shRNA against Jun increases the relative level of granulopoiesis. (A) Schematic representation of the LeGO-iG lentiviral vectors used to drive expression of GFP or Mef2c with and without shRNA against Jun (shJun). Mef2c expression is driven by the promoter of the SFFV enhancer/promoter, whereas the shRNA is driven by the murine H1 promoter. (B) Western blot analysis demonstrating the effective suppression of Jun expression in FDC-P1 cells infected with LeGO-iG shJun, as compared with control cells. (C) Bar graph demonstrating the percentage increase in the granulocytes (Gr1⁺ cells) in methycellulose cultures of BM cells receiving shJun, as compared with that of control infections (ie, without shJun, but otherwise under the same conditions and identical vectors). Values were calculated for cultures receiving GFP alone (+GFP) or Mef2c (+Mef2c), and with either a cytokine cocktail or M-CSF, as indicated. Shown is the average and standard error of 2 independent experiments. (D) Cytospins of cells stimulated with M-CSF in methylcellulose cultures show increased numbers of granulocytes in cultures expressing shJun.