FLOW CYTOMETRIC ANALYSIS OF CELLULAR-CHANGES IN MICE AFTER INTRADERMAL INOCULATION WITH A LIPOSOME-ISCOM ADJUVANTED VACCINE

As it is not known what changes to leucocyte homeostasis are mandatory for effective adjuvant action, the biological relevance of systemic c hanges elicited by different vaccine formulations can only be interpre ted in the context of the immunological outcomes. We used flow cytomet ry to quantify the changes in leucocyte subsets induced in mice intrad ermally immunized with SAMA4 (adjuvant group), outer membrane proteins (OMP) purified from Actinobacillus pleuropneumoniae (OMP antigen grou p), SAMA4 adjuvanted OMP (OMP vaccine group), or phosphate-buffered sa line (PBS: control group). This approach allowed direct comparisons to be made between the effects of antigen, adjuvant or antigen-adjuvant complexes on immune effector cell populations. Antigens complexed with the liposome-iscom hybrid adjuvant, SAMA4, generated strong antibody responses and cytotoxic T-cell activity in animals immunized intraderm ally, reflecting remobilization and recruitment of specific cell popul ations. Splenomegaly, due to granulocytosis, monocytosis and megakaryo cytosis, was most prominent in the OMP vaccine group. Histological exa mination of spleen sections confirmed that these changes were due prim arily to splenic haematopoiesis. Circulating numbers of granulocytes a nd monocytes increased significantly (P<0.05) in the blood of the OMP vaccine group, as did granulocyte numbers in the lungs (P<0.05). No ch anges in T-and B-cell numbers were detected by flow cytometry in the s pleens, lungs or blood over the 28-day period in any treatment group. Thymocyte numbers (predominantly CD4(+)CD8(+) cells) in the OMP vaccin e group fell by 95% within 3 days of immunization. Identical cellular responses were obtained when an innocuous antigen, ovalbumin, was comp lexed with SAMA4 instead of OMP, thus demonstrating that the adjuvant effects of SAMA4 were due to synergistic interaction between antigen a nd adjuvant and not due to the presence of toxic components. The assoc iation of strong adaptive immune responses with such complex changes i n leucocyte homeostasis induced by complexing adjuvant and antigen sug gested that the changes were important for effective vaccination and w ere not purely circumstantial.