Modifying macrophages at the periphery has the capacity to change microglial reactivity and increase ALS survival

Microglia and peripheral macrophages, combined, have been implicated in the motor neuron disease Amyotrophic Lateral Sclerosis (ALS), but without discriminating their respective roles. We now show that macrophages along peripheral motor neuron axons of ALS mice and patients react to neurodegeneration. In ALS mice, peripheral myeloid cell infiltration into the spinal cord was limited and disease duration dependent. Targeted gene modulation of the reactive oxygen species pathway in peripheral myeloid cells of ALS mice, using cell replacement, reduced both peripheral macrophage and microglial activation, delayed symptoms and increased ALS mouse survival. Transcriptomics revealed that sciatic nerve macrophages and microglia reacted very different to neurodegeneration, with abrupt temporal changes in macrophages and progressive, unidirectional activation in microglia. Modifying peripheral macrophages suppressed proinflammatory microglial responses, with a strong shift towards neuronal support. Thus, modifying macrophages at the periphery has the capacity to influence disease progression and is of therapeutic value for ALS. Overall design: GENERAL: The project has 2 parts: PART 1 (NON_GRAFTED) are non-grafted mutant hSOD1-G93A ALS mice (for a total of 51 RNAseq samples; all females), from which both spinal cord microglial cells (MG) (33 samples) and sciatic nerve peripheral macrophages (MP) (18 samples) were isolated for RNAseq analysis; PART 2 (GRAFTED) are mutant hSOD1-G93A ALS mice (for a total of 23 RNAseq samples; all females) that were grafted with bone-marrow (BM) at onset from either (1) mutant hSOD1-G93A ALS mice (as baseline disease control) (12 samples), or (2) wildtype hSOD1 mice (as experimental disease modifying condition) (11 samples). Like for Part 1, from these mice both spinal cord microglial cells (MG) (12 samples) and sciatic nerve peripheral macrophages (MP) (11 samples) were isolated for RNAseq analysis. DETAILED: both spinal cord microglial cells (MG) and sciatic nerve peripheral macrophages (MP), were isolated by anti-CD11b magnetic microbead-coupled antibody (MACS). For PART 1 (NON_GRAFTED): MG and MP were isolated at 4 disease stages from (non-grafted) mutant SOD1-G93A ALS mice. Disease stages were: pre-symptomatic, onset (weight peak), early symptomatic (10% weight loss) and end stage (see "Samples" for precise ages). For each diseae stage, 3-5 replicates (n=3-5) were done (1 mouse per sample/replica). For MG, one spinal cord/sample was used, for MP, two sciatic nerves/mouse/sample were used (see Publication for more details). As controls, both non-transgenic C57BL6/J mice (n=4) and wildtype hSOD1 transgenic mice (n=4) were used (at 2 ages corresponding to onset and end stage). For PART 2 (GRAFTED): MG and MP were isolated at 2 disease stages from (grafted) mutant SOD1-G93A ALS mice. Disease stages were: early symptomatic and end stage (see "Samples" for precise ages). For each diseae stage, 3 replicates (n=3) were done (1 mouse per sample/replica). For MG, one spinal cord/sample was used, for MP, two sciatic nerves/mouse/sample were used (see Publication for more details).