the second step, PRF is inserted into the canal space. PRF provides an additional supply of blood components and also acts as a scaffold that is solid and allows the growth of the tissue. Biodentine would be a better option for root canal capping as it has similar properties as human dentin.IIt has a very low cytotoxicityand eliminates the risk of cervical area discoloration.²

The studies conducted by Mishra et al., Shivashankar et al., Keswani et al and Nagaveni et al., on revascularization using PRF showed radiographically continued root lengthening, dentinal wall thickening, regression of periapical lesion, and apical closure.^16,41,42,53 Clinically there was no sensitivity to both palpation and percussion. Teeth showed a positive response to electric and cold pulp tests. The duration of studies on PRF ranged from 9 months to 2 years ( summarized from table 2)and was done on maxillary incisors except for one case report by Subhash et al., in which revascularization was done in a mandibular molar.⁴⁶ All the studies (summarized from table 2) have reported clinical success with no pain, absence of swelling and sinus, and no tenderness neither to percussion nor palpation (Table 2). Ulusoy et al., reported average time for sensitivity response in PRFtreated teeth was 5.27 months.⁵⁰ Isha et al., reported that 98% of the cases showed periapical healing and in 99% of the cases root lengthening occurred. They also reported that in 40% of the cases, good apical closure has occurred.⁴⁴ Ulusoy et al., reported 70.6% apical closure in their study.⁵⁰ Hazim et al., reported increased bone density and reduction in apical diameter after PRF revascularization.⁴⁹

Studies were done to compare revascularization via blood clots and PRF (Table 3). Isha et al., concluded that in the blood clot group, there was no apical closure when compared to the PRF group.⁴⁴ Studies revealed that in the PRF group, there was an accelerated apical closure and increased root thickness in comparison to the blood clot group.^{51, 52} Hazim et al., reported increased bone density in the PRF group than in the blood clot group.⁵² 

Many studies (Table 1 and 2) have shown radiographically continued root lengthening and thickening of dentinal walls, but the goal of treatment is the regeneration of functional pulp tissue and for that histological analysis is necessary (Table 4).

In histologic sections, hard tissues formed are found to be heterogeneous mineralized tissue that resembles bone or cementum and soft tissues were more likely to be periodontal ligaments than pulp tissue. In contrast, some human studies have shown the formation of pulp-liketissue after revascularization. The variability of tissues formed in the intracanal region is not yet understood, but it can be correlated to the unpredictable survival of cells of the apical papilla, apical pulpal tissue, or the Hertwig epithelial root sheath in the setting of trauma and inflammation.

Conclusion

Revascularization done using either blood clot or PRF in immature permanent teeth has shown to increase dentinal wall thickness and lengthening of the root, but to evaluate the success, more histological studies are required. Even though the goal is to regenerate functional pulp tissue, some of the histological analyses have shown cementum-like structures and no pulp-like vital tissue. Platelet rich Fibrin can be regarded as an ideal scaffold for the regeneration of vital tissue in necrotic immature teeth. Long-term clinical trials and studies are needed to compare the efficacy and superiority of PRF over blood clots in regenerative endodontic therapy.

Conflict of interest

None