Myeloid-derived suppressor cells (MDSCs) increase late sepsis immunosuppression and mortality in mice. We reported that microRNA (miR) 21 and miR-181b expression in Gr1+CD11b+ myeloid progenitors increase septic MDSCs in mice by arresting macrophage and dendritic cell differentiation. Here, we report how sepsis regulates miR-21 and miR-181b transcription. In vivo and in vitro binding studies have shown that C/EBPα transcription factor, which promotes normal myeloid cell differentiation, binds both miRNA promoters in Gr1+CD11b+ cells from sham mice. In contrast, in sepsis Gr1+CD11b+ MDSCs miR-21 and miR-181b promoters bind both transcription factors Stat3 and C/EBPβ, which co-imunoprecipitate as a single complex. Mechanistically, transcription factor Rb phosphorylation supports Stat3 and C/EBPβ accumulation at both miRNA promoters, and C/EBPβ or Stat3 depletion by siRNA in sepsis Gr1+CD11b+ MDSCs inhibits miR-21 and miR-181b expression. To further support this molecular path for MDSC accumulation, we found that Stat3 and C/EBP binding at miR-21 or miR-181b promoter was induced by IL-6, using a luciferase reporter gene transfection into naive Gr1+CD11b+ cells. Identifying how sepsis MDSCs are generated may inform new treatments to reverse sepsis immunosuppression.

Sepsis inflammation accelerates myeloid cell generation to compensate for rapid mobilization of the myeloid progenitors from bone marrow. This inflammation-driven myelopoiesis, however, generates myeloid progenitors with immunosuppressive functions that are unable to differentiate into mature, innate immune cells. The myeloid-derived suppressor cells (MDSCs) expand markedly in the later phases of sepsis, suppress both innate and adaptive immunity, and thus, elevate mortality. Using a murine model with myeloid-restricted deletion of the C/EBPβ transcription factor, we show that sepsis-induced generation of MDSCs depends on C/EBPβ. C/EBPβ myeloid cell-deficient mice did not generate MDSCs or develop immunosuppression and survived sepsis. However, septic mice still generated Gr1CD11b myeloid progenitors at the steady-state levels similar to the control sham mice, suggesting that C/EBPβ is not involved in healthy, steady-state myelopoiesis. C/EBPβ-deficient Gr1CD11b cells generated fewer monocyte- and granulocyte-like colonies than control mice did, indicating reduced proliferation potential, but differentiated normally in response to growth factors. Adoptive transfer of C/EBPβ-deficient Gr1CD11b cells from late septic mice exacerbated inflammation in control mice undergoing early sepsis, confirming they were not immunosuppressive. These results show that C/EBPβ directs a switch from proinflammatory to repressor myeloid cells and identifies a novel treatment target.

Myeloid progenitor-derived suppressor cells (MDSCs) arise from myeloid progenitors and suppress both innate and adaptive immunity. MDSCs expand during the later phases of sepsis in mice, promote immunosuppression, and reduce survival. Here, we report that the myeloid differentiation-related transcription factor nuclear factor I-A (NFI-A) controls MDSC expansion during sepsis and impacts survival. Unlike MDSCs, myeloid cells with conditional deletion of the Nfia gene normally differentiated into effector cells during sepsis, cleared infecting bacteria, and did not express immunosuppressive mediators. In contrast, ectopic expression of NFI-A in myeloid progenitors from NFI-A myeloid cell-deficient mice impeded myeloid cell maturation and promoted immune repressor function. Importantly, surviving septic mice with conditionally deficient NFI-A myeloid cells were able to respond to challenge with bacterial endotoxin by mounting an acute inflammatory response. Together, these results support the concept of NFI-A as a master molecular transcriptome switch that controls myeloid cell differentiation and maturation and that malfunction of this switch during sepsis promotes MDSC expansion that adversely impacts sepsis outcome.KEYWORDS inflammation, sepsis immunosuppression, MDSCs, NFI-A, sepsis M yeloid progenitor-derived suppressor cells (MDSCs) represent a heterogenous population of immature myeloid cells that includes progenitors and precursors of monocytes/macrophages, granulocytes, and dendritic cells (1-3). MDSCs are generated under a variety of inflammatory and infection conditions (1, 2) and are best characterized by their immunosuppressive functions (4-6). They may also promote persistent inflammation and chronic infection with catabolism during chronic sepsis (7). MDSCs suppress both innate and adaptive immunity via production of immunosuppressive mediators and inhibition of T cell proliferation and activation (3,4,8). Phenotypically, murine MDSCs coexpress the myeloid differentiation markers Gr1 and CD11b, similarly to the Gr1 ϩ CD11b ϩ myeloid progenitors that arise under normal physiological conditions (2, 9). Unlike the immunosuppressive Gr1 ϩ CD11b ϩ cells (i.e., MDSCs), normal Gr1 ϩ CD11b ϩ cells can differentiate into competent innate immune cells (3). Elimination of MDSCs in tumor-bearing mice enhances antitumor immunity (10). Since MDSCs are "immature" cells that deviate from the standard path of differentiation, it has been suggested that arrested myeloid cell differentiation and maturation may be responsible for MDSC generation and immunorepression (2, 6). How immature myeloid cells lose their ability to differentiate and instead generate MDSCs remains unclear. Some studies, however, have suggested that MDSCs retain their potential to differentiate and mature but are trapped in a MDSC phenotype in the environmental milieu of chronic inflammation or growing tumors (1). In support of this, Kusmartsev et al. (10) reported that MDSCs from tumor-bearing mice could differentiate into macrophages and...

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