Case Report/Clinical Techniques

Regenerative Endodontic Treatment of an Immature Permanent Tooth at an Early Stage of Root Development: A Case Report
Xu Chen, DDS, PhD,* Zhi-Fan Bao, MS, DDS,* Yao Liu, MS, DDS,* Ming Liu, MS, Xiao-Qing Jin, MS, and Xue-Bin Xu, MS, DDS*
Abstract
Introduction: Regenerative endodontic treatment (RET) has been used in treating nonvital immature permanent tooth whose root formation ranged from approximately two-thirds of the full root length to almost completely developed root with open apex at least 1.1 mm in diameter according to the reported cases. However, this case report was to introduce RET in an affected tooth at an early stage of root development. Methods: The premolar #29 in an 8-year-old girl had pulpal necrosis and apical periodontitis caused by the fracture of dens evaginatus. Its root was at the beginning of formation. Copious hemorrhagic drainage was observed after preparing of an access cavity. The canal was irrigated with 3% NaOCl solution, sterile normal saline, and chlorhexidine. Root dressing with triple antibiotic was then performed and left for 4 weeks. We used a K-le to create bleeding into the canal after ushing and drying the root canal. Mineral trioxide aggregate was carefully placed over the formed blood clot. Results: Clinical examination at 1, 3, 6, 9, and 12 months revealed an asymptomatic tooth. Radiographic examination revealed resolution of periapical radiolucency, increased thickening of the canal wall, and lengthening of the root, which demonstrated the continual development of the tooth root. Noticeably, the rst-month postoperative radiograph showed radiopaque image in the root canal like an isolated island, which was gradually obvious during follow-up. Cone-beam computed tomography revealed that the calcication was attached with dentin wall in buccolingual direction, and the root canal was not completely obliterated. Conclusions: RET is feasible for a tooth at an early stage of root development that has necrotic pulp and periapical lesion. (J Endod 2013;39:719722)

Key Words
Dens evaginatus, immature permanent tooth, regenerative endodontic treatment, root development

he completion of root development and closure of the apex occur up to 3 years after eruption of the tooth. Dental caries, trauma, and anomalous tooth morphology (ie, dens evaginatus) are potential causes of necrotic pulp of immature teeth, which will lead to the cessation of root formation. The endodontic treatment of teeth with immature root formation has always been a challenge for several reasons (1): 1. Mechanical instrumentation of the root canal is difcult because of the anatomic characteristics of the immature tooth. 2. It is difcult to seal the root canal by using traditional lling methods because of the absence of an apical stop. 3. Thin root canal walls of immature teeth are susceptible to fracture (2).

The conventional treatment of immature teeth with necrotic pulp is apexication (3). Apexication by using calcium hydroxide has some inherent disadvantages including variability of treatment time, unpredictability of apical closure, and difculty in patients follow-up and subsequent treatment (4). In addition, long-term root canal dressing by using calcium hydroxide weakens the root structure and may lead to future fracture of the root (1). One-visit apexication that is performed by placement of an apical barrier by using mineral trioxide aggregate (MTA) is an alternative to conventional long-term calcium hydroxide therapy and may shorten the treatment time between the patients rst appointment and the nal restoration (1, 5). A retrospective study indicates that the survival rate of MTA apexication is greater than that of calcium hydroxide apexication (6). Although MTA is effective in supporting new hard tissue formation in the apical area of the affected immature necrotic teeth, the risk of future fracture may still exist because the root width will not increase in MTA apexication-treated tooth (6). To achieve a better prognosis, regenerative endodontic treatment (RET), which is also named dental pulp revascularization, is another choice for treating immature teeth with necrotic pulp. Several case reports indicate that RET is an optional choice when treating nonvital immature teeth (2, 719). The term revascularization was rst introduced to describe the healing of the periapical abscess and continued root formation of a necrotic immature permanent tooth. Its root canal was not mechanically cleaned to its full length but was copiously irrigated and dressed with

From the *Department of Pediatric Dentistry and Department of Radiology, School of Stomatology, China Medical University, Shenyang; and Laboratory of Pediatric Dentistry, Liaoning Stomatology Research Institute, Shenyang, China. Supported by Liaoning Provincial Science and Technology Project funded by Science and Technology Department of Liaoning Province (2012225015). Address requests for reprints to Dr Xu Chen, Department of Pediatric Dentistry, School of Stomatology, China Medical University, No. 117 Nan-Jing North Street, Shenyang 110002, China. E-mail address: chenxu_cmu@sina.com 0099-2399/$ - see front matter Copyright 2013 American Association of Endodontists. http://dx.doi.org/10.1016/j.joen.2012.12.023

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antimicrobial agents (7). It is proposed that the term revascularization neither describes the technique of the treatment nor encompasses the actual healing and repair process that occurs in these cases (20). More and more scholars now use the term regenerative endodontic treatment instead. RET is performed by abundant irrigation, root canal dressing, and hemorrhage being induced into the root canal to form a blood clot. The main difference between RET and apexication is that RET involves irritation of the apical tissue to create bleeding into the root canal. The blood clot can provide growth factors for the cells and act as a scaffold for pulp regeneration (8, 21). The outcomes of dental pulp revascularization with blood clot in the canal space were better than those without blood clot in immature dog root canals that had been disinfected (21). The evoked-bleeding step in RET procedures triggers the signicant accumulation of undifferentiated stem cells into the canal space where these cells may contribute to the pulp regeneration (22). The exact types and sources of stem cells involved in RET are still unclear at present. There is no reference that describes the stage of root development indicated for using RET in immature permanent tooth. According to the reported cases, root formation of affected teeth ranged from two-thirds of the full root length to almost completely developed root with open apex at least 1.1 mm in diameter. However, we did use RET on a special case in which the affected tooth was at an early stage of root development.

Case Report
The patient was an 8-year-old girl with a history of swelling and pain of the right mandible for more than 6 months. She did not visit a dentist immediately because the symptoms were not continuous. After the periapical symptoms and swelling repeatedly occurred several times, she visited the Hospital of Stomatology afliated with China Medical University for treatment. On clinical examination, a sinus tract was found apically on the buccal aspect of the right mandibular second premolar. The premolar was free of caries, but fracture of the dens evaginatus of the second premolar was noted on visual inspection. The affected premolar #29 was sensitive to percussion and palpation with class III tooth mobility. The fractured surface of the affected premolar was explored, and the patient responded with no pain. Periapical radiographic examination revealed that tooth #29 was almost rootless with periapical radiolucency about 10 mm in diameter (Fig. 1A). It was diagnosed as having pulp necrosis and chronic apical abscess. The crown had formed, and the root had just started its formation. Considering the root was too short and needed further development to form a long and strong root to resist possible fracture, RET was proposed. Conventional calcium hydroxide apexication and RET were explained to the patient and her parents. RET was nally chosen, with expectation to achieve a better prognosis. After informed consent, the access was prepared under local anesthesia and rubber dam isolation. Copious hemorrhagic drainage was obtained after the access cavity was made. A size #15 K-le was introduced into the root canal to measure the tooth length by using radiographic examination. The working length was determined to be 8 mm, which was 1 mm less than the tooth length. A needle with a mark at 8 mm was placed into the canal, and the canal was slowly ushed in the sequence of 20 mL 3% NaOCl, 10 mL sterile normal saline, and 10 mL chlorhexidine. During the ushing with NaOCl, some black necrotic tissues were washed out of the canal. After the canal was dried, ciprooxacin, metronidazole, and minocycline powder were mixed with glycerol (23) and prepared into creamy mixture. A lentulo spiral instrument was used to deliver the antibiotic paste into the canal. The access cavity was then sealed with Caviton (GC Corp, Tokyo, Japan) and glass ionomer cement (3M ESPE, Seefeld, Germany). Oral hygiene instruction was provided, and the patient and her parents were told that they should come to the clinic immediately if there was pain or swelling. 720

Figure 1. (A) Preoperative periapical radiographic examination revealed that tooth #29 was almost rootless with periapical radiolucency about 10 mm in diameter; (B) structure with radiopaque image was found at the apical point 1 month after RET; (CF) at 3-, 6-, 9-, and 12-month recall, respectively, radiography showed continuous root extension and canal wall thickening of the affected tooth. The size of the deposited hard tissue in the canal space seemed to continue its growing during the follow-up. (GI) Periapical radiograph of the healthy contralateral premolar #20 was taken as control at 6, 9, and 12 months, respectively. The status of root development of tooth #29 after RET was almost the same as that of tooth #20.

After 2 weeks, the patient came for a second visit; she reported no pain postoperatively but felt uncomfortable when chewing on her right side. Clinical examination found that the sinus tract was much smaller. Tooth #29 was slightly sensitive to percussion and palpation with class II tooth mobility. Some reduction in the radiolucency was already evident as indicated by x-ray examination. Because the affected tooth with little root development still had abnormal mobility, a lower arch removable splint was applied to stabilize the affected mobile tooth and to eliminate the risk of avulsion caused by chewing.
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One month after the initial treatment, the tooth remained asymptomatic without obvious discoloration. The sinus tract was not present, and radiographic examination showed radiolucency had completely disappeared. After local anesthesia and rubber dam isolation, the access was opened again, and there was no sign of inammatory exudate. The antibiotic paste was ushed out with 10 mL 3% NaOCl and 10 mL sterile normal saline. The canal was then dried, and a size #15 K-le was inserted to irritate the periapical tissue gently to create some bleeding into the canal. The bleeding was stopped at the level of the cementoenamel junction by using a small cotton pellet soaked with sterile saline. After 15 minutes, the blood clot formed. At least 3-mm-thick MTA (Dentsply Tulsa Dental, Tulsa, OK) was carefully placed on the blood clot, followed by a wet cotton pellet. The cavity access was sealed with Caviton and glass ionomer cement. The lower arch splint was removed 4 weeks after applying, which was 6 weeks after initial treatment. At that time, the tooth was without abnormal mobility. During the clinical follow-up, the patient remained asymptomatic, the mobility of tooth #29 was within the normal limits, and the sinus tract did not reappear. The tooth had continuously erupted until reaching the occlusal plane. Noticeably, the rst-month postoperative radiograph showed structure with radiopaque image in the root canal like an isolated island (Fig. 1B), which was gradually obvious during follow-up (Fig. 1CF). Increased thickening of the root canal wall and lengthening of the root showed continual development of the root, which was accompanied by the signs of apical closure (Fig. 1F). We took x-rays of the healthy contralateral premolar #20 as control. The affected premolar was almost at the same developing speed as the healthy one (Fig. 1GI). At 12-month recall, cone-beam computed tomography (CBCT) was taken to evaluate the accurate position of the calcication formed in the root canal space. The result indicated the radiopaque image was attached with dentin wall in buccolingual direction like a bridge. There was space around the calcication tissue in the mesiodistal direction (Fig. 2). by dens evaginatus fracture while its root was at an early stage of development. Dental history provided by the patients parents mentioned that the second primary molar in the right mandible was exfoliated ahead of time because of severe dental caries. Because of early loss of the preceding tooth and without any clinical intervention, the successor, tooth #29 with dens evaginatus deformity, erupted early. This case was unique because the root of the affected tooth was at an early stage of development. We considered using apexication in this case; however, even if the apex was sealed, the root would not develop any extension, and long-term prognosis of this tooth with a very short root was not favorable. We tried using RET on the rootless tooth and found a new phenomenon after RET that has never been mentioned in previous reported cases. An X-ray examination showed a radiopaque image in the root canal from 4 weeks after applying MTA. Its size seemed to continue to increase until the last appointment, and the image of CBCT indicated that the root canal had not been completely obliterated yet. The nature of this radiopaque image and whether it would continue to form until the pulp canal was obliterated were still unknown. These problems were of great signicance to predict the long-term prognosis after RET. According to a series of clinical and histologic studies, regenerative tissues in the root canal are basically the following 4 types (25): 1. Revascularization of the pulp with accelerated dentin formation, leading to pulp canal obliteration 2. Ingrowth of cementum and periodontal ligament 3. Ingrowth of cementum, periodontal ligament, and bone 4. Ingrowth of bone and bone marrow The rst type was believed to have the best prognosis. Histologic study of the newly formed tissues in the canal space after RET of immature dog teeth revealed that the canal dentinal walls were thickened and extended by the apposition of newly generated cementum-like tissue, bone, or bone-like tissue; connective tissue similar to periodontal ligament was also observed in the canal space (21, 2628). The hard tissues newly formed on dentinal walls were distinct from dentin, bone, or bonelike tissue in the root canal space, although they resembled cementum but with signicantly different organization and maturation of collagen matrix (27, 28). Although the results of animal experiments showed that there were calcied tissues formed in the root canal space, the radiographs did not show any radiopaque image either in these experiments or in previous case reports. Wang et al (26) explained this phenomenon might be due to the angulation and image resolution of the radiographs, which did not provide sufcient information to reect accurately the newly grown hard tissues in the root canal.

Discussion
Dens evaginatus is an uncommon dental anomaly that exhibits by protrusion of a tubercle from occlusal surfaces of posterior teeth or lingual surfaces of anterior teeth. It occurs primarily in people of Asian descent and is also called central cusp in premolars (24). The greatest disadvantage of dens evaginatus is that these cusp-like tubercles are susceptible to pulp exposure from wear or fracture, leading to pulpal complications soon after eruption. In this unusual case, the pulpal necrosis and apical periodontitis of the early erupted tooth were caused

Figure 2. CBCT images in the axial section were examined by rolling the toolbar from the pulp chamber to the apex to assess the calcication in different levels in the root canal of tooth #29. (A) The layer of MTA; (B) the top of the calcication in the root canal was attached to the buccal aspect of the canal wall; (C) the calcication was attached with dentin wall in buccolingual direction like a bridge, and there was space around the calcication tissue in the mesiodistal direction; (D) there was no calcication close to the apical end. Arrows indicate the affected tooth #29.

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Because we performed an x-ray examination immediately after applying MTA on the blood clot, we could rule out the possibility of introduction of something calcied into the root canal articially. We speculated that the nature of the radiopaque image indicated by x-ray and CBCT in this case was bone-like tissue and/or cementum-like tissue. The nature of the newly formed hard tissues needs further research, and detection of mineral composition may be an effective method. Through the study of continuous x-ray lm before and after RET, we found that the position of the radiopaque image was just at the apical point, which was likely the position of Hertwigs epithelial root sheath for this rootless tooth. The size of the radiopaque image continued to increase, but its location did not change even after the root had extended (Fig. 1). On the basis of this phenomenon, it was hypothesized that after the infection was eradicated, the survived Hertwigs epithelial root sheath recovered its function and locally interacted with stem cells to generate new cementoblasts and/or osteoblasts to form cementum-like tissues and bone-like tissues that were shown to be the radiopaque image in xray lm. This hypothesis could also explain that the radiopaque image was not seen in the apical direction. Besides the hard tissues, the type of soft tissues newly formed in the root canal is also of interest. One patient who had received RET by using platelet-rich plasma got reversible pulpitis. Histologic examination of the tissue removed from the root canal during root canal treatment revealed that it was pulp-like connective tissue without odontoblasts (29). In the case report discussed here, the tooth root was at the beginning of its formation. The short root and severe apical abscess made the tooth extremely mobile. To eliminate the risk of avulsion caused by chewing, a lower arch removable splint was applied. Six weeks after initial treatment, the tooth mobility was within the normal limits, which could be due to healing of the periapical abscess and repair of alveolar bone. Jung et al (30) reported 2 cases of continued development of the apical end of the root that had been separated from the main root. As a result of the loose attachment between the apex of the developing root and apical papilla, the 2 parts can be easily detached by an external force (31) such as inappropriate operation in the apexication procedure or excessive tooth mobility during acute inammation. Therefore, stabilization of the loose tooth had a considerably positive effect in this case. RET has been proved to be a better choice when treating a nonvital immature permanent tooth because of the thickening of dentin wall and extension of root length after treatment. This would be very meaningful for the affected tooth with extremely short root. From this unique case, we brought forward the following viewpoints. The prognosis of RET for the affected immature permanent tooth might be related to the stage of root development. For the tooth with little root development, hard tissues seemed to form and deposit more easily because of more energetic stem cells. Although we did not know whether the calcication would completely obliterate the root canal in the future, we considered this case to be successful because the newly formed hard tissues on the root canal wall and apical end made the root stronger and longer. RET had more advantages than apexication when treating severe apical periodontitis of an immature permanent tooth at an early stage of root development.

References
1. Andreasen JO, Farik B, Munksgaard EC. Long-term calcium hydroxide as a root canal dressing may increase risk of root fracture. Dent Traumatol 2002;18:1347. 2. Thibodeau B, Trope M. Pulp revascularization of a necrotic infected immature permanent tooth: case report and review of the literature. Pediatr Dent 2007;29:4750. 3. Rafter M. Apexication: a review. Dent Traumatol 2005;21:18. 4. Shabahang S, Torabinejad M, Boyne PP, et al. A comparative study of root-end induction using osteogenic protein-1, calcium hydroxide, and mineral trioxide aggregate in dogs. J Endod 1999;25:15. 5. Steinig TH, Regan JD, Gutmann JL. The use and predictable placement of mineral trioxide aggregate in one-visit apexication cases. Aust Endod J 2003;29:3442. 6. Jeeruphan T, Jantarat J, Yanpiset K, et al. Mahidol study 1: comparison of radiographic and survival outcomes of immature teeth treated with either regenerative endodontic or apexication methodsa retrospective study. J Endod 2012;38:13306. 7. Iwaya SI, Ikawa M, Kubota M. Revascularization of an immature permanent tooth with apical periodontitis and sinus tract. Dent Traumatol 2001;17:1857. 8. Banchs F, Trope M. Revascularization of immature permanent teeth with apical periodontitis: new treatment protocol? J Endod 2004;30:196200. 9. Chueh LH, Huang GT. Immature teeth with periradicular periodontitis or abscess undergoing apexogenesis: a paradigm shift. J Endod 2006;32:120513. 10. Cotti E, Mereu M, Lusso D. Regenerative treatment of an immature, traumatized tooth with apical periodontitis: report of a case. J Endod 2008;34:6116. 11. Jung IY, Lee SJ, Hargreaves KM. Biologically based treatment of immature permanent teeth with pulpal necrosis: a case series. J Endod 2008;34:87687. 12. Shah N, Logani A, Bhaskar U, Aggarwal V. Efcacy of revascularization to induce apexication/apexogenesis in infected, nonvital, immature teeth: a pilot clinical study. J Endod 2008;34:91925. 13. Chueh LH, Ho YC, Kuo TC, et al. Regenerative endodontic treatment for necrotic immature permanent teeth. J Endod 2009;35:1604. 14. Ding RY, Cheung GS, Chen J, et al. Pulp revascularization of immature teeth with apical periodontitis: a clinical study. J Endod 2009;35:7459. 15. Reynolds K, Johnson JD, Cohenca N. Pulp revascularization of necrotic bilateral bicuspids using a modied novel technique to eliminate potential coronal discolouration: a case report. Int Endod J 2009;42:8492. 16. Petrino JA, Boda KK, Shambarger S, et al. Challenges in regenerative endodontics: a case series. J Endod 2010;36:53641. 17. Nosrat A, Sei A, Asgary S. Regenerative endodontic treatment (revascularization) for necrotic immature permanent molars: a review and report of two cases with a new biomaterial. J Endod 2011;37:5627. 18. Cehreli ZC, Isbitiren B, Sara S, Erbas G. Regenerative endodontic treatment (revascularization) of immature necrotic molars medicated with calcium hydroxide: a case series. J Endod 2011;37:132730. 19. Torabinejad M, Turman M. Revitalization of tooth with necrotic pulp and open apex by using platelet-rich plasma: a case report. J Endod 2011;37:2658. 20. Huang GT, Lin LM. Letter to the editor: comments on the use of the term "revascularization" to describe root regeneration. J Endod 2008;34:511. 21. Thibodeau B, Teixeira F, Yamauchi M, et al. Pulp revascularization of immature dog teeth with apical periodontitis. J Endod 2007;33:6809. 22. Lovelace TW, Henry MA, Hargreaves KM, Diogenes A. Evaluation of the delivery of mesenchymal stem cells into the root canal space of necrotic immature teeth after clinical regenerative endodontic procedure. J Endod 2011;37:1338. 23. Hoshino E, Kurihara-Ando N, Sato I, et al. In-vitro antibacterial susceptibility of bacteria taken from infected root dentine to a mixture of ciprooxacin, metronidazole and minocycline. Int Endod J 1996;29:12530. 24. Levitan ME, Himel VT. Dens evaginatus: literature review, pathophysiology, and comprehensive treatment regimen. J Endod 2006;32:19. 25. Andreasen JO, Bakland LK. Pulp regeneration after non-infected and infected necrosis, what type of tissue do we want? a review. Dent Traumatol 2012;28:138. 26. Wang X, Thibodeau B, Trope M, et al. Histologic characterization of regenerated tissues in canal space after the revitalization/revascularization procedure of immature dog teeth with apical periodontitis. J Endod 2010;36:5663. 27. Yamauchi N, Yamauchi S, Nagaoka H, et al. Tissue engineering strategies for immature teeth with apical periodontitis. J Endod 2011;37:3907. 28. Yamauchi N, Nagaoka H, Yamauchi S, et al. Immunohistological characterization of newly formed tissues after regenerative procedure in immature dog teeth. J Endod 2011;37:163641. 29. Torabinejad M, Faras H. A clinical and histological report of a tooth with an open apex treated with regenerative endodontics using platelet-rich plasma. J Endod 2012;38:8648. 30. Jung IY, Kim ES, Lee CY, Lee SJ. Continued development of the root separated from the main root. J Endod 2011;37:7114. 31. Sonoyama W, Liu Y, Yamaza T, et al. Characterization of the apical papilla and its residing stem cells from human immature permanent teeth: a pilot study. J Endod 2008;34:16671.

Acknowledgments
The authors thank Dr Song-Tao Shi (Associate Professor, Center for Craniofacial Molecular Biology, Ostrow School of Dentistry, University of Southern California) for giving some comments on the manuscript. They also thank Dr Carolyn W. Gibson (Professor, Department of Anatomy and Cell Biology, School of Dental Medicine, University of Pennsylvania) for English proong of the manuscript. The authors deny any conicts of interest related to this study.

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