IL-1R signaling affects AGM HSCs. (A) Flow cytometric analysis of freshly isolated E11 Il1r1^+/+ and Il1r1^−/− AGM cells showing the absolute number of c-kit⁺, CD45⁺, CD34⁺c-kit⁺, or CD34⁻Mac1⁺ cells per AGM (n = 3). (B) Flow cytometric analysis of E11 Il1r1^+/+ and Il1r1^−/− AGM explants showing absolute numbers of c-kit⁺, CD45⁺, or Mac1⁺ cells per AGM explant (n = 4-5). (C) Number of CFU total, CFU-M, BFU-E, CFU-G, CFU-GM, and CFU-GEMM per freshly isolated E11 Il1r1^+/+ and Il1r1^−/− AGM. Colonies from triplicate cultures were scored after 7 days of methylcellulose culture (n = 3, cells from 14 Il1r1^+/+ and 15 Il1r1^−/− AGMs). (D) Number of CFU total, CFU-M, BFU-E, CFU-G, CFU-GM, and CFU-GEMM per E11 Il1r1^+/+ and Il1r1^−/− AGM after 3 days of explant culture. Colonies from triplicate cultures were scored after 7 days of methylcellulose culture (n = 3, cells from 8 Il1r1^+/+ and 8 Il1r1^−/− AGMs). (E) Percentage of adult recipient mice repopulated with HSCs from Il1r1^+/+ and Il1r1^−/− AGM regions either directly transplanted (direct) or transplanted after 3 days of explant culture (explant). E11 AGM cells (1 and 0.3 or 1 and 0.2 embryo AGM tissue equivalents [ee]) were transplanted into irradiated adult recipients. At 4 months after transplantation, peripheral blood DNA of recipients was analyzed for donor hematopoietic chimerism by semiquantitative PCR. Only mice with more than 10% donor chimerism were considered repopulated. Each column represents the number of mice repopulated per number of recipients transplanted (the numbers for the Il1r1^+/+ and Il1r1^−/− columns are, respectively, 5 of 7 and 3 of 8 for 1 ee direct transplantation, 1 of 6 and 0 of 8 for 0.3 ee direct transplantation, 2 of 2 and 5 of 5 for 1 ee explant transplantation, and 8 of 10 and 2 of 10 for 0.2 ee explant transplantation). Combined results of 4 separate transplantation experiments. The error bars represent SEM.