Figure 7.

ProNGF causes protein interaction and nuclear translocation of p75^NTR and neurotrophin interacting factor (NRIF). Human retinal endothelial cells were treated with human mutant (hm-proNGF, 50 ng/ml) in the presence or absence of a selective p75^NTR antagonist (A, 20 μmol/l) for 48 hours. (a) Cell death was assessed by measuring activity of caspase enzyme and the results were presented as mean ± SD. n = 3–5 per group. *P < 0.05 versus other groups. (b) Representative images of immunolocalization showing that hm-proNGF induced nuclear translocation of NRIF (green) within the nucleus (blue) that was blocked by p75^NTR antagonist (A) (proNGF+A). (c) Enlarged micrographs of EC treated with hm-proNGF showing colocalization of p75^NTR (red), NRIF (green) within the nuclei (blue). Similar results were observed in two independent experiments. (d) Representative blots and statistical analysis showing that hmproNGF induced protein association between NRIF (90 kDa) and p75^ICD (17 kDa) in EC lysate immunoprecipitated with anti-p75^NTR and immunoblotted by anti-NRIF. Additional band for NRIF was detected at 100 kDa. Results presented as mean ± SD. n = 4 per group. *P < 0.05 versus all groups.