A Combination of Cytokines Rescues Highly Purified Leukemic CLL B-Cells from Spontaneous Apoptosis In Vitro
Figure 4
The pro-survival effect of PMA, IL-4 and a cytokine cocktail is sustained for 7 days of culture.
A. Apoptosis was evaluated after 24, 48, 72 and 168 hours of culture by annexin V–PE/7-AAD staining and flow cytometry. The survival of B-CLL cells in the presence of PMA or IL-4 or cytokine cocktail was greater than that of controls for up to 168 h. Lower panel: Cytometry plots from a representative patient at 168 h. Upper panel: The values reported are means ± SEM for 9 independent experiments, each performed in duplicate. B. 106 purified B-CLL cells/well were cultured in 24-well plates in 1 ml of RPMI 1640 complete medium in the presence of PMA or IL-4 or the cytokine cocktail. Changes in viable cell number were assessed (counted in duplicate) by a trypan blue exclusion method after 24, 48, 72 and 168 hours of culture. The values reported are means ± SEM for 9 independent experiments. The significance of differences was calculated with the Wilcoxon test: *p<0.05 **p<0.01. Red asterisks for PMA versus Medium, clear blue asterisks for Cc versus Medium and indigo asterisks for IL-4 versus Medium.