Bordetella pertussis filamentous hemagglutinin enhances the immunogenicity of liposome-delivered antigen administered intranasally

Abstract

In an attempt to increase the immunogenicity of mucosally delivered antigens, we incorporated the Bordetella pertussis filamentous hemagglutinin (FHA) adhesin into liposomes containing the glutathione S-transferase of Schistosoma mansoni (Sm28GST) as a model antigen. Outbred mice immunized twice intranasally with liposomes containing a constant suboptimal dose of Sm28GST and increasing doses of FHA produced anti-Sm28GST antibodies in a FHA dose-dependent manner. The addition of 3 microg of FHA to the liposomes induced more than 10-fold-higher anti-Sm28GST antibody titers, compared to those induced by liposomes without FHA. The presence of FHA did not alter the nature of the humoral immune response, and the sera contained anti-Sm28GST immunoglobulin G1 (IgG1), IgG2a, and IgG2b. However, anti-Sm28GST IgA was only detected when at least 3 microg of FHA was added to the preparation. These results show a promising potential for FHA to enhance the immunogenicity of mucosally administered antigens incorporated into liposomes.

FIG. 1

Immunoblot analysis of Sm28GST-FHA liposomes. Five microliters of each liposome suspension used for in vivo experiments (corresponding to 0.25 μmol of phospholipids) was analyzed by immunoblotting with rat hyperimmune polyclonal anti-FHA (left panel) and rat hyperimmune polyclonal anti-Sm28GST (right panel). The nitrocellulose sheets were then incubated with anti-rat antibodies coupled to alkaline phosphatase (Sigma) (1/4,000 in PBS containing 1% fat dry milk) and developed with 5-bromo-4-chloro-3-indolylphosphate–nitroblue tetrazolium (Promega, Madison, Wis.). Lanes 1 contained 3 μg of free rSm28GST, and lanes 2 contained 1.5 μg of free FHA. Lanes 3, 4, and 5 contained the three different Sm28GST-FHA liposome preparations (i.e., high, medium, and low concentrations of FHA, respectively). Lanes 6 contained the Sm28GST liposome preparation. The molecular mass markers, expressed in kilodaltons (kD), are given in the left margin.

FIG. 2

FACS analysis of Sm28GST-FHA liposomes. Sm28GST liposomes and Sm28GST-FHA liposomes were labeled with a rat anti-FHA polyclonal antibody followed by fluorescein isothiocyanate-labeled anti-rat antibody and then subjected to FACS analysis. The numbers (nb) of liposomes were plotted against fluorescence intensity as indicated.