Analysis of Collagen-induced Arthritis Model Mice Orally Infected with Porphyromonas gingivalis

Abstract

 Purpose: This study examined the impact of Porphyromonas gingivalis infection on rheumatoid arthritis (RA) in collagen-induced arthritis model mice (RA model mice).

 Materials and methods: We used eight-week old DBA/J1 mice as the RA model mice. We prepared antigen solution and adjuvant from bovine type II collagen as an emulsion and sensitized the model mice at 8 and 11 weeks to induce arthritis. For the experimental group, we used a group infected with P. gingivalis ATCC33277 strain (n=12). For the control group, we used a carboxy methylcellulose (CMC) administered group (n=12). We suspended P. gingivalis in 2.5% CMC, and administered a 0.1 ml aliquot directly inside of the mouth of the mice every other day at the concentration of 1×10⁹ CFU/ml. For the control group, we directly administered 0.1 ml of 2.5% CMC intraorally to the mice every other day. From the beginning of the experiment, we performed a clinical evaluation of arthritis every day using the method of Sarkar et al. Each week, we measured the body weight of the mice. On day 42, we sampled the mandible, joints from four limbs, and serum to examine the following items. To confirm P. gingivalis infection, we examined serum antibody titer with ELISA. We also analyzed MMP-3 and ACPA, which are clinical markers for rheumatoid arthritis, with ELISA. We evaluated micro CT images and the histological morphology of the mandibles and four limbs. The obtained results were statistically processed with Stat View software.

 Results: In the experimental group, serum antibody titer of P. gingivalis increased significantly, confirming the bacterial infection. The experimental group presented advanced redness and swelling in the extremities of four limbs compared to the control group. Forty-two days after the clinical evaluation of arthritis, the experimental group had an arthritis score that was 1.9 times greater than in the control group. The μCT analysis confirmed a significant increase in bone absorption in the experimental group compared to the control group through resorption images of alveolar bone. The experimental group exhibited swelling and bone deformation in the extremities of the four limbs, breaking of carpal cartilage, rough surface of knee joints, and rough surface of patella. MMP-3 values of the experimental group increased significantly compared to the control group. ACPA values of the experimental group also increased significantly compared to the control group. Knee joint tissues of the experimental group presented advanced invasion of inflammatory cells and damage to bones compared to the control group. MMP-13 immunostaining confirmed MMP-13 positive cells on the pannus and meniscus in the experimental group. The number of MMP-13 positive cells increased significantly compared to the control group.

 Conclusions: The results of this study suggested that oral infection of P. gingivalis accelerates destruction of the knee joints in collagen-induced arthritis model mice.