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. 2021 Dec 31;12(6):2859–2868. doi: 10.1016/j.apsb.2021.12.017

Figure 1.

Figure 1

NAMPT promotes the expansion of MDSCs independent of its enzymatic activity. (A) Immunoblot analysis of iNAMPT (intracellular NAMPT) and eNAMPT (extracellular NAMPT). (B) Cell growth of CT26 cells with stable knockdown of NAMPT. Left, CT26 scramble control (shNC) or shNAMPT cell confluency assessed by incuCyte proliferation assay; Right, immunoblot analysis of NAMPT. (C) Tumor growth curve. CT26 shNC control or shNAMPT cells were inoculated subcutaneously in BALB/c mice (n = 10). (D–H) Tumor infiltrating immune cells analyzed by flow cytometry. CT26 shNC control or shNAMPT cells were inoculated subcutaneously in BALB/c mice (n = 15). (D) Heatmap showing the proportion of the indicated immune cells in tumor infiltrating CD45+ cells. (E) MDSCs; (F) CD8+ T cells; (G) IFNγ+ CD8+ T cells; (H) Granzyme B+ CD8+ T cells. (I) In vitro MDSC differentiation assay. Left, the schematic illustration of co-culture experiment. MC38 scramble control (shNC) or shNAMPT cells were co-cultured with mice bone marrow. Right, normalized counts of GR1high CD11b+ cells analyzed by flow cytometry. (J) Signal transducer and activator of transcription 3 (STAT3) signaling change in MDSCs. MDSCs from murine bone marrow were treated with recombinant NAMPT (200 ng) for 2 h and immunoblotting analysis was performed. (K) Proportion of MDSCs in tumor infiltrating CD45+ cells. CT26 tumor-bearing mice were treated with vehicle control or FK866 (16 mg/kg, daily) for 5 days (n = 5). All data depict mean ± SEM. n.s., not significant. ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001.