ATP Is Required and Advances Cytokine-Induced Gap Junction Formation in Microglia In Vitro
Figure 7
Pro-inflammatory treatments upregulate Cx43 and Panx1 protein levels in microglia. (a) Confocal images show immunoreactivity for Cx43 (red) and Panx1 (green) in primary rat microglia under control conditions of after treatment with TNF-α plus ATP for 3.5 h or with TNF-α/IFN-γ for 9 h, in absence or presence of IL-6 (10 or 50 ng/mL, respectively). Arrows show microglia with segregation of Cx43 and Panx1. Scale bar: 10 μm. (b) Quantification of nonpolarized (black bars) versus polarized (dashed white bars) rat microglia under control conditions or after treatments shown in (a). Data are expressed as a percentage of the total number of cells per field, (up to 100 cells per field). versus control condition. (c) Representative Western blots from 3 independent experiments showing total protein levels of Cx43 and Panx1. Cell lysates were obtained from EOC20 cells under control conditions (lane C) or after the following treatments: TNF-α plus ATP (lane 1), IL-6/TNF-α plus ATP (lane 2), TNF-α/IFN-γ (lane 3), IL-6/TNF-α/IFN-γ (lane 4), TNF-α/IL-1β (lane 5), and IL-6/TNF-α/IL-1β (lane 6). Quantitation of Cx43 and Panx1 is shown; β-actin was used as a loading control for densitometric analysis.
