P-Glycoprotein Exacerbates Brain Injury Following Experimental Cerebral Ischemia by Promoting Proinflammatory Microglia Activation

Figure 5

P-glycoprotein silence or overexpress in endothelial cells influenced pro- and anti- inflammatory cytokines expression in cocultured microglia following oxygen glucose deprivation/reoxygenation. Endothelial cells (bEnd.3) were transfected with P-glycoprotein (P-gp) or negative control (NC) siRNA, P-gp or NC pcDNA3.1 plasmid, or untransfected, and then subjected to either oxygen glucose deprivation/reoxygenation (OGD/R) treatment or normal culture conditions. Following 24 hr coculture with microglia (BV2), bEnd.3 cells were harvested for western-blotting analyses, while BV2 cells were harvested for RT-PCR assay and medium was collected for ELISA assay. (a) Transwell coculture model for evaluating OGD/R-induced changes in bEnd.3 cells and BV2 cells. (b, c) Representative western-blotting images and quantification of P-gp levels (n = 3). (d) mRNA expression levels of IL-12, IL-6, IL-4, and YM-1 measured via RT-PCR assay as fold changes relative to control treatment (n = 4). (e) Contents of IL-12, IL-6, IL-4, and YM-1 determined by ELISA assay (n = 3). One-way ANOVA followed by the post hoc least significant difference test or Games Howell test for (b), (c), and (e). Mann–Whitney test for (d). All data are mean ± SD; , between two groups.

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