Modeling macrophage response to periodontal infections in vitro

Abstract

Periodontal disease (PD) is a chronic inflammatory disorder characterized by the destruction of connective tissue, tooth loss and systemic infections. Numerous virulence factors derived from chronic opportunistic pathogens such as Porphyromonas gingivalis are responsible for enhanced microbial colonization, stimulation of inflammatory responses, and suppression of host defense response. Individuals with PD present with a complex array of inflammatory mediators, and cellular infiltration by neutrophils, lymphocytes and monocytes/macrophages. Although macrophages comprise 5–30% of cells in the cellular infiltrate of human periodontal lesions, how macrophage skewing contributes to PD, and more specifically in the downstream development of a macrophage immune response to oral bacteria are unclear. We, including others have investigated the activation of monocytes/macrophages, and discovered the means of type I vs. type II inflammatory activation vs. deactivation of macrophages both in vitro and in vivo. We have shown previously that P. gingivalis infection induces M1 macrophage formation, whereas opsonized‐P. gingivalis challenge was able to switch to M2‐like phenotype demonstrating macrophage plasticity. However, the mechanism of modulation and kinetics of macrophage inflammatory reactions during the process is unclear. Prior research in the field has failed to appreciate that it is the same monocyte/macrophage cell population which is initially polarized towards an effector “inflammatory” program and subsequently re‐polarized to the deactivation program not just in PD but in any inflammation. The overarching goal of our research is to understand how macrophage skewing contributes to PD, and the downstream development of immune response to PD‐associated bacteria towards development of new macrophage‐based therapeutics. Since macrophage polarization dynamics cannot be studied in an animal model, a reliable in vitro system is needed to provide fundamental insights into macrophage biology. We have developed an in vitro model that recapitulates different phases of the inflammatory response (recruitment, initiation, development and resolution), based on purified human peripheral blood monocytes exposed to a battery of microenvironmental cues mimicking macrophage polarization/depolarization in vivo and we have created a kinetic profile of polarization by RNA analysis. Further we have applied our model to macrophage/gingival fibroblast coculture system to study kinetics of polarization. And by comparing gene clusters that are differentially expressed in M1 and M2, we have successfully recapitulated the macrophage polarization kinetics of acute inflammatory and resolution phases during periodontal inflammation and validated the key inflammatory and transcriptional factors using RT‐PCR at different phases. In conclusion, we developed an in vitro model which can be further employed to test novel macrophage‐based therapeutics to treat periodontal disease.

Expression of Macrophage Polarization Markers against the Most Prevalent Serotypes of Aggregatibacter actinomycetemomitans

“…Macrophage polarization-modulating drugs might be the future of the immune regulation for the prevention of, treatment, and reduction in patient susceptibility for periodontal diseases [ 45 , 46 , 47 , 48 ]. The complete study of the role of macrophages in periodontal disease could be a new area in the study of molecular diagnostics and therapeutic tools, the development of biomarkers, and upgrading the clinical protocols to treat periodontal diseases.…”