17β-Estradiol Abrogates TNF-α-Induced Human Brain Vascular Pericyte Migration by Downregulating miR-638 via ER-β

Pericytes (PCs) contribute to brain capillary/BBB integrity and PC migration is a hallmark for brain capillary leakage following pro-inflammatory insults. Estradiol promotes endothelial barrier integrity by inhibiting tumor necrosis factor-alpha (TNF-α)-induced PC migration. However, the underlying mechanisms remain unclear. Since micro-RNAs (miRs) regulate BBB integrity and increases in miR638 and TNF-α occur in pathological events associated with capillary leakage, we hypothesize that TNF-α mediates its capillary disruptive actions via miR638 and that estradiol blocks these actions. Using quantitative reverse transcription PCR, we first assessed the modulatory effects of TNF-α on miR638. The treatment of PCs with TNF-α significantly induced miR638. Moreover, transfection with miR638 mimic induced PC migration, whereas inhibitory miR638 (anti-miR) abrogated the pro-migratory actions of TNF-α, suggesting that TNF-α stimulates PC migration via miR638. At a molecular level, the pro-migratory effects of miR638 involved the phosphorylation of ERK1/2 but not Akt. Interestingly, estradiol downregulated the constitutive and TNF-α-stimulated expression of miR638 and inhibited the TNF-α-induced migration of PCs. In PCs treated with estrogen receptor (ER) ER-α, ER-β, and GPR30 agonists, a significant downregulation in miR638 expression was solely observed in response to DPN, an ER-β agonist. DPN inhibited the pro-migratory effects of TNF-α but not miR638. Additionally, the ectopic expression of miR638 prevented the inhibitory effects of DPN on TNF-α-induced PC migration, suggesting that interference in miR638 formation plays a key role in mediating the inhibitory actions of estradiol/DPN. In conclusion, these findings provide the first evidence that estradiol inhibits TNF-α-induced PC migration by specifically downregulating miR638 via ER-β and may protect the neurovascular unit during injury/stroke via this mechanism.