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. 2019 Apr 15;129(5):2094–2106. doi: 10.1172/JCI123955

Figure 3. Phenotypic and functional effects of NKG2A downregulation by PEBL.

Figure 3

(A) Downregulation of NKG2A is accompanied by decrease of surface CD94 expression. Flow cytometry dot plots show expression of NKG2A (CD159a PE) and of CD94 (anti-CD94 APC) in a representative sample of NK cells expanded by coculture with K562-mb15-41BBL and transduced with either anti-NKG2A PEBL-2 or GFP alone (Control). (B) Summary of CD94 versus NKG2A expression results obtained with NK cells from 7 donors. (C) NKG2C (CD159c APC) versus NKG2A expression obtained with NK cells from 8 donors. (D) Expression of CD25 in PEBL-transduced and control NK cells. Flow cytometry histograms show expression of CD25 (red, detected with anti–CD25 PE-Cy7); staining with isotype-matched nonreactive antibody (mouse IgG1 PE-Cy7) is shown in gray. (E) Survival and expansion of PEBL-transduced and control NK cells from 6 donors cultured with IL-2 (400 IU/ml). Data are shown as mean (± SD) of triplicate measurements at each time point. (F) Results of 4-hour cytotoxicity assays performed against luciferase-labeled K562 cells. BrightGlo was added after 4 hours of coculture, and luminescence was measured using a Flx 800 plate reader. Data are shown as mean (± SD) of triplicate measurements with NK cells from 8 donors. (G) Long-term cytotoxicity of PEBL-transduced and control NK cells against mCherry-transduced K562 cells at 1:8 E/T. K526 cell growth was measured with IncuCyte Zoom System. Data are shown as mean (± SD) of triplicate measurements with NK cells from 1 donor and of cultures without NK cells.