CSF-1R pTyr-721 signaling suppresses M1 responses and enhances M2 responses. (A) qRT-PCR results showing the enhancement by pTyr-721 signaling of the IFN-γ and LPS stimulation of iNOS, TNF-α, and IL-6 gene transcripts in cells stimulated with CSF-1. Cells were either starved for CSF-1 overnight, or constitutively grown in CSF-1 (120 ng/mL), then treated with either IFN-γ and LPS (+) or vehicle (−) for 18 hours. HPRT gene expression levels were used as endogenous control. mRNA levels are expressed as the log₂ fold-change relative to CSF-1–starved cells. (B) Nitrate/nitrite levels in cell lysates from equivalent cultures. (C) Relative CSF-1–induced surface expression of the M1 polarization marker MHC II on CD11b⁺ cells, determined by flow cytometry in CSF-1–treated (18 hours, +CSF-1) vs CSF-1–starved (−CSF-1) cells and expressed as a ratio of the +CSF-1 to −CSF-1 responses. (D) qRT-PCR results showing the enhancement by pTyr-721 signaling of the IL-4–stimulated expression of arginase-1, IL-4Rα, and mannose receptor-1 (MRC1) transcripts. Experiments were carried out as described in panel A. (E) Total arginase 1 activity of supernatants of equivalent cultures. (F) Stacked bar chart showing relative contribution of combined sets of macrophage M2 (CD206 and IL-4Rα) and M1 (MHC II) cell surface–specific polarization markers in CSF-1–treated cells. The relative CSF-1 response ratio for each set of cell surface markers was calculated as in panel C and expressed as a percentage for each cell line. Data are representative of 3 independent experiments (error bars, ±standard deviation (SD); *P ≤ .05).