Macrophage-mediated nanoparticle delivery to the periodontal lesions in established murine model via Pg-LPS induction Zhiwei Ma 1 , Frederik Dagnæs-Hansen 2 , Henrik Løvschall 3 , Wen Song 4 , Gitte K. Nielsen 2 , Chuanxu Yang 5 , Qintao Wang 1 , Jørgen Kjems 5 , Shan Gao 5 1 State Key Laboratory of Military Stomatology, Department of Periodontology, School of Stomatology, The Fourth Military Medical University, Xi-an, China; 2 Department of Biomedicine, Aarhus University, Aarhus C, Denmark; 3 Department of Dentistry, Health, Aarhus University, Aarhus C, Denmark; 4 State Key Laboratory of Military Stomatology, Department of Prothodontics, School of Stomatology, The Fourth Military Medical University, Xi-an, China; 5 Department of Molecular Biology and Genetics, Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Aarhus, Denmark We established a murine periodontitis model by local injection of lipopolysaccharide of Porphyromonas gingi- valis (Pg-LPS) into the gingival sulcus of mandibular left incisor four times with 48-h interval. The histological examination of the periodontal tissues demonstrated that significant loss of periodontal bone and ligaments was observed in the lesion side with abundant inflammatory cell infiltration. Two days after the last injection, Cy5-labelled siRNA/chitosan particles were injected intraperitoneally (ip). The chitosan/siRNA par- ticles were taken up by peritoneal macrophages, which subsequently migrated to the inflamed gingival area evaluated by in vivo imaging. The localization of mac- rophages in the inflamed region was further confirmed by immunofluorescent staining. The present report demonstrates that intragingival injection of Pg-LPS can be used to create an experimental model of periodon- tal inflammation in mice and that recruitment of macrophages with chitosan/siRNA nanoparticles to the inflamed area opens the possibility of an RNAi-based therapeutic approach using chitosan as a carrier in periodontitis. J Oral Pathol Med (2014) Keywords: chitosan/siRNA nanoparticle; macrophages; mouse model; periodontitis Introduction Periodontitis is a kind of highly prevalent chronic immuno- inflammatory disease that manifests progressive loss of gingival tissue, the periodontal ligament and supporting alveolar bone (1). To reveal the biological process of periodontitis, several animal models such as non-human primates, miniature pigs, dogs and rodent models have been developed over the years (2). The murine models are commonly accomplished by direct oral administration of bacteria (3) or by chemical induction (4). However, the microbiota of mouse is different from human, and it is difficult to select the optimal bacteria. In addition, the mouse can somehow resist bacterial infection and extremely high bacteria number is often required, which might cause microbiota hazards. Periodontitis is mostly associated with Porphyro- monas gingivalis (5); therefore, local injection of the lipo- polysaccharides from Porphyromonas gingivalis (Pg-LPS) may be a convenient method to induce periodontitis and is often used in rats and dogs (6, 7). However, to our knowledge, a Pg-LPS-induced mouse model has never been reported. The periodontal disease is closely related to oral hygiene and accumulation of bacterial deposits. Conse- quently, the mainly current therapeutic strategy of peri- odontitis in clinic is dental scaling, root planning and periodontal surgery. RNAi technique is a method of gene silencing and has been introduced in many disease research areas as a potentially therapeutic approach. In our previous studies, the chitosan-based systematic deliv- ery of siRNA has been successfully applied as an anti- inflammatory treatment in a murine arthritis model and radiation-induced fibrosis via ip injection (8, 9). The specific delivery of chitosan/siRNA nanoparticle is sup- posingly mediated by peritoneal macrophage’s uptake and accumulation in the inflamed region (10). In this study, we successfully established a murine periodontitis model by local injection of Pg-LPS and Correspondence: Qintao Wang, DDS, State Key Laboratory of Military Stomatology, Department of Periodontology, School of Stomatology, Fourth Military Medical University, Xi’an, China. Tel: +86 29 84776096, Fax: +86 29 83223047, E-mail: qintaowang@hotmail.com and Jørgen Kjems and Shan Gao, Department of Molecular Biology and Genetics, The Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Gustav Wiedsvej 14, 8000 Aarhus C, Denmark. Tel: +45 87155863, Fax: +45 87154041, E-mails: jk@mb.au.dk and shg@mb.au.dk Accepted for publication August 21, 2014 doi: 10.1111/jop.12269 J Oral Pathol Med © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd wileyonlinelibrary.com/journal/jop