Figure 5
IRF6 Inhibits PPARγ-Induced Repression of miR-106a Expression in Ischemic Cerebrovascular Endothelial Cells. (A,B) Control GFP and small-hairpin PPARγ RNA (PPARγ shRNA)-transfected murine cerebrovascular endothelial cells were pre-treated with pioglitazone prior to oxygen-glucose deprivation (OGD) exposure. Real-time PCR on the total RNA content from these cells revealed that OGD-induced miR-106a upregulation was reversed by pioglitazone, and this effect was dependent upon PPARγ. Cropped blots are displayed here. *P < 0.05 versus sham group, † P < 0.05 versus OGD group, ‡ P < 0.05 versus OGD + Piog group, # P < 0.05 versus OGD + Piog + Ad.GFP group. (C,D) Real-time PCR on the total RNA content from murine cerebrovascular endothelial cells transfected with adenoviral GFP, IRF6, PPARγ, or IRF6 + PPARγ revealed that PPARγ gain-of-function significantly reduced miR-106a expression. IRF6 gain-of-function alone had no significant impact upon miR-106a expression, but it significantly inhibited PPARγ suppression of miR-106a expression. Cropped blots are displayed here. *P < 0.05 versus Ad.GFP group, † P < 0.05 versus Ad.IRF6 group, ‡ P < 0.05 versus Ad.PPARγ group. (E,F) Wild-type mice or adenoviral-mediated IRF6 knockdown mice were treated with pioglitazone following middle cerebral artery (MCA) occlusion. After 24 hours of MCA reperfusion, cerebral microvessels were isolated. Real-time PCR on the total RNA content from these cerebral microvessels revealed that cerebral ischemia-induced miR-106a upregulation was reversed by pioglitazone. IRF6 knockdown alone had no significant impact upon miR-106a transcription, but IRF6 knockdown significantly promoted pioglitazone-driven PPARγ suppression of miR-106a transcription. Cropped blots are displayed here. *P < 0.05 versus sham group, † P < 0.05 versus Ischemia group, ‡ P < 0.05 versus Ischemia + Piog group, # P < 0.05 versus Ischemia + IRF6 KD group.
