Figure 10.

CCL2 is upregulated in Atf3 mutants and CCL2 overexpression reduces neurite growth. (a,b) Ccl2 (a) but not Ccl3 (b) mRNA was more strongly induced in injured Atf3 mutant FN compared with wt. (c) In ChIP experiments, ATF3 occupied ATF3 binding sites in the Ccl2 promoter. (d–f) In lesioned wt FN, CCL2-directed antibodies labelled FMNs at 8 d.p.l. (d). The number of CCL2 positive FMNs was elevated in injured ATF3-deficient mice (e). Higher magnification (inset in (e)) suggested CCL2 localization in cytoplasmic vesicles. (f) Quantification of CCL2 positive neurons per section at different times post-injury. (g–l) Wt DRG neurons were electroporated with expression vectors driving Cherry expression alone (g,j) or CCL2-Cherry (h,k). Cherry positive neurons were identified with anti-Cherry (red) and anti-βIII tubulin (green) directed antibodies. CCl2-Cherry positive neurons (arrows in h) and cultures (k) have decreased neurite growth and neuronal network density compared with control condition (g,j). (i,l) Quantification of neurite length of individual Cherry or CCL2-Cherry positive neurons (i) or of all neurites present on the entire coverslip regardless of Cherry expression (l). Numbers in bars indicate numbers of independent experiments or animals analysed. Data are presented as mean ± s.d. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001. Scale bar (d,e) = 100 µm; (g,h) = 50 µm; (j,k) = 1 µm.