Fig. 4

B-1P cells are recruited to cutaneous lesions in vivo. (A) Representative image of the lesions and a histological section of an infected footpad during the course of C57BL/6 mice infection with L. amazonensis. The inset image shows large Leishmania-PV of different cells within the infected tissue. (B) Normalized percentage of CD45⁺F4/80⁺ populations presenting B-1P cells (CD19⁺ cells) or macrophages (CD19⁻ cells) from infected wild-type (WT) mice with L. amazonensis. Different cell populations obtained from footpad lesions were analyzed by flow cytometry and represented as the percentage of total cells. Results correspond to the mean ± SD (n = 4). ***, p < 0.001 (Student’s t-test). (C) Normalized percentage of CD45⁺F4/80⁺ populations presenting B-1P cells (CD19⁺ cells) or macrophages (CD19⁻ cells) from infected TLR9^−/− mice with L. amazonensis. Results correspond to mean ± SD (n = 4). ***, p < 0.001 (Student’s t-test). (D) Total cell fluorescence intensity (CTCF) for the presence B-1P cells (IgM⁺) or PM (IgM⁻) cell populations in comparison with non-infected (NI) or infected (I) samples from the peritoneal cavity. At least 50 cells were analyzed by using Image J software. Results correspond to the mean ± SD. ****, p < 0.0001 (Student’s t-test). (E) Flow cytometry analysis of the B-1P cells (F4/80⁺CD19⁺) cell population 48 h after Leishmania infection in the peritoneal cavity of C57BL/6 mice. (F) Peritoneal cells were stained with AO, IgM-FITC, and Hoechst. B-1P cells (IgM⁺), macrophages (IgM⁻), and lymphocytes (IgM⁻ and small nuclei) were visualized in the representative image. Scale bar, 10 μm. (G) B-1P cells and (H) Ex-vivo analysis obtained from the peritoneal cavity of infected mice. The cells were stained and analyzed in Image J software measuring the corrected total cell fluorescence intensity (CTCF) and AO in different cell populations. At least 50 cells were analyzed. Results correspond to the mean ± SD. ***, p = 0.0003 (Student’s t-test). See also Fig. S4 for additional data.