Effects of human WT Ngb or E60Q Ngb on neuroprotective activities.

<p>(A) Western blot analyses of SH-SY5Y cell lysates after protein transduction. PBS, human WT Ngb or human E60Q Ngb was applied to differentiated SH-SY5Y cells with Chariot. The cells were then incubated for 3 h. Cell lysates were analyzed on 15.0% or 12.5% SDS/PAGE and by Western blot analyses using rabbit anti-Ngb polyclonal antibody or mouse anti-β-actin monoclonal antibody, respectively. (B,C) Effect of the E60Q mutation in human Ngb on SH-SY5Y cell death caused by the <i>in vitro</i> OGD model of ischemia. Human WT or E60Q Ngb was transfected into differentiated cells using Chariot. Following OGD/recovery, cell viability was measured by MTS (B) and TBE assays (C). All data are expressed as means ± SEM from three independent experiments, each performed in triplicate. Data were analyzed by one-way ANOVA followed by Tukey-Kramer post hoc tests. <i>*P</i><0.05, <i>**P</i><0.01. (D) Western blot analyses of SH-SY5Y cell lysates after transfection. Control vector, human WT or E60Q Ngb expression vector was transfected into differentiated SH-SY5Y cells with Lipofectamine. The cells were then incubated for 24 h. Cell lysates were analyzed on 15.0% or 12.5% SDS/PAGE and by Western blot analyses using rabbit anti-Ngb polyclonal antibody or mouse anti-β-actin monoclonal antibody, respectively. (E) Effect of the E60Q mutation in human Ngb on SH-SY5Y cell death caused by hydrogen peroxide. Differentiated SH-SY5Y cells transfected with control vector, human WT or E60Q HNgb expression vector with Lipofectamine were treated with hydrogen peroxide, and cell viability was measured by MTS assay. All data are expressed as means ± SEM from six independent experiments, each carried out in triplicate. Data were analyzed by one-way ANOVA followed by Tukey-Kramer post hoc tests. <i>**P</i><0.01.