Promotion of Endodontic Lesions in Rats by a Novel Extraradicular Biofilm Model Using Obturation Materials

Abstract

bAlthough extraradicular biofilm formation is related to refractory periapical periodontitis, the mechanism of extraradicular biofilm development, as well as its effect on periapical lesions, is unknown. Therefore, we aimed to develop an in vivo extraradicular biofilm model in rats and to identify and quantify extraradicular biofilm-forming bacteria while investigating the effect of extraradicular biofilms on periapical lesions. Periapical lesions were induced by exposing the pulpal tissue of the mandibular first molars of male Wistar rats to their oral environment. Four weeks later, gutta-percha points were excessively inserted into the mesial root canals of the right first molars (experimental sites) but not the left first molars (control sites). After 6 and 8 weeks of pulp exposure, the presence of extraradicular biofilms was confirmed histomorphologically, and biofilm-forming bacteria were identified by using classical culture methods. The biofilms were observed in the extraradicular area of the experimental sites. Similar species were detected both inside and outside the root canals. The bacterial count, quantified by real-time PCR assays, in the extraradicular area gradually increased in the experimental sites until 20 weeks after pulp exposure. After 8 weeks of pulp exposure, the periapical lesion volume that was measured by micro-computed tomography was significantly larger in the experimental sites than in the control sites (P < 0.05 by Welch's t test). These results suggest that we developed an extraradicular biofilm model in rats and that extraradicular biofilms affect developing periapical lesions. In clinical practice, we encounter patients in whom periapical periodontitis does not heal despite a root canal treatment following general dental procedures. The etiology is considered to be the result of residual bacteria in the lateral branch of the root canal, endodontic reinfection, or both. Recent studies have shown that biofilms that form outside the apical foramen, which are termed "extraradicular biofilm," are also involved in refractory periapical periodontitis (1-4). As the bacteria in biofilms are resistant to the host's immunoreaction and to antibiotics (5), dental biofilm diseases such as caries or periodontitis have been treated by mechanical removal. However, extraradicular biofilm is impossible to remove mechanically with nonsurgical endodontic treatments. As extraradicular biofilms cannot be accessed from the root canal for mechanical removal, the only treatment methods are endodontic surgery and tooth extraction.Current research is focused on developing methods to prevent or control biofilm formation (6, 7). We recently found three analogues of N-acyl homoserine lactone that participate in quorum sensing, a system of bacterial cell-to-cell communication, and inhibit Porphyromonas gingivalis biofilm formation (8). Furthermore, azithromycin, a 15-membered macrolide antibiotic, has an antibiofilm effect on P. gingivalis (9). For clinical applicability, these in vitro effects should be exami...

Dentinogenic effects of extracted dentin matrix components digested with matrix metalloproteinases

“…Pulp-capped teeth were removed en bloc with surrounding maxillary bone and immersed in the same fixative for an additional 24 h. The induced tertiary dentin (n = 6 in each group) was analysed by using a micro-CT scanner (R_mCT2, Rigaku, Tokyo, Japan) at settings of 90 kV and 160 µA with a scanning resolution of 20 μm intervals in individual image. After scanning, the area of induced tertiary dentin of each specimen was quantitatively analysed by using three-dimensional reconstruction images for bone (TRI/3D-BON; Ratoc System Engineering, Tokyo, Japan) with reference to previous report 67 , 68 . After confirming the consistency between micro-CT and histological images, image analysis was performed.…”

Micro‐CT analysis of chronic apical periodontitis induced by several specific pathogens

“…In another in vivo study, the volume of periapical lesion reached a plateau stage from 8 to 20 weeks after induction of CAP in rats (Kuremoto et al . ). In order to compare different CAP models at a relatively stable stage, the present study adopted the time point of 12 weeks (84 days) after CAP induction.…”

“…The data are the average AE SD. (Kuremoto et al 2014). In order to compare different CAP models at a relatively stable stage, the present study adopted the time point of 12 weeks (84 days) after CAP induction.…”

Options and Limitations in Clinical Investigation of Bacterial Biofilms

“…Likewise, a rat model of A. actinomycetemcomitans-induced periodontitis was combined with RT-PCR for versatile applications, ranging from genetic reduction to therapeutic efficacy evaluation according to exopolysaccharide variations (367). Another rat model, the in vivo extraradicular biofilm model, is more inclusive, as it may facilitate identification and quantification of biofilm-forming bacteria by use of real-time PCR (rt-PCR) and micro-computed tomography (368). Chinchillas (Chinchilla lanigera) have been employed for study of middle ear infections in an attempt to establish realistic infection models, to overcome limitations in biofilm analysis, and to correlate the sequence of events in polymicrobial infections with the host immune system.…”