Fig. 2

IL-5 is reduced in established human TH2 cells upon treatment with 4μ8c. a Blood was collected from seven individual volunteers in total. Cells were harvested from the blood using Ficoll, and CD4⁺ cells were isolated using Dynabeads. The cells were activated with plate-bound α-CD3 and α-CD28 for eleven days under TH2 conditions (IL-2, IL-4, α-IFNγ, and α-IL-12). The cells were rested for 24 h and then re-stimulated with plate-bound antibodies for 24 h in the presence or absence (−) of 4μ8c. An ELISA was performed on the supernatants. Of note, for TH2 cells differentiated from one individual, we were unable to detect IL-5 after differentiating the cells for 11 days, and that sample was removed from the IL-5 analysis, leaving us with an N of six. One of the samples from an individual was lost prior to running the ELISA for IL-4, leaving us with an N of six. The data shown are the results of six individual human samples for IL-4 and IL-5 and seven individual human samples for IL-13. The standard error, upper and lower bars, and the mean, middle bar, is shown in all graphs. Hypothesis testing was done by Student’s T test unpaired, Welch’s correction (p value < 0.05). b CD4⁺ cells were isolated from human blood as in A, activated under TH2 or TH1 conditions (IL-2, IL-12, and α-IL-4) for three days, and then stimulated with PMA and ionomycin in the presence of monensin for four hours. Intracellular staining was performed. The results are representative of six samples for IL-4 and three samples for IL-5. c The results for the percent positive and mean fluorescence intensity (MFI) d for IL-4 and IL-5 in treated and untreated cells differentiated three days in the presence of 4μ8c is shown for all intracellular flow experiments performed