Effects of MR16-1 treatment on macrophage polarization after spinal cord injury (SCI), as delineated by double immunolocalization. (A) At 3 days post-injury, large amounts of inducible nitric oxide synthase (iNOS) colocalized with CD11b (merged)-positive cells were found inthe rat IgG control group), whereas only a few were found in (B) the MR16-1-treated group. (C) The differences in the presence of merged double-immunopositive cells between the two groups were significant from 1 to 7 days after injury. (D) Scarce numbers of arginase 1 colocalized with CD11b-positive (merged) cells were found in the rat IgG control group, in contrast to (E) the MR16-1-treated group; (F) analysis showed predominance of merged double-immunopositive cells in the MR16-1-treated group from 1 to 7 days after injury. (G) At 7 days post-SCI, a larger number of cells immunopositive for CD16/32 and CD11b (merged) was found in the injury epicenter in the rat IgG control group than in (H) the MR16-1-treated group, and (I) these differences were significant from 3 to 14 days after injury. (K) Cells immunopositive for CD206 and CD11b (merged) were prevalent in the MR16-1-treated group, but barely present in (J) the rat IgG control group. (L) The population of CD206/CD11b-immunopositive cells became significantly greater in the MR16-1-treated group compared with the control group from 3 to 14 days after injury. CD11b conjugated to (A, B, D, E, G, H, J, K); Alexa fluor 568 (red) (A, B) iNOS, (D, E) arginase 1, (G, H) CD16/32 and (J, K) CD206 conjugated to Alexa fluor 488 (green). Scale bar = 50 μm. (C, F, I, L) Data are expressed as mean ± SD; n = 3 for each group. *P <0.05, **P <0.01 by paired t-test.