Comparative outcome of revascularization in bilateral, non-vital, immature maxillary anterior teeth supplemented with or without platelet rich plasma: A case series.

Loss of pulp vitality in an immature permanent tooth arrests root development. This leads to tooth with open apex and weak lateral dentinal walls. Management of such necrotic teeth with immature roots poses several treatment challenges. The documented study was performed to evaluate and compare apexogenesis induced by revascularization, with and without platelet rich plasma (PRP) in non-vital, immature anterior teeth. Three patients having bilateral, non-vital, immature maxillary central incisors with apical periodontitis were recruited after institutional ethical clearance. Subsequent to chemo-mechanical preparation, revascularization with and without PRP was randomly induced in either of the tooth. The cases were followed-up clinically and radiographically at 6 and 12 months. There was a marked difference in periapical healing, apical closure and dentinal wall thickening of teeth treated by revascularization with PRP.

INTRODUCTION

In young children, anterior teeth by virtue of their position in the dental arch are prone to trauma. In cases of complicated tooth fracture with the loss of pulp vitality, cessation of root development with an open apex is a serious sequel. Management of such immature, non-vital teeth is a challenge. Until date, treatment protocols mentioned were surgery and retrograde sealing, apical barrier formation with calcium hydroxide[1] or mineral trioxide aggregate (MTA)[2] and customized or thermoplasticized gutta-percha obturation technique.

Recently, there has been a paradigm shift in the management of these teeth based on the “regenerative concept.” Case series/reports have documented the efficacy of revascularization. This involves intentional induction of bleeding from the peri-apex and formation of an intra-canal blood clot, which acts as a scaffold. Platelet rich plasma (PRP), an autologous first generation platelet concentrate with a rich source of growth factors has been proposed as a potential addendum/substitute scaffold to improve the outcome of this procedure.[34] Until date only few case reports on the use of PRP in the revascularization are cited in the literature.[5] This case series comparatively evaluates the outcome of revascularization with and without PRP in three cases of bilateral, non-vital, immature, anterior teeth in terms of periapical healing (PAH), apical closure (AC), lateral dentinal wall thickening and root lengthening (RL).

CASE REPORTS

Case 1

A healthy 10-year-old boy was referred for evaluation and management of his broken upper anterior teeth, subsequent to a fall 3-year prior. Root canal therapy was initiated, but not completed by a general dental practitioner. Intraoral examination revealed Ellis class III fracture and a well-defined, localized swelling in relation to both the upper central incisors. Radiographic examination showed immature open apices with thin dentinal walls in relation to both the teeth [Figure 1a]. Based on clinical and radiographic examination, a diagnosis of acute periapical abscess in relation to both the central incisor was established. Taking into consideration the stage of root development, the maturation of the dentinal walls and the wide-open apices, revascularization procedure with and without PRP was planned. The risks, complications and possible outcome of this treatment were explained and parental informed consent was obtained.

Figure 1

(a) Intaroral periapical radiograph of maxillary central incisors showing open apices with thin dentinal walls. Revascularization with and without platelet rich plasma performed in 21 and 11 respectively. (b) At 6-month, 21 and 11 showing progressive dentinal wall thickening. (c) At 1 year, marked maturation and root lengthening in 21

Teeth were isolated under rubber dam and re-accessed with a round diamond and an endo-Z bur (Dentsply Maillefer, Tulsa, OK). Purulent discharge was observed in both the teeth. The canals were copiously irrigated with normal saline. Working length was established with a paper point (Dentsply Maillefer, Tulsa, OK) and confirmed radiographically by placing a large file in the canal. Minimal mechanical instrumentation with an International Organization for Standardization number 60H-file (Dentsply Maillefer, Tulsa, OK) and irrigation with 20 mL 2.5% sodium hypochlorite (NaOCl, Cmident, India) was performed. The canal was dried with paper points and an inter-appointment medication of triple antibiotic paste (as per the protocol mentioned by Hoshino et al. 1996) was applied with a sterile number 30 hand lentulo spiral (Dentsply Maillefer, Tulsa, OK). Teeth were temporarily restored with intermediate restorative material (Caulk Dentsply, Milford, DE). This was the chemo-mechanical and disinfection protocol followed for all cases. However, in cases of weeping canals, another inter-appointment dressing was given until the tooth was symptom-free and the canal was dry. Patients were recalled after 4-week. Teeth were re-accessed under rubber dam and revascularization with and without PRP were randomly induced in upper left and right central incisors respectively [Figure 1b].

Local anesthetic solution without adrenaline (LOX 2% Neon Lab, India) was infiltrated around the apices of both the central incisors. A sterile endodontic file with a rubber stopper set at 2 mm beyond the established working length was taken. With sharp, fine strokes, the file was pushed past the confines of the canal into the periapical tissue. When frank bleeding was evident, a dry cotton pellet was inserted 3-4 mm into the canal and held there for 5-7 min to allow blood clot formation in the apical third. This revascularization protocol was followed for both the teeth.

PRP preparation was carried out in a simple, table top laboratory centrifugation machine (Remi model no. — R–8C, India). A total volume of 8.5 mL of whole blood was drawn by venipuncture of antecubital vein. It was then collected in a 10-mL sterile glass tube coated with an anticoagulant (acid citrate dextrose). Whole blood was initially centrifuged (soft spin at 2400 rpm for 10 min) to separate PRP and platelet-poor plasma (PPP) portions from the red blood cell fraction. PRP and PPP portions were again centrifuged (hard spin at 3600 rpm for 15 min) to separate the PRP from the PPP.

Intra-canal bleeding was induced as described above. Freshly prepared PRP, soaked on a 1 mm2 × 1 mm2 sterile collagen sponge (Metrogene, Septodont, France) was introduced into the root canal of maxillary left central incisor with cotton pliers and carried to the middle third with a size 30 finger plugger (Sybronendo, CA, USA). Access openings were restored with resin modified glass ionomer cement (Photac-Fill, 3MESPE, Minnesota). Standardized intraoral periapical baseline and subsequent follow-up radiographs at 6 and 12 months were taken with a Rinn positioning device (Dentsply, Elgin, IL).

Case 2

A healthy 23-year-old man was referred with the complaint of lingering pain in upper front teeth for the past 1 week. Patient had a history of trauma 15-year ago, for which no treatment was sought. Intraoral examination revealed discolored both upper central incisors and a sinus tract on palatal aspect of upper left lateral incisor. Radiographic examination revealed well-defined, bilateral periapical radiolucency involving both upper centrals and upper left lateral incisor [Figure 2a]. Both the central incisors had open apices and thin lateral dentinal walls. Based on clinical and radiographic examination, a diagnosis of pulpal necrosis with chronic apical abscess in relation to all three teeth was established. Upper left lateral incisor was managed by conventional endodontic therapy. Revascularization procedure with and without PRP was randomly induced in upper left and right central incisors respectively [Figure 2b]. Final esthetic rehabilitation was carried out with porcelain fused to metal crowns.

Figure 2

(a) Intaroral periapical radiograph showing open apices and periapical lesions associated with both the maxillary central incisors. (b) At 6-month, 21 treated by revascularization + platelet rich plasma shows complete healing of periapical lesion and good root lengthening. Tooth number 11 treated by revascularization shows satisfactory healing. (c) At 1-year, further narrowing of canal space in apical one-third and thickening of the lateral walls is evident in 21

Case 3

A healthy, 13-year-old girl was referred with the chief complain of dull, continuous pain for past 3 months in her upper front teeth. Intraoral soft-tissue and radiographic examination revealed a well-defined, localized swelling and a bilateral periapical radiolucency involving upper central incisors. Both teeth had open apices and thin lateral dentinal walls [Figure 3a]. Based on clinical and radiographic examination, a diagnosis of chronic abscess in relation to both the teeth was established [Figure 3b]. After the infection control revascularization, with and without PRP was randomly induced in upper left and right central incisors respectively.

Figure 3

(a) Intaroral radiograph reveals open apices and periapical lesions associated with 11 and 21. (b) At 6-month follow-up, 21 treated by revascularization + platelet rich plasma and 11 by revascularization showed good healing. (c) At 1-year, narrowing of canal space in apical one-third and thickening of the lateral walls and root lengthening is quite evident in 21 with complete healing of periapical lesions in both teeth

RESULTS

The cases were followed-up clinically and radiographically at 6 and 12 months. Clinically, all the three cases were asymptomatic with complete resolution of pain, swelling and intraoral sinus. Two endodontists (who were not involved in the study) were trained to interpret the intraoral periapical radiographs for PAH, AC, dentinal walls thickening and RL by rigorous, multiple training sessions until the consensus was reached between them. The X-rays were interpreted by both the trained endodontists independently and then jointly to arrive at a consensus. X-rays were scored either as satisfactory (+), good (++) or excellent (+++) as presented in Table 1. PAH was excellent in group II (two out of three cases) compared with group I in which it was good (all three cases). AC was excellent in one case in group II compared with group I in which it was satisfactory in all three cases. RL was good to excellent in group II compared to group I in which it was satisfactory. Dentinal wall thickening was good in group II compared to group I in which it was satisfactory. Overall, on three parameters, PRP was found to enhance the regenerative and healing response in immature teeth. However, RL was comparable in both groups [Table 2].

Table 1

Radiographic findings at 1-year follow-up

Table 2

Number of cases showing radiographic findings at 1-year follow-up

DISCUSSION

The present case series compared the clinical and radiographic treatment outcome of revascularization with and without PRP in immature, non-vital anterior teeth. Revascularization can be achieved successfully if a suitable matrix is provided in a disinfected root canal for tissue in-growth and a coronal bacteria-tight seal is provided. Mechanical instrumentation of the immature teeth with blunderbuss canal is difficult as it often leads to fracture of thin, lateral dentinal walls.[6] Therefore, minimal mechanical instrumentation is recommended for the disinfection of such teeth. Chemical disinfection with various concentrations of sodium hypochlorite (NaOCl) and intracanal placement of triple antibiotic paste has been proposed.[78] In this study, a triple antibiotic paste containing equal proportions of bactericidal metronidazole (Albert David, Kolkata, India), ciprofloxacin (Sifam Pharmaceuticals, Gujarat, India) and bacteriostatic minocycline (Aurobindo, Andhra Pradesh, India) components were ground and mixed with distilled water to a thick paste consistency.

In revascularization, the in-growth of new tissue from the periapical area needs a matrix for its support. A matrix of blood clot provides not only a base for stem cell adhesion, but also for its growth, differentiation and migration. Thus, in the revascularization if an intra-canal matrix is provided, undifferentiated mesenchymal cells can proliferate and differentiate under the organizing influence of Hertwig's epithelial root sheath, thus re-establishing the pulp vitality. These cells can be recruited from the apical pulp tissue remnants,[910] periodontal ligaments,[1112] apical papilla or the bone marrow.[1314] Till date, blood clot has been used commonly as a matrix.[1516] In revascularization cases, the bleeding is induced intentionally by over-instrumentation in the periapical area. The clot formed consists of cross-linked fibrin, which serves as a pathway for the migration of the stem cells from the periapical area.[1117] Furthermore, this blood clot contains different cells that are enriched with several growth factors, important for the wound healing process.[18] The bleeding induced angiogenesis also helps in recruiting stem cells necessary for a successful outcome.[3]

However, formation of an intracanal blood clot is not always predictable. Bleeding may be reduced when an inter-appointment medication of calcium hydroxide is given, as it can cause periapical coagulation necrosis. Clot formation may be compromised if vasoconstrictor (adrenaline) containing local anesthetic is used. The concentration of growth factors in the blood clot is unpredictable and limited. Furthermore, after clot formation, erythrocytes undergo necrosis, affecting the properties of the matrix.[3]

To overcome these drawbacks and to augment the healing process, use of PRP is highly desirable. It is a first generation, autologous platelet concentrate containing different growth factors such as platelet derived growth factor, transforming growth factors β, insulin like growth factor, vascular endothelial growth factor, epidermal growth factor, epithelial cell growth factor. These growth factors are released when platelets are de-granulated, which can be carried out by various methods; addition of thrombin, calcium containing products (e.g., — calcium chlorite, calcium sulfate etc.,) or even shaking the platelets.

In the present study, PRP supplement was found to improve the outcome of revascularization [Figure 1c, 2c and 3c]. The PRP scaffold could have helped to stabilize the already existing blood clot and enhance angiogenesis. It could have also led to sustained release of growth factors, which play an important role in recruitment, retention and proliferation of stem cells, which in turn may have led to hastened maturogenesis. However, the use of PRP has few disadvantages; need to draw blood in young patients and requirement for special equipment and reagents to prepare PRP.

Apart from the root canal disinfection and use of a suitable scaffold, the quality of the coronal restoration is also very important to achieve success of revascularization treatment. This critical requirement of a bacterial-tight coronal seal can be met with the use of composite, MTA, cavit, glass ionomer or their combinations. In the reported cases, the access openings were restored with resin modified glass ionomer cement (Photac-Fill, 3M, ESPE, Minnesota).

In the present study, bleeding induced blood clot served as a matrix, PRP as an addendum and collagen as a carrier for PRP. Nevertheless, each of them can act as a matrix on their own merit; limitations of one can be compensated by the other, by their combined use.

CONCLUSION

Revascularization is an effective method for inducing maturogenesis in non-vital, immature teeth. Supple-mentations with PRP can potentially improve and hasten the desired biological outcome of this regenerative technique. However, randomized prospective clinical trials are needed to establish PRP supplements in revascularization cases routinely and its impact on the final outcome.