RNA Interference-Mediated Silencing of Atp6i Prevents Both Periapical Bone Erosion and Inflammation in the Mouse Model of Endodontic Disease

Abstract

dDental caries is one of the most prevalent infectious diseases in the United States, affecting approximately 80% of children and the majority of adults. Dental caries may lead to endodontic disease, where the bacterial infection progresses to the root canal system of the tooth, leading to periapical inflammation, bone erosion, severe pain, and tooth loss. Periapical inflammation may also exacerbate inflammation in other parts of the body. Although conventional clinical therapies for this disease are successful in approximately 80% of cases, there is still an urgent need for increased efficacy of treatment. In this study, we applied a novel gene-therapeutic approach using recombinant adeno-associated virus (AAV)-mediated Atp6i RNA interference (RNAi) knockdown of Atp6i/TIRC7 gene expression to simultaneously target periapical bone resorption and periapical inflammation. We found that Atp6i inhibition impaired osteoclast function in vitro and in vivo and decreased the number of T cells in the periapical lesion. Notably, AAV-mediated Atp6i/TIRC7 knockdown gene therapy reduced bacterial infection-stimulated bone resorption by 80% in the mouse model of endodontic disease. Importantly, Atp6i؉/؊ mice with haploinsufficiency of Atp6i exhibited protection similar to that in mice with bacterial infection-stimulated bone erosion and periapical inflammation, which confirms the potential therapeutic effect of AAV-small hairpin RNA (shRNA)-Atp6i/TIRC7. Our results demonstrate that AAV-mediated Atp6i/TIRC7 knockdown in periapical tissues can inhibit endodontic disease development, bone resorption, and inflammation, indicating for the first time that this potential gene therapy may significantly improve the health of those who suffer from endodontic disease.

A Small Molecule, Odanacatib, Inhibits Inflammation and Bone Loss Caused by Endodontic Disease

“…Thus, it is important to determine the changes in the trends of the cells (number change) and the inflammation (more severe or not) in the periapical lesion area by inhibition of Ctsk and its possible mechanism at different time points. To test the possible role during the progress of periapical lesions mediated by Ctsk, we applied the periapical mouse model established by our previous study (10,17). Samples from the normal group with or without inhibitor (only images of day 42 are shown in Fig.…”

“…Exposure of the left and right first molar pulp was performed as described previously (10,17). In brief, mice were anesthetized with peritoneal injection of 62.5 mg/kg ketamine and 12.5 mg/kg xylazine according to body weight.…”

“…Bacterial culture and infection procedure protocols were conducted as described previously (10,17). In brief, exposed pulps were infected with a mixture of four common human endodontic pathogens, including Prevotella intermedia (ATCC 25611; American Type Culture Collection [ATCC], Manassas, VA), Fusobacterium nucleatum (ATCC 25586), Peptostreptococcus micros (ATCC 33270), and Streptococcus intermedius (ATCC 27335).…”

“…Micro-computed tomography (micro-CT) scans were evaluated for bone loss as described previously, with modifications (10,17). Briefly, the two-dimensional picture that showed the largest target root canal was selected as the baseline image.…”

“…P values of Ͻ0.05 and U values of Ͼ1.96 were considered significant. Data quantification analyses were performed using the NIH Image J program as described previously (10,17).…”