(4) microRNAs in Resolution of Periodontal Inflammation:Therapeutic, Non-Surgical Intervention in Humans

Introduction: While significant progress has been made in elucidating cellular and molecular mechanisms associated with resolution of periodontal inflammation, very little is known regarding the role of miRNAs in this process. Given that clinical and radiologic determinants of periodontal disease i.e. probing depths, clinical attachment levels, and crestal bone loss are measured retrospectively, identification for accurate biomarkers in the diagnosis of periodontitis, at earlier inflammatory stages of disease is paramount. This ex-vivo, case-control study focused on comprehensive miRNA profiling before and after non-surgical periodontal therapy in humans.

Methods: Gingival biopsies were collected from age and gender-matched subjects with periodontal disease (n = 10) before/after non-surgical therapy and from periodontally healthy (n = 11) subjects. Differential miRNA expression quantified using Taqman miRNA arrays. Bioinformatic global pathway analysis and miRNA target prediction was performed for differentially expressed miRNAs. RT-qPCR was used to validate human miRNA data in murine model of ligature-induced periodontitis. Primary human macrophages were transfected with miR-29c mimic/inhibitors and supernatant cytokine levels quantified by multiplex bead assays after challenge with live P. gingivalis (Pg) or A. actinomycetecomitans (Aa). Wound closure assays were performed to monitor miRNA impact on cell migration.

Results: 46 miRNAs were responsive to non-surgical periodontal therapy suggesting a role in the resolution of periodontal inflammation. Of importance, miR-29 family (miR-29a, -b, -c) expression was upregulated in diseased biopsies and restored to levels approximating healthy biopsies post-therapy. These findings corroborated with the murine periodontitis model. In-vitro functional analysis revealed reduced secretion of anti-inflammatory cytokines (IL-10, IL1RA) in miR-29c transfected macrophages challenged with live Pg/Aa. miR-29c overexpression attenuated macrophage migration in wound closure assays. Impaired cytokine secretion and cell motility by miR-29c is regulated by its putative targets- IL10RB and TGF-β.

Conclusions: Elevation of miR-29c in diseased gingiva and its repression post-therapy indicates miR-29c may play a key role in pathogen/antigen clearance and its downregulation is required for wound healing and/or clinical improvements in periodontal parameters. This is the first study to comprehensively identify changes in miRNA profiles during the resolution phase of periodontal inflammation and highlight the therapeutic potential of miR-29c in promoting pro-resolution environment and periodontal destruction.
Clinical Relevance: Characterization of miRNA as potential biomarkers for early detection of periodontitis can prove to be greatly effective for diagnosis and management of oral diseases.

Lead Author's Contributions to the Research Project. : Lead author was responsible for writing the IRB proposal, recruiting subjects, data collection, performing the data analysis, and the composition of the abstract