Extracellular Vesicles Promote Apical Periodontitis.

is an important contributor to the persistence of chronic apical periodontitis. However, the mechanism by which infection in the root canals and dentinal tubules affects periapical tissue remains unclear. Bacterial extracellular vesicles (EVs) act as natural carriers of microbe-associated molecular patterns (MAMPs) and have recently attracted considerable attention. In this study, we investigated the role of EVs derived from in the pathogenesis of apical periodontitis. We observed that EVs can induce inflammatory bone destruction in the periapical areas of mice. Double-labeling immunofluorescence indicated that M1 macrophage infiltration was increased by EVs in apical lesions. Moreover, in vitro experiments demonstrated the internalization of EVs into macrophages. Macrophages tended to polarize toward the M1 profile after treatment with EVs. Pattern recognition receptors (PRRs) can recognize MAMPs of bacterial EVs and, in turn, trigger inflammatory responses. Thus, we performed further mechanistic exploration, which showed that EVs considerably increased the expression of NOD2, a cytoplasmic PRR, and that inhibition of NOD2 markedly reduced macrophage M1 polarization induced by EVs. RIPK2 ubiquitination is a major downstream of NOD2. We also observed increased RIPK2 ubiquitination in macrophages treated with EVs, and EV-induced macrophage M1 polarization was notably alleviated by the RIPK2 ubiquitination inhibitor. Our study revealed the potential for EVs to be considered a virulence factor of and found that EVs can promote macrophage M1 polarization via NOD2/RIPK2 signaling. To our knowledge, this is the first report to investigate apical periodontitis development from the perspective of bacterial vesicles and demonstrate the role and mechanism of EVs in macrophage polarization. This study expands our understanding of the pathogenic mechanism of and provides novel insights into the pathogenesis of apical periodontitis.