Emmanuel Albuquerque-Souza
| Emmanuel Albuquerque-Souza |
Emmanuel Albuquerque-Souza1, Katie Crump2, Kantapon Rattanaprukskul3, Yajie Li3, Benjamin Shelling3, Xia-Juan Xia3, Sinem Esra Sahingur3
1Endodontics, University of Pennsylvania, School of Dental Medicine; 2Department of Biological Sciences, Nova Southeastern University; 3Periodontics, University of Pennsylvania, School of Dental Medicine
Introduction
TLR9 is a critical nucleic acid sensing receptor in mediating periodontitis and periodontitis-associated comorbidities. Emerging evidence implicates TLR9 as a key sensor during aging, although its role in the pathophysiology of periodontitis in aged organisms is unexplored. Here, we investigated whether TLR9-mediated host responses can regulate the pathogenesis of periodontitis during aging by utilizing a multipronged approach comprised of clinical and pre-clinical studies.
Methods
In a case-control model, we assessed TLR9 expression in gingival tissues from older (≥55 years) and younger (<55 years old) periodontitis (10-younger and 25-older) and healthy (17-younger and 10-older) individuals by qPCR. In an in vivo longitudinal model, using young (8-10-weeks) and aged (18-22-months) wild-type and TLR9-/- mice, we measured alveolar bone levels, gingival expression of inflammatory markers (Cxcl8/Rankl/Il-6) and danger signals (S100A8/S100A9), and the pro-senescence balance (p16INK4a:p19ARF) by using microtomography, qPCR, and immunofluorescence, respectively. Ex-vivo experiments with WT and TLR9-/- bone-marrow derived macrophages (BMDMs) primed by TLR9 ligands evaluated inflammaging/senescence markers (IL-6/S100A8-A9/β-galactosidase) by ELISA; p16INK4a:p19ARF balance by immunofluorescence; and osteoclast differentiation by TRAP staining.
Results
Increased gingival TLR9 expression was noted in periodontitis affected older subjects compared to older or younger healthy individuals. Mechanistically, this finding was supported by our in vivo model whereby aged-WT mice developed severe alveolar bone resorption when compared to their younger counterpart, whereas aged-TLR9-/- animals presented insignificant bone loss when compared to the younger groups. In parallel, a boosted inflammaging milieu exhibiting higher expression of inflammatory/osteoclast mediators and danger signals was noted in gingival tissues of aged-WT mice compared to aged-TLR9-/- mice. Consistently, WT-aged mice displayed an increased p16INK4a: p19ARF deleterious pro-aging balance at both gene and protein levels when compared to the younger groups and aged-TLR9-/- animals. Ex-vivo experiments with BMDMs further corroborated in vivo and clinical data and showed enhanced inflammatory-senescence circuit followed by increased osteoclast differentiation.
Conclusion
Together our findings provided the first systematic evidence indicating TLR9 as one of the key drivers of periodontitis during aging through boosting a deleterious inflammaging/senescence environment.