Spatiotemporal quantification of recruit and resident macrophages after crush nerve injury utilizing immunohistochemistry.

The purpose of this study was to investigate quantitatively the temporal and spatial regulation and the morphological changes of the recruit and resident macrophages in the sciatic nerve during Wallerian degeneration and the following regeneration using immunohistochemistry. Sciatic nerves in Sprague-Dawley (SD) rats were examined after nerve crush. The rats were anesthetized with 100 mg of ketamine and 20 mg of xylazine in a dose of 1 ml/kg by intraperitoneal injection. Anti-ED-1 antibody was used to detect phagocytic macrophage and anti-OX-6 antibody was used to detect MHC class II cells. Few ED-1-immunopositive cells were seen within the normal sciatic nerve. After crush injury the number and the size of ED-1-immunopositive cells started to increase in all the segments distal to the crush site 3 days after injury and the number and size reached its peak on day 14 when the population of macrophage was 150 times higher in all the segments compared to controls. However, the number of ED-1-immunopositive cells and the size of the cells remains significantly high even after day 56 when functional recovery and axonal regeneration were complete. OX-6-immunopositive cells were observed within the control sciatic nerves. The number decreases significantly 3 days after injury in all the segments distal to the crush site but showed no significant difference thereafter. There were also no significant differences in the cell areas. ED-1-immunopositive phagocytic macrophages show significant differences temporally in both the cell number and the size even after axonal regeneration.