Fig. 5.

ADAM10 controls splenic cDC2 homeostasis via Notch2-dependent but FLT3- and LT-βR–independent pathways. (A) FACS plot shows expression of cell-surface FLT3 on splenic SIRP-α⁺ cDC2 from control (black) and ADAM10^ΔCD11c (blue) mice. Filled gray histogram: isotype control. Bar graph represents quantification of FLT3 expression (mean fluorescent intensities, MFI) on splenic XCR1⁺ cDC1 and SIRP-α⁺ cDC2 from control and ADAM10^ΔCD11c mice. (B) Canonical pathway analysis (Ingenuity) associated with significantly changed genes between bulk-sorted ADAM10-deficient/control CD4⁺ cDC2A. Control and ADAM10^ΔCD11c mice were treated with recombinant FLT3 ligand (rFLT3L) or saline as control. (C) Absolute number of splenic cDC in saline and rFLT3L treated control and ADAM10^ΔCD11c mice. (D) Absolute number of splenic cDC1 (Left) and Langerin⁺ cDC1 (Right) in saline and rFLT3L-treated control and ADAM10^ΔCD11c mice. (E) Absolute number of splenic cDC2 (Left), ESAM^hi cDC2A (Middle), and CX₃CR1⁺ cDC2B (Right) in saline and rFLT3L-treated control and ADAM10^ΔCD11c mice. (F) Ltbr, Nik, and Relb mRNA expression in bulk-sorted control or ADAM10-deficient splenic CD8-α⁺ cDC1, CD4⁺ cDC2, and CD4^– cDC2 subsets. (G) Control and ADAM10^ΔCD11c mice were treated with agonistic anti–LT-βR antibody or saline as control. Bar graphs shows the frequency of ESAM^hi cells among SIRP-α⁺ cDC2 (Left) or the frequency of apoptotic cells within this population (Right) in saline-treated and antibody-treated control versus ADAM10^ΔCD11c mice. (H) Flow cytometric analysis of Notch2 expression on splenic CD11c⁺MHCII⁺SIRP-α⁺ cDC2 from control (black line) and ADAM10^ΔCD11c (blue line) mice. Filled gray histogram: isotype control. (I) Venn diagram summarizing the pair-wise overlap of significant differentially expressed genes (DE genes) between ADAM10-deficient/control CD11c⁺MHCII⁺CD4⁺ cDC2A (purple), ADAM10-deficient/control CD4^– cDC2B (green), and Notch2-KO/control CD11b⁺ cDC2 (yellow). Data from Notch2-KO/control CD11b⁺ cDC2 are retrieved from GEO, accession no. GSE45698 (44). (J) Venn diagram summarizing the pair-wise overlap of significantly changed genes between ADAM10-deficient/control CD8-α⁺ cDC2 (orange) and Notch2-KO/control CD205⁺ cDC1 (yellow). Data from Notch2-KO/control CD205⁺ cDC1 are retrieved from GEO, accession no. GSE45698 (44). *P < 0.05, **P < 0.01, and ***P < 0.001 (Student’s t test); data are pooled from two or more experiments (n = 3 to 6 mice/experiment); FACS plots show one representative mouse/group with mean frequencies or mean fluorescent intensity ± SEM.