Figure 4. ST6Gal-1 suppresses neutrophilic, but not monocytic, differentiation of GMPs.

FcγRII/III⁺ progenitors (0.75 × 10⁵) enriched from pooled WT marrow were expanded in culture in the presence of murine G-CSF, SCF, and IL-3. ST6Gal-1-treated cells were subjected to 3 h of ex vivo sialylation before expansion. At days 4 and 6, samples were counted and analyzed by flow cytometry for lineage markers. Cells undergoing apoptosis were enumerated on d 7, with annexin V and PI staining. In a separate experiment, GMPs enriched from pooled WT marrow were expanded in culture in the presence of murine G-CSF, M-CSF, SCF, and IL-3. After 3 d, samples were analyzed by flow cytometry for lineage markers. (A) Total cells per well recovered after 4 and 6 d in culture. Dashed line: day 0 cell counts (n = 4). *P < 0.05. (B) Quantification of total CD11b⁺ Gr-1⁺ neutrophils per well recovered after 6 d in culture (n = 4). **P < 0.01. (C) Apoptotic index of treated vs. untreated cells. Early apoptotic cells were defined as annexin V^hiPI^lo; late apoptotic cells were defined as annexin V^hiPI^hi. NS, not significant. (D) Representative flow plots showing GMP neutrophil differentiation. Gate indicates CD11b⁺ Ly6C low Ly6G⁺ neutrophils. Percentages are of total cells recovered. (E) Quantification of total CD11b⁺ Ly6C low Ly6G⁺ neutrophils and CD11b⁺ Ly6C⁺ monocytes per well (n = 5). **P < 0.01.