Figure 4: Mutator erythrocytes are prematurely destroyed by the spleen macrophages.

(a) FACS analysis and quantification of phosphatidylserine (PS) exposure in peripheral blood erythrocytes at the age of 8 months (two-tailed P=0.001, Student’s t-test). WT: n=7, Mutator: n=15 (30,000 cells analyzed per sample). (b) FACS analysis and quantification of total amount of erythroid cells (Ter119+) in the spleen (two-tailed P<0.0001, Student’s t-test); (c) phosphatidylserine exposure in splenic erythroid cells in Mutator versus WT at the age of 8 months (two-tailed P<0.0001, Student’s t-test). WT: n=9, Mutator: n=9 (30,000 cells analyzed per sample). (d) The amount of activated macrophages (CD45+, mac3+) in the spleen at the age of 8 months (two-tailed P=0.0001, Student’s t-test). WT: n=9, Mutator: n=9 (30,000 cells analyzed). (e) Histological analysis of iron (blue) in the spleen red pulp (RP, WP=white pulp) at the age of 8 months (WT: n=5, Mutator: n=5, six to nine histological sections were analyzed per sample). Solid line in inset: individual macrophage. Scale bars, 50 μm, inset scale bars, 25 μm. (f) Histological analysis of iron (blue) in the bone marrow at the age of 10 months (WT: n=5, Mutator: n=5, six to nine histological sections were analyzed per sample). Scale bars, 25 μm. (g) In vivo erythrocyte lifespan in 6-month-old Mutators and WT: 30% less Mutator erythrocytes survived 21 days in circulation when compared with WT cells (two-tailed P=0.03, linear regression analysis). WT: n=3, Mutator: n=3 (FACS analysis; 30,000 cells analyzed per sample per timepoint). For b–d,g, data are shown as mean±s.d.