Histologic Examinations of Teeth Treated with 2 Scaffolds: A Pilot Animal Investigation Mahmoud Torabinejad, DMD, MSD, PhD, Hadi Faras, BDM, MSD, Robert Corr, DDS, MSD, Kenneth R. Wright, PhD, and Shahrokh Shabahang, DDS, MS, PhD Abstract Introduction: A growing body of evidence is building a case for the possibility of tissue regeneration within the root canal of necrotic teeth, allowing for continued root development. However, it remains unknown what type of tissue is produced after regenerative endodontics. The purpose of this study was to use blood clots and platelet-rich plasma (PRP) as scaffolds in regenerative endodontics under ideal conditions in a ferret model to examine the tissues generated within the root canals. Methods: The pulps of 21 canine teeth from 7 young ferrets were extirpated using broaches without filing the canal walls. Bleeding was stimulated from the peri- apical tissues, and a blood clot was induced in the canal space to the level of the cementoenamel junction in 12 teeth. PRP was prepared and placed in the canals to the level of the cementoenamel junction in 9 teeth. The cor- onal access was sealed with mineral trioxide aggregate. Seven canines were not operated on and served as con- trols. Three months later, block sections including each canine and its surrounding tissues were removed for his- tologic evaluation. The tissues found in the canals of experimental teeth were compared with those in the control teeth. Results: Almost all of the experimental teeth showed the presence of intracanal bonelike tissue. No evidence of dentinal wall thickening or apical nar- rowing was noted in the experimental teeth. Conclu- sions: In this experimental model, the use of either PRP or blood clots during regenerative endodontics leads to the formation of intracanal bonelike tissue without continual root maturation. (J Endod 2014;40:515–520) Key Words Bone, cementum, dentin, ferret, regeneration, revascu- larization T he potential for dental tissue regeneration and continued root development of re- planted teeth is reported in several publications in the dental literature. However, the presence of infection may interfere with the healing process (1–3). Traditionally, it has been thought that successful tissue regeneration cannot be an expected outcome after a tooth has become infected. Throughout the years, several authors have suggested that regenerative endodontics may be possible in teeth with immature apices despite the presence of pulpal necrosis and apical pathosis. Several case reports and case series have been published showing radiographic continuation of root development in teeth with necrotic pulps, apical lesions, and open apices, as evidenced by the deposition of hard tissue within the canals of these teeth (4–16). Although the radiographic appearance of apical closure and canal wall thickening can give the impression that a normal, functional pulp has regenerated, there is no his- tologic evidence to support these findings. There are several animal studies examining the nature of tissues present in the root canals of teeth treated with procedures that are supposed to result in regeneration of the dental pulp (4, 17, 18). Connective tissue, cementum, and bone have been reported in the root canals of the experimental teeth (4, 17, 18). Torabinejad and Turman used platelet-rich plasma (PRP) instead of whole blood in a maxillary premolar with necrotic pulp and an open apex (19). The tooth was clin- ically asymptomatic 5 1/2 months later and radiographically showed resolution of the periapical lesion, further root development, and continued apical closure. Vitality tests at that time revealed the presence of sensitivity to cold, and an electric pulp test elicited a positive response similar to those found in the adjacent premolar. Fourteen months af- ter the procedure, a biopsy of the intracanal tissue was obtained during the root canal treatment (4). Histologic examination revealed the presence of pulplike vital connective tissue with blood vessels and few inflammatory cells at the periphery. There was no ev- idence of odontoblasts or bone in the examined tissue. The purpose of this pilot study was to use blood clots or PRP in regenerative endodontics under ideal conditions in a ferret model as scaffolds and examine the tissues generated within the root canals of the experimental teeth. Materials and Methods All animal experiments were performed in accordance with protocols approved by the Loma Linda University Animal Research Committee. Seven 70-day-old male ferrets with incompletely developed canines were used in this experiment. One canine from each animal was randomly chosen to serve as the control, and the 3 remaining canines served as experimental teeth, providing 21 experimental and 7 control teeth. Preoper- ative radiographs were obtained to confirm the presence of an open apex and the absence of pre-existing pathosis in each cuspid tooth. All procedures were conducted under a clean protocol with the use of sterile ma- terials and equipment. For general anesthesia, anesthesia was induced in the ferrets with 3% isoflurane gas (Phoenix Pharmaceuticals, Inc, St Joseph, MO). The animals were then intubated with a size 2.5-mm endotracheal tube and maintained under 2%– 2.5% isoflurane gas. Each experimental tooth was then decoronated with a sterile dia- mond bur approximately 3 mm above the gingival crest to expose its pulp. The access was refined with sterile flame-shaped diamond and #4 round carbide burs. The entire pulp was extirpated with sterile barbed broaches. The canal was then irrigated with approximately 10 mL sterile saline and dried with sterile paper points. One milliliter From the Loma Linda University, Loma Linda, California. Address requests for reprints to Dr Mahmoud Torabinejad, Loma Linda University, Loma Linda, CA 92350. E-mail address: mtorabinejad@llu.edu 0099-2399/$ - see front matter Copyright ª 2014 American Association of Endodontists. http://dx.doi.org/10.1016/j.joen.2013.12.025 Basic Research—Biology JOE — Volume 40, Number 4, April 2014 Teeth Treated with 2 Scaffolds 515