Figure 7.

Effects of NFI-A knockout on the expression of miR-223, M-CSFr and G-CSFr. (a) miR-223 is expressed in late sepsis Gr1⁺CD11b⁺ cells from NFI-A conditional knockout mice. Bone marrow Gr1⁺CD11b⁺ cells were isolated from sham, early and late septic mice. miRNA-enriched RNA was isolated, and levels of miR-21 and miR-181b were measured by RT-qPCR. Values were normalized to U6 RNA as an internal control. (b) Expression of M-CSFr and G-CSFr is increased in late sepsis Gr1⁺CD11b⁺ cells from NFI-A conditional knockout mice. Portion of the late sepsis Gr1⁺CD11b⁺ cells described in (a) was used for RNA extraction and determination of M-CSFr and G-CSFr mRNA levels by RT-qPCR. Values were normalized to GAPDH as an internal control. Data in (a) and (b) are expressed as mean ± SD (*^/#P < 0.05) of five mice per group and are presented relative the values from sham mice (set at onefold). Data represent one of two experiments. *Compared with control mice; ^#compared with early sepsis mice. (c) Ectopic expression of NFI-A in late sepsis Gr1⁺CD11b⁺ cells from knockout mice abolishes miR-223 expression. Late sepsis Gr1⁺CD11b⁺ cells from NFI-A conditional knockout mice were transfected with empty vector or NFI-A expression plasmid for 36 h. RNA was isolated, and levels of miR-223 were measured as in (a). (d) Ectopic expression of NFI-A in late sepsis Gr1⁺CD11b⁺ cells from knockout mice inhibits M-CSFr and G-CSFr. Portion of the late sepsis Gr1⁺CD11b⁺ cells described in (c) was used to determine M-CSFr and G-CSFr mRNA levels by RT-qPCR as in (b). Data in (c) and (d) are expressed as mean ± SD (*P < 0.05) of five mice per group and are presented relative the values from control, sham mice as in (a) and (b) (set at onefold). Data represent one of two experiments. *Compared with vector. cKO: conditional knockout; r: receptor.