Stat3 and C/EBPβ synergize to induce miR-21 and miR-181b expression during sepsis

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/EBP alpha 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/EBP beta, which co-imunoprecipitate as a single complex. Mechanistically, transcription factor Rb phosphorylation supports Stat3 and C/EBP beta accumulation at both miRNA promoters, and C/EBPO 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.