FIGURE 7.

Heat shock-induced phosphorylated NF-H is reduced by Pin1 knockdown. A, 5 DIC cortical neurons were transfected with either control siRNA (a-d and e-h) or Pin1-siRNA (i-l) and then exposed to 44 °C ina5%CO₂ incubator for 30 min (heat shock, e-h and i-l). Neurons were immunostained, p-NF-H was detected using RT-97 (red), and Pin1 was detected by using Pin1 antibody. Scale bar, 20 μm. B, representative neurons are subjected to heat shock as indicated. Intensity of RT-97 immunoreactivity (p-NF-H) in neuronal cell bodies was analyzed with the ImageJ histogram. The mean signal intensity (total pixel density/number pixels) is shown. Values in graphs are means ± fluorescence intensity (in arbitrary densitometric units) in perikarya from four different experiments. Note the accumulation of p-NF-H in perikarya in heat shock-treated neurons and restoration of normal levels of p-NF-H in Pin1 siRNA-treated cortical neurons.