Fig. 7

The protective immune response induced by a divalent vaccine in a F. nucleatum and P. gingivalis mixed-infection model. a Experimental schedule. Mice were intranasally immunized with PBS, 5.1 μg FlaB-LK1-tFomA (BtA), 8 μg Hgp44-LK3-FlaB (HB) or a mixture of 5.1 μg BtA and 8 μg HB (BtA + HB) three times at a two-week interval. Naive mice were used as controls. Two weeks after the final immunization, the immunized mice were orally challenged with F. nucleatum (ATCC 10953) and P. gingivalis (ATCC 33277) for 3 successive days as described in the Methods section. b Determination of inflammatory cytokines and MMP9 mRNA expressions in the gingival tissues. Three days after the final challenge, gingival tissues were prepared and cytokines and MMP9 mRNA expression levels were determined by qRT-PCR. c Determination of bone density and distance from the CEJ to the ABC. Forty days after the final challenge, alveolar bone loss was determined by measuring the bone volume density (bone volume/tissue volume; BV/TV) and distance from the CEJ to the ABC using micro-CT. The region of interest (ROI) was defined as a cuboidal bone body encompassing the roots of the first molar (M1) and second molar (M2) of the maxillae. The data are presented as the mean ± SEM for each group. N = 8, *P < 0.05, **P < 0.01, *** P < 0.001, NS non-significant