RNAi-mediated silencing of Atp6i and Atp6i haploinsufficiency prevents both bone loss and inflammation in a mouse model of periodontal disease.

Hongbing Jiang,Junqing Ma,Lijie Zhang,Shengmei Feng,Guochun Zhu,Byron Tucker,Hongliang Ci,Yi-Ping Li,Lin Wang,Philip Stashenko,Wei Chen,Liang Hao,Michael Glogauer

doi:10.1371/journal.pone.0058599

Abstract

Periodontal disease affects about 80% of adults in America, and is characterized by oral bacterial infection-induced gingival inflammation, oral bone resorption, and tooth loss. Periodontitis is also associated with other diseases such as rheumatoid arthritis, diabetes, and heart disease. Although many efforts have been made to develop effective therapies for this disease, none have been very effective and there is still an urgent need for better treatments and preventative strategies. Herein we explored for the first time the possibility that adeno-associated virus (AAV)-mediated RNAi knockdown could be used to treat periodontal disease with improved efficacy. For this purpose, we used AAV-mediated RNAi knockdown of Atp6i/TIRC7 gene expression to target bone resorption and gingival inflammation simultaneously. Mice were infected with the oral pathogen Porphyromonas gingivalis W50 (P. gingivalis) in the maxillary periodontium to induce periodontitis. We found that Atp6i depletion impaired extracellular acidification and osteoclast-mediated bone resorption. Furthermore, local injection of AAV-shRNA-Atp6i/TIRC7 into the periodontal tissues in vivo protected mice from P. gingivalis infection-stimulated bone resorption by >85% and decreased the T-cell number in periodontal tissues. Notably, AAV-mediated Atp6i/TIRC7 knockdown also reduced the expression of osteoclast marker genes and inflammation-induced cytokine genes. Atp6i+/− mice with haploinsufficiency were similarly protected from P. gingivalis infection-stimulated bone loss and gingival inflammation. This suggests that AAV-shRNA-Atp6i/TIRC7 therapeutic treatment may significantly improve the health of millions who suffer from P. gingivalis-mediated periodontal disease.

Highlights

Periodontitis is a common chronic inflammatory disease that is induced by polymicrobial infection, with a prominent pathogen being Porphyromonas gingivalis
In order to determine the ability of the associated virus (AAV)-shAtp6i and the AAV-sh-luc-yellow fluorescent protein (YFP) vectors to transduce target cells, we examined the expression of EGFP or YFP in target cells
Mouse bone marrow (MBM) isolated from wild-type BALB/cJ mice was cultured with M-CSF and receptor activator of nuclear factor kappa-B ligand (RANKL) to generate osteoclasts, that were transduced with AAV-shAtp6i or AAV-sh-luc-YFP

Summary

Introduction

Periodontitis is a common chronic inflammatory disease that is induced by polymicrobial infection, with a prominent pathogen being Porphyromonas gingivalis. An investigation conducted by Kawai et al that activated T- and B-cells in the gingival tissues are the primary sources of receptor activator of nuclear factor kappa-B ligand (RANKL) that induce osteoclastogenesis, osteoclast activation, and bone loss in periodontitis [1]. Inhibiting the function of RANKL produced by activated T-cells can prevent alveolar bone loss [2]. These studies provide strong evidence that T-cell activation mediates bone loss via recruitment and activation of osteoclasts. Functional studies of TIRC7 in vitro and in vivo via TIRC72/ 2 null mice have indicated that TIRC7 has a significant association with the regulation of T- and B-cell activation [7]. Several studies have demonstrated increased survival of organ allograft transplants with anti-TIRC7 mAb therapy [8,9]

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Results

Conclusion