Figure 2.

CX3CR1 and CCR2 expression in the lumbar spinal cord and sciatic nerve. (a) Low magnification fluorescence images of CCR2^RFP/WT/CX3CR1^GFP/WT (non-mSOD1 type) and SOD1^G93A/CCR2^RFP/WT/CX3CR1^GFP/WT (mSOD1-Tg) mouse spinal cord. At 12 weeks of age, CX3CR1⁺ (green) but not CCR2⁺ (red) cells were sparsely visible only in the spinal anterior horns (arrowhead). CX3CR1⁺ cells markedly increased as the disease progressed from 16 to 20 weeks of age, whereas CCR2⁺ peripheral immunocytes rarely infiltrated at 20 weeks of age. Infiltration of CX3CR1⁺ cells along the intramedullary lower motoneuron axonal tracts was prominent at 16 weeks of age (arrow). (b) Low magnification fluorescence images of CCR2^RFP/WT/CX3CR1^GFP/WT (non-mSOD1 type) and SOD1^G93A/CCR2^RFP/WT/CX3CR1^GFP/WT (mSOD1-Tg) mouse sciatic nerves. In the sciatic nerves of a mSOD1-Tg mouse, CX3CR1⁺ cell infiltrations were visible as early as 4 weeks of age and CX3CR1⁺CCR2⁺ cells appeared at 8 weeks of age (pre-symptomatic stage). These infiltrated cells showed a foamy appearance (inset in the 8 weeks merged image). Such inflammatory infiltrates were not seen in non-mSOD1 type mice. (c) The quantitative analysis of CCR2⁺ CX3CR1⁻ (red), CCR2⁻CX3CR1⁺ (green), and CX3CR1⁺CCR2⁺ (yellow) areas (%) in sciatic nerves. In SOD1^G93A mice, numbers of all types of macrophages (CCR2⁻CX3CR1⁺ green, and CCR2⁺CX3CR1⁺ yellow macrophages) increased as the disease progressed, whereas CCR2⁺CX3CR1⁻ red immune cells did not. Scale bars: (a) 500 μm; (b) 50 μm and 10 μm (inset). n.s. = not significant. *p < 0.05; **p < 0.001; ***p < 0.0001.