Figure 2.
FL macrophage development requires Xpr1 expression in hematopoietic cells. (A) Representative flow cytometry plots and total cell numbers (mean ± SD) of FL macrophages (Mac) extracted from E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl embryos (pre-gated on CD45+Lin− [CD19−CD3−B220−Ter119−CD49b−CD90.2−] cells). Data show n = 4–8 per group and are pooled from two independent experiments. Statistical analysis was performed using the Student’s t test. (B) Graphs showing embryo and liver weights of E18.5 Xpr1fl/+, Xpr1fl/fl, Vav1iCreXpr1fl/+, and Vav1iCreXpr1fl/fl embryos. Data show n = 5–9 per group and were pooled from three independent experiments. Statistical analysis was performed using one-way ANOVA with Tukey’s multiple comparison test. (C) Representative flow cytometry plots and total cell numbers (mean ± SD) of liver Ly6Chi monocytes (Mo) extracted from E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl embryos (pre-gated on CD45+Lin−F4/80−CD117lo-intCD48− cells). Data show n = 4–8 per group and are pooled from two independent experiments. Statistical analysis was performed using the Student’s t test. (D) UMAP plots of myeloid cells extracted from livers of E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl embryos (pre-gated on CD45+Lin−CD64+F4/80+MHCII−Ly6G− cells) and analyzed by flow cytometry. Data were generated from n = 2–4 samples per group and are representative of at least four independent experiments. (E) Graphs showing frequencies of cell clusters defined among CD45+ cells (D). Data show n = 2 per group and are representative of at least four independent experiments. (F) Heatmap of cell surface marker expression on cell clusters defined in D. Data were transformed and percentile normalized. (G) Immunohistochemistry of E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl livers. DAPI (blue), CD64 (red), and F4/80 (green). Insets: Magnifications of the outlined regions showing expression of all markers, CD64/DAPI, and F4/80/DAPI (from left to right). Images are representative of at least five embryos per group. Scale bar: 40 µm. (H) Immunohistochemistry of E15.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl livers, stained for Hoechst (blue), IBA1 (cyan), and TIM4 (white). Insets: Single stainings. Images are representative of three embryos per group. Scale bar: 100 µm in overview, 50 µm in inset. **P < 0.01, ***P < 0.001, and ****P < 0.0001. ns, not significant.

FL macrophage development requires Xpr1 expression in hematopoietic cells. (A) Representative flow cytometry plots and total cell numbers (mean ± SD) of FL macrophages (Mac) extracted from E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl embryos (pre-gated on CD45+Lin− [CD19−CD3−B220−Ter119−CD49b−CD90.2−] cells). Data show n = 4–8 per group and are pooled from two independent experiments. Statistical analysis was performed using the Student’s t test. (B) Graphs showing embryo and liver weights of E18.5 Xpr1fl/+, Xpr1fl/fl, Vav1iCreXpr1fl/+, and Vav1iCreXpr1fl/fl embryos. Data show n = 5–9 per group and were pooled from three independent experiments. Statistical analysis was performed using one-way ANOVA with Tukey’s multiple comparison test. (C) Representative flow cytometry plots and total cell numbers (mean ± SD) of liver Ly6Chi monocytes (Mo) extracted from E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl embryos (pre-gated on CD45+Lin−F4/80−CD117lo-intCD48− cells). Data show n = 4–8 per group and are pooled from two independent experiments. Statistical analysis was performed using the Student’s t test. (D) UMAP plots of myeloid cells extracted from livers of E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl embryos (pre-gated on CD45+Lin−CD64+F4/80+MHCII−Ly6G− cells) and analyzed by flow cytometry. Data were generated from n = 2–4 samples per group and are representative of at least four independent experiments. (E) Graphs showing frequencies of cell clusters defined among CD45+ cells (D). Data show n = 2 per group and are representative of at least four independent experiments. (F) Heatmap of cell surface marker expression on cell clusters defined in D. Data were transformed and percentile normalized. (G) Immunohistochemistry of E16.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl livers. DAPI (blue), CD64 (red), and F4/80 (green). Insets: Magnifications of the outlined regions showing expression of all markers, CD64/DAPI, and F4/80/DAPI (from left to right). Images are representative of at least five embryos per group. Scale bar: 40 µm. (H) Immunohistochemistry of E15.5 Xpr1fl/fl and Vav1iCreXpr1fl/fl livers, stained for Hoechst (blue), IBA1 (cyan), and TIM4 (white). Insets: Single stainings. Images are representative of three embryos per group. Scale bar: 100 µm in overview, 50 µm in inset. **P < 0.01, ***P < 0.001, and ****P < 0.0001. ns, not significant.

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