Abstract
BackgroundCoccidioidomycosis or Valley fever is caused by a highly virulent fungal pathogen: Coccidioides posadasii or immitis. Vaccine development against Coccidioides is of contemporary interest because a large number of relapses and clinical failures are reported with antifungal agents. An efficient Th1 response engenders protection. Thus, we have focused on developing a dendritic cell (DC)-based vaccine for coccidioidomycosis. In this study, we investigated the immunostimulatory characteristics of an intranasal primary DC-vaccine in BALB/c mouse strain that is most susceptible to coccidioidomycosis. The DCs were transfected nonvirally with Coccidioides-Ag2/PRA-cDNA. Expression of DC-markers, Ag2/PRA and cytokines were studied by flow cytometry, dot-immunoblotting and cytometric bead array methods, respectively. The T cell activation was studied by assessing the upregulation of activation markers in a DC-T cell co-culture assay. For trafficking, the DCs were co-transfected with a plasmid DNA encoding HSV1 thymidine kinase (TK) and administered intranasally into syngeneic mice. The trafficking and homing of TK-expressing DCs were monitored with positron emission tomography (PET) using 18F-FIAU probe. Based on the PET-probe accumulation in vaccinated mice, selected tissues were studied for antigen-specific response and T cell phenotypes using ELISPOT and flow cytometry, respectively.ResultsWe found that the primary DCs transfected with Coccidioides-Ag2/PRA-cDNA were of immature immunophenotype, expressed Ag2/PRA and activated naïve T cells. In PET images and subsequent biodistribution, intranasally-administered DCs were found to migrate in blood, lung and thymus; lymphocytes showed generation of T effector memory cell population (TEM) and IFN-γ release.ConclusionsIn conclusion, our results demonstrate that the intranasally-administered primary DC vaccine is capable of inducing Ag2/PRA-specific T cell response. Unique approaches utilized in our study represent an attractive and novel means of producing and evaluating an autologous DC-based vaccine.
Highlights
Coccidioidomycosis or Valley fever is caused by a highly virulent fungal pathogen: Coccidioides posadasii or immitis
There was no difference between the primary dendritic cell (DC) originating from either C57BL6 or BALB/c mouse strains
When the cells were allowed to continue differentiating for 13 days in presence of GM-CSF and IL-4, the DCs developed typical morphology of mature DCs characterized by the appearance of dendrite structures on their cell surface
Summary
Coccidioidomycosis or Valley fever is caused by a highly virulent fungal pathogen: Coccidioides posadasii or immitis. We have focused on developing a dendritic cell (DC)-based vaccine for coccidioidomycosis. Based on the PET-probe accumulation in vaccinated mice, selected tissues were studied for antigen-specific response and T cell phenotypes using ELISPOT and flow cytometry, respectively. As is evident from the studies in patients with disseminated coccidioidomycosis and animal models, the susceptibility to the disease is related to defective T cellimmune responses [5]. Unlike other antigen-presenting immune cells, DCs migrate to lymph nodes, and activate naïve immune cells including T cells. Based on this property, DC-based vaccines have been evaluated in animal models of a variety of infections as well as cancer. Our laboratory’s focus is on developing a DC-based vaccine for coccidioidomycosis [8,22,23]
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