Expression of miR-155 and SHIP1 in CD16-activated NK cells, and the role of miR-155 in regulating IFN-γ. (A) Primary human NK cells were activated with immobilized human IgG and/or IL-12 (10 ng/mL) for 24 hours and quantified for miR-155 and (B) BIC mRNA expression by real-time RT-PCR. (C) Supernatants were analyzed for IFN-γ production by ELISA. (D) SHIP1 mRNA expression was quantified by real-time RT-PCR and (E) protein expression by Western blot. (F) CD56⁺ primary human NK cells were infected using Lenti or Lenti–miR-155 vectors, sorted for GFP, and left unstimulated or stimulated with immobilized human IgG and/or IL-12 (10 ng/mL) for IFN-γ secretion. This experiment is representative of 2 experiments performed with identical results. (G) NK cells were isolated from spleens of Bic^+/+ and Bic^−/− mice, cultured 8 days in medium containing IL-2 (900 IU/mL), stimulated for 24 hours in vitro with immobilized anti-CD16 Ab and/or IL-12 (20 ng/mL) and assayed for IFN-γ.