American Society of Hematology

Figure 3.

$Figure 3. Effect of megakaryocytes on OM function. (A) Representative flow cytometric analysis of OMs and macrophages (MΦ) cultured for 1 week with megakaryocytes. NCCs and megakaryocytes were isolated from CD45.2 mice, and BM-derived macrophages and CD45+F4/80+ cells were collected from CD45.1 mice. (B) Cocultures of NCC-derived OBs (CD45−F4/80− cells) mixed with either the original number of OMs contained in freshly isolated NCCs (OM+OB) or a complimentary number (equivalent to the original number of OMs in NCCs) of BM-derived CD45+F4/80+ macrophages (MΦ+OB) were established. LSK cells were added with or without MKs, and LSK progeny were assessed for CFU content on day 7. CFU fold increase was calculated relative to that obtained from 250 fresh LSK cells. *P < .001 vs LSK, OB, MΦ+OB; +P < .001 vs MK, OB+MK, MΦ+OB+MK; 1-way ANOVA). (C) Fresh NCCs or fractions of NCCs (OBs or OMs) cultured with or without MKs (as indicated on the x-axis) were seeded with LSK cells then assayed 7 days later for their CFU content. CFU fold increase was calculated relative to that obtained from 250 fresh LSK cells. *P < .05 vs LSK, OM, and OB groups; +P < .005 vs MK, OM+MK, and OB+MK groups; #P < .05 vs 1w OM+OB and 1wNCC groups; one-way ANOVA). Please note that data in panels B and C are reported as fold increase rather than absolute numbers of CFUs to highlight the magnitude 1wNCC of CFU increase in these cultures. MK, megakaryocyte; OB, osteoblast.$

Effect of megakaryocytes on OM function. (A) Representative flow cytometric analysis of OMs and macrophages (MΦ) cultured for 1 week with megakaryocytes. NCCs and megakaryocytes were isolated from CD45.2 mice, and BM-derived macrophages and CD45⁺F4/80⁺ cells were collected from CD45.1 mice. (B) Cocultures of NCC-derived OBs (CD45⁻F4/80⁻ cells) mixed with either the original number of OMs contained in freshly isolated NCCs (OM+OB) or a complimentary number (equivalent to the original number of OMs in NCCs) of BM-derived CD45⁺F4/80⁺ macrophages (MΦ+OB) were established. LSK cells were added with or without MKs, and LSK progeny were assessed for CFU content on day 7. CFU fold increase was calculated relative to that obtained from 250 fresh LSK cells. *P < .001 vs LSK, OB, MΦ+OB; ⁺P < .001 vs MK, OB+MK, MΦ+OB+MK; 1-way ANOVA). (C) Fresh NCCs or fractions of NCCs (OBs or OMs) cultured with or without MKs (as indicated on the x-axis) were seeded with LSK cells then assayed 7 days later for their CFU content. CFU fold increase was calculated relative to that obtained from 250 fresh LSK cells. *P < .05 vs LSK, OM, and OB groups; ⁺P < .005 vs MK, OM+MK, and OB+MK groups; ^#P < .05 vs 1w OM+OB and 1wNCC groups; one-way ANOVA). Please note that data in panels B and C are reported as fold increase rather than absolute numbers of CFUs to highlight the magnitude 1wNCC of CFU increase in these cultures. MK, megakaryocyte; OB, osteoblast.

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