Fig. 4

M1 to M2a conversion of microglia with cyclic AMP and IL-4 can be used in vivo after SCI and significantly attenuates the production of tissue-damaging free radicals. a, b Immunoblotting of tissue homogenates from the injury epicenter at 24 h after contusive SCI in rat for the prototypical M2 marker Arginase-1 shows an increase in expression after SCI that is further potentiated by the co-delivery of db-cyclic AMP (50 mg/kg) and IL-4 (30 μg/Kg) administered <30 min post-injury. Statistical significance indicated at **p < 0.01 or *p < 0.05 versus uninjured controls or ^# p < 0.05 versus SCI only controls. Compared to SCI controls, the administration of db-cAMP and IL-4 produced a dramatic reduction in the numbers of Iba1⁺ (c, g) and ED1⁺ (green, d, h) microglia and macrophages within the lesion epicenter at 24 h post-SCI. While few ED1⁺ microglia and macrophages were Arginase-1⁺ (red, white arrows) in SCI-only controls (d, f), almost all ED1⁺ microglia and macrophages found in db-cyclic AMP and IL-4-treated animals co-expressed Arginase-1 (h, j; white arrows). The nuclei of the cells have been counterstained with Hoechst (blue). Scale bar = 20 μm. k Compared to SCI only controls, a dramatic reduction in the total level of free radicals within the injury epicenter was observed with the systemic administration of db-cyclic AMP and IL-4. Statistical significance indicated at ^# p < 0.05 versus SCI only controls. Four animals were used in each group. Following contusive SCI in mouse, compared to injury-only controls (l–n), daily treatment initiating at <30 min post-SCI induction with IL-4 (30 μg/kg) and db-cyclic AMP (50 mg/kg) for seven consecutive days post-SCI (o–q) increased the number of Isolectin-IB4 positive immune cells (red) that were immunoreactive for Arginase-1 (green) within the lesion site. The nuclei of the cells have been counterstained with Hoechst (blue). Scale bar = 40 μm. r Quantitative assessment of the numbers of Isolectin-IB4+ cells immunoreactive for Arginase-1 showed a significant increase with db-cyclic AMP and IL-4 after SCI. Statistical significance indicated at ^### p < 0.001 versus SCI-only controls