The percentage of IL-34⁺ cells in healthy individuals was evaluated in CD4⁺ or CD8⁺ CD45RC^lo or CD45RC^hi T cells (A) or in FOXP3⁺ versus FOXP3^– CD45RC^lo CD8⁺ or CD4⁺ T cells (B and C). (A) The mean ± SEM of 27 healthy individuals is represented. Mann-Whitney U test, *P < 0.05, ***P < 0.001. Representative histogram and plot of healthy individuals (B) showing the mean of expression ± SEM of 10 healthy individuals (C). (D) Soluble IL-34 was tested for suppression of CD4⁺CD25^– T cell proliferation in response to allogeneic T cell–depleted PBMCs and analyzed by flow cytometry for CFSE dilution after 5 days of culture (n = 2–5 experiments performed in duplicate). Results are expressed as the mean ± SEM of the relative proportion of dividing CD4⁺CD25^– T cells. Representative histogram of 1 experiment: Proliferation of CFSE-labeled CD4⁺CD25^– T cells cocultured with T cell–depleted PBMCs with increased concentrations of IL-34. Two-way ANOVA with Bonferroni’s post test versus M-CSF, ***P < 0.001. (E) The relative proportion of CFSE-labeled dividing CD4⁺CD25^– T cells stimulated with allogeneic T cell–depleted PBMCs was analyzed after 5 days of culture, in the presence of CD8⁺CD45RC^lo or CD4⁺CD25^hiCD127^– Tregs at 1:1 effector/suppressor ratios. The proliferation after addition of anti–IL-34–blocking Ab was evaluated and compared with isotypic control (n = 5–6 experiments performed in triplicate). The proportion of dividing CD4⁺CD25^– T cells in the control proliferation condition with allogeneic T cell–depleted PBMCs only represented approximately 60% of the cells on day 5 and was given a value of 100 in each experiment. Results are expressed as the mean ± SEM of the relative proportion of dividing CD4⁺CD25^– T cells. A representative raw CFSE profile is shown in the right panel. Wilcoxon test (versus proliferation without Tregs = 100), **P < 0.01.