Methods: The plasmid encoding CCL19-GFP fusion protein, designated as pCCL19/GFP was constructed by inserting murine ccl19 gene into the GFP-expressing vector pAcGFP1-N1. Chemotactic effect of the CCL19-GFP fusion protein expressed by pCCL19/GFP on dendritic cells (DCs) was assessed in vitro and in vivo. BALB/c mice were intramuscularly immunized with anti-caries DNA vaccine pCIA-P or with pCIA-P plus pCCL19/GFP. Serum samples were collected at 0, 4, 6, 8, 10, 12, 14 and 16 weeks after the first immunization and analyzed for anti-PAc IgG level by ELISA. Splenocytes derived from immunized mice were restimulated with rPAc protein for 48 h, and the production of IFN-γ and IL-4 was determined by ELISA. Additionally, the presence and protein expression of pCCL19/GFP in spleen and draining lymph nodes following intramuscular immunization were assessed by PCR and immunofluorescence techniques.
Results: CCL19-GFP fusion protein showed strong chemotactic activity on DCs in vitro and in vivo. Genetic co-transfer of CCL19 increased the levels of serum PAc-specific IgG by 2-fold approximately. Splenocytes from pCIA-P plus pCCL19/GFP vaccinated mice produced significantly higher level of IFN-γ than thoes from pCIA-P vaccinated mice, suggesting induction of a Th1-biased immune response in the presence of pCCL19/GFP. Moreover, following muscular immunization, pCCL19/GFP was shown to exist and express CCL19-GFP fusion protein in spleen and draining lymph nodes, which might explain the increased number of DCs in these tissues.
Conclusion: These findings indicate that CCL19 co-immunization can enhance the immunogenicity of anti-caries DNA vaccine by inducing chemotactic migration of DCs to secondary lymphoid tissues.