Fig. 4: R-P4 is non-toxic to T and B cells.

a Primary human CD4⁺ T cells (left) and B cells (right) were incubated with R-P4 (20 μM) or granadaene (0.5 μM) for 1 h at 37 °C, and cytotoxicity was measured by LDH release into cell supernatant relative to 100% lysis (Triton X-100 (0.1%)) and 0% lysis (PBS) controls. Mean and standard error from three independent experiments are shown. Groups were compared with a two-tailed unpaired t test. CD4⁺ T cells: p = 0.0024, B cells: p = 0.0084. b Primary human CD4⁺ T cells were treated with PMA (10 ng/mL) and anti-CD3ε (100 μg/mL) (stimulated) or media alone (unstimulated). Then, cells were treated with either PBS or R-P4 (20 μM) and incubated at 37 °C for 48 h. Cells were stained for CD69, resuspended in DAPI (0.5 μM), and analyzed by flow cytometry. Percent CD69⁺ of DAPI− cells from three independent experiments are represented with mean and standard error. Treatment groups were compared using one-way ANOVA with Tukey’s post test. PBS (unstimulated) vs. PBS (stimulated): p < 0.0001, PBS (unstimulated) vs. R-P4 (stimulated): p < 0.0001, PBS (stimulated) vs. R-P4 (stimulated): p = 0.8151. c Primary human B cells were treated with IL-4 (20 ng/mL) and anti-CD40 (5 μg/mL) (stimulated) or media alone (unstimulated). Then, cells were treated with either PBS or R-P4 (20 μM) and incubated at 37 °C for 48 h. Cells were stained for CD69, resuspended in DAPI (0.5 μM), and analyzed by flow cytometry. Percent CD69⁺ of DAPI− cells from three independent experiments are represented with mean and standard error. Treatment groups were compared using one-way ANOVA with Tukey’s post test. PBS (unstimulated) vs. PBS (stimulated): p = 0.0091, PBS (unstimulated) vs. R-P4 (stimulated): p = 0.0453, PBS (stimulated) vs. R-P4 (stimulated): p = 0.3893. *, p < 0.05. **, p < 0.01. ****, p < 0.0001. n.s., Not significant.