The role of chemokines and dendritic cells in regulation of IL-4 and fungal immunity.

Histoplasma capsulatum is a dimorphic fungus that can cause lethal infection in immunocompromised hosts. In order to resolve infection, the host must mount a TH1 immune response that is accompanied by an influx of inflammatory cells to the primary site of infection, the lung. Chemokines regulate cell recruitment and TH1/TH2 polarization; however, the role of chemokines in control of H. capsulatum infection has not been explored. We have demonstrated that the chemokine receptor CCR2 is requisite for protective immunity to H. capsulatum. H. capsulatum infected CCR2 mice manifested defects in inflammatory cell recruitment, increased interleukin (IL)-4, and progressive infection. Increased IL-4 in CCR2 mice diminished host resistance as demonstrated by the ability of IL-4-neutralized CCR2 mice to resolve infection without altering cell recruitment. In contrast, mice lacking CCL2, a major CCR2 ligand, did not exhibit elevated IL-4 and resolved infection. Lack of both CCL2 and the CCR2 ligand CCL7 produced a phenotype similar to that observed in CCR2 mice demonstrating for the first time that CCL2 and CCL7 mutually regulate IL-4. We examined the cellular sources that contributed to IL-4 production in lungs of CCR2 mice. Surprisingly, numerous alveolar macrophages and dendritic cells (DC) secreted IL-4 in addition to CD4 T cells at the peak of infection. We explored the mechanism by which IL-4 exacerbated infection. Elevations of IL-4 in CCR2 mice were associated with alternatively activated M which exhibited increased arginase-1. Administration of L-arginine, an arginase-1 substrate, or BEC ((S)-(2-Boronoethyl)-L-cysteine, HCl), an arginase-1 inhibitor, partially decreased fungal burden in CCR2 mice indicating that increased arginase-1 expression is deleterious to host defense. We investigated the mechanism by which IL-4 becomes elevated in lungs of CCR2 mice. Infected CCR2 or CCL2-CCL7 deficient mice exhibited a marked decrement in the number of CD8 conventional DC in lungs, therefore we hypothesized that these cells restrict IL-4 generation. Adoptive transfer of antigen (Ag)-loaded WT bone marrow-derived DC (Ag-BMDC) to WT or CCR2 mice resulted in suppression of IL-4 whereas adoptive transfer of BMDC that were not exposed to Ag had no effect. Transfer of Ag-BMDC lacking CCR2 also limited IL-4 generation. We asked if DCmediated MHCII-Ag presentation to CD4 T cells was necessary for IL-4 restriction. MHCII Ag-BMDC did not limit IL-4 when transferred to CCR2 mice and depletion of CD4 T cells prior to, but not at the initiation of infection, resulted in decreased IL-4 indicating that AgBMDC suppression of IL-4 is dependent on limiting production by CD4 T cells early during infection. Fungal burden was not reduced in CD4-depleted CCR2 mice but CCR2 mice that received Ag-BMDC in addition to depletion of CD4 T cells were able to control infection. Together, the data suggest that elevated IL-4 in CCR2 mice arises from decreased recruitment of DC capable of presenting antigen. Furthermore, CD4 T cells dampen immunity in H. capsulatum infected-CCR2 mice. Acknowledgements I would like to thank my mentor Dr. George S. Deepe, Jr. Dr. Deepe allowed me to take my research in any direction that I found interesting and has guided me in every way possible throughout this journey. Because of his mentoring I feel independent and prepared to leave the lab to pursue new research projects. I must also thank him for the endless amount of manuscript editing he has performed. Although he could have saved much time for himself by rewriting my original drafts, he spent a great amount of time teaching me how to write scientifically. I would also like to thank the past and present members of the laboratory who have given their scientific and emotional support over the years.