Journal: Cancer research

Article Title: GSK 3 inhibition drives maturation of NK cells and enhances their antitumor activity

doi: 10.1158/0008-5472.CAN-17-0799

Figure Lengend Snippet: CD3−CD56dimCD57− NK cells and CD3−CD56dimCD57+ NK cells were sorted from donor PBMCs (n = 4) by FACS, labeled with CellTrace dye and cultured for 7 days at a 2:1 ratio with sorted, autologous CD14+ monocytes with 10 ng/ml IL-15 and either DMSO or 5 μM CHIR99021. Representative FACS histogram plots of CellTrace, CD57 and CD62L from each sorted population after culture are shown (upper panel). Cumulative (n = 4) data showing the percentages of CD57+ NK cells and CD62L+ NK cells within the total CD3−CD56+ NK cell population before and after culture (lower panel). Results are from 3 independent experiments. All cumulative data is shown as mean ± SEM. Paired Student’s t tests were used for statistical analyses. *p ≤ 0.05, **p ≤ 0.01.

Article Snippet: In some experiments, NK cells were labeled with CellTrace proliferation dye (Thermo-Fisher) prior to culture.

Techniques: Labeling, Cell Culture

Journal: Cancer research

Article Title: GSK 3 inhibition drives maturation of NK cells and enhances their antitumor activity

doi: 10.1158/0008-5472.CAN-17-0799

Figure Lengend Snippet: CD3−CD56+ NK cells and CD14+ monocytes were isolated from PBMCs and co-cultured together or separated by transwells for 7 days with 10 ng/ml IL-15 and either DMSO or 5 μM CHIR99021. A, Representative FACS scatter plots of CD57 and NKG2C expression on gated CD3−CD56+ NK cells before and after culture in each condition are shown (upper panel). Cumulative data (n = 6) of the percentage of CD57+ NK cells and the mean fluorescence intensity of CD57 gated on CD57+ NK cells within the total CD3−CD56+ NK cell population before and after culture (lower panel). Results are from 3 independent experiments. B, CD3−CD56dim NK cells were sorted from PBMCs and cultured for 7 days with 10 ng/ml IL-15 and either DMSO or 5 μM CHIR99021. Shown are representative FACS histogram plots of CellTrace proliferation dye dilution in gated CD3−CD56+CD57− NK cells and CD3−CD56+CD57+ NK cells (upper panel). Cumulative data (n = 5) showing the frequencies of CD3−CD56+CD57− and CD3−CD56+CD57+ NK cells that have undergone the indicated number of divisions (lower panel). C, Representative FACS scatter plots of CD57 and NKG2C expression on sorted CD3−CD56dim NK cells before and after culture in each condition (upper panel). Cumulative data (n = 5) of the percentage of CD57+ NK cells and the mean fluorescence intensity of CD57 gated on CD57+ NK cells within the total CD3−CD56+ NK cell population before and after culture (lower panel). Results are from 2 independent experiments. D, The indicated NK cell subsets were sorted from PBMCs and cultured 7 days with 10 ng/ml IL-15 and either DMSO or 5 μM CHIR99021. Shown are representative FACS scatter plots of CD56 and CD57 expression on NK cells from each culture condition (left panel). Cumulative data (n = 3) of the percentage of CD57+ NK cells from each culture condition (right panel). Results are from 3 independent experiments. All cumulative data is shown as mean ± SEM. Paired Student’s t tests were used for statistical analyses. *p ≤ 0.05, **p ≤ 0.01. Two-way ANOVA was used to determine statistical significance for CellTrace proliferation dye assays.

Article Snippet: In some experiments, NK cells were labeled with CellTrace proliferation dye (Thermo-Fisher) prior to culture.

Techniques: Isolation, Cell Culture, Expressing, Fluorescence