Research Article

Lipopolysaccharide Stimulates p62-Dependent Autophagy-Like Aggregate Clearance in Hepatocytes

Figure 4

Nrf2-mediates p62-LC3 aggregate formation in hepatocytes. (a) Confocal imaging of Nrf2 (red) and quantification of Nrf2 fluorescence in the nucleus (right hand graph). Data represent mean ± SEM; versus control (time 0) level. (b) Immunoblot for Nrf2 in whole cell lysates from WT hepatocytes pretreated with either control siRNA or siRNA targeting Nrf2 followed by LPS stimulation (100 ng/mL) for up to 24 h. (c) Confocal imaging and quantification of GFP-LC3II puncta (right hand graph) in WT hepatocytes pretreated with control or Nrf2-siRNA prior to LPS stimulation for up to 16 h with LPS (100 ng/mL). (d) Confocal immunofluorescence of GFP-LC3 (green) with p62 (red) showing colocalization (merge = yellow) in WT hepatocytes pretreated with either control or Nrf2 siRNA prior to LPS stimulation for 4 h. Data represent mean ± SEM; versus Nrf2 siRNA LPS control (time 0); images representative of at least three separate experiments. Confocal microscopy at 60x magnification and 1.5x zoom. Arrows indicate p62/LC3 aggregates. LC3 puncta count was normalized by number of nuclei.
267350.fig.004a
(a)
267350.fig.004b
(b)
267350.fig.004c
(c)
267350.fig.004d
(d)