Experimental models

Abstract

Introduction and Objectives:The paracoccidioidomycosis (PCM) is characterized by a chronic inflammatory granulomatous reaction. Experimental and clinical evidences indicate that cellular, rather than humoral immunity, is the effective mechanism that controls the pathogenesis and the evolution of PCM. One of the main question regard this infection is related to the selection of antigens that could induce protection in susceptible hosts. In this work, our aim was to evaluate the potential of the use of monophosphoryl lipid A (MPL-SE) as adjuvant mixed with surface antigens from P. brasiliensis (sPbAg), in order to develop an effective immune response against the infection with the pathogenic fungus. Methods and Results: C57BL/6 and MyD88KO mice were immunized three times by subcutaneous route (three weeks apart) with sPbAg mixed with MPL-SE (according to Corixa Corporation protocol). Yeast cells (Pb18) were shook vigorously (in PBS) for 30s in vortex and the supernatant (sPbAg) was used in the assays. Control mice were injected with MPL-SE with PBS or with sPbAg only. Two weeks after the last boost, the animals were challenged (iv) with viable yeast forms of P. brasiliensis. The results showed absence of lesions in lungs, liver and spleen of mice immunized with sPbAg plus MPL-SE. Moreover, the colony-forming units (CFUs) was almost absent in the organs. The protection is supposed to be mediated by IFN-γ, given that this cytokine was detected in homogenates of lungs, liver and spleen. Moreover, the levels of specific antifungal IgG2a antibodies in the sera and the DTH reaction were significantly higher when compared to the animals immunized with MPL-SE, sPbAg, or only infected. The mechanism of protection is dependent of the cell activation through Toll-like receptors. Indeed, when MyD88KO mice were immunized with MPL-SE plus sPbAg or MPL-SE plus PBS the animals were not able to control fungal growth and dissemination. Surprisingly, the inoculation of sPbAg prior to infection resulted in a severe granulomatous lesions and fungal dissemination to the all organs analyzed. Clearly, increased fungal growth after sPbAg inoculation was dependent of IL-4, since high level of this cytokine was found in the homogenates of lungs, spleen and liver of mice injected with sPbAg, but not in the other groups of mice (only infected, injected with MPL-SE, or with MPL-SE plus sPbAg). Moreover, the injection of sPbAg did not affect the course of infection in mice deficient of IL-4, suggesting that the exacerbation of PCM after inoculation of sPbAg is mediated by IL-4. Conclusion: Our study indicates that the formulation of Th2-inducing antigens from P. brasiliensis with MPL-SE, a naturally derived disaccharide adjuvant of Salmonella minnesota, is able to elicit a protective immunity against P. brasiliensis infection. The protection of immunized mice is mediated by a Th1-type immune response, whose induction is dependent of Toll-like receptor activation. Financial support: CNPq, FAPESP, FAEPA -PHAGOCYTOSIS, EXPRESSION OF CO-STIMULATOR...