RESUMO
OBJECTIVES: Outer inflammatory protein A (OipA) is an essential adhesin of Helicobacter pylori. We aimed to evaluate the effects of a recombinant OipA in the induction of crucial cytokines as a vaccine candidate and propolis as an adjuvant in C57BL/6 mice. MATERIALS AND METHODS: C57BL/6 mice were divided into nine groups according to the disposition of antigen and adjuvant and route of administration: subcutaneous (sc) or gavage. The administrated recombinant purified OipA and propolis concentrations were 10 µg/ml and 40 µg/ml, respectively. After vaccination, we measured expression levels of IFN-γ and IL-4 cytokine genes in the spleen cells of mice by real-time PCR. RESULTS: All results were contrasted with the negative sample. By sc injection, the expression of INF-γ was increased 3.5 and 2.9-fold for OipA and OipA plus propolis, respectively. By gavage 4.4 and 11-fold increase was found for OipA and OipA plus propolis, respectively. The administration of propolis by gavage showed more increase than Sc injection concerning the production of INF-γ. The 11-fold increase for injection of OipA plus propolis by gavage was comparable OipA plus Freund's adjuvant injected subcutaneously. This result suggested an excellent immunological response toward OipA concerning the production of INF-γ in mice. In all cases there were no notable IL-4 production increases. CONCLUSION: The results confirm the efficiency of OipA in induction of IFN-γ production, and thereby the cellular immune response. Propolis could be a suitable adjuvant.
RESUMO
BACKGROUND: Regarding the important role of proinflammatory outer membrane protein (OipA) in the pathogenesis of Helicobacter pylori infection and immunomodulatory activity of propolis, we aimed to evaluate the immunogenicity effect of a purified recombinant OipA protein and propolis in the induction of two cytokines, interferon-gamma (IFN-γ) and interleukin-4 (IL-4), in a macrophage cell model. MATERIALS AND METHODS: The recombinant protein used in the present study corresponding to the oipA expressing a 34-35 kDa protein. OipA protein was purified by Ni-NTA affinity chromatography. The purified OipA protein (2.5- 40 µg /mL) and the propolis ethanolic extract (5-40 µg/mL) were incubated with phorbol 12-myristate 13-acetate-treated human myelomonocytic cell line U937 cells. IL-4 and IFN-γ levels were measured after 48 hours of incubation using enzyme-linked immunosorbent assay. RESULTS: The amounts of IL-4 and IFN-γ were significantly increased. The optimum concentration of OipA for the secretion of IL-4 was 5 µg/ml (P<0.0001). At higher concentrations, the amount of IL-4 diminished until suppression at 40 µg/mL. The optimum concentration of propolis, resulting in the most significant increased secretion of both IL-4 and IFN-γ was 40 µg/mL (P=0.0001 and P=0.0004). CONCLUSION: We found that an OipA concentration of 10 µg/mL was more effective for IFN-γ production; however, it was not effective for the high production of IL-4. Therefore, it is postulated that the OipA could mainly induce a Th1 response through the production of IFN-γ. We also observed propolis's capability to induce IFN-γ production; however, the effective concentration for this was the same as for IL-4. Therefore, as an adjuvant, proper concentration of propolis is required for OipA to give the optimum response.
