Neuroprotective effect of G-CSF depends on NEP.

<p>A–F, Fluorescent microscopic images of mouse primary cortical neuron cultures. A, Untreated neurons. B, Neurons treated with 5 µM oAβ_1–42. C, Neurons treated with 5 µM oAβ_1–42 and 100 ng/ml G-CSF. D, Neurons treated with 5 µM oAβ_1–42, 100 ng/ml G-CSF and 0.1 ng/ml thiorphan. E, Neurons treated with 5 µM oAβ_1–42, 100 ng/ml G-CSF and 1 ng/ml thiorphan. F, Neurons treated with 5 µM oAβ_1–42, 100 ng/ml G-CSF and 10 ng/ml thiorphan. The NEP inhibitor canceled the neuroprotective effects of G-CSF. Neurons were stained with anti–MAP-2 antibodies (green) and Aβ was stained with 4G8 antibodies (red). Scale bar: 50 µm. G, Relative neuronal survival. The number of viable neurons (MAP-2–positive neurons) was quantified relative to results observed with untreated neurons. The NEP inhibitor dose-dependently suppressed the neuroprotective effects of G-CSF. *, <i>p</i><0.001; †, <i>p</i><0.001; ‡, <i>p</i><0.05; §, <i>p</i><0.01. Values are means ± SEM (n = 3). H, WST-8 assay. The NEP inhibitor reversed the neuroprotective effects of G-CSF in a dose-dependent manner. *, <i>p</i><0.001; †, <i>p</i><0.001; ‡, <i>p</i><0.05; §, <i>p</i><0.01. Values are means ± SEM (n = 6).