mDia1 knockout (KO) mice develop neutropenia with increased expression levels of CD11b on neutrophils. (A) Absolute neutrophil count of 20-week old mDia1 KO (n = 13) and their littermate wild-type (WT) control mice (n = 13). (B) mDia1 WT (n = 8) and KO (n = 8) mouse bone marrow cells (CD45.2⁺) were retro-orbitally injected into lethally irradiated CD45.1⁺ congenic female mice. Absolute neutrophil count was analyzed at 3 months posttransplantation. (C) Bone marrow cells from CD45.1⁺ wild-type mice were transplanted into lethally irradiated CD45.2⁺ mDia1 WT (n = 5) or KO (n = 7) mice. Absolute neutrophil count was analyzed 3 months after transplantation. (D) Relative percentage and MFI of CD11b expression levels on the peripheral blood (PB) neutrophils and bone marrow (BM) granulocytes in mDia1 WT (n = 8) and KO (n = 6) mice. Data were obtained from 4 independent experiments. (E) CD11b expression levels on premature (CD11b^low) and mature (CD11b^high) bone marrow granulocytes. Data were obtained from 4 independent experiments (WT mice, n = 8; KO mice, n = 6). (F) Correlation analysis of neutrophil counts and CD11b expression levels (WT mice, n = 12; KO mice, n = 11). (G-H) CD11b expression levels on bone marrow granulocytes and peripheral blood neutrophils of the receipt mice, as described in panels B and C, were analyzed by flow cytometric analysis and presented in panels G and H, respectively. All data are shown as mean ± standard error of the mean. (I) Schematic illustration of the competitive bone marrow transplantation assay (left) and the levels of CD11b in the recipient mice (n = 3) from wild-type and knockout donor cells. (J) The absolute cell numbers for total bone marrow cells and granulocytes were quantified by flow cytometer (n = 3 in each group). (K) The percentage of apoptotic granulocytes in peripheral blood were assayed by annexin V (AnV) staining (WT mice, n = 9; KO mice, n = 11). (L) Relative mRNA levels of CD11b in the granulocytes from bone marrow and peripheral blood were assayed by quantitative polymerase chain reaction (WT mice, n = 7; KO mice, n = 5). (M). In vitro endocytosis assay of CD11b with peripheral blood neutrophils from mDia1 wild-type and knockout mice. Data were obtained from 3 independent experiments. (N) Representative flow cytometry results of the relative CD11b expression levels in a normal patient and a patient with del(5q) MDS. The cells were analyzed from the gated granulocytic population. (O) Quantitative analysis of data from panel N (del(5q) MDS patients, n = 8; normal patients, n = 14). *P < .05; **P < .01. FACS, fluorescence-activated cell sorting; NE, neutrophils; rRNA, ribosomal RNA; SSC-A, side scatter pulse area.