Immune Regulation by Regulatory T Cells Regulatory T cells (Tregs) play an important role in modulating immune response to self-antigens and infectious agents. Multiple regulatory mechanisms have been assigned to Tregs but how they modulate discrete immune functions has not been well defined. We are interested in determining how Tregs block cytokine production by CD4 T cells and how effector T cells may in turn modulate Treg function. Analysis of NK-kB localization in T cells relative to nuclear stain by Amnis Imagestream. To address the consequences of Treg encounter on CD4 T cell function we have performed detailed kinetic analysis in vitro and in vivo on changes in the CD4 targets. Regulatory T cells appear to abort the activation program of CD4+ T cells within 6 hours of Treg encounter (Sojka et.al. 2005, JI; Sukiennicki & Fowell 2006, JI). Using the advanced technology of the Amnis, which combines microscopy and flow cytometry, we have identified Treg-abrogation of specific signaling pathways in the target CD4 T cells and we are currently determining the functional consequence of the altered signaling and how Tregs might be mediating such effects. We are also interested in how Tregs are themselves modulated at sites of inflammation (Lazarski et.al. 2009, Immunity Correspondence). Using in vivo models of tissue inflammation (ear dermis) including the diabetic pancreas of the NOD mouse we are defining the cellular and molecular changes to Tregs as they enter different types of inflammation and how this impacts on their survival and regulatory function.