Duque‐Correa, Maria A.

Abstract Macrophages with suppressive characteristics such as Tumor associated macrophages or Myeloid derived suppressor cells have been defined showing regulatory function under inflammatory or polarizing conditions. Until now, there are no reports indicating that under normal, non-inflammatory or non-polarizing conditions, macrophages could have immunoregulatory functions. Utilizing knock-in Foxp3-GFP mice, we identified a new population of Foxp3 positive cells that were CD11b+F4/80+ cells. These cells were observed in spleen, lymph nodes, bone marrow and thymus. Similar to Tregs, the CD11b+F4/80+Foxp3+ macrophages inhibited T cell proliferation, whereas Foxp3neg macrophages did not. The Foxp3neg macrophages could be converted to express Foxp3. These induced-CD11b+F4/80+Foxp3+ cells displayed inhibitory properties similar to natural Foxp3pos macrophages. Based on the immunoregulatory properties of these cells, we termed them Macrophage regulatory cells (Mac-regs). These Mac-regs express CD68hiF4/80hiCTLA4hiGITRhiIL-4Rhi. Interestingly, GR-1 is not a differentiation marker. The cytokine, chemokine, growth factor and genomic profiles of Mac-regs are different compared to CD11b+F4/80+Foxp3-cells. For the first time, we demonstrated the occurrence of a natural population of macrophages displaying regulatory function which may contribute to the homeostatic balance of the immune system. This research was supported by Grants from the NIH and AFAR.

Regulatory T cells (T reg cells) are characterized by the expression of the forkhead lineage-specific transcription factor Foxp3, and their main function is to suppress T cells. While evaluating T reg cells, we identified a population of Foxp3-positive cells that were CD11b+F4/80+CD68+, indicating macrophage origin. These cells were observed in spleen, lymph nodes, bone marrow, thymus, liver, and other tissues of naive animals. To characterize this subpopulation of macrophages, we devised a strategy to purify CD11b+F4/80+Foxp3+ macrophages using Foxp3-GFP mice. Analysis of CD11b+F4/80+Foxp3+ macrophage function indicated that these cells inhibited the proliferation of T cells, whereas Foxp3− macrophages did not. Suppression of T cell proliferation was mediated through soluble factors. Foxp3− macrophages acquired Foxp3 expression after activation, which conferred inhibitory properties that were indistinguishable from natural Foxp3+ macrophages. The cytokine and transcriptional profiles of Foxp3+ macrophages were distinct from those of Foxp3− macrophages, indicating that these cells have different biological functions. Functional in vivo analyses indicated that CD11b+F4/80+Foxp3+ macrophages are important in tumor promotion and the induction of T reg cell conversion. For the first time, these studies demonstrate the existence of a distinct subpopulation of naturally occurring macrophage regulatory cells in which expression of Foxp3 correlates with suppressive function.

Abstract T cell responses are compromised in the elderly. The B7-CD28 family receptors are critical in the regulation of immune responses. We evaluated whether the B7-family and CD28-family receptors were differentially expressed in dendritic cells, macrophages, and CD4+ and CD8+ T cells from young and old mice, which could contribute to the immune dysfunction in the old. Although most of the receptors were equally expressed in all cells, >85% of the old naive CD8+ T cells expressed B7-H1 compared with 25% in the young. Considering that B7-H1 negatively regulates immune responses, we hypothesized that expression of B7-H1 would downregulate the function of old CD8+ T cells. Old CD8+ T cells showed reduced ability to proliferate, but blockade of B7-H1 restored the proliferative capacity of old CD8+ T cells to a level similar to young CD8+ T cells. In vivo blockade of B7-H1 restored antitumor responses against the B7-H1− BM-185–enhanced GFP tumor, such that old animals responded with the same efficiency as young mice. Our data also indicate that old CD8+ T cells express lower levels of TCR compared with young CD8+ T cells. However, following antigenic stimulation in the presence of B7-H1 blockade, the levels of TCR expression were restored in old CD8+ T cells, which correlated with stronger T cell activation. These studies demonstrated that expression of B7-H1 in old CD8+ T cells impairs the proper activation of these cells and that blockade of B7-H1 could be critical to optimally stimulate a CD8 T cell response in the old.