Time course of inflammatory and anti-inflammatory macrophage markers in cells examined by flow cytometry. Cells that stained for markers associated with macrophagic and microglial phenotypes were detected by flow cytometry of cells isolated from the lesion and/or perilesion area. (A) and (B) Comparison of cells stained with (A) general macrophage marker CD68 and anti-inflammatory macrophage (M2)-associated marker CD163 and (B) CD68 versus inflammatory macrophage (M1)-associated marker CD40 demonstrates a change in population occurring between 3 days and 1 week postinjury. (C) Statistically significant changes in the percentage of cells relative to controls that stained positive for the M2-associated marker CD163 were detected at 3 and 5 days postinjury (CD163: control = 0.9 ± 0.2%, 3 days = 12 ± 3%, 5 days = 20 ± 2%; F = 55.70). No significant differences were observed regarding the percentages of cells that stained for CD40 and CD68 over the time course. *P < 0.05. (D) Normalization of CD40 or CD163 to CD68 for flow cytometry data at each time point relative to controls allows better visualization of the degree of changes in M1 versus M2 ratios of macrophages. Data for CD163 are statistically significant at 1, 3 and 5 days postinjury by one-way analysis of variance (CD163: control = 120 ± 30%, 1 day = 350 ± 20%, 3 days = 1,000 ± 100%, 5 days = 1,500 ± 100%; F = 92.61). Error bars indicate standard errors of the mean. *P < 0.05.