Fig. 2

LAP modulates lymphocyte proliferation in a dependent pyrimidine biosynthesis manner. Murine CD4 T cells were purified from lymph nodes of naive C57BL/6 male mice and labeled with 1 μM Dye Efluor® 670 for 15 min at 37 °C and stimulated for 4 days in the presence of anti-CD3 (3 μg/ml) and anti-CD28 (1.5 μg/ml). Cells were concomitantly incubated in the presence of lapachol (Lap) or leflunomide (Lef) (10, 30 and 100 μM). a The percentage of suppression was assessed by the proliferation of murine CD4 T cells assessing dye dilution in flow cytometry analysis. Human CD4 T cells were purified from blood of healthy volunteers and labeled with 1 μM Dye Efluor® 670 for 15 min at 37 °C and stimulated for 4 days in the presence of anti-CD3 (3 μg/ml) and anti-CD28 (1.5 μg/ml). Cells were concomitantly incubated in the presence of LAP or LEF (10, 30 and 100 μM). b The percentage of suppression was assessed by the proliferation of human CD4 T cells assessing dye dilution in flow cytometry analysis. Human CD4 T cells were purified from blood of healthy volunteers and labeled with 1 μM Dye Efluor® 670 for 15 min at 37 °C and stimulated for 4 days in the presence of anti-CD3 (3 μg/ml) and anti-CD28 (1.5 μg/ml). Cells were concomitantly incubated or not with LAP (10, 30, and 100 μM) and/or uridine (30, 100, and 300 μM). c The percentage of suppression was assessed by the proliferation of human CD4 T cells assessing dye dilution in flow cytometry analysis. The results were expressed using the following formula: [proliferation of CD4 T cells only – (proliferation of CD4 T cells with LEF or LAP)/proliferation of CD4 T cells only] × 100. Data are shown as mean ± SEM, n = 5 per group. *P < 0.05, **P < 0.01, ***P < 0.001